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Keke Dong, Xiangfei Meng, Hongyi Diao, Bing Qi, Zhuangzhi Chen, Wei Ma, Yihang Zhang, Minmin Yang, Jing Zhao, Liu Liu J Chem Inf Model DOI: 10.1021/acs.jcim.5c00816 Abstract Existing tools for analyzing next-generation sequencing (NGS) data from DNA-encoded library (DEL) screens are often limited to custom internal methods, focusing narrowly on protein–ligand interactions and lacking standardization in compound selection. These tools typically ignore sequencing depth, error rates, and quality control, leading to time-consuming and subjective analysis. To address these issues, we developed PB-DELASA, a fully automated, standardized, and accurate DEL data analysis workflow. It incorporates AI, computational analysis, and medicinal chemistry expertise to generate 2D/3D visualizations and ranked compound lists. Validated through a screen against CDK9, PB-DELASA identified potent and selective hits with minimal synthesis effort. The source code is publicly available. Summary PB-DELASA is a novel, automated platform for analyzing DEL screening data. It integrates QC, enrichment analysis, and compound prioritization into a single workflow. The tool uses AI and empirical rules to reduce false positives and recommend high-quality compounds for off-DNA synthesis. It was validated using a CDK9 screening campaign, resulting in the discovery of novel, potent, and selective inhibitors. The platform improves efficiency, reduces bias, and supports medicinal chemists in early-stage drug discovery. Highlights 1. Automated, end-to-end DEL data analysis workflow 2. Combines AI, computational tools, and medicinal chemistry expertise 3. 2D/3D visualization and compound prioritization 4. Validated with CDK9 target; identified potent and selective hits 5. Reduces synthetic workload and improves hit confidence 6. Open-source and user-friendly for non-bioinformaticians Conclusion PB-DELASA addresses major limitations in current DEL data analysis by providing a standardized, automated, and accurate workflow. It enhances the reliability of hit identification, reduces manual intervention, and supports efficient compound prioritization. The successful validation against CDK9 demonstrates its potential to accelerate early-stage drug discovery. By making the tool open-source, the authors aim to promote broader adoption and further development in the DEL community.
Suraj Kanoo,∥ Eduardo de Pedro Beato,∥ Tim Schulte, Lara Vogelsang, Luca Torkowski, Felix Waldbach, Philipp Hartmann, Riya Kayal, Karl-Josef Dietz, and Tobias Ritter* J. Am. Chem. Soc. 2025, https://doi.org/10.1021/jacs.5c11842 Abstract C–N cross coupling reactions are widely employed for the construction of carbon–nitrogen bonds. However, control of chemoselectivity in the presence of the amino functionality in oligonucleotides remains a challenge. Here, we report the development of a new ruthenium reagent that enables the chemoselective N-arylation of amine–DNA conjugates with distinct chemoselectivity when compared to conventional palladium-based C–N bond-forming catalysts. The ruthenium reagent activates commercially available haloarenes in situ via η6 π-arene coordination for subsequent SNAr with the amine. The method is compatible with various commercially available haloarenes and aliphatic amines, and the reaction proceeds under mild conditions.
Marc Flajolet; Yashoda SUNKARI WO2024097744A2 Link: https://patents.google.com/patent/WO2024097744A2/en? Abstract The present invention provides methods of generating diverse chemical structures on DNA through Wittig olefination of novel on-DNA phosphorane ylides and Homer- Wadsworth-Emmons reaction of on-DNA β-keto phosphonates. The methods of this invention provide access to DNA-encode libraries (DELs) of diverse peptides, peptidomimetics, chalcone-based molecules, and the like.
Developing new medicines demands smart technology and powerful tools. DNA-encoded library (DEL) technology, with its unique ability to rapidly screen billions of compounds to identify small molecule hits, has transformed drug discovery. HitGen Inc. is raising the bar with the introduction of OpenDEL™ 5.0 – featuring a total of 4 billion diverse compounds designed to tackle challenging biological targets. OpenDEL™ 5.0 comes in a convenient kit format that is accessible to scientists in industry and academia alike. Key Improvements: ✔ Expanded Chemical Diversity: 4 billion Compounds • Increased Library Size: Now with 25% more compounds, OpenDEL™ 5.0 offers 3.8 billion small molecules compounds and 200 million peptide compounds to further expand chemical space for targeting protein-protein-interactions (PPI). • Encompassing Library Design: 59 distinct libraries provide a broad range of chemistries to address different types of targets and discovery challenges. ✔ Optional Macrocycle Library: Unlocking Challenging Targets • A dedicated macrocycle library: Available as an add-on, featuring 4-10 amino acids in the ring. Comes with a linear peptide control library. • Dual-Kit Flexibility: Choose between OpenDEL™-Small Molecules or OpenDEL™-Macrocycle – or combine both for maximum coverage. ✔ Streamlined Workflow & Faster Access • Redesigned protocols: Intuitive visual guides simplify key steps, reducing onboarding time. • Expedited delivery: Next-day shipping available to minimize wait time OpenDEL™ Physicochemical Properties Distribution OpenDEL™ compounds are designed with balanced physicochemical properties, ensuring high-quality hits from screening to development. OpenDEL™ in the Literature 1. Grogan A, Ahn S, Israel D, et al. Abstract P2154: A novel allosteric modulator of the β1AR identified by DNA-encoded small molecule library screening demonstrates unique pharmacology and function. Circ Res. 2023;133(Suppl 1):AP2154. doi:10.1161/res.133.suppl_1.P2154 2. Zhang, C.; Pitman, M.; Dixit, A.; Leelananda, S.; Palacci, H.; Lawler, M.; Belyanskaya, S.; Grady, L.; Franklin, J.; Tilmans, N.; Mobley, D. L. Building Block-Based Binding Predictions for DNA-Encoded Libraries. J. Chem. Inf. Model. 2023, 63 (16), 5120– 5132, DOI: 10.1021/acs.jcim.3c00588 3. Brooun A, Fagan P, Bergqvist S, et al. Identification and characterization of inhibitors of SHOC2-MRAS-PP1C complex assembly [published online ahead of print April 2025]. Cancer Res. 2025;85(8_Supplement_1):3152. doi:10.1158/1538-7445.AM2025-3152 4. Wellnitz, J. et al. Enabling open machine learning of DNA encoded library selections to accelerate the discovery of small molecule protein binders. Preprint at https://doi.org/10.26434/chemrxiv-2024-xd385 (2024). OpenDEL™ Use and Customer Testimonials Begin Your Exploration Today OpenDEL™ 5.0 is now available worldwide. Let HitGen help you with your next drug discovery breakthrough.
In the pursuit of new therapeutics, the initial phase of identifying chemical starting points—or "hits"—is crucial. Among the most prominent strategies employed in modern drug discovery are DNA-Encoded Libraries (DEL), *High-Throughput Screening (HTS), and Fragment-Based Drug Discovery (FBDD). Each offers a distinct philosophy and set of advantages for navigating the vastness of chemical space. The following table provides a high-level overview of their core characteristics, placing emphasis on the innovative DEL approach. Feature/Aspect DNA-Encoded Libraries (DEL) High-Throughput Screening (HTS) Fragment-Based Drug Discovery (FBDD) Philosophy Massive parallel interrogation Automated individual testing Efficient binding & optimization Library Scale 10⁸ – 10¹¹ compounds 10⁵ – 10⁶ compounds 10² – 10⁴ fragments Key Strength Unprecedented scale and diversity at a low cost per compound tested. A proven, direct path to identifying drug-like molecules. High hit rate and superior ligand efficiency of starting points. Main Consideration Requires off-DNA synthesis and validation of hits; DNA-compatible chemistry needed. High infrastructure cost; limited by the actual diversity of the physical library. Requires sensitive biophysical methods (SPR, NMR); optimization can be lengthy. Best Suited For Novel targets, rapid exploration of chemical space, and projects seeking novel chemotypes. Targets with established assay formats and organizations with large, diverse compound collections. Challenging targets with well-defined pockets, where high-quality leads are a priority. Summary and Strategic Outlook The choice between DEL, HTS, and FBDD is rarely a question of which technology is superior, but rather which is the most appropriate for a specific project's goals and constraints. * DEL has emerged as a powerful tool for its ability to screen enormous chemical space in a single experiment, offering a highly cost-effective method for generating novel starting points, especially for emerging or undrugged targets. * HTS remains the established and reliable workhorse for many organizations, providing a direct route to potent hits from existing libraries. * FBDD is often lauded for its efficiency and the high quality of its lead compounds, which typically exhibit excellent optimization potential. A forward-looking R&D strategy often involves integrating these approaches. For instance, a novel hit discovered through a DEL screen can be optimized using the principles of FBDD, while HTS libraries can be augmented with novel chemotypes identified from DELs. This complementary use of technologies leverages the unique strengths of each, creating a more robust and effective drug discovery pipeline.
Gavin W. Collie*, Bryony Ackroyd, Catriona Corbishley, Daniel H. O’Donovan, Alex Edwards, Andrea Gohlke, Xiaoxiao Guo, Bethan Howells, Yuliang Li, Andrew Madin, Alexander G. Milbradt, Emma L. Rivers*,Sandeep K. Talapatra, Elizabeth Underwood, and Alice Webb ACS Med. Chem. Lett. DOI: 10.1021/acsmedchemlett.5c00396 Abstract NSD2 is a key epigenetic regulator and has received considerable attention as a drug target due to its well-documented role in tumorigenesis. We report here a DNA-encoded library screen targeting the PWWP1 domain of NSD2 from which we discovered novel, potent, and selective binders. Furthermore, these compounds were used to develop a novel crystal system, increasing our understanding of the folding of this domain. Together, these results provide a solid molecular and structural basis for the further study of the PWWP1 domain of NSD2 as a cancer drug target.
Ariane F. Frey; Merlin Schwan; Annabelle C. Weldert; Valerie Kadenbach; Jürgen Kopp; Zarina Nidoieva; Robert A. Zimmermann; Lukas Gleue; Collin Zimmer; Marko Jörg; Kristina Friedland; Mark Helm; Irmgard Sinning; Fabian Barthels bioRxiv, 2025 https://doi.org/10.1101/2025.01.15.632061AbstractThe human RNA methyltransferase DNMT2 is thought to be involved in various pathophysiological processes, yet, a major challenge in drug targeting DNMT2 is given by the fact that current SAH-derived inhibitors have poor target selectivity and limited cellular permeability. In this study, we have performed a DNA-encoded library (DEL) screening on DNMT2 yielding five non-SAH-like hit structures, three of which feature a peptidomimetic scaffold. All DEL hits could be validated by orthogonal biophysical and biochemical assays for DNMT2 binding. At the same time, the lead structure did not interact with related methyltransferases from the DNMT and NSUN families highlighting an unmatched DNMT2-targeting selectivity profile. Subsequent crystallographic studies revealed the unique ligand binding mode including an active site loop rearrangement and the formation of a cryptic allosteric binding pocket able to modulate the enzymatic activity by non-covalent DNMT2 dimerization. Based on the crystallographic results, we performed a structure-activity relationship study around the inhibitor lead structure resulting in an optimized DNMT2 inhibitor (KD=3.04 µM), which was able to reduce m5C levels in MOLM-13 tRNA.
CHENGDU, China, July 4, 2025 – Shanghai Stock Exchange listed company HitGen Inc. ("HitGen", SSE: 688222.SH) today announced the release of its inaugural sustainability report, aiming to present the company's philosophies and policies in environmental, social, and governance ("ESG") areas, as well as its sustainability practices and performance in the year of 2024, systematically addressing stakeholders' concerns. Dr. Jin Li, Chairman of the Board and CEO of HitGen Inc., commented: "It is my great pleasure to share with you HitGen's inaugural Sustainability Report. As an enterprise at the forefront of drug discovery research, HitGen aspires to advance innovative drug discovery for the healthcare industry. We are committed to original innovation of drug discovery, continuously delivering new molecular entities to the industry, and contributing to improved human health. In 2024, while continuing to enhance our competitiveness in biopharmaceutical R&D services, we systematically integrated sustainability concepts throughout our operational processes, striving to achieve synergistic development in governance efficiency, low-carbon transmission, talent development, and community well-being." Sustainable Corporate Governance A robust governance framework and capabilities form the solid foundation for sustainable corporate development. HitGen continuously enhances its governance structure to ensure transparent and scientific decision-making. Adhering to compliance and ethical standards as the baseline, we operate with integrity to establish industry benchmarks. We are committed to building a comprehensive risk management system, driving long-term and stable corporate growth, and creating sustainable value for all stakeholders. Responsible Value Chain HitGen advances sustainable development across its entire value chain from supply, operations to client engagement by establishing sustainable supply chain system, industry-academia-research collaborative innovation networks, full-cycle quality control mechanisms, and robust data security safeguards, for the purpose of continuously building a resilient and responsible value ecosystem to fulfill its long-term commitment to human health. Harmonious Internal and External Ecosystem HitGen takes social responsibility as its fundamental duty, demonstrating support through safeguarding employee rights and interests, creating an inclusive atmosphere with diversity and equality, empowering personal growth through training and development programs, protecting employee health and safety, and actively engaging in community contributions – all these actions exemplify the Company’s commitment to society. Eco-Conscious Environmental Management Adhering to the "Eco-Conscious, Planet-Responsible" philosophy, HitGen is dedicated in minimizing adverse impacts on the environment during R&D and operations, conserving natural resources, reducing greenhouse gas emissions, and protecting biodiversity and clean water sources. Dr. Jin Li said, "Guided by our value-creation-driven sustainability vision, we firmly believe our fundamental mission is to create value for society, value for development, and value for the future – thereby propelling technological innovation and business expansion. On this journey, we will persistently leverage market needs as our compass and technological breakthroughs as our engine, continuously optimizing our innovative drug R&D service system to build an ecosystem of positive reciprocity where patients benefit, the industry advances, employees grow, and all stakeholders achieve shared success." For more information on HitGen's Sustainability Report, please visit https://www.hitgen.com/en/sustainability.html
Tony Georgiev, Francesca Migliorini, Andrea Ciamarone, Marco Mueller, Ilaria Biancofiore, Pinuccia Faviana, Francesco Bartoli, Young Seo Park Kim, Lucrezia Principi, Ettore Gilardoni, Gabriele Bassi, Nicholas Favalli, Emanuele Puca, Dario Neri, Sebastian Oehler & Samuele Cazzamalli Nature Biomedical Engineering (2025) DOI: 10.1038/s41551-025-01432-6 Abstract Improving the specificity of prostate cancer treatment requires ligands that bind selectively and with ultra-high affinity to tumour-associated targets absent from healthy tissues. Prostatic acid phosphatase has emerged as an alternative target to prostate-specific membrane antigen, as it is expressed in a broader subset of prostate cancers and is not detected in healthy organs such as the salivary glands and kidneys. Here, to discover selective binders to prostatic acid phosphatase, we constructed two DNA-encoded chemical libraries comprising over 6.7 million small molecules based on proline and phenylalanine scaffolds. Screening against the purified human prostatic acid phosphatase yielded OncoACP3, a small organic ligand with picomolar binding affinity. When radiolabelled with lutetium-177, OncoACP3 selectively accumulated in enzyme-expressing tumours with a long residence time (biological half-life greater than 72 h) and a high tumour-to-blood ratio (>148 at 2 h after administration). Lutetium-177-labelled OncoACP3 cured tumours in mice at low, well-tolerated doses. Its conjugation to the cytotoxic agent monomethyl auristatin E facilitated tumour-selective payload deposition, resulting in potent anti-tumour activity. The modular structure of OncoACP3 supports flexible payload delivery for the targeted treatment of metastatic prostate cancer. Summary Philochem’s research team successfully identified high-affinity ligands targeting prostate-specific membrane antigen (ACP3) using DNA-encoded library (DEL) technology. The study demonstrates a rapid and efficient path from hit identification to preclinical validation, highlighting DEL’s utility in accelerating radioligand therapy development. Highlights 1. High-Affinity Ligand Discovery - Two phosphonate-focused DELs were screened against ACP3, yielding enriched hits with strong binding motifs. - Optimized compounds achieved sub-nanomolar inhibition (SPR-confirmed) and >100-fold improved affinity versus the original ligand. - Fluorophore-conjugated ligands selectively stained ACP3-expressing prostate cancer cells, confirming target engagement. 2. Therapeutic Efficacy in Preclinical Models - 177Lu-labeled conjugates showed ~70 %ID/g tumor uptake in xenografts with minimal off-target accumulation and slow washout. - Significant tumor regression was observed, outperforming a reference radioligand derived from earlier inhibitors. - Small-molecule drug conjugates (cleavable linker + MMAE payload) also demonstrated potent antitumor activity. Conclusion Philochem’s work delivers a robust pipeline of ACP3-targeting ligands with translational potential in radioligand therapy and antibody-free drug conjugates. It validates DEL as a key enabling technology for accelerating cancer therapeutic discovery.
From June 16th to 19th (US time), the 2025 BIO International Convention will take place at the Boston Convention & Exhibition Center in Massachusetts, USA. As the largest and most comprehensive event for biotechnology, the BIO International Convention represents the full ecosystem of biotech with 20,000 industry leaders from across the globe. HitGen Inc. looks forward to meeting you at Booth 1851. We also encourage scheduling meetings in advance through the BIO Partnering™ system to explore collaboration opportunities.
Anjali Dixit & Brian M. Paegel Nature Protocols (2025) DOI: 10.1038/s41596-025-01190-4 Abstract This protocol details the construction of solid-phase DNA-encoded libraries (DELs) that merge one-bead–one-compound (OBOC) combinatorial chemistry with high-throughput DNA sequencing. Photocleavable linkers allow library members to be released from individual polystyrene beads while the encoding DNA tag remains attached, enabling both affinity and activity-based screening. Step-by-step instructions cover bifunctional linker synthesis, split-and-pool combinatorial cycles, enzymatic DNA ligation, quality control by single-bead qPCR/NGS, and validation by MALDI-TOF MS. The entire workflow is completed in ~4 weeks without specialized automation and furnishes 10⁴–10⁶ photocleavable, off-DNA-screenable compounds ready for microfluidic or FACS-based functional assays. Summary Solid-phase DEL technology overcomes the limitations of conventional on-DNA DELs by physically isolating each library member on a bead and permitting photochemical release for cell-free or cell-based activity assays. The protocol begins with synthesis of a dual-scale bead set (160 µm QC beads and 10 µm screening beads) displaying a coumarin/arginine-modified linker terminated with a propargylglycine alkyne and a photocleavable o-nitrobenzyl bromide handle. Copper-catalyzed azide–alkyne cycloaddition (CuAAC) installs an azide-modified hairpiece DNA (N₃-HDNA) to create bifunctional beads. Combinatorial three-cycle chemistry (diamine displacement → Fmoc-amino acid acylation → carboxylic acid acylation) is performed in 96-well filter plates with alternating aqueous DNA-ligation steps that append unique oligonucleotide codes for every building block (BB). A final bead-specific barcode (BSB) ligation increases sequence diversity to >3,000 unique codes per compound. Quality control is rigorous: every cycle is monitored by qPCR quantification of DNA molecules per bead (MPB), LC–MS of TFA-cleaved QC beads, and single-bead Sanger sequencing matched to MALDI-TOF MS. Baseline NGS of 1,500-bead aliquots establishes BB frequency thresholds for hit calling. A 300-mg synthesis routinely yields 6 × 10⁸ beads, equivalent to ~65,000 copies of each compound in a 9,216-member two-cycle library—sufficient for >2,000 microfluidic droplet screens or 50 min FACS analysis. Data analysis scripts (Python/Figshare) decode reads via UMI aggregation and Hamming-distance error correction, enabling structure–activity relationship (SAR) extraction and machine-learning training sets. Highlights 1. Photocleavable, off-DNA screening: Library members are released from beads under UV light while DNA tags remain attached, enabling dose–response and cellular phenotypic assays impossible with conventional on-DNA DELs. 2. One-bead–one-compound plus DNA encoding: Combines OBOC spatial isolation with massively parallel sequence deconvolution, eliminating tedious hit resynthesis and providing direct SAR data. 3. Modular split-and-pool chemistry: Three-cycle protocol (amine→amide→amide) uses 96-well plates and standard peptide-coupling reagents; readily reconfigurable to other medicinally relevant reactions validated by DNA-encoded reaction rehearsal. 4. Built-in quality control: Every synthetic and enzymatic step is tracked by single-bead qPCR, PAGE, LC–MS, and MALDI-TOF MS; ≥90 % of QC beads must yield correct m/z and clean Sanger reads for library release. 5. Scalable & accessible: All reagents are commercially available; no automation required; 4-week timeline from bead swelling to NGS-validated library; yields enough material for thousands of screens at 10–30 compound equivalents per assay. Conclusion The solid-phase DEL protocol described here delivers a versatile, quality-controlled route to 10⁴–10⁶ photocleavable small molecules that can be interrogated in both biochemical and cell-based assays. By integrating OBOC spatial segregation with DNA encoding, researchers gain the throughput advantages of DEL selection while preserving the pharmacological fidelity of off-DNA compound evaluation. Rigorous QC checkpoints at every cycle ensure that libraries entering screening are chemically and physically intact, minimizing false positives and maximizing SAR information richness. The workflow is intentionally manual and low-cost, making functional DEL screening accessible to any laboratory equipped for routine solid-phase peptide synthesis and qPCR.
Xiaoyun Wang; Jianxian Sun; Shabbir Ahmad; Diwen Yang; Fengling Li; U. Hang Chan; Hong Zeng; Conrad V. Simoben; Scott Houliston; Aiping Dong; Albina Bolotokova; Elisa Gibson; Maria Kutera; Pegah Ghiabi; Ivan Kondratov; Tetiana Matviyuk; Alexander Chuprina; Danai Mavridi; Christopher Lenz; Andreas C. Joerger; Benjamin D. Brown; Richard B. Heath; Wyatt W. Yue; Lucy K. Robbie; Tyler S. Beyett; Susanne Müller; Stefan Knapp; Rachel Harding; Matthieu Schapira; Peter J. Brown; Vijayaratnam Santhakumar; Suzanne Ackloo; Cheryl H. Arrowsmith; Aled M. Edwards; Hui Peng; Levon Halabelian bioRxiv, 2025 https://doi.org/10.1101/2025.01.17.633682AbstractWe report an enantioselective protein affinity selection mass spectrometry screening approach (EAS-MS) that enables the detection of weak binders, informs about selectivity, and generates orthogonal confirmation of binding. After method development with control proteins, we screened 31 human proteins against a designed library of 8,210 chiral compounds. 16 binders to 12 targets, including many proteins predicted to be “challenging to ligand”, were discovered and confirmed in orthogonal assays. 7 binders to 6 targets bound in an enantioselective manner, with KDs ranging from 3 to 20 µM. Binders for four targets (DDB1, WDR91, WDR55, and HAT1) were selected for in-depth characterization using X-ray crystallography. In all four cases, the mechanism for enantioselective selectivity was readily explained. EAS-MS can be used to identify and characterize selective and weakly-binding ligands for novel protein targets with unprecedented throughput and sensitivity.
Sumaiya Iqbal, Wei Jiang, Eric Hansen, Tonia Aristotelous, Shuang Liu, Andrew Reidenbach, Cerise Raffier, Alison Leed, Chengkuan Chen, Lawrence Chung, Eric Sigel, Alex Burgin, Sandy Gould, Holly H. Soutter npj Drug Discovery 2, 5 (2025) https://doi.org/10.1038/s44386-025-00007-4 Abstract DNA-encoded library (DEL) technology enables ultra-high-throughput screening of millions to billions of compounds in a cost-effective manner. Coupled with machine learning (ML), DEL data can be used to train predictive models for virtual screening of drug-like compounds. This study presents a systematic evaluation of 15 DEL+ML combinations—three DELs and five ML models—for hit discovery against two therapeutic targets, CK1α and CK1δ. Among 808 predicted binders tested, 10% were confirmed as true binders in biophysical assays, including two nanomolar hits. Additionally, 94% of predicted non-binders were confirmed as true negatives. The study highlights the importance of chemical diversity in training data and model generalizability over accuracy. All models and data have been made publicly available for further use. Summary This study introduces a five-module DEL+ML pipeline: DEL screening, data preparation, ML model development, hit prediction, and experimental validation. Three DELs (MS10M, HG1B, DD11M) and five ML models (MLP, SVM, RF, XGB, ChemProp) were evaluated for their ability to identify orthosteric binders of CK1α/δ. The HG1B DEL-trained ChemProp model achieved the best performance, with a 15% hit rate and identification of two nanomolar binders. The results demonstrate that chemical diversity of the DEL training set is more critical than library size, and that deep learning-based models outperform traditional ML algorithms. Highlights 1. First systematic evaluation of 15 DEL+ML combinations for hit discovery. 2. HG1B DEL-trained ChemProp model achieved the highest hit rate (15%). 3. Two nanomolar binders were identified (CK1α: 308 nM; CK1δ: 187 nM and 69.6 nM). 4. 94% of predicted non-binders were experimentally confirmed as true negatives. 5. Chemical diversity of training data is more important than library size. 6. Neural network models (ChemProp, MLP) outperformed traditional models (RF, SVM, XGB). All models and training data are publicly available for community use. Conclusion This study demonstrates the effectiveness of integrating DEL screening with machine learning for efficient and scalable hit discovery. By systematically comparing different DELs and ML models, we found that the chemical diversity, drug-like properties, and balanced representation of binders and non-binders in the training data are key determinants of model performance. The HG1B DEL-trained ChemProp model emerged as the top performer, successfully identifying multiple potent binders, including nanomolar-level hits. The DEL+ML pipeline not only accelerates hit identification but also effectively filters out non-binders, reducing experimental costs. We recommend using chemically diverse and drug-like DELs for training and leveraging deep learning models for optimal performance. All models and data have been released as open-source resources to support further research and development.
Jagrit Grover; Amal Tom Sebastian; Siddhartha Maiti; Alex C. Bissember; Debabrata Maiti Chem. Soc. Rev., 2025https://doi.org/10.1039/D0CS00488J Abstract In organic synthesis, C(sp3)–H functionalization is a revolutionary method that allows direct alteration of unactivated C–H bonds. It can obviate the need for pre-functionalization and provides access to streamlined and atom economical routes for the synthesis of complex molecules starting from simple starting materials. Many strategies have evolved, such as photoredox catalysis, organocatalysis, non-directed C–H activation, transiently directed C–H activation, and native functionality directed C–H activation. Together these advances have reinforced the importance of C(sp3)–H functionalization in synthetic chemistry. C(sp3)–H functionalization has direct applications in pharmacology, agrochemicals, and materials science, demonstrating its ability to transform synthetic approaches by creating new retrosynthetic disconnections and boost the efficiency of chemical processes. This review aims to provide an overview of current state of C(sp3)–H functionalization, focusing more on recent breakthroughs and associated mechanistic insights.
Yalan Tu; Qi An; Liang Huang; Chuan Wang; Dongmei Wang Front. Pharmacol., 2025, 16https://doi.org/10.3389/fphar.2025.1550158 Abstract Drug discovery plays a crucial role in medicinal chemistry, serving as the cornerstone for developing new treatments to address a wide range of diseases. This review emphasizes the significance of advanced strategies, such as Click Chemistry, Targeted Protein Degradation (TPD), DNA-Encoded Libraries (DELs), and Computer-Aided Drug Design (CADD), in boosting the drug discovery process. Click Chemistry streamlines the synthesis of diverse compound libraries, facilitating efficient hit discovery and lead optimization. TPD harnesses natural degradation pathways to target previously undruggable proteins, while DELs enable high-throughput screening of millions of compounds. CADD employs computational methods to refine candidate selection and reduce resource expenditure. To demonstrate the utility of these methodologies, we highlight exemplary small molecules discovered in the past decade, along with a summary of marketed drugs and investigational new drugs that exemplify their clinical impact. These examples illustrate how these techniques directly contribute to advancing medicinal chemistry from the bench to bedside. Looking ahead, Artificial Intelligence (AI) technologies and interdisciplinary collaboration are poised to address the growing complexity of drug discovery. By fostering a deeper understanding of these transformative strategies, this review aims to inspire innovative research directions and further advance the field of medicinal chemistry.
Leidy D. Caraballo G; Inci Cevher Zeytin; Purva Rathi; Che-Hsing Li; Ai-Ni Tsao; Yaery J. Salvador L; Manish Ranjan; Brendan Magee Traynor; Andras A. Heczey ACS Synth. Biol., 2025, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acssynbio.4c00649 Abstract DNA modification and synthesis are fundamental to genetic engineering, and systems that enable time- and cost-effective execution of these processes are crucial. Iteration of genetic construct variants takes significant time, cost and effort to develop new therapeutic strategies to treat diseases including cancer. Thus, decreasing cost and enhancing simplicity while accelerating the speed of advancement is critical. We have developed a PCR-based platform that allows for deletion, replacement, insertion, mutagenesis, and synthesis of DNA (DRIMS). These modifications rely on the recA-independent recombination pathway and are carried out in a single amplification step followed by DpnI digestion and transformation into competent cells. DNA synthesis is accomplished through sequential PCR amplification reactions without the need for a DNA template. Here, we provide proof-of-concept for the DRIMS platform by performing four deletions within DNA fragments of various sizes, sixty-four replacements of DNA binding sequences that incorporate repeat sequences, four replacements of chimeric antigen receptor components, fifty-one insertions of artificial microRNAs that form complex tertiary structures, five varieties of point mutations, and synthesis of eight DNA sequences including two with high GC content. Compared to other advanced cloning methods including Gibson and “in vivo assembly”, we demonstrate the significant advantages of the DRIMS platform. In summary, DRIMS allows for efficient modification and synthesis of DNA in a simple, rapid and cost-effective manner to accelerate the synthetic biology field and development of therapeutics.
Quan Zuo; Xinyi Song; Jie Yan; Guangjun Bao; Yiping Li; Jieting Shen; Zeyuan He; Kuan Hu; Wangsheng Sun; Rui Wang J. Am. Chem. Soc., 2025, XXXX, XXX, XXX-XXXhttps://doi.org/10.1021/jacs.4c17615 Abstract Modular chemical postmodification of peptides is a promising strategy that supports the optimization and innovation of hit peptide therapeutics by enabling rapid derivatization. However, current methods are primarily limited to traditional bio-orthogonal strategies and chemical ligation techniques, which require the preintroduction of non-natural amino acids and impose fixed methods that limit peptide diversity. Here, we developed the Tyrosine-1,2,3-Triazine Ligation (YTL) strategy, which constructs novel linkages (pyridine and pyrimidine) through a “one-pot, two-step” process combining SNAr and IEDDA reactions, promoting modular post modification of Tyr-containing peptides. After optimizing the YTL strategy and establishing standard procedures, we successfully applied it to the solid-phase postmodification of various biorelated peptides, such as the synthesis of dual-mode imaging probes and long-acting GLP-1 analogs. As a proof of concept, a library of 384 amphipathic peptides was constructed using YTL based on 96-well microfiltration plates. Modular modifications were then performed on the screened template tripeptide RYR, leading to the generation of 20 derivatives. The antibacterial activity of these derivatives was systematically characterized, identifying Z8 as a potential antibacterial candidate.
R. Edward Richard; Boris Belotserkovskii ChemRxiv, 2025 https://doi.org/10.26434/chemrxiv-2025-k5fc7AbstractThis study presents a theoretical framework for understanding the binding behavior of bivalent molecules—entities with two linked ligands—to immobilized targets. Bivalent molecules demonstrate increased binding compared to monovalent counterparts due to an avidity effect arising from the enhanced local concentration of the second ligand upon the first ligand's attachment. Our findings indicate that a shorter tether between ligands increases equilibrium bivalent binding, provided the short tether can span adjacent binding sites without strain. However, shorter tethers may also prolong the search for closely spaced targets, delaying equilibrium to an impractical degree. We propose a theoretical model to evaluate the equilibrium and kinetic parameters of bivalent binding, to enable optimal design of bivalent DNA-Encoded Libraries, and to effect highly efficient DNA-Encoded library (DEL) selection processes.
Yaëlle Fischer; Ruel Cedeno; Dhoha Triki; Bertrand Vivet; Philippe Schambel ACS Med. Chem. Lett., 2025, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsmedchemlett.4c00505AbstractDELs enable efficient experimental screening of vast combinatorial libraries, offering a powerful platform for drug discovery. Apart from ensuring the druglike physicochemical properties, other key parameters to maximize the success rate of DEL designs include the scaffold diversity and target addressability. While several tools exist to assess chemical diversity, a dedicated computational approach combining both parameters is currently lacking. Here, we present a cheminformatics tool leveraging scaffold analysis and machine learning to evaluate both scaffold diversity and target-orientedness. Using two in-house libraries as a case study, we demonstrate the workflow’s ability to distinguish between generalist and focused libraries. This capability can guide medicinal chemists in selecting libraries tailored for specific objectives, such as hit-finding or hit-optimization.
Kaili Zhang; Yi Wang; Shuze Jiang; Yifan Li; Pan Xiang; Yuxuan Zhang; Yongzi Chen; Min Chen; Weijun Su; Liren Liu; Shuai Li Int. J. Biol. Macromol., 2025, 298, 140089https://doi.org/10.1016/j.ijbiomac.2025.140089 Abstract DNA-encoded libraries are invaluable tools for high-throughput screening and functional genomics studies. However, constructing high-abundance libraries in mammalian cells remains challenging. Here, we present dsDNA-assembly-PCR (dsDAP), a novel Gibson-assembly-PCR strategy for creating DNA-encoded libraries, offering improved flexibility and efficiency over previous methods. We demonstrated this approach by investigating the impact of translation initiation sequences (TIS) on protein expression in HEK293T cells. Both CRISPR-Cas9 and piggyBac systems were employed for genomic integration, allowing comparison of different integration methods. Our results confirmed the importance of specific nucleotides in the TIS region, particularly the preference for adenine at the -3 position in high-expression sequences. We also explored the effects of library dilution on genotype-phenotype correlations. This Gibson-assembly-PCR strategy overcomes limitations of existing methods, such as restriction enzyme dependencies, and provides a versatile tool for constructing high-abundance libraries in mammalian cells. Our approach has broad applications in functional genomics, drug discovery, and the study of gene regulation.
Qigui Nie; Xianfu Fang; Jiale Huang; Tingting Xu; Yangfeng Li; Gong Zhang; Yizhou Li Small Methods, 2025, 2401631https://doi.org/10.1002/smtd.202401631 Abstract Deoxyribonucleic acid (DNA), a fundamental biomacromolecule in living organisms, serves as the carrier of genetic information. Beyond its role in encoding biological functions, DNA's inherent ability to hybridize through base pairing has opened new avenues for its application in biological sciences. This review introduces DNA nanotechnology and DNA-encoded library (DEL), and highlights their shared design principles related to DNA assembly. First, a foundational overview of structural DNA nanotechnology, including its design strategies and historical development is provided. Subsequently, various approaches are examined to dynamic DNA nanotechnology, from strand displacement reactions to DNA-templated polymer synthesis. Second, how the principle of DNA assembly has facilitated the development of diverse formats of self-assembly-based DEL synthesis, DNA-template reactions (DTS), and DNA template-mediated proximity induction effects are examined. These advancements are all underpinned by the unique property of DNA assembly. Finally, this review summarizes the common principles shared by DNA nanotechnology and DEL in terms of methodology and design. Additionally, the potential synergies are explored between these two technologies, envisioning future applications where they can be combined to create more versatile and exquisite functionalities.
Brian Belmontes; Katherine K. Slemmons; Chun Su; Siyuan Liu; Antonia N. Policheni; Jodi Moriguchi; Hong Tan; Fang Xie; Daniel Andrew Aiello; Yajing Yang; Raul Lazaro; Famke Aeffner; Matthew G. Rees; Melissa M. Ronan; Jennifer A. Roth; Mikkel Vestergaard; Sanne Cowland; Jan Andersson; Ian Sarvary; Qing Chen; Pooja Sharma; Patricia Lopez; Nuria Tamayo; Liping H. Pettus; Sudipa Ghimire-Rijal; Susmith Mukund; Jennifer R. Allen; Jason DeVoss; Angela Coxon; Jordi Rodon; François Ghiringhelli; Nicolas Penel; Hans Prenen; Sanne Glad; Chen-Hua Chuang; Kiana Keyvanjah; Danielle M. Townsley; John R. Butler; Matthew P. Bourbeau; Sean Caenepeel; Paul E. Hughes Cancer Discov., 2025, 139-161https://doi.org/10.1158/2159-8290.CD-24-0887 Abstract One of the most robust synthetic lethal interactions observed in multiple functional genomic screens has been the dependency on protein arginine methyltransferase 5 (PRMT5) in cancer cells with MTAP deletion. We report the discovery of the clinical stage MTA-cooperative PRMT5 inhibitor AMG 193, which preferentially binds PRMT5 in the presence of MTA and has potent biochemical and cellular activity in MTAP-deleted cells across multiple cancer lineages. In vitro, PRMT5 inhibition induces DNA damage, cell cycle arrest, and aberrant alternative mRNA splicing in MTAP-deleted cells. In human cell line and patient-derived xenograft models, AMG 193 induces robust antitumor activity and is well tolerated with no impact on normal hematopoietic cell lineages. AMG 193 synergizes with chemotherapies or the KRAS G12C inhibitor sotorasib in vitro and combination treatment in vivo substantially inhibits tumor growth. AMG 193 is demonstrating promising clinical activity, including confirmed partial responses in patients with MTAP-deleted solid tumors from an ongoing phase 1/2 study.Significance: AMG 193 preferentially inhibits the growth of MTAP-deleted tumor cells by inhibiting PRMT5 when in complex with MTA, thus sparing MTAP wild-type normal cells. AMG 193 shows promise as a targeted therapy in a clinically defined patient population.
Yuhan Gui; Rui Hou; Yuchen Huang; Yu Zhou; Shihao Liu; Ling Meng; Ying Li; Fong Sang Lam; Ruoyun Ding; Yan Cao; Gang Li; Xiaojie Lu; Xiaoyu Li Angew. Chem. Int. Ed. Engl., 2025, e202421172https://doi.org/10.1002/anie.202421172 Abstract Small molecules that can bind to specific cells have broad application in cancer diagnosis and treatment. Screening large chemical libraries against live cells is an effective strategy for discovering cell-targeting ligands. The DNA-encoded chemical library (DEL or DECL) technology has emerged as a robust tool in drug discovery and has been successfully utilized in identifying ligands for biological targets. However, nearly all DEL selections have predefined targets, while target-agnostic DEL selections interrogating the entire cell surface remain underexplored. Herein, we systematically optimized a cell-based DEL selection method against cancer cells without predefined targets. A 104.96-million-member DEL was selected against MDA-MB-231 and MCF-7 breast cancer cells, representing high and low metastatic properties, respectively, which led to the identification of cell-specific small molecules. We further demonstrated cell-targeting applications of these ligands in cancer photodynamic therapy and targeted drug delivery. Finally, leveraging the DNA tag of DEL compounds, we identified α-enolase (ENO1) as the cell surface receptor of one of the ligands targeting the more aggressive MDA-MB-231 cells. Overall, this work offers an efficient approach for discovering cell-targeting small molecule ligands by using DELs and demonstrates that DELs can be a useful tool to identify specific surface receptors on cancer cells.
Xianfu Fang; Xianguo Ning; Yangfeng Li; Gong Zhang; Xiaohong Fan; Yizhou Li Org. Biomol. Chem., 2025https://doi.org/10.1039/d4ob02089h Abstract Synthesis of chemically diverse heterocyclic scaffolds in DNA-encoded libraries is highly demanded. We herein reported a convenient one-pot multi-component on-DNA synthetic strategy to afford multi-substituted 2,3-dihydrofuran scaffolds via pyridinium ylide-mediated cyclization. This reaction exhibited modest to excellent conversions for a broad range of DNA-conjugated aldehydes, β-ketonitriles and pyridinium salts under mild reaction conditions. Furthermore, the compatibility of this strategy with DEL construction was verified by enzymatic DNA ligation, PCR amplification and mock library synthesis.
Xiansheng Zhang; Xinyuan Zhu; Yong Li; Yan Li; Wen Luo; Maaz Khan; Jiamin Pan; Hong Pan; Hua Xie; Guilong Zhao Curr. Med. Chem., 2025https://doi.org/10.2174/0109298673320136241024054435 Abstract Branched-chain amino acids (BCAAs) are essential amino acids for humans and play an indispensable role in many physiological and pathological processes. Branched-chain amino acid aminotransferase (BCAT) is a key enzyme that catalyzes the metabolism of BCAAs. BCAT is upregulated in many cancers and implicated in the development and progress of some other diseases, such as metabolic and neurological diseases; and therefore, targeting BCAT might be a potential therapeutic approach for these diseases. There are two isoforms of BCAT, i.e., cytoplasmic BCAT1 (or BCATc) and mitochondrial BCAT2 (or BCATm). The discovery of BCAT inhibitors was initiated by Warner-Lambert, a subsidiary of Pfizer, in 2000, followed by many other pharmaceutical companies, such as GlaxoSmithKline (GSK), Ergon, Icagen, Agios, and Bayer. Strategies of high-throughput screening (HTS), DNA-Encoded library technology (ELT), and fragment-based screening (FBS) have been employed for hit identification, followed by structural optimization. Despite low selectivity, both BCAT1 and BCAT2 selective inhibitors were individually developed, each with a few chemical structural classes. The most advanced BCAT1 inhibitor is BAY-069, discovered by Bayer, which has a potent enzymatic inhibitory activity against BCAT1 and a decent in vitro and in vivo pharmacokinetic profile but displayed weaker cellular inhibitory activity and almost no anti-proliferative activity. There are no BCAT inhibitors currently under investigation in clinical trials. Further studies are still needed to discover BCAT inhibitors with a more druggable profile for proof of concept. This review focuses on the latest progress of studies on the understanding of the physiology and pathology of BCAT and the discovery and development of BCAT inhibitors. The structure-activity relationship (SAR) and the druggability, and the challenges of BCAT inhibitors are discussed, with the aim of inspiring the discovery and development of BCAT inhibitors in the future.
Yi Wang; Xiushan Chen In Silico Pharmacol., 2025, 13, 11 https://doi.org/10.1007/s40203-024-00299-wAbstractMatrix metalloproteinase-8 (MMP-8), a type II collagenase, is a key enzyme in the degradation of collagens and is implicated in various pathological processes, making it a promising target for drug discovery. Despite advancements in the development of MMP-8 inhibitors, concerns over potential adverse effects persist. This study aims to address these concerns by focusing on the development of novel compounds with improved safety profiles while maintaining efficacy. In this study, we employed a computational approach to screen potent and safe inhibitors of MMP-8 from the Natural Product Activity and Species Source Database (NPASS). Initially, we constructed a pharmacophore model based on the crystal structure of the MMP-8-FIN complex (PDB ID: 4EY6) utilizing the Pharmit tool. This model then guided the selection of 44 promising molecules from NPASS, setting the stage for further analysis and evaluation. We comprehensively evaluated their drug-likeness and toxicity profiles. Molecules 21, 4, and 44 were identified as potentially effective MMP-8 inhibitors through a robust pipeline that included ADMET profiling, molecular docking, and molecular dynamics simulations. Notably, molecule 21 stood out for its low toxicity, high binding stability, and favorable ADMET profile, while molecule 44 demonstrated excellent affinity. These compounds offer structural novelty compared to known MMP-8 inhibitors. These computational results can be combined with in vitro experiments in the future to validate their activity and safety. These findings provide an important reference for drug design of MMP-8 inhibitors.
Matthew F. Calabrese J. Med. Chem., 2025, 68(2), 1090–1091 https://doi.org/10.1021/acs.jmedchem.4c03091AbstractThe Target 2035 initiative is an ambitious proposal to discover pharmacological tools across the human proteome. This Viewpoint summarizes an approach inspired by that goal leveraging DNA-encoded library coupled with machine learning approaches to assess the ligandability of the WD40 repeat target class of proteins.
Rhian S. Holvey; Daniel A. Erlanson; Iwan J. P. de Esch; Barbara Farkaš; Wolfgang Jahnke; Tsuyoshi Nishiyama; Andrew J. Woodhead J. Med. Chem., 2025, 68(2), 986–1001https://doi.org/10.1021/acs.jmedchem.4c02593 Abstract This Perspective summarizes successful fragment-to-lead (F2L) studies that were published in 2023 and is the ninth installment in an annual series. A tabulated summary of the relevant articles published in 2023 is provided (17 entries from 16 articles), and a comparison of the target classes, screening methods, and overall fragment or lead property trends for 2023 examples and for the combined entries over the years 2015–2023 is discussed. In addition, we identify several trends and innovations in the 2023 literature that promise to further increase the success of fragment-based drug discovery (FBDD), particularly in the areas of NMR and virtual screening, fragment library design, and fragment linking.
Jing Wang; Fan Zhou; Yuping Xu; Lei Zhang Chem. Asian J., 2025, e202401114 https://doi.org/10.1002/asia.202401114 Abstract In this review, we aim to provide an overview of the organic photocatalyst-promoted carbohydrate synthesis and modification under light irradiation, as well as discuss the benefits and drawbacks, pro...
Zher Yin Tan; Joel K. A. Adade; Xiebin Gu; Cody J. S. Hecht; Michael Salcius; Bingqi Tong; Shuang Liu; Seungmin Hwang; Frédéric J. Zécri; Daniel B. Graham; Stuart L. Schreiber; Ramnik J. Xavier Cell Chem. Biol.,2025https://doi.org/10.1016/j.chembiol.2024.12.002 Abstract Chemical inducers of proximity (CIPs) are molecules that recruit one protein to another and introduce new functionalities toward modulating protein states and activities. While CIP-mediated recruitment of E3 ligases is widely exploited for the development of degraders, other therapeutic modalities remain underexplored. We describe a non-degrader CIP-DNA-encoded library (CIP-DEL) that recruits FKBP12 to target proteins using non-traditional acyclic structures, with an emphasis on introducing stereochemically diverse and rigid connectors to attach the combinatorial library. We deployed this strategy to modulate ATG16L1 T300A, which confers genetic susceptibility to Crohn's disease (CD), and identified a compound that stabilizes the variant protein against caspase-3 (Casp3) cleavage in a FKBP12-independent manner. We demonstrate in cellular models that this compound potentiates autophagy, and reverses the xenophagy defects as well as increased cytokine secretion characteristic of ATG16L1 T300A. This study provides a platform to access unexplored chemical space for CIP design to develop therapeutic modalities guided by human genetics.
Hengrui Liu; Arie Zask; Farhad Forouhar; Sho Iketani; Alana Williams; Daniel R. Vaz; Dahlya Habashi; Karenna Choi; Samuel J. Resnick; Seo Jung Hong; David H. Lovett; Tian Bai; Alejandro Chavez; David D. Ho; Brent R. Stockwell Nat. Commun., 2025, 16, 152https://doi.org/10.1038/s41467-024-55421-5 Abstract Variants of SARS-CoV-2 have continued to emerge across the world and cause hundreds of deaths each week. Due to the limited efficacy of vaccines against SARS-CoV-2 and resistance to current therapies, additional anti-viral therapeutics with pan-coronavirus activity are of high interest. Here, we screen 2.8 billion compounds from a DNA-encoded chemical library and identify small molecules that are non-covalent inhibitors targeting the conserved 3CL protease of SARS-CoV-2 and other coronaviruses. We perform structure-based optimization, leading to the creation of a series of potent, non-covalent SARS-CoV-2 3CL protease inhibitors, for coronavirus infections. To characterize their binding mechanism to the 3CL protease, we determine 16 co-crystal structures and find that optimized inhibitors specifically interact with both protomers of the native homodimer of 3CL protease. Since 3CL protease is catalytically competent only in the dimeric state, these data provide insight into the design of drug-like inhibitors targeting the native homodimer state. With a binding mode different from the covalent 3CL inhibitor nirmatrelvir, the protease inhibitor in the COVID drug Paxlovid, these compounds may overcome resistance reported for nirmatrelvir and complement its clinical utility.
Asitha Adrian Abeywardane; Tushar Dattu Apsunde; Gianluca Nicolas Etienne; Alessia Gambardella; James Kevin Mainquist; Andrew J. Meyer; Brian Minie; Steven Paula; Dwayne A. L. Vickers; Ken Yamada; Docket Central US20240183076A1 https://www.freepatentsonline.com/y2024/0183076.htmlAbstractThe present disclosure generally relates to compositions, methods, sorters, systems, devices and uses for screening for bioactive substances in emulsion droplets. In some embodiments, the compositions are a continuous phase formulation for stable emulsions, In some embodiments, the methods are for preparing a monodisperse polyethylene glycol acrylamide (PEGA) co-polymer resin, or for preparing a core-shell bead. In some embodiments, the systems and devices include a sorter, comprising an inlet channel, first and second outlet channels meeting the inlet channel at a junction, and first and second electrodes proximate respective first and second sides of the junction sorter. In some embodiments, the systems and devices include a sorter, comprising a microwell array plate configured to host one microdroplet per microwell, a fluorescence microscope, an imager configured to automatically image assay droplets and identify desired droplets, and an automated microcapillary-based droplet sampling device configured to continuously deposit desired droplets to hit wells.
Benson Chen; Mohammad M. Sultan; Theofanis Karaletsos J. Chem. Inf. Model., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jcim.3c01699AbstractDNA-encoded library (DEL) has proven to be a powerful tool that utilizes combinatorially constructed small molecules to facilitate highly efficient screening experiments. These selection experiments, involving multiple stages of washing, elution, and identification of potent binders via unique DNA barcodes, often generate complex data. This complexity can potentially mask the underlying signals, necessitating the application of computational tools, such as machine learning, to uncover valuable insights. We introduce a compositional deep probabilistic model of DEL data, DEL-Compose, which decomposes molecular representations into their monosynthon, disynthon, and trisynthon building blocks and capitalizes on the inherent hierarchical structure of these molecules by modeling latent reactions between embedded synthons. Additionally, we investigate methods to improve the observation models for DEL count data, such as integrating covariate factors to more effectively account for data noise. Across two popular public benchmark data sets (CA-IX and HRP), our model demonstrates strong performance compared to count baselines, enriches the correct pharmacophores, and offers valuable insights via its intrinsic interpretable structure, thereby providing a robust tool for the analysis of DEL data.
Yanrui Suo; Xu Qian; Zhaoping Xiong; Xiaohong Liu; Chao Wang; Baiyang Mu; Xinyuan Wu; Weiwei Lu; Meiying Cui; Jiaxiang Liu; Yujie Chen; Mingyue Zheng; Xiaojie Lu J. Med. Chem., 2024, 67(21), 18969–18980 https://doi.org/10.1021/acs.jmedchem.4c01416AbstractDNA-encoded library (DEL) technology is an effective method for small molecule drug discovery, enabling high-throughput screening against target proteins. While DEL screening produces extensive data, it can reveal complex patterns not easily recognized by human analysis. Lead compounds from DEL screens often have higher molecular weights, posing challenges for drug development. This study refines traditional DELs by integrating alternative techniques like photocross-linking screening to enhance chemical diversity. Combining these methods improved predictive performance for small molecule identification models. Using this approach, we predicted active small molecules for BRD4 and p300, achieving hit rates of 26.7 and 35.7%. Notably, the identified compounds exhibit smaller molecular weights and better modification potential compared to traditional DEL molecules. This research demonstrates the synergy between DEL and AI technologies, enhancing drug discovery.
Antonia F. Stepan; Alistair D. Richardson Synfacts, 2025, 21(01), 87 https://doi.org/10.1055/a-2467-5525AbstractDNA-encoded library (DEL) platforms have emerged as a powerful tool for drug discovery. Combinatorial chemical synthesis, paired with the unique DNA identifiers, enable researchers to quickly generate large libraries and easily identify the subsequent hits. One of the major challenges facing DEL technology is the lack of DNA-compatible chemical reactions. Although much progress has been made in this area, expanding the DEL toolbox is imperative for continued growth. Building blocks with primary amine functionality are common in DEL synthesis. Thus, developing new transformations capable of on-DNA manipulation of amines is attractive. On-DNA diazo-transfer was initially reported in 2011, however the need for a more general procedure persists. This report outlines the advantages of using fluorosulfuryl azide (FSO2N3) for on-DNA diazo-transfer with high conversion and DNA fidelity. In addition to DNA-compatibility, successful DEL methods must also tolerate the functional groups needed for subsequent transformations. High conversion was achieved on substrates containing halides, alcohols, protected amines, nitro groups, nitriles, and heterocycles. Successful diazo-transfer was demonstrated with aliphatic amines as well as anilines, which were previously challenging substrates. Electron-deficient heteroaromatic anilines, such as 2-aminopyridines, were unsuccessful. Both dsDNA- and ssDNA-conjugated amines were utilized. DNA integrity was established through ligation of a DNA tag and subsequent gel electrophoresis. Quantitative assessment of DNA damage indicated ~94 % remaining amplifiable material. Further functionalization of the DNA-conjugated azide was also demonstrated. The authors include safety information in the Supporting Information.
Zian Liao; Diana Monsivais; Martin M. Matzuk J. Control Release, 2024, 376, 429-440 https://doi.org/10.1016/j.jconrel.2024.10.036AbstractEndometriosis, defined by the growth of endometrial tissues outside of the uterine cavity, is a global health burden for ∼200 million women. Patients with endometriosis usually present with chronic pain and are often diagnosed with infertility. The pathogenesis of endometriosis is still an open question; however, tissue stemness and immunological and genetic factors have been extensively discussed in the establishment of endometriotic lesions. Current treatments for endometriosis can be categorized into pharmacological management of hormone levels and surgical removal of the lesions. Both approaches have limited efficacy, with recurrences often encountered; thus, there is no complete cure for the disease or its symptoms. We review the current knowledge of the etiology of endometriosis and summarize the advancement of pharmacological management of endometriosis. We also discuss our efforts in applying DNA-encoded chemistry technology (DEC-Tec) to identify bioactive molecules for the treatment of endometriosis, offering new avenues for developing non-hormonal treatment options for those patients who seek spontaneous pregnancies.
Colin S. Swenson; Gunasheil Mandava; Deborah M. Thomas; Raymond E. Moellering Chem. Rev., 2024, 124, 22, 13020–13093 https://doi.org/10.1021/acs.chemrev.4c00423AbstractThe development of potent, specific, and pharmacologically viable chemical probes and therapeutics is a central focus of chemical biology and therapeutic development. However, a significant portion of predicted disease-causal proteins have proven resistant to targeting by traditional small molecule and biologic modalities. Many of these so-called “undruggable” targets feature extended, dynamic protein–protein and protein–nucleic acid interfaces that are central to their roles in normal and diseased signaling pathways. Here, we discuss the development of synthetically stabilized peptide and protein mimetics as an ever-expanding and powerful region of chemical space to tackle undruggable targets. These molecules aim to combine the synthetic tunability and pharmacologic properties typically associated with small molecules with the binding footprints, affinities and specificities of biologics. In this review, we discuss the historical and emerging platforms and approaches to design, screen, select and optimize synthetic “designer” peptidomimetics and synthetic biologics. We examine the inspiration and design of different classes of designer peptidomimetics: (i) macrocyclic peptides, (ii) side chain stabilized peptides, (iii) non-natural peptidomimetics, and (iv) synthetic proteomimetics, and notable examples of their application to challenging biomolecules. Finally, we summarize key learnings and remaining challenges for these molecules to become useful chemical probes and therapeutics for historically undruggable targets.
Martin P. Schwalm; Johannes Dopfer; Adarsh Kumar; Francesco A. Greco; Nicolas Bauer; Frank Löhr; Jan Heering; Sara Cano-Franco; Severin Lechner; Thomas Hanke; Ivana Jaser; Viktoria Morasch; Christopher Lenz; Daren Fearon; Peter G. Marples; Charles W. E. Tomlinson; Lorene Brunello; Krishna Saxena; Nathan B. P. Adams; Frank von Delft; Susanne Müller; Alexandra Stolz; Ewgenij Proschak; Bernhard Kuster; Stefan Knapp; Vladimir V. Rogov Nat. Commun., 2024, 15, 10204 https://doi.org/10.1038/s41467-024-54409-5AbstractRecent successes in developing small molecule degraders that act through the ubiquitin system have spurred efforts to extend this technology to other mechanisms, including the autophagosomal-lysosomal pathway. Therefore, reports of autophagosome tethering compounds (ATTECs) have received considerable attention from the drug development community. ATTECs are based on the recruitment of targets to LC3/GABARAP, a family of ubiquitin-like proteins that presumably bind to the autophagosome membrane and tether cargo-loaded autophagy receptors into the autophagosome. In this work, we rigorously tested the target engagement of the reported ATTECs to validate the existing LC3/GABARAP ligands. Surprisingly, we were unable to detect interaction with their designated target LC3 using a diversity of biophysical methods. Intrigued by the idea of developing ATTECs, we evaluated the ligandability of LC3/GABARAP by in silico docking and large-scale crystallographic fragment screening. Data based on approximately 1000 crystal structures revealed that most fragments bound to the HP2 but not to the HP1 pocket within the LIR docking site, suggesting a favorable ligandability of HP2. Through this study, we identified diverse validated LC3/GABARAP ligands and fragments as starting points for chemical probe and ATTEC development.
Zachary Pastorel; Juliette Zanzi; Yves Canac; Olivier Baslé; Michael Smietana Eur. J. Org. Chem., 2024, e202401297 https://doi.org/10.1002/ejoc.202401297AbstractPhotochemistry has provided a powerful alternative to generate reactive intermediates under mild reaction conditions and photobiocatalysis is currently widely applied in organic synthesis. While enzyme-based photocatalysis has attracted a lot of attention, light-driven DNA-based biohybrid systems are also increasingly being applied to a large variety of applications from molecular imaging to enantioselective catalysis, and other fields such as DNA-encoded libraries (DELs). This review gives insights into these research fields by classifying the existing light-driven DNA-based biohybrid systems developed to date.
Byeong Jun Koo; Yonghwan Kwon; Ah Hyeon Son; Dong Hyeon Kim; Soo Bok Kim; Soyeon Kim; Woojin Jeon; Min Sang Kwon; Han Yong Bae Cell Rep. Phys. Sci., 2024, 5, 102279 https://doi.org/10.1016/j.xcrp.2024.102279Abstract<h2>Summary</h2><p>Sulfur(VI)-fluoride exchange (SuFEx) has emerged as a strategy to capitalize on intermolecular conjugation possible using sulfonyl fluorides. The facile synthesis of alkylated sulfur(VI) species is crucial, and their water-compatible nature unlocks a myriad of potential applications. Here, we report an aquaphotocatalysis approach for efficiently synthesizing open-chain sulfonyl fluorides facilitated by bulk water. A range of amines and carboxylic acids is efficiently converted to alkyl sulfonyl fluoride products under mild aqueous conditions, using 0.1–1 mol% of cyanoarene organic photocatalyst (1,3-dicyano-2,4,5,6-tetrakis(diphenylamino)-benzene, 4DP-IPN). Detailed mechanistic investigations suggest a single-electron transfer photochemical pathway. Sulfonyl fluoride products are used in SuFEx click chemistry for intermolecular conjugation with diverse small molecules and a DNA scaffold. This water-accelerated single-electron transfer aquaphotocatalytic strategy provides a platform for accessing alkyl sulfonyl fluoride libraries for potential drug discovery.</p>
Sean M. McKenna; Chiara Borsari; Sean Chin Chan; Etienne J. Donckele; Franco F. Faucher; Melissa C. Grenier-Davies; Krupanandan Haranahalli; Brieuc Matagne; Andrii Monastyrskyi; Rita Petracca ChemMedChem, 2024, e202400853 https://doi.org/10.1002/cmdc.202400853AbstractThe annual EFMC-ACS MEDI Medicinal Chemistry Frontiers was held in Utrecht in Spring 2024. Summarized here are some of the major themes from the event, including the enriching research contributions ...
Seungyoon Kang; Gyung A. Kim; Myo Naing Win; Yeongcheol Ki; Hohjai Lee; Min Su Han Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.4c00483AbstractThe dsDNA-selective fluorescent-dye-based DNA damage assay was developed for DNA-encoded library (DEL) synthesis. For the various DEL synthesis conditions, the assay was validated through cross-checking with high-performance liquid chromatography (HPLC) analysis, and the fact was confirmed that the usage of a specific ratio of organic solvent can critically induce DNA damage. Also, the applicability of the assay was confirmed through the screening of the DNA-damaging condition of the on-DNA amide coupling reaction and Pd-catalyzed on-DNA N-arylation reaction.
Xiang Fu; Yaxin Li; Mingjin He; Junxin Ren; Zinan Wei; Zhenghui Kang; Wenhao Hu Org. Lett., 2024, 26(48), 10376–10381 https://doi.org/10.1021/acs.orglett.4c04011AbstractA DNA-compatible three-component reaction is disclosed for the synthesis of on-DNA polysubstituted isoxazolidines that serve as privileged core scaffolds in numerous natural products and bioactive molecules. This one-pot approach involves the 1,3-dipolar cycloaddition of DNA-tagged styrenes with diazo compounds and nitrosoarenes in an aqueous solution of KOAc. The reaction demonstrates excellent functional group compatibility, providing a conventional protocol for the construction of a DNA-labeled isoxazolidine library.
Tynan Kelly; Xiaolong Yang Biosensors, 2024, 14, 570 https://doi.org/10.3390/bios14120570AbstractRecent advances in drug discovery have established biosensors as indispensable tools, particularly valued for their precision, sensitivity, and real-time monitoring capabilities. The review begins with a brief overview of cancer drug discovery, underscoring the pivotal role of biosensors in advancing cancer research. Various types of biosensors employed in cancer drug discovery are then explored, with particular emphasis on fluorescence- and bioluminescence-based technologies such as FRET, TR-FRET, BRET, NanoBRET, and NanoBiT. These biosensors have enabled breakthrough discoveries, including the identification of Celastrol as a novel YAP-TEAD inhibitor through NanoBiT-based screening, and the development of TR-FRET assays that successfully identified Ro-31-8220 as a SMAD4R361H/SMAD3 interaction inducer. The integration of biosensors in high throughput screening and validation for cancer drug compounds is examined, highlighting successful applications such as the development of LATS biosensors that revealed VEGFR as an upstream regulator of the Hippo signaling pathway. Real-time monitoring of cellular responses through biosensors has yielded invaluable insights into cancer cell signaling pathways, as demonstrated by NanoBRET assays detecting RAF dimerization and HiBiT systems monitoring protein degradation dynamics. The review addresses challenges linked to biosensor applications, such as maintaining stability in complex tumor microenvironments and achieving consistent sensitivity in HTS applications. Emerging trends are discussed, including integrating artificial intelligence and advanced nanomaterials for enhanced biosensor performance. In conclusion, this review offers a comprehensive analysis of fluorescence- and bioluminescence-based biosensor applications in the dynamic cancer drug discovery field, presenting quantitative evidence of their impact and highlighting their potential to revolutionize targeted cancer treatments.
Yaelle Fischer; Ruel Cedeno; Dhoha Triki; Bertrand Vivet; Philippe Schambel ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-9fm01AbstractDNA-encoded libraries (DELs) enable efficient experimental screening of vast combinatorial molecular libraries, making it a powerful platform for drug discovery. Apart from ensuring the druglike physicochemical properties, other key parameters to maximize the success rate of DEL designs include the scaffold diversity and target addressability. While several computational tools have been developed to evaluate DEL chemical diversity, a dedicated tool that combines both parameters is currently lacking. In this work, we developed a computational approach to systematically evaluate both the scaffold diversity and target-orientedness of DELs using Bemis-Murcko (BM) scaffold analysis and machine learning. To demonstrate the utility of this approach, we present a case study using two of our in-house produced libraries. We show that our workflow can effectively distinguish between a generalist and a focused library. Furthermore, we show that although focused libraries tend to have higher compound-based addressability, they could suffer from lower scaffold-based addressability relative to a generalist library. Consequently, we illustrate how our computational tool can guide medicinal chemists in deciding which library to screen as a function of the objective, whether it is hit-finding or hit-optimization. To facilitate utilization, this tool is freely available both as a web application and as a Python script at https://github.com/novalixofficial/NovaWebApp.
James Wellnitz; Shabbir Ahmad; Nabin Begale; Jermiah Joseph; Hong Zeng; Albina Bolotokova; Aiping Dong; Shaghayegh Reza; Pegah Ghiabi; Gibson Elisa; Xuemin Cheng; Guiping Tu; Xianyang Li; Jian Liu; Dengfeng Dou; Jin Li; Rachel J. Harding; Aled M. Edwards; Benjamin Haibe-Kains; Levon Halabelian; Alexander Tropsha; Rafael Couñago ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-xd385AbstractRecent advances in DNA-encoded library (DEL) screening have created bioactivity datasets containing billions of molecules, unlocking new opportunities for machine learning (ML) in drug discovery. However, most ultra-large DEL libraries are proprietary, limiting the advancement of ML tools for big chemical data analytics and hindering the democratization of DEL-ML technology. We address this gap by developing an open, end-to-end DEL-ML framework using public datasets, where enriched binders are represented by common chemical fingerprints, ensuring proprietary data protection. We demonstrate that ML models can be built and validated on fingerprinted DEL data and then applied to virtual screening (VS) of billion-sized, publicly accessible chemical libraries. As a proof-of-concept, we screened the human protein WDR91 using the HitGen OpenDEL library (3 billion molecules) and trained ML models, which were used to screen the Enamine REAL Space library (37 billion molecules). Fifty potential binders were identified, 48 of which were tested, and seven were confirmed as novel binders with dissociation constants (KD) from 2.7 to 21 μM that were successfully co-crystalized with WDR91. This fully automated, open-source workflow demonstrates the potential of DEL-ML models in discovering novel binders and promotes the use of open chemical bioactivity datasets and ML to accelerate drug discovery.
Qingyi Zhao; Hanqing Zhao; Shijie Zhang; Weiwei Lu; Yujun Zhao; Xiaojie Lu; Xuan Wang ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-f03hnAbstractDNA-encoded library technology (DELT) facilitates the generation of billions of DNA-tagged macrocycles containing noncanonical amino acids, synthesized through diverse cyclization strategies. The varied appended residues and ring conformations expand chemical space, enabling the identification of hit compounds with improved properties such as higher binding affinity, enhanced metabolic stability, and increased oral bioavailability. This study expands the on-DNA chemical toolkit by introducing a DNA-compatible C(sp3)-C(sp3) bond formation method via a visible-light-mediated desulfurative macrocyclization. This reaction proceeds efficiently under mild conditions, exhibiting broad substrate scope and good conversions. Notably, the reaction condition is com-patible with free amines, allowing for late-stage modifications and the preparation of covalent DNA-encoded macrocyclic libraries.
Soya Koremura; Akihiro Sugawara; Yohei Morishita; Taro Ozaki; Teigo Asai Org. Lett., 2024, 26(42), 9151–9156 https://doi.org/10.1021/acs.orglett.4c03588AbstractPresented herein is a synthetic biological method using genome mining and heterologous expression systems that provides access to natural products with desirable structural features as building blocks. In this investigation, we synthesized polyketide-peptide hybrid macrocycles with DNA tags, which have the potential to access a DNA-encoded library containing over one million compounds. This study demonstrates that synthetic biology offers a tool for expanding the diversity of building blocks, facilitating the exploration of unexplored chemical space.
Cong Wang; Yinghui Lu; Zhiyuan Peng; Chi Zhang; Qingchao Liu; Xue-Qiang Wang; Tiezheng Jia CCS Chem., 2024 https://doi.org/10.31635/ccschem.024.202404703AbstractWith the blossom of DNA encoded library (DEL) and nucleic acid aptamer techniques, DNA-compatible reactions draw increasing attention in both the pharmaceutical industry and academia. In this regard, the incorporation of organosulfur scaffolds on DNA via ...
Xudong Wang; Liwei Xiong; Ying Zhu; Sixiu Liu; Wenfeng Zhao; Xinyuan Wu; Mengnisa Seydimemet; Linjie Li; Peiqi Ding; Xian Lin; Jiaxiang Liu; Xuan Wang; Zhiqiang Duan; Weiwei Lu; Yanrui Suo; Mengqing Cui; Jinfeng Yue; Rui Jin; Mingyue Zheng; Yechun Xu; Lianghe Mei; Hangchen Hu; Xiaojie Lu J. Am. Chem. Soc., 2024, 146(49), 33983–33996 https://doi.org/10.1021/jacs.4c12992AbstractThe COVID-19 pandemic, exacerbated by persistent viral mutations, underscored the urgent need for diverse inhibitors targeting multiple viral proteins. In this study, we utilized covalent DNA-encoded libraries to discover innovative triazine-based covalent inhibitors for the 3-chymotrypsin-like protease (3CLpro, Nsp5) and the papain-like protease (PLpro) domains of Nsp3, as well as novel non-nucleoside covalent inhibitors for the nonstructural protein 12 (Nsp12, RdRp). Optimization through molecular docking and medicinal chemistry led to the development of LU9, a nonpeptide 3CLpro inhibitor with an IC50 of 0.34 μM, and LU10, whose crystal structure showed a distinct binding mode within the 3CLpro active site. The X-ray cocrystal structure of SARS-CoV-2 PLpro in complex with XD5 uncovered a previously unexplored binding site adjacent to the catalytic pocket. Additionally, a non-nucleoside covalent Nsp12 inhibitor XJ5 achieved a potency of 0.12 μM following comprehensive structure-activity relationship analysis and optimization. Molecular dynamics revealed a potential binding mode. These compounds offer valuable chemical probes for target validation and represent promising candidates for the development of SARS-CoV-2 antiviral therapies.
John C. Taylor; Carmen Gu Liu; James D. Chang; Brianna E. Thompson; Anthony W. Maresso Microbiol. Spectr., 2024, 12(11) https://doi.org/10.1128/spectrum.00365-24AbstractReactive oxygen species (ROS) pose a lethal risk for all life forms by causing damage to cell processes, genome-wide DNA damage-driving mutation, replicative instability, and death. Thus, the development of mechanisms to resist or repair ROS-induced DNA damage is critical for the reliable replication of nucleic acids. DNA repair and protection mechanisms have been discovered in all forms of life. However, the vast array of microbes that may harbor novel repair or protection mechanisms, especially bacterial viruses, have not been adequately assessed. Here, we screened a microbial gene library composed primarily of phage open reading frames (ORFs) to uncover elements that overcome a DNA damage blockade. We report the discovery of one such protein, termed F21, which promotes bacterial survival by possibly repairing or protecting DNA in the face of ROS-induced DNA damage.IMPORTANCEDiscovery of proteins that promote DNA damage repair and protection in the face of reactive oxygen species (ROS) is of vital importance. Our group is in possession of a unique microbial DNA library with which we can screen for undiscovered genes that encode novel proteins with DNA damage repair and protective functions. This library is composed of diverse DNA from a variety of sources, namely bacteriophages, which must be assessed for their novel functions. This work focuses on the discovery of DNA damage repair and protection, but the possibilities for discovery are endless, thus highlighting the significance of this work.
Genhong Zhang; Ruoxi Yu; Yuegang Chen Chin. J. Org. Chem., 2024, 44, XXXX https://doi.org/10.6023/cjoc202408001AbstractDeveloping economical, efficient, and practical methods to construct C(sp2)-C(sp3) bonds has always been a central issue in synthetic chemistry. In recent years, the use of transition metal-catalyzed C(sp2)-C(sp3) cross coupling has become a powerful synthesis method in photochemical and electrochemical synthesis due to its high functional group compatibility, high reaction selectivity and environmental friendliness. Alcohols, as one of the most widely distributed and naturally abundant organic compounds, has great value in organic chemistry in utilizing alcohol deoxygenation to produce alkyl radicals. In this perspective, we have briefly summaried the C(sp2)-C(sp3) coupling reactions of alcohol derivatives with aryl (pseudo) halides under photo/electro catalysis, including esters, N-heterocyclic carbene adducts, phosphine-oxide adducts, bromides and acetals as alcohol derivatives.
Arthur Gonse; Jelena Gajić; Jean-Pierre Daguer; Sofia Barluenga; Robbie Loewith; Nicolas Winssinger ACS Chem. Biol., 2024, 19(12), 2502–2514 https://doi.org/10.1021/acschembio.4c00597AbstractPleckstrin homology (PH) domains are structural motifs critical for cellular processes, such as signal transduction and cytoskeletal organization. Due to their involvement in various diseases, PH domains are promising therapeutic targets, yet their highly charged and hydrophobic binding sites are not ideal for traditional small drugs. In this study, we designed a DNA-encoded library (DEL) mimicking phospholipids to identify novel modulators targeting PH domains with uncharted chemical properties. Screening against several PH domains led to the discovery of 2DII, a small molecule that selectively binds to mSin1PH. This compound can modulate mTORC2 activity by impairing mTORC2's membrane interactions, resulting in reduced AKT1 phosphorylation. A micromapping via Dexter energy transfer based on 2DII bearing an iridium catalyst (2DII-Ir), along with a biotin-diazirine small molecule was used for target identification by proteomics, which confirmed mSin1 as the primary intracellular target of 2DII, demonstrating its potential for selective mTORC2 pathway modulation. These findings introduce a novel strategy for targeting PH domains and provide a foundation for the development of therapeutic interventions that modulate PH-domain-dependent signaling pathways.
Xianfu Fang; Yunzhu Ju; Jiayou Wang; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett., 2024, 26(41), 8916–8921 https://doi.org/10.1021/acs.orglett.4c03369AbstractThiohydantoin represents a significant class of biologically active privileged heterocyclic scaffolds. Herein, we present a convenient and robust DNA-compatible method for constructing a thiohydantoin-focused DNA-encoded library. This reaction can be applied to a wide variety of isothiocyanate partners, arylamine feedstocks, and diverse α-amine acid derivatives, exhibiting excellent conversions, high functional group tolerance, and preservation of DNA tag integrity. Our method allows for easy access to a valuable three-cycle thiohydantoin-focused DNA-encoded library.
Yun Hu; Hillary Dequina; Minxue Huang; Yun Ding; Martha Vestling; Jing Chai; Jennifer Schomaker ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-42fn7AbstractThe synthesis of DNA-encoded libraries (DELs) is a critical drug discovery technology that enables the rapid generation of billions of DNA-tagged compounds. However, a barrier to on-DNA reaction development in academic laboratories is the need for dedicated instrumentation for ion-pairing reversed-phased liquid chromatography-mass spectrometry (IP-RP LC-MS), which is required for reaction monitoring and analysis. In this work, we report the use of a hydrophilic interaction chromatography (HILIC) LC-MS method with an MS-friendly mobile phase as a convenient way to desalt and resolve crude mixtures. This approach allows academic labs to easily translate reaction and sample preparation protocols directly from the literature. HILIC-LC-MS analysis was employed in diverse oxidative three-component click reactions on-DNA that display broad substrate tolerance and observable damage to the DNA. This work not only provides access to new heterocyclic compounds libraries, but offers new tools and inspiration for continued on-DNA reaction development by academic labs due to its ease of setup and analysis.
Masatoshi Niwa; Jun Hayashida; Munefumi Tokugawa; Takeshi Nanya; Masako Tanabe; Naoko Honda; Takehiko Inohana; Hajime Fukano; Yukihiro Shigeta; Takeshi Kuboyama; Shin Itoh Chem., 2024, e202403233 https://doi.org/10.1002/chem.202403233AbstractDNA-encoded library (DEL) technology is a crucial tool in pharmaceutical research, rapidly identifying compounds that bind to a target of interest from an extensive pool of compounds. In this study, we propose a new method for generating single-stranded DELs (ssDELs) with compounds at the 3' end. The introduction of uniquely designed hairpin-shaped headpieces containing deoxyuridine (NC-HP) and the use of a cleavage enzyme facilitate the conversion from double-stranded DELs (dsDELs) to such ssDELs. Moreover, Klenow fill-in provides the dsDELs with photo-crosslinkers covalently linked to the coding region, which exhibit durability even under stringent washing conditions and enable photo-crosslinking with a high signal-to-noise ratio, as also confirmed in cell-based photo-crosslinking selections.
Dario Akaberi; Monireh Pourghasemi Lati; Janina Krambrich; Julia Berger; Grace Neilsen; Emilia Strandback; S. Pauliina Turunen; Johan Wannberg; Hjalmar Gullberg; Martin Moche; Praveen Kumar Chinthakindi; Tomas Nyman; Stefan G. Sarafianos; Anja Sandström; Josef D. Järhult; Kristian Sandberg; Åke Lundkvist; Oscar Verho; Johan Lennerstrand Antimicrob. Agents Chemother., 2024, 68, 10 https://doi.org/10.1128/aac.00909-24AbstractIn vitro screening of large compound libraries with automated high-throughput screening is expensive and time-consuming and requires dedicated infrastructures. Conversely, the selection of DNA-encoded chemical libraries (DECLs) can be rapidly performed with routine equipment available in most laboratories. In this study, we identified novel inhibitors of SARS-CoV-2 main protease (Mpro) through the affinity-based selection of the DELopen library (open access for academics), containing 4.2 billion compounds. The identified inhibitors were peptide-like compounds containing an N-terminal electrophilic group able to form a covalent bond with the nucleophilic Cys145 of Mpro, as confirmed by x-ray crystallography. This DECL selection campaign enabled the discovery of the unoptimized compound SLL11 (IC50 = 30 nM), proving that the rapid exploration of large chemical spaces enabled by DECL technology allows for the direct identification of potent inhibitors avoiding several rounds of iterative medicinal chemistry. As demonstrated further by x-ray crystallography, SLL11 was found to adopt a highly unique U-shaped binding conformation, which allows the N-terminal electrophilic group to loop back to the S1' subsite while the C-terminal amino acid sits in the S1 subsite. MP1, a close analog of SLL11, showed antiviral activity against SARS-CoV-2 in the low micromolar range when tested in Caco-2 and Calu-3 (EC50 = 2.3 µM) cell lines. As peptide-like compounds can suffer from low cell permeability and metabolic stability, the cyclization of the compounds will be explored in the future to improve their antiviral activity.
Shengdong Wang; Haoran Li; Yuwei Liu; Silin Zhang; Lei Ma; Zhongyi Zeng; Zhi Zhou; Hongtao Xu; Wei Yi PREPRINT available at Research Square, 2024 https://doi.org/10.21203/rs.3.rs-4944895/v1AbstractThe development of practical and selective deuterium incorporation strategies for building deuterated molecules, especially deuterium-labeled amino acids, has become an increasing focus in basic research, but still remains challenging. We herein report a bioinspired calcium(II)/HFIP-catalyzed site-selective modular reductive deuteration amination of simple keto acids/esters with amines by using d2-Hantzsch ester as the deuterium source, delivering N-α-deuterated amino acid motifs in a remarkable deuteration efficiency (&gt;99 deuterated ratio) and with diverse functionality (&gt;130 examples), for which various substrates, especially including amino acids, peptides, drug molecules and natural products bearing different substituted forms, was found to be well compatible. Besides, the application of this strategy in the on-DNA synthesis of DNA-tagged N-α-deuterated amino acids/peptides has been exemplified. The work provides an efficient and innovative solution for deuterated amino acid chemistry and should have a robust application potential in organic synthesis, medicinal chemistry and chemical biology.
Xu Wang; Dimitar Gotchev; Kristi Yi Fan; Marvin M. Vega; Nagraj Mani; Kayleigh McGovern-Gooch; Andrea Cuconati; Breanna Tercero; Xiaohe Wang; Philip Carpino; Klaus Maskos; Paolo A. Centrella; Andreas Schmitt; Franziska Preuss; Archna Prasad; Chia-Yi Chen; Matthew A. Clark; John P. Guilinger; Shawn Johnstone; Konstanze von König; Anthony D. Keefe; Jenny Liu; Stéphane Turcotte; Ying Zhang; Debora L. Konz Makino; Angela M. Lam; Andrew G. Cole; Michael J. Sofia J. Med. Chem., 2024, 67(21), 19623–19667 https://doi.org/10.1021/acs.jmedchem.4c02009AbstractThe recent global COVID-19 pandemic has highlighted treatments for coronavirus infection as an unmet medical need. The main protease (Mpro) has been an important target for the development of SARS-CoV-2 direct-acting antivirals. Nirmatrelvir as a covalent Mpro inhibitor was the first such approved therapy. Although Mpro inhibitors of various chemical classes have been reported, they are generally less active against nirmatrelvir-resistant variants and have limited pan-coronavirus potential, presenting a significant human health risk upon future outbreaks. We here present a novel approach and utilized DNA-encoded chemical library screening to identify the noncovalent Mpro inhibitor 5, which demonstrated a distinct binding mode to nirmatrelvir. A macrocyclization strategy designed to lock the active conformation resulted in lactone 12 with significantly improved antiviral activity. Further optimization led to the potent lactam 26, which demonstrated exceptional potency against nirmatrelvir-resistant variants as well as against a panel of viral main proteases from other coronaviruses.
S. Neha Ashraf; J. Henry Blackwell; Geoffrey A. Holdgate; Simon C. C. Lucas; Alisa Solovyeva; R. Ian Storer; Benjamin C. Whitehurst Drug Discov. Today, 2024, 29(10), 104143 https://doi.org/10.1016/j.drudis.2024.104143AbstractIdentification of high-quality hit chemical matter is of vital importance to the success of drug discovery campaigns. However, this goal is becoming ever harder to achieve as the targets entering the portfolios of pharmaceutical and biotechnology companies are increasingly trending towards novel and traditionally challenging to drug. This demand has fuelled the development and adoption of numerous new screening approaches, whereby the contemporary hit identification toolbox comprises a growing number of orthogonal and complementary technologies including high-throughput screening, fragment-based ligand design, affinity screening (affinity-selection mass spectrometry, differential scanning fluorimetry, DNA-encoded library screening), as well as increasingly sophisticated computational predictive approaches. Herein we describe how an integrated strategy for hit discovery, whereby multiple hit identification techniques are tactically applied, selected in the context of target suitability and resource priority, represents an optimal and often essential approach to maximise the likelihood of identifying quality starting points from which to develop the next generation of medicines.
Danila Ryzhikh; Hyewon Seo; Jihoon Lee; Jieon Lee; Myung Hee Nam; Minsoo Song; Gil Tae Hwang J. Org. Chem., 2024, 89(22), 16957–16963 https://doi.org/10.1021/acs.joc.4c02098AbstractThe β-amino ketones produced through the Mannich reaction hold significant potential as candidates for various drugs. In this study, we optimized on-DNA Mannich reaction conditions and applied them to investigate the reactions of DNA-conjugated aldehydes with various amine and ketone building blocks. The developed on-DNA Mannich reaction preserved the DNA integrity and established viable routes for library production. These results underscore the potential of the Mannich reaction in DNA-encoded library (DEL) synthesis.
Grant Koch; Alexander Engstrom; Jaru Taechalertpaisarn; Justin Faris; Satoshi Ono; Matthew R. Naylor; R. Scott Lokey J. Med. Chem., 2024, 67, 21, 19612–19622 https://doi.org/10.1021/acs.jmedchem.4c01956AbstractHydrocarbon-determined shake-flask measurements have demonstrated great utility for optimizing lipophilicity during early drug discovery. Alternatively, chromatographic methods confer reduced experimental error and improved handling of complex mixtures. In this study, we developed a chromatographic approach for estimating hydrocarbon–water shake-flask partition coefficients for a variety of macrocyclic peptides and other bRo5 molecules including PROTACs. The model accurately predicts experimental shake-flask measurements with high reproducibility across a wide range of lipophilicities. The chromatographic retention times revealed subtle conformational effects and correlated with the ability to sequester hydrogen bond donors in low dielectric media. Estimations of shake-flask lipophilicity from our model also accurately predicted trends in MDCK passive cell permeability for a variety of thioether-cyclized decapeptides. This method provides a convenient, high-throughput approach for measuring lipophilic permeability efficiency and predicting passive cell permeability in bRo5 compounds that is suitable for multiplexing pure compounds or investigating the properties of complex library mixtures.
Millicent Dockerill; Pramod M. Sabale; Francesco Russo; Sofia Barluenga; Nicolas Winssinger JACS Au, 2024, 4(10), 4013–4022 https://doi.org/10.1021/jacsau.4c00738AbstractDNA-encoded libraries connect the phenotypes of synthetic molecules to a DNA barcode; however, most libraries do not tap into the potential of Darwinian evolution. Herein, we report a DNA-templated synthesis (DTS) architecture to make peptides that are stabilized into α-helical conformations via head-to-tail supramolecular cyclization. Using a pilot library targeting MDM2, we show that repeated screening can amplify a binder from the lowest abundance in the library to a ranking that correlates to binding affinity. The study also highlights the need to design libraries such that the chemistry avoids biases from the heterogeneous yield in DTS.
Višnja Stepanić; Dalibor Hršak; Renata Kobetić Croat. Chem. Acta, 2024, 97(4) https://doi.org/10.5562/cca4130AbstractMolecules that act on the biological target at micromolar level at least are called hits. The usual method for identifying hits is high-throughput screening (HTS) of chemical libraries in relevant in vitro assays. An even more efficient, cost-effective and faster method for identifying hits is to perform virtual pre-screening, where the top scoring hits are validated in appropriate in vitro assays. Both wet HTS and virtual screening using structure- or ligand-based approaches utilise large libraries containing millions to billions of drug-like compounds. In this paper, we provide an insight into the state of the art in large collections of small molecular weight molecules, i) public databases for synthetic compounds (PubChem, ChEMBL) and natural products (COCONUT, LOTUS) and commercial ultra-large chemical libraries, ii) make-on-demand virtual libraries (Enamine, Galaxi®, ZINC-22) and iii) wet DNA-encoded libraries (DELs). Machine learning methods for characterising and visualising molecular diversity in screening collections are also described.
Wei Hou; Shaoneng Hou; Yuang Gu; Shuning Zhang; Peixiang Ma; Hai-Yu Hu; Hongtao Xu ChemBioChem, 2024, 25(24), e202400641 https://doi.org/10.1002/cbic.202400641AbstractThe continuous development of click reactions with new connecting linkage is crucial for advancing the frontiers of click chemistry. Selenium-nitrogen exchange (SeNEx) chemistry, a versatile chemistry in click chemistry, represents an all-encompassing term for nucleophilic substitution events that replace nitrogen at an electrophilic selenium(II) center, enabling the flexible and efficient assembly of linkages around a Se(II) core. Several SeNEx chemistries have been developed inspired by the biochemical reaction between Ebselen and cysteine residue, and demonstrated significant potential in on-plate nanomole-scale parallel synthesis, selenium-containing DNA-encoded library (SeDEL) synthesis, as well as peptide and protein bioconjugation. This concept aims to present the origins, advancements, and applications of selenium(II)-nitrogen exchange (SeNEx) chemistry while also outlining the potential directions for future research in this field.
Mengmeng Zheng; Lingchao Kong; Jianmin Gao Chem. Soc. Rev., 2024, 53, 11888 https://doi.org/10.1039/D4CS00750FAbstractNovel bioconjugation reactions have been heavily pursued for the past two decades. A myriad of conjugation reactions have been developed for labeling molecules of interest in their native context as well as for constructing multifunctional molecular entities or stimuli-responsive materials. A growing cluster of bioconjugation reactions were realized by tapping into the unique properties of boron. As a rare element in human biology, boronic acids and esters exhibit remarkable biocompatibility. A number of organoboron reagents have been evaluated for bioconjugation, targeting the reactivity of either native biomolecules or those incorporating bioorthogonal functional groups. Owing to the dynamic nature of B–O and B–N bond formation, a significant portion of the boron-enabled bioconjugations exhibit rapid reversibility and accordingly have found applications in the development of reversible covalent inhibitors. On the other hand, stable bioconjugations have been developed that display fast kinetics and significantly expand the repertoire of bioorthogonal chemistry. This contribution presents a summary and comparative analysis of the recently developed boron-mediated bioconjugations. Importantly, this article seeks to provide an in-depth discussion of the thermodynamic and kinetic profiles of these boron-enabled bioconjugations, which reveals structure–reactivity relationships and provides guidelines for bioapplications.
Huda Barhoosh; Anjali Dixit; Wesley G. Cochrane; Valerie Cavett; Robin N. Prince; Brooke O. Blair; Fred R. Ward; Kim F. McClure; Phillip A. Patten; Margot G. Paulick; Brian M. Paegel ACS Cent. Sci., 2024, 10(10), 1960–1968 https://doi.org/10.1021/acscentsci.4c01218AbstractSmall molecule probes exist for only ∼2% of human proteins because most lack functional binding pockets or cannot be assayed for high-throughput screening. Selective translation modulation circumvents canonical druggability and assay development constraints by using in vitro transcription-translation (IVTT) as a universal biochemical screening assay. We developed an IVTT activity assay by fusing a GFP reporter to various target gene sequences and screened the target sequences for inhibitors in microfluidic picoliter-scale droplets using a 5,348-member translation inhibitor DNA-encoded library (DEL). Screening a proof-of-concept PCSK9-GFP reporter yielded many hits; 6/7 hits inhibited PCSK9-GFP IVTT (IC50 1-20 μM), and the lead hit reduced PCSK9 levels in HepG2 cells. Preliminary selectivity was informed by counterscreening the DEL against a frameshift mutant PCSK9-GFP reporter. A plug-and-play approach to assay development and screening was demonstrated by scouting the DEL for activity using reporter genes of targets with difficult-to-assay or even unknown function (RPL27, KRASG12D, MST1, USO1). This microfluidic IVTT DEL screening platform could scale probe discovery to the human proteome and perhaps more broadly across the tree of life.
Sumaiya Iqbal; Wei Jiang; Eric Hansen; Tonia Aristotelous; Shuang Liu; Andrew Reidenbach; Cerise Raffier; Alison Leed; Chengkuan Chen; Lawrence Chung; Eric Sigel; Alex Burgin; Sandy Gould; Holly Soutter ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-2xrx4AbstractDNA-Encoded Library (DEL) technology allows the screening of millions, or even billions, of encoded compounds in a pooled fashion which is faster and cheaper than traditional approaches. These massive amounts of data related to DEL binders and not-binders to the target of interest enable Machine Learning (ML) model development and screening of large, readily accessible, drug-like libraries in an ultra-high-throughput fashion. Here, we report a comparative assessment of the DEL+ML pipeline for hit discovery using three DELs and five ML models (fifteen DEL+ML combinations using two different feature representations). Each ML model was used to screen a diverse set of drug-like compound collections to identify orthosteric binders of two therapeutic targets, Casein kinase 1𝛼/δ (CK1𝛼/δ). Overall, 10% and 94% of the predicted binders and not-binders were confirmed in biophysical assays, including two nanomolar binders (187 and 69.6 nM affinity for CK1𝛼 and CK1δ, respectively). Our study provides insights into the DEL+ML paradigm for hit discovery: the importance of an ensemble ML approach in identifying a diverse set of confirmed binders, the usefulness of large training data and chemical diversity in the DEL, and the significance of model generalizability over accuracy. We shared our results via an open-source repository for further use and development of similar efforts.
Regina Pikalyova; Tagir Akhmetshin; Dragos Horvath; Alexandre Varnek ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-qh3bnAbstractVisualization of the combinatorial library chemical space provides a comprehensive overview of available compound classes, their diversity, and physicochemical property distribution - key factors in drug discovery. Typically, this visualization requires time- and resource-consuming compound enumeration, standardization, descriptor calculation, and dimensionality reduction. In this study, we present the Combinatorial Library Neural Network (CoLiNN) designed to predict the projection of compounds on a 2D chemical space map using only their building blocks and reaction information, thus eliminating the need for compound enumeration. Trained on 2.5K virtual DNA-Encoded Libraries (DELs), CoLiNN demonstrated high predictive performance, accurately predicting the compound position on Generative Topographic Maps (GTMs). GTMs predicted by CoLiNN were found very similar to the maps built for enumerated structures. In the library comparison task, we compared the GTMs of DELs and the ChEMBL database. The similarity-based DELs / ChEMBL rankings obtained with “true” and CoLiNN predicted GTMs were consistent. Therefore, CoLiNN has the potential to become the go-to tool for combinatorial compound library design – it can explore the library design space more efficiently by skipping the compound enumeration.
Xin Jin; Ning Ding; Hong-Yu Guo; Qing Hu Eur. J. Med. Chem., 2024, 116733 https://doi.org/10.1016/j.ejmech.2024.116733AbstractMacrocyclic compounds have emerged as potent tools in the field of drug design, offering unique advantages for enhancing molecular recognition, improving pharmacokinetic properties, and expanding the chemical space accessible to medicinal chemists. This review delves into the evolutionary trajectory of macrocyclic-based strategies, tracing their journey from laboratory innovations to clinical applications. Beginning with an exploration of the defining structural features of macrocycles and their impact on drug-like characteristics, this discussion progresses to highlight key design principles that have facilitated the development of diverse macrocyclic drug candidates. Through a series of illustrative representative case studies from approved macrocyclic drugs and candidates spanning various therapeutic areas, particular emphasis is placed on their efficacy in targeting challenging protein-protein interactions, enzymes, and receptors. Additionally, this review thoroughly examines how macrocycles effectively address critical issues such as metabolic stability, oral bioavailability and selectivity. Valuable insights into optimization strategies employed during both approved and clinical phases underscore successful translation of promising leads into efficacious therapies while providing valuable perspectives on harnessing the full potential of macrocycles in drug discovery and development endeavors.
Pratik R. Chheda; Nicholas Simmons; Zhicai Shi Org. Lett., 2024, 26, 31, 6754–6759 https://doi.org/10.1021/acs.orglett.4c02474AbstractOn-DNA carboxylic acids are important synthetic intermediates in the synthesis of DNA-encoded library (DEL) structures. Herein, we report an oxoammonium salt-mediated, room temperature, solution-phase oxidation of DNA-linked primary alcohols into carboxylic acids. This method exhibits a wide substrate scope, encompassing aliphatic, benzylic, and heterobenzylic alcohols, and is compatible with DEL encoding strategies. This advancement facilitates a DEL strategy to utilize unprotected alcohols as inert, masked carboxylic acids and enables access to noncommercial bifunctional carboxyl intermediates to enhance the accessible chemical diversity within DELs.
Juraj Velcicky; Matthias R. Bauer; Achim Schlapbach; Guillaume Lapointe; Arndt Meyer; Markus Vögtle; Ernst Blum; Estelle Ngo; Catherine Rolando; Pierre Nimsgern; Sylvie Teixeira-Fouchard; Hansjoerg Lehmann; Pascal Furet; Frédéric Berst; Jens Schümann; Rowan Stringer; Patrice Larger; Cindy Schmid; Catriona T. Prendergast; Simone Riek; Patrick Schmutz; Sylvie Lehmann; Jörg Berghausen; Clemens Scheufler; Jean-Michel Rondeau; Christoph Burkhart; Thomas Knoepfel; Nina Gommermann J. Med. Chem., 2024, 67, 18, 16692–16711 https://doi.org/10.1021/acs.jmedchem.4c01520AbstractIL-17, a pro-inflammatory cytokine produced mainly by Th17 cells, is involved in the immune response to fungal and bacterial infections, whereas its aberrant production is associated with autoimmune and inflammatory diseases. IL-17 blocking antibodies like secukinumab (Cosentyx) have been developed and are used to treat conditions like psoriasis, psoriatic arthritis, and ankylosing spondylitis. Recently, the low molecular weight IL-17 inhibitor LY3509754 entered the clinic but was discontinued in Phase 1 due to adverse effects. In this study, we explored the replacements of furazan moiety posing a potential toxicology risk in LY3509754. By exploring replacements such as heterocycles as amide-isosteres as well as α-F-acrylamides, two compounds (18 and 26) were identified. Both compounds effectively reduced knee swelling in a rat arthritis model. However, early rat and dog toxicity studies revealed adverse findings, preventing their further development and indicating that furazan might not be responsible for the adverse effects of LY3509754.
Yanrui Suo; Kaige Li; Xing Ling; Kenian Yan; Weiwei Lu; Jinfeng Yue; Xiaohua Chen; Zhiqiang Duan; Xiaojie Lu Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.4c00307AbstractThe DNA-encoded library (DEL) is a robust tool for chemical biology and drug discovery. In this study, we developed a DNA-compatible light-promoted reaction that is highly efficient and plate-compatible for DEL construction based on the formation of the indazolone scaffold. Employing this high-efficiency approach, we constructed a DEL featuring an indazolone core, which enabled the identification of a novel series of ligands specifically targeting E1A-binding protein (p300) after DEL selection. Taken together, our findings underscore the feasibility of light-promoted reactions in DEL synthesis and unveil promising avenues for developing p300-targeting inhibitors.
Ravikumar Jimmidi; Diana Monsivais; Hai Minh Ta; Kiran L. Sharma; Kurt M. Bohren; Srinivas Chamakuri; Zian Liao; Feng Li; John M. Hakenjos; Jian-Yuan Li; Yuji Mishina; Haichun Pan; Xuan Qin; Matthew B. Robers; Banumathi Sankaran; Zhi Tan; Suni Tang; Yasmin M. Vasquez; Jennifer Wilkinson; Damian W. Young; Stephen S. Palmer; Kevin R. MacKenzie; Choel Kim; Martin M. Matzuk Proc. Natl. Acad. Sci., USA, 2024, 121(47), e2413108121 https://doi.org/10.1073/pnas.2413108121AbstractActivin receptor type 1 (ACVR1; ALK2) and activin receptor like type 1 (ACVRL1; ALK1) are transforming growth factor beta family receptors that integrate extracellular signals of bone morphogenic proteins (BMPs) and activins into Mothers Against Decapentaplegic homolog 1/5 (SMAD1/SMAD5) signaling complexes. Several activating mutations in ALK2 are implicated in fibrodysplasia ossificans progressiva (FOP), diffuse intrinsic pontine gliomas, and ependymomas. The ALK2 R206H mutation is also present in a subset of endometrial tumors, melanomas, non-small lung cancers, and colorectal cancers, and ALK2 expression is elevated in pancreatic cancer. Using DNA-encoded chemistry technology, we screened 3.94 billion unique compounds from our diverse DNA-encoded chemical libraries (DECLs) against the kinase domain of ALK2. Off-DNA synthesis of DECL hits and biochemical validation revealed nanomolar potent ALK2 inhibitors. Further structure-activity relationship studies yielded center for drug discovery (CDD)-2789, a potent [NanoBRET (NB) cell IC50: 0.54 μM] and metabolically stable analog with good pharmacological profile. Crystal structures of ALK2 bound with CDD-2281, CDD-2282, or CDD-2789 show that these inhibitors bind the active site through Van der Waals interactions and solvent-mediated hydrogen bonds. CDD-2789 exhibits high selectivity toward ALK2/ALK1 in KINOMEscan analysis and NB K192 assay. In cell-based studies, ALK2 inhibitors effectively attenuated activin A and BMP-induced Phosphorylated SMAD1/5 activation in fibroblasts from individuals with FOP in a dose-dependent manner. Thus, CDD-2789 is a valuable tool compound for further investigation of the biological functions of ALK2 and ALK1 and the therapeutic potential of specific inhibition of ALK2.
Ingo Muegge; Jörg Bentzien; Yunhui Ge Expert Opin. Drug Discov., 2024 https://doi.org/10.1080/17460441.2024.2390511AbstractABSTRACT Introduction For the past two decades, virtual screening (VS) has been an efficient hit finding approach for drug discovery. Today, billions of commercially accessible compounds are routinely screened, and many successful examples of VS have been reported. VS methods continue to evolve, including machine learning and physics-based methods. Areas covered The authors examine recent examples of VS in drug discovery and discuss prospective hit finding results from the critical assessment of computational hit-finding experiments (CACHE) challenge. The authors also highlight the cost considerations and open-source options for conducting VS and examine chemical space coverage and library selections for VS. Expert opinion The advancement of sophisticated VS approaches, including the use of machine learning techniques and increased computer resources as well as the ease of access to synthetically available chemical spaces, and commercial and open-source VS platforms allow for interrogating ultra-large libraries (ULL) of billions of molecules. An impressive number of prospective ULL VS campaigns have generated potent and structurally novel hits across many target classes. Nonetheless, many successful contemporary VS approaches still use considerably smaller focused libraries. This apparent dichotomy illustrates that VS is best conducted in a fit-for-purpose way choosing an appropriate chemical space. Better methods need to be developed to tackle more challenging targets.
Huiya Zhang; Yuyu Xing; Yixuan Yang; Bixi Tang; Zhaoyun Zong; Jia Li; Yi Zang; Matthew Bogyo; Xiaojie Lu; Shiyu Chen Chembiochem, 2024, 25(23), e202400559 https://doi.org/10.1002/cbic.202400559AbstractProfiling the substrate sequence preferences of proteases is important for understanding both biological functions as well as for designing protease inhibitors. Several methods are available for profiling the sequence specificity of proteases. However, there is currently no rapid and high-throughput method to profile specificity of proteases for noncanonical substrates. In this study, we described a strategy to use a DNA-encoded noncanonical substrate library to identify the protease substrates composed of both canonical and noncanonical amino acids. This approach uses a DNA-encoded peptide library and introduces a biotin molecule at the N-terminus to immobilize the library on a solid support. Upon protease hydrolysis, the released DNA tag of the substrate peptides can be sequenced to identify the substrate structures. Using this approach, we profiled trypsin and fibroblast activation protein α and discovered noncanonical substrates that were more efficiently cleaved than the commonly used substrates. The identified substrates of FAP were further used to design corresponding covalent inhibitors containing non-canonical sequences with high potency for the target protease. Overall, our approach can aid in the development of new protease substrates and inhibitors.
Yiran Huang; Rui Hou; Fong Sang Lam; Yunxuan Jia; Yu Zhou; Xun He; Gang Li; Feng Xiong; Yan Cao; Dongyao Wang; Xiaoyu Li J. Am. Chem. Soc., 2024, 146(35), 24638–24653 https://doi.org/10.1021/jacs.4c08624AbstractIdentifying biologically active ligands for membrane proteins is an important task in chemical biology. We report an approach to directly identify small molecule agonists against membrane proteins by selecting DNA-encoded libraries (DELs) on live cells. This method connects extracellular ligand binding with intracellular biochemical transformation, thereby biasing the selection toward agonist identification. We have demonstrated the methodology with three membrane proteins: epidermal growth factor receptor (EGFR), thrombopoietin receptor (TPOR), and insulin receptor (INSR). A ∼30 million and a 1.033 billion-compound DEL were selected against these targets, and novel agonists with subnanomolar affinity and low micromolar cellular activities have been discovered. The INSR agonists activated the receptor by possibly binding to an allosteric site, exhibited clear synergistic effects with insulin, and activated the downstream signaling pathways. Notably, the agonists did not activate the insulin-like growth factor 1 receptor (IGF-1R), a highly homologous receptor whose activation may lead to tumor progression. Collectively, this work has developed an approach toward "functional" DEL selections on the cell surface and may provide a widely applicable method for agonist discovery for membrane proteins.
Qianping Huang; Yuang Gu; An Qin; Peixiang Ma; Hongtao Xu; Shuning Zhang ACS Med. Chem. Lett., 2024, 15(9), 1591–1597 https://doi.org/10.1021/acsmedchemlett.4c00307AbstractDNA-encoded library (DEL) is a powerful hit selection technique in both basic science and innovative drug discovery. In this study, we report a robust and straightforward DNA-compatible diazo-transfer reaction utilizing FSO2N3 as the diazo-transfer reagent in solution. This reaction demonstrates high conversions and facile operation while being metal-free and maintaining high levels of DNA fidelity. It is also compatible with a wide range of substrates, allowing for convenient access to both aliphatic and aromatic amines. Consequently, it will further enrich the DEL chemistry toolbox.
Chuan Liu; Zhonghui Chen; Tao Chen; Hongmei Song; Jianbo Shen; Xiaoxi Yuan; Shuai Xia; Qian Liu; Qiuxia Chen; Qiang Tian; Xiaoyun Meng; Zhu Han; Xiaofei Dong; Yu Yang; Longying Cai; Xuemin Cheng; Yangyang Jia; Guansai Liu; Jin Li; Junyou Ge; Dengfeng Dou J. Med. Chem., 2024, 67(17), 15780–15795 https://doi.org/10.1021/acs.jmedchem.4c01412AbstractPeptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1) has emerged as a promising therapeutic target for cancer treatment. However, the current PIN1 inhibitors have shown limited efficacy in animal models, leaving the question of whether PIN1 is a proper oncologic target still unanswered. By screening a 1 trillion DNA-encoded library (DEL), we identified novel nonacidic compounds. Among resynthesized DEL compounds, DEL1067-56-469 (A0) is the most potent one (KD = 430 nM, IC50 = 420 nM). Further optimization of A0 resulted in compound C10 with much improved potency (KD = 25 nM, IC50 = 150 nM). As an alternative approach, C10 was then converted into proteolysis targeting chimeras (PROTACs) in order to achieve deeper downregulation of the PIN1 protein in cancer cell lines. Unfortunately, neither PIN1 inhibitors nor PIN1 PROTACs demonstrated meaningful antiproliferation activity. In addition, siRNA knock-down experiments provided unfavorable evidence of PIN1 as an oncologic target. Our findings highlight the complexity of targeting PIN1 for cancer therapy.
Yi Zhao; Meixian Liu; Tian Qin; Yongqiang Peng; Guang Lin; Chao Che; Zhendong Zhu SLAS Tech., 2024 https://doi.org/10.1016/j.slast.2024.100174AbstractAn optimized Affinity Selection-Mass Spectrometry (AS-MS) workflow has been developed for the efficient identification of potent USP1 inhibitors. USP1 was immobilized on agarose beads, ensuring low small molecule retention, efficient protein capture, and protein stability. The binding affinity of 49 compounds to USP1 was evaluated using the optimized AS-MS method, calculating binding index (BI) values for each compound. Biochemical inhibition assays validated the AS-MS results, revealing a potential correlation between higher BI values and lower IC50 values. This optimized workflow enables rapid identification of high-quality USP1 inhibitor hits, facilitating structure-activity relationship studies and accelerating the discovery of potential cancer therapeutics.
Michelle Keller; Dimitar Petrov; Andreas Gloger; Bastien Dietschi; Kilian Jobin; Timon Gradinger; Adriano Martinelli; Louise Plais; Yuichi Onda; Dario Neri; Jörg Scheuermann Science, 2024, 384(6701), 1259-1265 https://doi.org/10.1126/science.adn3412AbstractThe first drugs discovered using DNA-encoded chemical library (DEL) screens have entered late-stage clinical development. However, DEL technology as a whole still suffers from poor chemical purity resulting in suboptimal performance. In this work, we report a technique to overcome this issue through self-purifying release of the DEL after magnetic bead–based synthesis. Both the first and last building blocks of each assembled library member were linked to the beads by tethers that could be cleaved by mutually orthogonal chemistry. Sequential cleavage of the first and last tether, with washing in between, ensured that the final library comprises only the fully complete compounds. The outstanding purity attained by this approach enables a direct correlation of chemical display and encoding, allows for an increased chemical reaction scope, and facilitates the use of more diversity elements while achieving greatly improved signal-to-noise ratios in selections. DNA-encoded chemical libraries enable screening of an enormous number of prospective drug compounds in parallel. The idea is that the functional components of each prospective drug are encoded with a DNA sequence that can be amplified later to identify the structure of a hit. However, incomplete syntheses can introduce errors in matching the tag to the functionality. Keller et al. devised a dual linker technique to ensure fidelity. Orthogonally cleavable linkers tether the first and last introduced components to a bead. Cutting both linkers successively, with washing in between, release only complete structures. —Jake S. Yeston
Pieter Bos; Fabio Ranalli; Emelie Flood; Shawn Watts; Daigo Inoyama; Jennifer Knight; Anthony Clark; Andrew Placzeck; Jiashi Wang; Aleksey Gerasyuto; Sarah Silvergleid; Wu Yin; Shaoxian Sun; Robert Abel; Sathesh Bhat ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-nxrg6AbstractThe hit identification stage of a drug discovery program generally involves the design of novel chemical scaffolds with desired biological activity against the target(s) of interest. One common approach is scaffold hopping, which is the manual design of novel scaffolds based on known chemical matter. One major limitation of this approach is narrow chemical space exploration, which can lead to difficulties in maintaining or improving biological activity, selectivity, and favorable property space. Another limitation is the lack of preliminary structure-activity relationship (SAR) data around these designs, which could lead to selecting suboptimal scaffolds to advance lead optimization. To address these limitations, we propose AutoDesigner - Core Design (CoreDesign), a de novo scaffold design algorithm. Our approach is a cloud-integrated, de novo design algorithm for systematically exploring and refining chemical scaffolds against biological targets of interest. The algorithm designs, evaluates, and optimizes a vast range - from millions to billions - of molecules in silico, following defined project parameters encompassing structural novelty, physicochemical attributes, potency, and selectivity. In this manner, CoreDesign can generate novel scaffolds and also explore preliminary SAR around each scaffold using FEP+ potency predictions. CoreDesign requires only a single ligand with quantifiable binding affinity and an initial binding hypothesis, making it especially suited for the hit-identification stage where experimental data is often limited. To validate CoreDesign in a real-world drug discovery setting, we applied it to the design of novel, potent Wee1 inhibitors with improved selectivity over PLK1. Starting from a single known ligand, CoreDesign rapidly explored over 23 billion molecules to identify 1,342 novel chemical series with a mean of 4 compounds per scaffold. Importantly, all chemical series met the predefined property space requirements. To rapidly analyze this large amount of data and prioritize chemical scaffolds for synthesis, we utilize t-Distributed Stochastic Neighbor Embedding (t-SNE) plots of in silico properties. The chemical space projections allowed us to rapidly identify a structurally novel 5-5 fused core meeting all the hit-identification requirements. Several compounds were synthesized and assayed from the scaffold, displaying good potency against Wee1 and excellent PLK1 selectivity. Our results suggest that CoreDesign can significantly speed up the hit-identification process and increase the probability of success of drug discovery campaigns by allowing teams to bring forward high-quality chemical scaffolds de-risked by the availability of preliminary SAR.
Hyemi Lee; Jihye Choe; Min-Hee Son; In-Hyun Lee; Min Ju Lim; Jimin Jeon; Siyoung Yang Antioxidants, 2024, 13(8), 943 https://doi.org/10.3390/antiox13080943AbstractBromodomain and extra-terminal domain (BET) family proteins regulate transcription and recognize lysine residues in histones. Selective BET inhibitors targeting one domain have attracted attention because they maintain normal physiological activities, whereas pan (nonselective) BET inhibitors do not. Osteoarthritis (OA) is a joint disorder characterized by cartilage degeneration for which no treatment currently exists. Here, we investigated whether the selective inhibition of BET proteins is an appropriate therapeutic strategy for OA. We focused on the development and characterization of 2-(4-(2-(dimethylamino)ethoxy)-3,5-dimethylphenyl)-5,7-dimethoxyquinazolin-4(3H)-one (BBC0906), a novel bromodomain 2 (BD2)-specific inhibitor designed to suppress OA progression. Using a DNA-encoded chemical library (DEL) screening approach, BBC0906 was identified because of its high affinity with the BD2 domain of BET proteins. BBC0906 effectively reduced reactive oxygen species (ROS) production and suppressed catabolic factor expression in chondrocytes in vitro. Moreover, in an OA mouse model induced by the destabilization of the medial meniscus (DMM), BBC0906 intra-articular injection attenuated cartilage degradation and alleviated OA. Importantly, BBC0906 selectively inhibits the BD2 domain, thus minimizing its potential side effects. We highlighted the therapeutic potential of targeting BET proteins to modulate oxidative stress and suppress cartilage degradation in OA. BBC0906 is a promising candidate for OA treatment, offering improved safety and efficacy.
Thomas X. Garcia; Martin M. Matzuk Mol. Reprod. Dev., 2024, 91(10), e70000 https://doi.org/10.1002/mrd.70000AbstractThe development of novel non-hormonal male contraceptives represents a pivotal frontier in reproductive health, driven by the need for safe, effective, and reversible contraceptive methods. This comprehensive review explores the genetic underpinnings of male fertility, emphasizing the crucial roles of specific genes and structural variants (SVs) identified through advanced sequencing technologies such as long-read sequencing (LRS). LRS has revolutionized the detection of structural variants and complex genomic regions, offering unprecedented precision and resolution over traditional next-generation sequencing (NGS). Key genetic targets, including those implicated in spermatogenesis and sperm motility, are highlighted, showcasing their potential as non-hormonal contraceptive targets. The review delves into the systematic identification and validation of male reproductive tract-specific genes, utilizing advanced transcriptomics and genomics studies with validation using novel knockout mouse models. We discuss the innovative application of small molecule inhibitors, developed through platforms like DNA-encoded chemistry technology (DEC-Tec), which have shown significant promise in preclinical models. Notable examples include inhibitors targeting serine/threonine kinase 33 (STK33), soluble adenylyl cyclase (sAC), cyclin-dependent kinase 2 (CDK2), and bromodomain testis associated (BRDT), each demonstrating nanomolar affinity and potential for reversible and specific inhibition of male fertility. This review also honors the contributions of Dr. David L. Garbers whose foundational work has paved the way for these advancements. The integration of genomic, proteomic, and chemical biology approaches, supported by interdisciplinary collaboration, is poised to transform male contraceptive development. Future perspectives emphasize the need for continued innovation and rigorous testing to bring these novel contraceptives from the laboratory to clinical application, promising a new era of male reproductive health management.
Yange Wei; Bo Wu; Mingqiu Liu; Chun-Ping Cui Int. J. Mol. Sci., 2024, 25(16), 8870 https://doi.org/10.3390/ijms25168870AbstractBone homeostasis relies on the delicate balance between osteoblast-mediated bone formation and osteoclast-mediated bone resorption. The casein kinase 2 interacting protein-1 (CKIP-1), a specific CK2α subunit-interacting protein, has been documented as one of the crucial negative regulators of bone formation. CKIP-1 siRNA therapy has constraints that limit its use in clinical applications. Therefore, it is necessary to explore effective targeting strategies for CKIP-1. In this study, we observed an upregulation of CKIP-1 protein expression in the microgravity environment, while its ubiquitination levels decreased. We further investigated the interaction between CKIP-1 and VHL and found that VHL enhanced CKIP-1 degradation through the ubiquitylation–proteasome system (UPS). Additionally, we discovered a small molecule ligand, named C77, through DNA-encoded library (DEL) screening, which binds to CKIP-1 both in vivo and in vitro, as confirmed by Surface Plasmon Resonance (SPR) and the Cellular Thermal shift assay (CETSA), respectively. Our findings demonstrated the potential of VHL and C77 as guiding factors in the development of CKIP-1-based Proteolysis-Targeting Chimeras (PROTACs), which could be future therapeutic interventions in disuse osteoporosis.
Shashiprabha Dubey; Zanjila Azeem; Pintu Kumar Mandal Adv. Synth. Catal., 2024 https://doi.org/10.1002/adsc.202400695AbstractAbstract. In synthetic carbohydrate chemistry, the modification of glycosyl radicals pathway stands as a central area of focus. The radical-based reactions often demonstrate remarkable compatibility with various functional groups owing to the mild initiation conditions. In particular, the identification of novel glycosyl radical precursors, combined with advanced reaction techniques, has substantially broadened the scope of glycosyl compound synthesis. Despite the presence of versatile donors, the synthesis of noble donors is still addressed as a synthetic need and challenges associated with sugar chemistry. Currently, a new class of glycosyl radical precursors has been developed which enables the production of C-, S-, O-, and N-glycosides efficiently. In this light, we highlight strategies towards bench-stable glycosyl sulfoxides, sulphone, and sulfite donors that can enable the site-, regio- and stereoselective transformation of protected or naked sugar synthons in synthetic carbohydrate chemistry. Here, this review article covers the recent developments in the selective glycosyl radical diversification such as glycosyl alkylation, arylation, alkenylation, sulfuration, C-H activation, and DNA conjugation via the bench-stable donors along with mechanistic aspects, challenges, and future directions. 1. Introduction 2. Generation of glycosyl radical from glycosyl sulfoxide, glycosyl sulfone, and glycosyl sulfinate 3. Glycosyl radicals and the stereoselectivity 4. Early history of glycosyl sulfoxides, sulfones, and sulfinate donors 5. Generation of glycosyl radicals from glycosyl sulfoxides 6. Generation of glycosyl radicals from glycosyl sulfones 6.1 Allyl glycosyl sulfones 6.2 Heteroaryl glycosyl sulfones 7. Generation of glycosyl radicals from glycosyl sulfinates 8. Summary and Outlook
Laura J. Byrnes; Won Young Choi; Paul Balbo; Mary Ellen Banker; Jeanne Chang; Shi Chen; Xuemin Cheng; Yang Cong; Jeff Culp; Hongxia Di; Matt Griffor; Justin Hall; Xiaoyun Meng; Barry Morgan; James J. Mousseau; Jennifer Nicki; Thomas O'Connell; Simeon Ramsey; Alex Shaginian; Suman Shanker; John Trujillo; Jinqiao Wan; Fabien Vincent; Stephen W. Wright; Felix Vajdos ACS Chem. Biol., 2024, 19(10), 2186–2197 https://doi.org/10.1021/acschembio.4c00397AbstractPeptidyl arginine deiminases (PADs) are important enzymes in many diseases, especially those involving inflammation and autoimmunity. Despite many years of effort, developing isoform-specific inhibitors has been a challenge. We describe herein the discovery of a potent, noncovalent PAD2 inhibitor, with selectivity over PAD3 and PAD4, from a DNA-encoded library. The biochemical and biophysical characterization of this inhibitor and two noninhibitory binders indicated a novel, Ca2+ competitive mechanism of inhibition. This was confirmed via X-ray crystallographic analysis. Finally, we demonstrate that this inhibitor selectively inhibits PAD2 in a cellular context.
Jens Carlsson; Andreas Luttens Curr. Opin. Chem. Biol., 2024, 87, 102829 https://doi.org/10.1016/j.sbi.2024.102829AbstractStructure-based virtual screening aims to find molecules forming favorable interactions with a biological macromolecule using computational models of complexes. The recent surge of commercially available chemical space provides the opportunity to search for ligands of therapeutic targets among billions of compounds. This review offers a compact overview of structure-based virtual screens of vast chemical spaces, highlighting successful applications in early drug discovery for therapeutically important targets such as G protein-coupled receptors and viral enzymes. Emphasis is placed on strategies to explore ultra-large chemical libraries and synergies with emerging machine learning techniques. The current opportunities and future challenges of virtual screening are discussed, indicating that this approach will play an important role in the next-generation drug discovery pipeline.
Huihong Wang; Teng Chen; Xiaohong Fan; Yangfeng Li; Wei Fang; Gong Zhang; Yizhou Li Chem. Commun., 2024 https://doi.org/10.1039/D4CC02547DAbstractThe versatile reactivity of isothiocyanate intermediates enabled the diversity-oriented synthesis (DOS) of N-heterocycles in a DNA-compatible manner. We first reported a mild in situ conversion of DNA-conjugated amines to isothiocyanates. Subsequently, a set of diverse transformations was successfully developed to construct 2-thioxo-quinazolinones, 1,2,4-thiadiazoles, and 2-imino thiazolines. Finally, the feasibility of these approaches in constructing DELs was further demonstrated through enzymatic ligation and mock pool preparation. This study demonstrated the advantages of combining in situ conversion strategies with DOS, which effectively broadened the chemical and structural diversity of DELs.
Jerrett Holdaway; Gunda I. Georg Science, 2024, 384(6698), 849-850 https://doi.org/10.1126/science.adp6432AbstractDespite the current availability of contraceptive methods, nearly half of all pregnancies worldwide are unintended (1), resulting in over 70 million abortions annually and imposing considerable emotional, physical, and financial burdens on individuals and health care systems. Hormonal-based approaches for male contraception are being investigated in clinical trials, but none have been approved (2). Nonhormonal targets present a promising avenue with potentially fewer side effects, but few are pursued for drug discovery. On page 885 of this issue, Ku et al. (3) report a drug screen of multibillion compounds encoded with DNA that enabled the discovery and development of a potent inhibitor of serine/threonine-protein kinase 33 (STK33). This testis-enriched kinase is indispensable for male fertility in humans and mice. The STK33 inhibitor CDD-2807 induced reversible male infertility without measurable toxicity in mice. These findings provide substantial support that targeted nonhormonal approaches can be effective in the development of a male contraceptive pill.
Angela F. Ku; Kiran L. Sharma; Hai Minh Ta; Courtney M. Sutton; Kurt M. Bohren; Yong Wang; Srinivas Chamakuri; Ruihong Chen; John M. Hakenjos; Ravikumar Jimmidi; Katarzyna Kent; Feng Li; Jian-Yuan Li; Lang Ma; Chandrashekhar Madasu; Murugesan Palaniappan; Stephen S. Palmer; Xuan Qin; Matthew B. Robers; Banumathi Sankaran; Zhi Tan; Yasmin M. Vasquez; Jian Wang; Jennifer Wilkinson; Zhifeng Yu; Qiuji Ye; Damian W. Young; Mingxing Teng; Choel Kim; Martin M. Matzuk Science, 2024, 384(6698), 885-890 https://doi.org/10.1126/science.adl2688AbstractMen or mice with homozygous serine/threonine kinase 33 (STK33) mutations are sterile owing to defective sperm morphology and motility. To chemically evaluate STK33 for male contraception with STK33-specific inhibitors, we screened our multibillion-compound collection of DNA-encoded chemical libraries, uncovered potent STK33-specific inhibitors, determined the STK33 kinase domain structure bound with a truncated hit CDD-2211, and generated an optimized hit CDD-2807 that demonstrates nanomolar cellular potency (half-maximal inhibitory concentration = 9.2 nanomolar) and favorable metabolic stability. In mice, CDD-2807 exhibited no toxicity, efficiently crossed the blood-testis barrier, did not accumulate in brain, and induced a reversible contraceptive effect that phenocopied genetic STK33 perturbations without altering testis size. Thus, STK33 is a chemically validated, nonhormonal contraceptive target, and CDD-2807 is an effective tool compound.
Sara Puglioli; Mosè Fabbri; Claudia Comacchio; Laura Alvigini; Roberto De Luca; Sebastian Oehler; Ettore Gilardoni; Gabriele Bassi; Samuele Cazzamalli; Dario Neri; Nicholas Favalli Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.4c00233AbstractDNA-Encoded Libraries (DELs) allow the parallel screening of millions of compounds for various applications, including de novo discovery or affinity maturation campaigns. However, library construction and HIT resynthesis can be cumbersome, especially when library members present an unknown stereochemistry. We introduce a permutational encoding strategy suitable for the construction of highly pure single-stranded single-pharmacophore DELs, designed to distinguish isomers at the sequencing level (e.g., stereoisomers, regio-isomers, and peptide sequences). This approach was validated by synthesizing a mock 921,600-member 4-amino-proline single-stranded DEL (“DEL1”). While screening DEL1 against different targets, high-throughput sequencing results showed selective enrichment of the most potent stereoisomers, with enrichment factors that outperform conventional encoding strategies. The versatility of our methodology was additionally validated by encoding 24 scaffolds derived from different permutations of the amino acid sequence of a previously described cyclic peptide targeting Fibroblast Activation Protein (FAP-2286). The resulting library (“DEL2”) was interrogated against human FAP, showing selective enrichment of five cyclic peptides. We observed a direct correlation between enrichment factors and on-DNA binding affinities. The presented encoding methodology accelerates drug discovery by facilitating library synthesis and streamlining HIT resynthesis while enhancing enrichment factors at the DEL sequencing level. This facilitates the identification of HIT candidates prior to medicinal chemistry and affinity maturation campaigns.
Hongtao Zhao; Eva Nittinger; Christian Tyrchan bioRxiv, 2024 https://doi.org/10.1101/2024.05.16.594622AbstractChemical space exploration has gained significant interest with the increase in available building blocks, which enables the creation of ultra-large virtual libraries containing billions or even trillions of compounds. However, the challenge of selecting most suitable compounds for synthesis arises, and one such challenge is hit expansion. Recently, Thompson sampling, a probabilistic search approach, has been proposed by Walters et al. to achieve efficiency gains by operating in the reagent space rather than the product space. Here, we aim to address some of its shortcomings and propose optimizations. We introduce a warmup routine to ensure that initial probabilities are set for all reagents with a minimum number of molecules evaluated. Additionally, a roulette wheel selection is proposed with adapted stop criteria to improve sampling efficiency, and belief distributions of reagents are only updated when they appear in new molecules. We demonstrate that a 100% recovery rate can be achieved by sampling 0.1% of the fully enumerated library, showcasing the effectiveness of our proposed optimizations.
Stephen O. Duke; Alyssa Twitty; Claire Baker; David Sands; Louis Boddy; María Lucía Travaini; Gustavo Sosa; Alexander L. A. Polidore; Amit J. Jhala; Jack M. Kloeber; Xavier Jacq; Lucas Lieber; Maria Celeste Varela; Martina Lazzaro; Ana P. Alessio; Christopher C. Ladner; Denis Fourches; Itai Bloch; Maayan Gal; Jonathan Gressel; Karthik Putta; Yael Phillip; Ifat Shub; Eyal Ben-Chanoch; Franck E. Dayan Weed Sci., 2024, 72, 444–464 https://doi.org/10.1017/wsc.2024.54AbstractDuring the past 30 yr an impasse has developed in the discovery and commercialization of synthetic herbicides with new molecular targets and novel chemistries. Similarly, there has been little success with bioherbicides, both microbial and chemical. These bioherbicides are needed to combat fast-growing herbicide resistance and to fulfill the need for more environmentally and toxicologically safe herbicides. In response to this substantial and growing opportunity, numerous start-up companies are utilizing novel approaches to provide new tools for weed management. These diverse new tools broaden the scope of discovery, encompassing advanced computational, bioinformatic, and imaging platforms; plant genome–editing and targeted protein degradation technologies; and machine learning and artificial intelligence (AI)-based strategies. This review contains summaries of the presentations of 10 such companies that took part in a symposium held at the WSSA annual meeting in 2024. Four of the companies are developing microbial bioherbicides or natural product–based herbicides, and the other six are using advanced technologies, such as AI, to accelerate the discovery of herbicides with novel molecular target sites or to develop non-GMO, herbicide-resistant crops.
Takashi Osawa; Ryosuke Kita; Yuuya Kasahara; Harumi Yamaguma; Taisuke Nakayama; Haruhiko Kamada; Satoshi Obika Bioorg. Med. Chem., 2024, 110, 117814 https://doi.org/10.1016/j.bmc.2024.117814AbstractOligonucleotide therapeutics, particularly antisense oligonucleotides (ASOs), have emerged as promising candidates in drug discovery. However, their effective delivery to the target tissues and cells remains a challenge, necessitating the development of suitable drug delivery technologies for ASOs to enable their practical application. In this study, we synthesized a library of chemically modified dipeptide-ASO conjugates using a recent synthetic method based on the Ugi reaction. We then conducted in vitro screening of this library using luciferase-expressing cell lines to identify ligands capable of enhancing ASO activity. Our findings suggest that N-(4-nitrophenoxycarbonyl)glycine may interact with the thiophosphate moiety of the phosphorothioate-modification in ASO. Through our screening efforts, we identified two ligands that modestly reduced luciferase luminescence in a cell type-selective manner. Furthermore, quantification of luciferase mRNA levels revealed that one of these promising dipeptide-ASO conjugates markedly suppressed luciferase RNA levels through its antisense effect in prostate-derived DU-145 cells compared to the ASOs without ligand modification.
Raghunath Bag; Malobika Kar; Nagendra K. Sharma Eur. J. Org. Chem., 2024, e202400513 https://doi.org/10.1002/ejoc.202400513AbstractAn overview of the recent advances on the selective C(sp2)- and C(sp3)−H chalcogenation of α-amino acids and peptides using various chalcogenides has been demonstrated. As a result, the evolved methodologies, encompassing transition-metal, metal-free, and photocatalytic protocols, exhibit superior efficiency compared to conventional chalcogenation methods. Natural α-amino acids are building blocks of biomacromolecules proteins and peptides, which are essential for living organisms. Recently, structurally modified amino acids and their peptides have received high demand for the development of various types of biomaterials, including therapeutic drug candidates. For example, the chalcogenated amino acids and their peptides serve as privileged structural scaffolds, derived from native amino acids. Given the widespread utility of sulfur/selenium-containing compounds in biological, pharmaceutical, and medicinal chemistry, efficient chalcogenation protocol has been accomplished through various catalytic methods such as transition-metal, metal-free, and photocatalytic strategies. Among them, transition-metal based strategies through C−H activation have been extensively explored for the chalcogenation of amino acids and peptides. Concurrently, metal-free and photocatalytic strategies offer sustainable and step-economically viable pathways for achieving successful chalcogenation. In this review, we have focused on the recent achievements of both C(sp2)−H and C(sp3)−H chalcogenation of α-amino acids and peptides with chalcogenides along with their substrate scopes, mechanisms, advantages, and limitations.
Xiaojie Lu; Hangchen Hu; Lianghe Mei; Xudong Wang; Liwei Xiong; Ying Zhu; Sixiu Liu; Wenfeng Zhao; Seydimemet Mengnisa; Linjie Li; Xian Lin; Jiaxiang Liu; Xuan Wang; Zhiqiang Duan; Weiwei Lu; Yanrui Suo; Xinyuan Wu; Mengqing Cui; Jinfeng Yue; Yechun Xu ChemRxiv, 2024 https://doi.org/10.1101/2024.05.08.593105AbstractThe global coronavirus disease 2019 (COVID-19) pandemic persists, with the ongoing mutation of the virus. Consequently, the development of inhibitors with diverse binding modes and mechanisms of action, along with the elucidation of novel binding sites is of paramount importance. The 3-chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro) are two validated cysteine proteases that cleave the viral polyprotein and are essential for viral replication. In this study, we utilized covalent DNA-Encoded libraries (CoDELs) workflow to identify two series of triazine-based covalent inhibitors targeting 3CLpro and PLpro. Molecular docking facilitated the identification of optimization pathways, further refined through medicinal chemistry efforts, leading to the development of the non-peptide 3CLpro inhibitor LU9, which exhibited an IC50 value of 0.34 μM, and crystal structure of LU10 revealed a unique binding mode within the active site. Additionally, the X-ray cocrystal structure of SARS-CoV-2 PLpro with XD5 uncovered a previously unexplored binding site, adjacent to the catalytic pocket, providing an opportunity for further development of PLpro inhibitors. Overall, these novel compounds serve as valuable chemical probes for target validation and represent promising drug candidates for the continued development of SARS-CoV-2 antivirals.
Marion Gras; Pauline Adler; Michael Smietana Chem., 2024, e202401069 https://doi.org/10.1002/chem.202401069AbstractPeptide-oligonucleotide conjugates (POCs) are covalent architectures composed of a DNA or RNA molecules linked to a peptide. These constructs have found widespread applications ranging from hybrid nanomaterials to gene-targeted therapies. Considering the important role of POCs, a new catalytic approach for their preparation is reported here, that could be applied either on solid support in anhydrous media, or post-synthetically in aqueous buffer. Single amino acids, peptides and cell penetrating peptides (CPPs) were conjugated to various oligo(ribo)nucleotides with high conversions and good isolated yields. The applicability of the method was demonstrated on more than 35 examples including an analogue of a commercial therapeutic oligonucleotide. Other conjugation partners, such as deoxycholic acid and biotin were also successfully conjugated to oligonucleotides. To highlight the potential of this catalytic approach, these conditions have been applied to iterative processes, which is of high interest for the development of DNA-Encoded Libraries.
Cristina Martini; Muhammad Idham Darussalam Mardjan; Andrea Basso Beilstein J. Org. Chem., 2024, 20, 1839-1879 https://doi.org/10.3762/bjoc.20.162AbstractThe Groebke–Blackburn–Bienaymé (GBB) three-component reaction, discovered in 1998, is a very efficient strategy to assemble imidazo[1,2-a]-heterocycles starting from amidines, aldehydes and isocyanides. This review aims to exhaustively describe innovative aspects of this reaction achieved during the last five years, and classifies them into five categories: synthetic methods, building blocks, scaffolds, biological activities and physical properties.
Stephanie Anne Robinson; Jennifer Anne Co; Steven Mark Banik Cell Chem. Biol., 2024 https://doi.org/10.1016/j.chembiol.2024.04.001AbstractSmall molecule molecular glues can nucleate protein complexes and rewire interactomes. Molecular glues are widely used as probes for understanding functional proximity at a systems level, and the potential to instigate event-driven pharmacology has motivated their application as therapeutics. Despite advantages such as cell permeability and the potential for low off-target activity, glues are still rare when compared to canonical inhibitors in therapeutic development. Their often simple structure and specific ability to reshape protein-protein interactions pose several challenges for widespread, designer applications. Molecular glue discovery and design campaigns can find inspiration from the fields of synthetic biology and biophysics to mine chemical libraries for glue-like molecules.
Yuhan Lv; Min Wu; Weiwei Lu; Meiying Cui; Jiaxiang Liu; Wei Yi; Xuan Wang; Xiaojie Lu Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.4c01553AbstractInspired by previous selection outcomes, we investigated and developed a rhodium-promoted C-H activation/annulation reaction of DNA-linked terminal alkynes and aromatic acids. This reaction exhibits excellent efficiency with high conversions and a broad substrate scope. Most importantly, the unique DEL-compatible conditions provide a better scenario for yielding an isocoumarin scaffold compared to conventional organic reaction conditions, and this newly developed on-DNA method has confirmed its feasibility in preparing DNA-encoded libraries.
Sophia You; Glen McIntyre; Toby Passioura Expert Opin. Drug Discov., 2024 https://doi.org/10.1080/17460441.2024.2367024AbstractCyclic peptides are an established class of pharmaceuticals, with the ability to bind to a broader range of protein targets than traditional small molecules while also being capable of oral availability and cell penetration. Historically, cyclic peptide drugs have been discovered almost exclusively through natural product mining approaches; however, the last two decades have seen the development of display screening approaches capable of rapidly identifying de novo (i.e. not natural product derived) cyclic peptide ligands to targets of interest. In this review, the authors describe the current clinical landscape for cyclic peptide pharmaceuticals. This article focuses on the discovery approaches that have led to the development of different classes of molecules and how the development of newer technologies, particularly phage and mRNA display, has broadened the clinical applicability of such molecules. The field of de novo cyclic peptide drug discovery is reaching maturity, with the first drugs identified through display screening approaches reaching the market in recent years. Many more are in clinical trials; however, significant technical challenges remain. Technological improvements will be required over the coming years to facilitate the identification of membrane permeable cyclic peptides capable of oral availability and targeting intracellular proteins.
Pinwen Cai; Basilius Sauter; Saule Zhanybekova; Dennis Gillingham ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-8v8kmAbstractAlthough using DNA encoded libraries (DELs) to find small molecule binders of target proteins is well-established, identifying DEL hits for functions other than binding remains challenging. We demonstrate here a technique where pools of DNA-linked small molecules are mixed with pools of DNA-linked protein targets and optimal small/protein pairs are identified based on their ability to catalyze the transfer of ubiquitin (Ub) onto the target proteins. Since the transfer of Ub is the first step in the tagging of proteins for proteasomal destruction, finding small molecules that can selectively reprogram Ub-transfer is one of the great challenges in contemporary drug development. Our work provides the framework for a new type of functional DEL screen that matches small molecule Ub-transfer catalysts with their optimal protein substrates. We believe the technology could be especially useful in discovering and optimizing molecular glue degraders.
Kshitij Rai; Yiduo Wang; Ronan W. O’Connell; Ankit B. Patel; Caleb J. Bashor Curr. Opin. Biomed. Eng., 2024, 100553 https://doi.org/10.1016/j.cobme.2024.100553AbstractEngineering synthetic regulatory circuits with precise input-output behavior—a central goal in synthetic biology—remains encumbered by the inherent molecular complexity of cells. Non-linear, high-dimensional interactions between genetic parts and host cell machinery make it difficult to design circuits using first principles biophysical models. We argue that adopting data-driven approaches that integrate modern machine learning (ML) tools and high-throughput experimental approaches into the synthetic biology design/build/test/learn process could dramatically accelerate the pace and scope of circuit design, yielding workflows that rapidly and systematically discern design principles and achieve quantitatively precise behavior. Current applications of ML to circuit design are occurring at three distinct scales: 1) learning relationships between part sequence and function; 2) determining how part composition determines circuit behavior; 3) understanding how function varies with genomic/host-cell context. This work points toward a future where ML-driven genetic design is used to program robust solutions to complex problems across diverse biotechnology domains.
Weijie You; Alba L. Montoya; Srikanta Dana; Raphael M. Franzini; Clemens Steegborn ChemMedChem, 2024, e202400273 https://doi.org/10.1002/cmdc.202400273AbstractSirtuin 6 (Sirt6), an NAD+-dependent deacylase, has emerged as a promising target for aging-related diseases and cancer. Advancing the medicinal chemistry of Sirt6 modulators is crucial for the development of chemical probes aimed at unraveling the intricate biological functions of Sirt6 and unlocking its therapeutic potential. A proprietary DNA-encoded library yielded Sirt6 inhibitor 2-Pr, displaying remarkable inhibitory activity and isoform-selectivity, and featuring a chemical structure distinct from reported Sirt6 modulators. In this study, we explore the inhibitory mechanism of 2-Pr, evaluating the impact of chemical modifications and presenting a crystal structure of the Sirt6/ADP-ribose/2-Pr complex. Notably, co-crystal structure analysis reveals an unexpected and unprecedented binding mode of Sirt6, with 2-Pr spanning the acyl channel of the enzyme, extending into the acetyl-lysine binding pocket, and reaching toward the C-site. This unique binding mode guides potential avenues for developing potent and selective Sirt6 inhibitors.
Pratik R. Chheda; Nicholas Simmons; Zhicai Shi Org. Lett., 2024, 26(20), 4365-4370 https://doi.org/10.1021/acs.orglett.4c01398AbstractDNA-encoded libraries (DELs) are a key technology for identifying small-molecule hits in both the pharmaceutical industry and academia, but their chemical diversity is largely limited to water-compatible reactions to aid in the solubility and integrity of encoding DNA tags. To broaden the DEL chemical space, we present a workflow utilizing DNA–cationic surfactant complexation that enables dissolution and reactions on-DNA in anhydrous organic solvents. We demonstrate its utility by developing DEL-compatible photoredox decarboxylative C(sp2)–C(sp3) coupling under water-free conditions. The workflow is optimized for the 96-well format necessary for large-scale DEL productions, and it enables screening and optimization of DEL-compatible reactions in organic solvents.
Michał Koziarski; Mohammed Abukalam; Vedant Shah; Louis Vaillancourt; Doris Alexandra Schuetz; Moksh Jain; Almer van der Sloot; Mathieu Bourgey; Anne Marinier; Yoshua Bengio arXiv, 2024, 2404.10094 https://doi.org/10.48550/arXiv.2404.10094AbstractDNA-encoded libraries (DELs) are a powerful approach for rapidly screening large numbers of diverse compounds. One of the key challenges in using DELs is library design, which involves choosing the building blocks that will be combinatorially combined to produce the final library. In this paper we consider the task of protein-protein interaction (PPI) biased DEL design. To this end, we evaluate several machine learning algorithms on the PPI modulation task and use them as a reward for the proposed GFlowNet-based generative approach. We additionally investigate the possibility of using structural information about building blocks to design a hierarchical action space for the GFlowNet. The observed results indicate that GFlowNets are a promising approach for generating diverse combinatorial library candidates.
Rupali J. S.; Vidya Madhuri E.; Gundreddy Raja Reddy; Voodikala S. Akhil; Ramya N.; Sagar D. J. Adv. Biol. Biotechnol., 2024, 27(7), 709-720 https://doi.org/10.9734/jabb/2024/v27i71030AbstractSpecies identification is essential for recognizing and describing biodiversity. Traditionally, this process has relied on morphological diagnosis through taxonomic studies, which have certain constraints such as subjectivity and time-consuming processes. With the advancement of modern molecular techniques, DNA barcoding has gained global attention. The term "DNA barcoding" refers to the technique of establishing species-level identification by sequencing a short fragment of the mitochondrial cytochrome c oxidase subunit I (COI) gene, the "DNA barcode," from a specimen that is taxonomically unknown and comparing it to a reference library of barcodes from known species. This review article explores the evolution of DNA barcoding, its universal marker, and its application in insect taxonomy, emphasizing its role in accelerating species discovery and biodiversity documentation. In India, DNA barcoding initiatives have made considerable progress, yet there remains a vast opportunity to barcode the country's rich insect diversity. Overall, DNA barcoding emerges as a powerful tool to address the urgent need for efficient species identification and biodiversity conservation in an ever-changing world.
Nannan Diao; Jianing Hou; Xinyu Peng; Yaru Wang; Axin He; Haiyan Gao; Linlin Yang; Pei Guo; Junyan Wang; Da Han Angew. Chem. Int. Ed. Engl., 2024, e202406330 https://doi.org/10.1002/anie.202406330AbstractAmplifying DNA conjugated affinity ligands can improve the sensitivity and multiplicity of cell imaging and play a crucial role in comprehensively deciphering cellular heterogeneity and dynamic changes during development and disease. However, the development of one-step, controllable, and quantitative DNA amplification methods for multiplexed imaging of live-cell membrane proteins is challenging. Here, we introduce the template adhesion reaction (TAR) method for assembling amplifiable DNA sequences with different affinity ligands, such as aptamers or antibodies, for amplified and multiplexed imaging of live-cell membrane proteins with high quantitative fidelity. The precisely controllable TAR enables proportional amplification of membrane protein targets with variable abundances by modulating the concentration ratios of hairpin templates and primers, thus allowing sensitive visualization of multiple membrane proteins with enhanced signal-to-noise ratios (SNRs) without disturbing their original ratios. Using TAR, we achieved signal-enhanced imaging of six proteins on the same live-cell within 1-2 h. TAR represents an innovative and programmable molecular toolkit for multiplexed profiling of membrane proteins in live-cells.
Xiaochen Tang; Wenxing Li; Tianshu Chen; Runchi Zhang; Yilin Yan; Chenbin Liu; Hongquan Gou; Fanping Zhang; Qiuhui Pan; Dongsheng Mao; Xiaoli Zhu Small Methods, 2024, 2400505 https://doi.org/10.1002/smtd.202400505Abstracto-DAExM is a versatile expansion and signal labeling method, showing signal amplification efficacy, enhancement of SNR, and expandable multiplexing capability, which reveals heterogeneous information...
Shea L. Johnson; Galen Missig; Minghua Wang; Kosalvisal Ouk; Kushali Gupta; Hanh Nho Nguyen; May Fern Toh; Tammy Szu-Yu Ho; David Gray; Hongjun Zhang; Yong Mi Choi-Sledeski; Claude Barberis; David J. Stone; Sokhom Pin; Jongwon Lim Bioorg. Med. Chem. Lett., 2024, 110, 129889 https://doi.org/10.1016/j.bmcl.2024.129889AbstractStudies have shown that disrupting the formation of the ligand-RET-GFRα complex could be an effective way of treating pain and itch. Compared to traditional high-throughput screens, DNA encoded libraries (DELs) have distinguished themselves as a powerful technology for hit identification in recent years. The present work demonstrates the use of DEL technology identifying compound 16 as the first GFRa2/GFRa3 small molecule inhibitor (0.1/0.2 μM respectively) selective over RET. This molecule represents an opportunity to advance the development of small-molecule inhibitors targeting the GFRα-RET interface for the treatment of pain and itch.
Stuart Schreiber ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-q1n61AbstractThis Perspective explores molecular glues and bifunctional compounds – proximity-inducing compounds – and offers a framework to understand and exploit their similarity to hot spots, missense mutations, and posttranslational modifications (PTMs).
Zhaomei Sun; Ying Zhong; Yahui Chen; Lingqian Xiao; Jiangying Wang; Fanming Zeng; Kexin Yang; Nicolas Duchemin; Yun Jin Hu Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.4c01138AbstractDNA-encoded library (DEL) technologies enable the fast exploration of gigantic chemical space to identify ligands for the target protein of interest and have become a powerful hit finding tool for drug discovery projects. However, amenable DEL chemistry is restricted to a handful of reactions, limiting the creativity of drug hunters. Here, we describe a new on-DNA synthetic pathway to access sulfides and sulfoximines. These moieties, usually contemplated as challenging to achieve through alkylation and oxidation, can now be leveraged in routine DEL selection campaigns.
Lewis Mervin; Alexey Voronov; Mikhail Kabeshov; Ola Engkvist ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-2rlk7-v2AbstractMachine-learning (ML) and Deep-Learning (DL) approaches to predict the molecular properties of small molecules are increasingly deployed within the design-make-test-analyse (DMTA) drug design cycle to predict molecular properties of interest. Despite this uptake, there are only a few automated packages to aid their development and deployment that also support uncertainty estimation, model explainability and other key aspects of model usage. This represents a key unmet need within the field and the large number of molecular representations and algorithms (and associated parameters) means it is non-trivial to robustly optimise, evaluate, reproduce, and deploy models. Here we present QSARtuna, a molecule property prediction modelling pipeline, written in Python and utilising the Optuna, Scikit-learn, RDKit and ChemProp packages, which enables the efficient and automated comparison between molecular representations and machine learning models. The platform was developed considering the increasingly important aspect of model uncertainty quantification and explainability by design. We provide details for our framework and provide illustrative examples to demonstrate the capability of the software when applied to simple molecular property, reaction/reactivity prediction and DNA encoded library enrichment analyses. We hope that the release of QSARtuna will further spur innovation in automatic ML modelling and provide a platform for education of best practises in molecular property modelling. The code to the Qptuna framework is made freely available via GitHub.
Anrong Chen; Yang Han; Rongfeng Wu; Bo Yang; Lijuan Zhu; Feng Zhu Nat. Commun., 2024, 15, 5228 https://doi.org/10.1038/s41467-024-49547-9AbstractC-1 Glycals serve as pivotal intermediates in synthesizing diverse C-glycosyl compounds and natural products, necessitating the development of concise, efficient and user-friendly methods to obtain C-1 glycosides is essential. The Suzuki-Miyaura cross-coupling of glycal boronates is notable for its reliability and non-toxic nature, but glycal donor stability remains a challenge. Herein, we achieve a significant breakthrough by developing stable glycal boronates, effectively overcoming the stability issue in glycal-based Suzuki-Miyaura coupling. Leveraging the balanced reactivity and stability of our glycal boronates, we establish a robust palladium-catalyzed glycal-based Suzuki-Miyaura reaction, facilitating the formation of various C(sp2)-C(sp), C(sp2)-C(sp2), and C(sp2)-C(sp3) bonds under mild conditions. Notably, we expand upon this achievement by developing the DNA-compatible glycal-based cross-coupling reaction to synthesize various glycal-DNA conjugates. With its excellent reaction reactivity, stability, generality, and ease of handling, the method holds promise for widespread appication in the preparation of C-glycosyl compounds and natural products.
Chang Liu; Hui Zhang Expert Opin. Drug Discov., 2024 https://doi.org/10.1080/17460441.2024.2354871AbstractHigh-throughput mass spectrometry that could deliver > 10 times faster sample readout speed than traditional LC-based platforms has emerged as a powerful analytical technique, enabling the rapid analysis of complex biological samples. This increased speed of MS data acquisition has brought a critical demand for automatic data processing capabilities that should match or surpass the speed of data acquisition. Those data processing capabilities should serve the different requirements of drug discovery workflows. This paper introduced the key steps of the automatic data processing workflows for high-throughput MS technologies. Specific examples and requirements are detailed for different drug discovery applications. The demand for automatic data processing in high-throughput mass spectrometry is driven by the need to keep pace with the accelerated speed of data acquisition. The seamless integration of processing capabilities with LIMS, efficient data review mechanisms, and the exploration of future features such as real-time feedback, automatic method optimization, and AI model training is crucial for advancing the drug discovery field. As technology continues to evolve, the synergy between high-throughput mass spectrometry and intelligent data processing will undoubtedly play a pivotal role in shaping the future of high-throughput drug discovery applications.
Yuhan Lv; Min Wu; Weiwei Lu; Meiying Cui; Jiaxiang Liu; Wei Yi; Xuan Wang; Xiaojie Lu ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-xw963AbstractAs discovered by the previous selection outcomes, we developed a Rhodium-promoted C-H activation/ annulation reaction of DNA-linked terminal alkyne and aromatic acid. This reaction exhibits excellent efficiency with high conversions and a wide broad sub-strate scope. Most importantly, the unique DEL-compatible condition provides a better scenario to yield an isocoumarin scaffold compared to conventional organic reaction condition, and this newly developed on-DNA method has confirmed its feasibility in preparing DNA-encoded libraries.
Catherine A. Hurd; Jacob T. Bush; Andrew J. Powell; Louise J. Walport bioRxiv, 2024 https://doi.org/10.1101/2024.03.20.585962AbstractmRNA display is a powerful technology to screen libraries of >1012 cyclic peptides against a protein target, enabling the rapid discovery of high affinity ligands. These cyclic peptides are particularly well suited to challenging protein targets that have been difficult to drug with small molecules. However, target choice can still be limited as screens are typically performed against purified proteins which often demands the use of isolated domains and precludes the use of aggregation-prone targets. Here, we report a method to perform mRNA display selections in mammalian cell lysates without the need for prior target purification, vastly expanding the potential target scope of mRNA display. We have applied the methodology to identify low to sub-nanomolar peptide binders for two targets; a NanoLuc subunit (LgBiT) and full-length bromodomain-containing protein 3 (BRD3). Our cyclic peptides for BRD3 were found to bind to the extraterminal (ET) domain of BRD3 and the closely related BRD proteins, BRD2 and BRD4. While many chemical probes exist for the bromodomains of BRD proteins, the ET domain is relatively underexplored, making these peptides valuable additions to the BRD toolbox.
Dongliang Guan; Jiaxiang Liu; Feifei Chen; Jian Li; Xiaowen Wang; Weiwei Lu; Yanrui Suo; Feng Tang; Lefu Lan; Xiaojie Lu; Wei Huang J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.4c00607AbstractPage 3784. An inadvertent error during the proofreading process resulted in the unintentional deletion of activity data for compounds listed in Table 1. The complete Table 1 is presented here.
Lewis Mervin; Alexey Voronov; Mikhail Kabeshov; Ola Engkvist J. Chem. Inf. Model., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jcim.4c00457AbstractMachine-learning (ML) and deep-learning (DL) approaches to predict the molecular properties of small molecules are increasingly deployed within the design–make–test–analyze (DMTA) drug design cycle to predict molecular properties of interest. Despite this uptake, there are only a few automated packages to aid their development and deployment that also support uncertainty estimation, model explainability, and other key aspects of model usage. This represents a key unmet need within the field, and the large number of molecular representations and algorithms (and associated parameters) means it is nontrivial to robustly optimize, evaluate, reproduce, and deploy models. Here, we present QSARtuna, a molecule property prediction modeling pipeline, written in Python and utilizing the Optuna, Scikit-learn, RDKit, and ChemProp packages, which enables the efficient and automated comparison between molecular representations and machine learning models. The platform was developed by considering the increasingly important aspect of model uncertainty quantification and explainability by design. We provide details for our framework and provide illustrative examples to demonstrate the capability of the software when applied to simple molecular property, reaction/reactivity prediction, and DNA encoded library enrichment classification. We hope that the release of QSARtuna will further spur innovation in automatic ML modeling and provide a platform for education of best practices in molecular property modeling. The code for the QSARtuna framework is made freely available via GitHub.
Shuai Han; Xinyun Guo; Min Wang; Huan Liu; Yidan Song; Yunyun He; Kuang-Lung Hsueh; Weiren Cui; Wenji Su; Letian Kuai; Jason Deng ACS Med. Chem. Lett., 2024, 15(9), 1456–1466 https://doi.org/10.1021/acsmedchemlett.4c00121AbstractDNA-encoded library (DEL) technology, especially when combined with machine learning (ML), is a powerful method to discover novel inhibitors. DEL-ML can expand a larger chemical space and boost cost-effectiveness during hit finding. Heme oxygenase-1 (HO-1), a heme-degrading enzyme, is linked to diseases such as cancer and neurodegenerative disorders. The discovery of five series of new scaffold HO-1 hits is reported here, using a DEL-ML workflow, which emphasizes the model's uncertainty quantification and domain of applicability. This model exhibits a strong extrapolation ability, identifying new structures beyond the DEL chemical space. About 37% of predicted molecules showed a binding affinity of K D < 20 μM, with the strongest being 141 nM, amd 14 of those molecules displayed >100-fold selectivity for HO-1 over heme oxygenase-2 (HO-2). These molecules also showed structural novelty compared to existing HO-1 inhibitors. Docking simulations provided insights into possible selectivity rationale.
Kelly A. McCarthy; Douglas J. Marcotte; Sangram Parelkar; Crystal L. McKinnon; Lindsay E. Trammell; Eric L. Stangeland; Rachael R. Jetson ChemMedChem, 2024, e202400093 https://doi.org/10.1002/cmdc.202400093AbstractInhibition of poly (ADP-ribose) polymerase-1 (PARP1), a DNA repair enzyme, has proven to be a successful strategy for the treatment of various cancers. With the appropriate selection conditions and protein design, DNA-encoded library (DEL) technology provides a powerful avenue to identify small molecules with the desired mechanism of action towards a target of interest. However, DNA-binding proteins, such as PARP1, can be challenging targets for DEL screening due to non-specific protein-DNA interactions. To overcome this, we designed and screened a PARP1 catalytic domain construct without the autoinhibitory helical domain. This allowed us to interrogate an active, functionally-relevant form of the protein resulting in the discovery of novel isoindolinone PARP1 inhibitors with single-digit nanomolar potency. These inhibitors also demonstrated little to no PARP1-DNA trapping, a property that could be advantageous in the clinic.
Rick A. Homan; John D. Lapek; Christina M. Woo; Sherry Niessen; Lyn H. Jones; Christopher G. Parker Nat. Rev. Methods Primers, 2024, 4, 30 https://doi.org/10.1038/s43586-024-00308-4AbstractSmall molecules can serve as leads for new therapeutics as well as powerful tools to investigate biological processes. Understanding the interactions of these molecules, particularly in native biological environments, is fundamentally critical to their utility. Photoaffinity labelling (PAL) represents one of the few strategies that enable the direct mapping of interactions of small molecules with proteins. PAL uses latent functional groups that form reactive intermediates only upon exposure to light of specific wavelengths that, subsequently, covalently adduct to proximal biomolecules, allowing for their enrichment and identification. Although the original applications of PAL date to six decades ago, the more recent integration with powerful mass spectrometry-based proteomic methods has profoundly impacted the ability to illuminate molecular interactions on a global scale. In this Primer, we discuss the current state-of-the-art of PAL-based strategies for studying molecular interactions in native systems, with a focus on investigations of small molecule–protein interactions. We cover topics including the basic principles of PAL chemistries, PAL probe design, experimental considerations, data analysis and applications of PAL illustrated by recent examples. Finally, we provide our perspective on persistent challenges and our outlook on the field.
Yuhan Lv; Baiyang Mu; Shaozhao Qin; Xinyuan Wu; Weiwei Lu; Meiying Cui; Jiaxiang Liu; Xuan Wang; Xiaojie Lu Tetrahedron Lett., 2024, 142, 155076 https://doi.org/10.1016/j.tetlet.2024.155076AbstractEmploying pre-existing functionalized scaffolds to generate focused libraries exemplifies an expedited approach in terms of synthetic feasibility and efficiency. This strategic synthesis is particularly well-suited for scaffolds that are unattainable with DNA-compatible chemical tools. This manuscript utilizes pyridazinone as a paradigmatic scaffold to elucidate the synthetic workflow. Three functionalized pyridazinone reagents were obtained by conventional organic chemistries and then applied to prepare corresponding DNA-encoded libraries (DELs) by robust DNA-compatible reactions and abundant building blocks (BBs), ensuring both diversity and quality in the final libraries. The feasibility of these focused DELs were confirmed by subsequent affinity selection and off-DNA validation against the BRD4 target.
Javiera Baeza; Pablo Cruz; Paola Ojeda; Francisco Adasme-Carreño; Mauricio Bedoya; Oscar Cerda; Wendy González ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-6pdxrAbstractProtein-protein interactions (PPIs) regulate crucial physiological and pathological processes. PPIs are considered a class of biological targets almost infeasible for small molecules because the binding surfaces are usually large and shallow. Peptides are molecules able to bind to these drug targets; they can be used as modulators and mimic one of the interaction partners. This review details the advances in in silico peptide design and experimental approaches for the evaluation of PPI-based peptides.
Shuai Han; Xinyun Guo; Min Wang; Huan Liu; Yidan Song; Yunyun He; Kuang-Lung Hsueh; Weiren Cui; Wenji Su; Letian Kuai; Jason Deng ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-k5v0sAbstractDNA-encoded library (DEL) technology has proven to be a powerful method for discovering novel inhibitors for diverse targets. Particularly when combined with machine learning (ML), the DEL-ML workflow expands the chemical space and enhances cost-effectiveness, offering new opportunities to find desired hit molecules. Heme oxygenase-1 (HO-1), primarily a heme-degrading enzyme, has been identified as a potential therapeutic target in diseases such as cancer and neurodegenerative disorders. Despite years of study, the HO-1 inhibitor toolbox remains limited. Here, we report the discovery of five series of novel scaffold HO-1 inhibitors using a DEL-ML workflow that emphasizes the model’s uncertainty quantification and its domain of applicability. The DEL-ML model demonstrated a strong ability to extrapolate to novel chemical spaces by identifying new structures. Approximately 33% of the predicted molecules, validated by biophysical assays, had a binding affinity of K¬D < 15 µM, with the strongest affinity being 141 nM. Fourteen tested molecules showed over 100-fold selectivity towards HO-1 over Heme oxygenase-2 (HO-2). These molecules are also structurally novel compared to the reported HO-1 inhibitors. Further, binding mode simulations via docking provided insights into the possible selectivity rationale of some selective series.
Simon C. C. Lucas; J. Henry Blackwell; Ulf Börjesson; David Hargreaves; Alexander G. Milbradt; Samiyah Ahmed; Mark J. Bostock; Carine Guerot; Andrea Gohlke; Olaf Kinzel; Michelle L. Lamb; Nidhal Selmi; Christopher J. Stubbs; Nancy Su; Qibin Su; Haiou Luo; Ting Xiong; Xiaoqian Zuo; Sana Bazzaz; Corey Bienstock; Paolo A. Centrella; Kyle E. Denton; Diana Gikunju; Marie-Aude Guié; John P. Guilinger; Christopher Hupp; Anthony D. Keefe; Takashi Satoh; Ying Zhang; Emma L. Rivers ACS Med. Chem. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsmedchemlett.4c00113AbstractBfl-1 is overexpressed in both hematological and solid tumors; therefore, inhibitors of Bfl-1 are highly desirable. A DNA-encoded chemical library (DEL) screen against Bfl-1 identified the first known reversible covalent small-molecule ligand for Bfl-1. The binding was validated through biophysical and biochemical techniques, which confirmed the reversible covalent mechanism of action and pointed to binding through Cys55. This represented the first identification of a cyano-acrylamide reversible covalent compound from a DEL screen and highlights further opportunities for covalent drug discovery through DEL screening. A 10-fold improvement in potency was achieved through a systematic SAR exploration of the hit. The more potent analogue compound 13 was successfully cocrystallized in Bfl-1, revealing the binding mode and providing further evidence of a covalent interaction with Cys55.
Yiqing Zhang; Suzanne Blum ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-zjhtgAbstractPerforming organic reactions in water provides environmental and toxicity advantages compared to in organic solvents, but such strategies are still evolving, limited by current understanding of these multiphasic systems. Here, a fluorescence lifetime imaging microscopy (FLIM) technique characterizes surfactant-dependent partitioning of organics in a system that mimics the early, zinc-related steps of a Negishi-like cross-coupling reaction in water, under synthetically relevant high concentrations of substrate (0.18 M), where large emulsion droplets (5–20 um) form an organic phase. Brij-30 resulted in the highest degree of partitioning of organics in the aqueous phase, which was not predictable from simple hydrophilic–lipophilic balances (HLB). The ionic surfactant CTAC suppressed reactivity of the metallic zinc surface with both neutral (TMEDA) and anionic (carboxylate) reactants, presumably through competitive chloride binding and concurrent cetrimonium coating, a finding that may contribute to the reduced performance of ionic surfactant in the bench-scale coupling reaction.
Suzanne Ackloo; Fengling Li; Magda Szewczyk; Almagul Seitova; Peter Loppnau; Hong Zeng; Jin Xu; Shabbir Ahmad; Yelena A Arnautova; AJ Baghaie; Serap Beldar; Albina Bolotokova; Paolo A Centrella; Irene Chau; Matthew A Clark; John W Cuozzo; Saba Dehghani-Tafti; Jeremy S Disch; Aiping Dong; Antoine Dumas; Jianwen A. Feng; Pegah Ghiabi; Elisa Gibson; Justin Gilmer; Brian Goldman; Stuart R Green; Marie-Aude Guié; John P Guilinger; Nathan Harms; Oleksandra Herasymenko; Scott Houliston; Ashley Hutchinson; Steven Kearnes; Anthony D Keefe; Serah W Kimani; Trevor Kramer; Maria Kutera; Haejin A Kwak; Cristina Lento; Yanjun Li; Jenny Liu; Joachim Loup; Raquel AC Machado; Christopher J Mulhern; Sumera Perveen; Germanna L Righetto; Patrick Riley; Suman Shrestha; Eric A Sigel; Madhushika Silva; Michael D. Sintchak; Belinda L Slakman; Rhys D Taylor; James Thompson; Wen Torng; Carl Underkoffler; Moritz von Rechenberg; Ian Watson; Derek J Wilson; Esther Wolf; Manisha Yadav; Aliakbar K Yazdi; Junyi Zhang; Ying Zhang; Vijayaratnam Santhakumar; Aled M Edwards; Dalia Barsyte-Lovejoy; Matthieu Schapira; Peter J Brown; Levon Halabelian; Cheryl H Arrowsmith bioRxiv, 2024 https://doi.org/10.1101/2024.03.03.583197AbstractProtein class-focused drug discovery has a long and successful history in pharmaceutical research, yet most members of druggable protein families remain unliganded, often for practical reasons. Here we combined experiment and computation to enable discovery of ligands for WD40 repeat (WDR) proteins, one of the largest human protein families. This resource includes expression clones, purification protocols, and a comprehensive assessment of the druggability for hundreds of WDR proteins. We solved 21 high resolution crystal structures, and have made available a suite of biophysical, biochemical, and cellular assays to facilitate the discovery and characterization of small molecule ligands. To this end, we use the resource in a hit-finding pilot involving DNA-encoded library (DEL) selection followed by machine learning (ML). This led to the discovery of first-in-class, drug-like ligands for 9 of 20 targets. This result demonstrates the broad ligandability of WDRs. This extensive resource of reagents and knowledge will enable further discovery of chemical tools and potential therapeutics for this important class of proteins.Competing Interest StatementBLS, BG, JSD, JZ, JWC, MvR, PR, SK, and TK are employees and shareholders of Relay Therapeutics.BLIBiolayer InterferometryDELDNA Encoded LibraryDLIDdrug-like densityDSFDifferential Scanning FluorimetryFPFluorescence PolarizationGCNNGraph Convolutional Neural NetworkHDXHydrogen-Deuterium eXchangeHTSHigh-Throughput ScreeningMLMachine LearningNanoBRETNanoLuciferase Bioluminescence Resonance Energy TransferPPIProtein-Protein InteractionPROTACProteolysis Targeting ChimeraSPRSurface Plasmon ResonanceTmProtein melting temperatureWDRTryptophan-Aspartate Repeat
Guy Durant; Fergus Boyles; Kristian Birchall; Charlotte M. Deane Nat. Comput. Sci., 2024, 4, 735–743 https://doi.org/10.1038/s43588-024-00699-0AbstractMany studies have prophesied that the integration of machine learning techniques into small-molecule therapeutics development will help to deliver a true leap forward in drug discovery. However, increasingly advanced algorithms and novel architectures have not always yielded substantial improvements in results. In this Perspective, we propose that a greater focus on the data for training and benchmarking these models is more likely to drive future improvement, and explore avenues for future research and strategies to address these data challenges.
Xing Ling; Sixiu Liu; Yixuan Yang; Qian Dong; Lisa A. Marcaurelle; Wei Huang; Yun Ding; Xuan Wang; Xiaojie Lu Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.4c00085AbstractWe correct the structures of trehalose and GalNAc in Scheme 1, the structure of DEL-B in Scheme 4, and additional information in the Acknowledgment. A revised Supporting Information file with corrected captions and corrected structures.
Hongtao Zhao ACS Med. Chem. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsmedchemlett.4c00093AbstractStructure-based virtual screening has gained momentum again as the high attrition rate at every stage of drug discovery drives the need to explore a greater chemical space. From the Bayesian perspective, its shortcomings as a viable strategy for sustainable hit discovery are discussed, with regard to the prior hit rates of screening libraries and the performance of computational methods. Lessons are shared in selecting virtual hits for experimental validation learned from a series of eight successful campaigns, one of which impacted the discovery of a drug candidate currently in clinical trials.
Yuhan Gui; Rui Hou; Yuchen Huang; Yu Zhou; Shihao Liu; Ling Meng; Ying Li; Fong Sang Lam; Clara Shania Wong; Chan Tat Yin; Gang Li; Xiaojie Lu; Xiaoyu Li ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-3mrz3AbstractSmall molecule ligands that can specifically recognize the surface of cancer cells have wide utilities in cancer diagnosis and treatment. Screening large combinatorial libraries against live cells is an effective approach to discover cell-targeting ligands. In the past decade, DNA-encoded chemical library (DEL or DECL) has become a powerful technology in drug discovery and been successfully used in ligand discovery against numerous biological targets. However, nearly all DEL selections had predefined targets, whereas completely unbiased DEL selections interrogating the entire cell surface remain underexplored. In this report, we systematically optimized cell-based DEL selection method to perform unbiased selections against cancer cells without predefined targets. A 104.96-million-member DEL was selected against MDA-MB-231 and MCF7, a pair of breast cancer cell lines with high and low metastatic properties, respectively, and cell-specific small molecule ligands and ligand combinations (“clusters”) have been identified. We further show that the ligand cluster could be optimized to improve the binding affinity and applied in cell-targeting applications including cancer photodynamic therapy and targeted drug delivery. Finally, we leveraged the DNA tag of the DEL compounds and identified the cell surface receptor of an individual ligand targeting MDA-MB-231 cells. Overall, this work provides an efficient method for discovering cell-targeting small molecules and demonstrated the potential of DELs as a tool for cancer biomarker discovery.
Hugo Belda; David Bradley; Evangelos Christodoulou; Stephanie D. Nofal; Malgorzata Broncel; David Jones; Heledd Davies; M. Teresa Bertran; Andrew G. Purkiss; Roksana W. Ogrodowicz; Dhira Joshi; Nicola O’Reilly; Louise Walport; Antoine Claessens; Andrew Powell; David House; Svend Kjaer; Christian R. Landry; Moritz Treeck bioRxiv, 2024 https://doi.org/10.1101/2024.02.22.581535AbstractAmong the ∼200 Plasmodium species that infect vertebrates, six infect humans. Of these, P. falciparum causes >95% of all ∼500,000 annual fatalities. Phylogenetically, P. falciparum belongs to the Laverania subgenus, a group of Plasmodium species that infect great apes. Common to Laverania species is the family of FIKK kinases. One million years ago, a single FIKK kinase conserved in all Plasmodium species gained an export element in the Laverania subgenus and expanded into the family of ∼20 atypical FIKK kinases, most of which are exported into the host cell. The fikk genes are conserved in syntenic loci across the Laverania, arguing for a rapid expansion controlling important functions in host cell remodelling and pathogenesis. We provide evidence that the FIKK paralogues evolved specific and mutually exclusive phosphorylation motif preferences, conserved across their Laverania orthologues, in a short evolutionary timeframe. Surprisingly, we find that FIKK13 has evolved exclusive tyrosine-phosphorylation preference, which was thought to be absent in Plasmodium species. Combining a crystal structure with AlphaFold2 predictions, we identify residues that determine kinase-specificity within the FIKK family in a fast-evolving flexible loop. Finally, we show that all expressed members of the FIKK kinase family can be chemically inhibited in vitro using a single compound. Such a pan-specific inhibitor of this kinase family important for virulence could reduce the ability of the parasite to gain escape-mutations and resistance.
An Le; Eden Teclemichael; Svetlana Krylova; Sergey Krylov ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-rt7bsAbstractHigh stringency (e.g., low target concentration) is key for obtaining high-affinity aptamers in SELEX; however, excessive stringency greatly increases the probability of SELEX failure. The control of stringency in SELEX remains a technical art based solely on intuition. A major reason for this is the lack of a measure of stringency. Here we introduce the Binder-to-Nonbinder Ratio at the output of partitioning (BNR), a parameter that characterizes stringency quantitatively: increasing stringency leads to decreasing BNR. BNR is determined experimentally by simply measuring by qPCR the quantities of oligonucleotides after partitioning in the presence and absence of target. The theory suggests, and our SELEX experiments with two targets confirm, that BNR must be kept statistically significantly greater than zero to avoid SELEX failure due to excessive stringency. Using BNR will help experimenters to rationalize the choice of conditions which the stringency depends on, e.g., target concentration and time of partitioning.
Maëva Pichon; Marcel Hollenstein Commun. Chem., 2024, 7, 138 https://doi.org/10.1038/s42004-024-01216-0AbstractOligonucleotides are advancing as essential materials for the development of new therapeutics, artificial genes, or in storage of information applications. Hitherto, our capacity to write (i.e., synthesize) oligonucleotides is not as efficient as that to read (i.e., sequencing) DNA/RNA. Alternative, biocatalytic methods for the de novo synthesis of natural or modified oligonucleotides are in dire need to circumvent the limitations of traditional synthetic approaches. This Perspective article summarizes recent progress made in controlled enzymatic synthesis, where temporary blocked nucleotides are incorporated into immobilized primers by polymerases. While robust protocols have been established for DNA, RNA or XNA synthesis is more challenging. Nevertheless, using a suitable combination of protected nucleotides and polymerase has shown promises to produce RNA oligonucleotides even though the production of long DNA/RNA/XNA sequences (>1000 nt) remains challenging. We surmise that merging ligase- and polymerase-based synthesis would help to circumvent the current shortcomings of controlled enzymatic synthesis.
Mirela Delibegović; Sergio Dall’Angelo; Ruta Dekeryte Nat. Rev. Endocrinol., 2024 https://doi.org/10.1038/s41574-024-00965-1AbstractProtein tyrosine phosphatase 1B (PTP1B), a non-transmembrane phosphatase, has a major role in a variety of signalling pathways, including direct negative regulation of classic insulin and leptin signalling pathways, and is implicated in the pathogenesis of several cardiometabolic diseases and cancers. As such, PTP1B has been a therapeutic target for over two decades, with PTP1B inhibitors identified either from natural sources or developed throughout the years. Some of these inhibitors have reached phase I and/or II clinical trials in humans for the treatment of type 2 diabetes mellitus, obesity and/or metastatic breast cancer. In this Review, we summarize the cellular processes and regulation of PTP1B, discuss evidence from in vivo preclinical and human studies of the association between PTP1B and different disorders, and discuss outcomes of clinical trials. We outline challenges associated with the targeting of this phosphatase (which was, until the past few years, viewed as difficult to target), the current state of the field of PTP1B inhibitors (and dual phosphatase inhibitors) and future directions for manipulating the activity of this key metabolic enzyme.
Yusong Ye; Madeline Berry; William J. Bock; Kunpeng Cheng; Sajiv K. Nair; Christiana S. Park; Ryan L. Patman; Sylvie Sakata; Michelle Tran-Dubé; Joyann S. Donaldson; Guanyu Yang; Guansai Liu Org. Lett. 2024, 26, 16, 3338–3342 https://doi.org/10.1021/acs.orglett.4c00604AbstractIsoquinolone is one of the most common heterocyclic core structures in countless natural products and many bioactive compounds. Here, a highly efficient approach to synthesize isoquinolone scaffolds on DNA via rhodium(III)-catalyzed C–H activation has been described. This chemistry transformation is robust and has shown good compatibility with DNA, which is suitable for DNA-encoded library synthesis.
Lewis Mervin; Alexey Voronov; Mikhail Kabeshov; Ola Engkvist ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-2rlk7AbstractMachine-learning (ML) and Deep-Learning (DL) approaches to predict the molecular properties of small molecules are increasingly deployed within the design-make-test-analyse (DMTA) drug design cycle to predict molecular properties of interest. Despite this uptake, there are only a few automated packages to aid their development and deployment that also support uncertainty estimation, model explainability and other key aspects of model usage. This represents a key unmet need within the field and the large number of molecular representations and algorithms (and associated parameters) means it is non-trivial to robustly optimise, evaluate, reproduce, and deploy models. Here we present Qptuna, a molecule property prediction modelling pipeline, written in Python and utilising the Optuna, Scikit-learn, RDKit and ChemProp packages, which enables the efficient and automated comparison between molecular representations and machine learning models. The platform was developed considering the increasingly important aspect of model uncertainty quantification and explainability by design. We provide details for our framework and provide illustrative examples to demonstrate the capability of the software when applied to simple molecular property, reaction/reactivity prediction and DNA encoded library enrichment analyses. We hope that the release of Qptuna will further spur innovation in automatic ML modelling and provide a platform for education of best practises in molecular property modelling. The code to the Qptuna framework is made freely available via GitHub.
Cen Huang; Xingyuan Liu; Xinyuan Wu; Ling Meng; Xiaojie Lu; Xiaoyu Li ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-rmgtvAbstractWe report a selection method of DNA-encoded libraries (DELs) that can identify the compounds able to induce ubiquitination of the protein of intertest (POI). Since the selection readout of the method is based on POI ubiquitination, rather than POI binding or ternary complex formation, the identified compounds are more functionally relevant and more predicative of active protein degraders. In this study, by selecting a DEL of 950 different combinations of POI ligands, linkers, and E3 ligase ligands against the BD1 domain of bromodomain-containing protein 4 (BRD4-BD1) in the presence and absence of ATP, we have identified a potent BRD4-BD1 degrader (DC50: ~9.7 nM) and also a short-isoform-selective BRD4 degrader (DC50: 0.26 μM). Furthermore, we show that the selection based on POI binding or ternary complex formation identified compounds that may have induced stable complexes but are inactive degraders. This approach may be an efficient and broadly applicable method for discovering functional protein degraders, as well as E3 ligase ligands, by harnessing the vast chemical diversity of DELs.
Yunyuan Huang; Yanxi Li; Jinrong Min Trends Pharmacol. Sci., 2024 https://doi.org/10.1016/j.tips.2024.01.007AbstractThe PWWP domain binds to both histone and DNA of a nucleosome in a bivalent way. PWWP domain-containing proteins are involved in different biological processes, and their aberrant expression is implicated in various human diseases. Here, we discuss the recent developments and challenges in targeting the PWWP domain for therapeutic intervention.
Patrick R. Fitzgerald; Anjali Dixit; Chris Zhang; David L. Mobley; Brian M. Paegel J. Chem. Inf. Model., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jcim.4c00232AbstractDNA-encoded library technology grants access to nearly infinite opportunities to explore the chemical structure space for drug discovery. Successful navigation depends on the design and synthesis of libraries with appropriate physicochemical properties (PCPs) and structural diversity while aligning with practical considerations. To this end, we analyze combinatorial library design constraints including the number of chemistry cycles, bond construction strategies, and building block (BB) class selection in pursuit of ideal library designs. We compare two-cycle library designs (amino acid + carboxylic acid, primary amine + carboxylic acid) in the context of PCPs and chemical space coverage, given different BB selection strategies and constraints. We find that broad availability of amines and acids is essential for enabling the widest exploration of chemical space. Surprisingly, cost is not a driving factor, and virtually, the same chemical space can be explored with "budget" BBs.
Xudong Wang; Ying Zhu; Qingyi Zhao; Weiwei Lu; Yechun Xu; Hangchen Hu; Xiaojie Lu ACS Med. Chem. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsmedchemlett.4c00069AbstractDNA-encoded library (DEL) technology is gaining attention for its rapid construction and deconvolution capabilities. Our study explored a novel strategy using rational DELs tailored for the SARS-CoV-2 papain-like protease, which revealed new fragments. Structural changes post-DEL screening mimic traditional medicinal chemistry lead optimization. We unveiled unique aromatic structures offering an alternative optimization path. Notably, we identified superior binding fragments targeting the BL2 groove. Derivative 16 emerged as the most promising by exhibiting IC50 values of 0.25 μM. Derivative 6, which features an aromatic fragment capped with a naphthalene moiety, showed IC50 values of 2.91 μM. Molecular modeling revealed hydrogen bond interactions with Lys157 residue and potential covalent interactions with nearby amino acid residues. This research underscored DEL’s potential for fragment-based drug discovery against SARS-CoV-2 protease.
Zijian Liu; Baiyang Mu; Zhiqiang Duan; Xuan Wang; Xiaojie Lu Synth. Bio. J., 2024, 5, DOI: 10.12211/2096-8280.2024-008 https://doi.org/10.12211/2096-8280.2024-008AbstractDNA-Encoded Library (DEL) technology, as an emerging means of small molecule drug screening, has become an important and indispensable technology platform for new drug discovery and development. The technology incorporates many advantages from combinatorial chemistry, molecular biology, and chemical bioinformatics, which greatly improve the efficiency of compound library synthesis and screening. Meanwhile, driven by the development of nucleic acid-compatible chemical reactions and high-throughput sequencing technology, DEL technology has made remarkable progress and gradually become a fast, economical, and efficient high-throughput screening platform, and has been more and more widely used in seedling compounds screening by schools, research institutes, and large pharmaceutical companies. The success of a DEL screening relies heavily on the chemical space and structural diversity of the compound libraries, both of which are directly affected by the number of chemical reactions compatible with nucleic acids. Therefore, developing the on-DNA chemical reactions to continuously enrich the chemical toolbox for DEL synthesis and thus enhance the structural diversity and drug potential of the molecules in the libraries has been the focus in this field. In recent years, the number of on-DNA chemical reactions has increased significantly, greatly broadening the scope of chemical reactions available for DEL construction. Meanwhile, a series of novel reaction methods, such as photocatalysis, electrocatalysis, and biosynthesis, have also emerged in the application of on-DNA chemical reactions and further expanded the field that on-DNA chemical reactions can reach. In this paper, we systematically review the metal-catalyzed on-DNA chemical reactions in recent years, including C(sp2)-C(sp2) bond-formation reactions, C(sp3)-C(sp3) bond-formation reactions, C(sp2)-C(sp3) bond-formation reactions, and C(sp2)-X bond-formation reactions; the synthesis of on-DNA privileged heterocycles with single-ring, fused-ring, and spirocyclic rings that using target-oriented synthetic and diversity-oriented synthetic strategies; the research progress of photocatalytic and enzyme-catalyzed on-DNA chemical reactions. However, the currently developed on-DNA reactions also have problems, such as compatibility with nucleic acids and substrate suitability. In the future, it is important and promising to exploit more robust on-DNA reactions that can proceed under mild conditions, new types of on-DNA reactions, and the combination of high-throughput screening and computer-assisted on-DNA reactions.
Kisanet Tadesse; Raphael I. Benhamou Non-Coding RNA, 2024, 10(2), 17 https://doi.org/10.3390/ncrna10020017AbstractMicroRNAs (miRs) have been implicated in numerous diseases, presenting an attractive target for the development of novel therapeutics. The various regulatory roles of miRs in cellular processes underscore the need for precise strategies. Recent advances in RNA research offer hope by enabling the identification of small molecules capable of selectively targeting specific disease-associated miRs. This understanding paves the way for developing small molecules that can modulate the activity of disease-associated miRs. Herein, we discuss the progress made in the field of drug discovery processes, transforming the landscape of miR-targeted therapeutics by small molecules. By leveraging various approaches, researchers can systematically identify compounds to modulate miR function, providing a more potent intervention either by inhibiting or degrading miRs. The implementation of these multidisciplinary approaches bears the potential to revolutionize treatments for diverse diseases, signifying a significant stride towards the targeting of miRs by precision medicine.
Nanna Birkmose; Emilie U. Frydendahl; Charlotte R. Knudsen Biochemistry, 2024, 63(24), 3273–3286 https://doi.org/10.1021/acs.biochem.4c00013AbstractDNA-encoded libraries hold great potential for discovering small, cyclized peptides with drug potential. Split-intein circular ligation of peptides and proteins (SICLOPPS) is a well-established method for in vivo selection of cyclic peptides targeting specific intracellular components. However, the method has mainly been used in prokaryotic cells. In contrast, selection studies performed directly in eukaryotic cells allow for the identification of cyclic peptides promoting a functional outcome, without the need to define a specific cellular target. Here, we report the construction of a Saccharomyces cerevisiae-specific SICLOPPS library of 80 million members, via careful optimization of several steps to increase the size of the library. Individual library members were shown to be correctly expressed and processed in yeast. High-throughput sequencing was conducted on the randomized primer used for library construction and the pure yeast SICLOPPS library isolated from Escherichia coli. A distinct guanine insertion bias was observed in the peptide-encoding, randomized sequence, which was primarily attributed to the degenerate primer used to introduce the randomized sequence. Moreover, high-throughput sequencing was performed on the library before and after the induction of cyclic peptide expression in yeast. Importantly, expression of the SICLOPPS library in S. cerevisiae caused only a marginal further sequence bias. Our work paves the way for selection studies using a large and diverse library to identify cyclic peptides of therapeutic interest that promote a specific phenotypic outcome in eukaryotic organisms, with yeast representing a beneficial model system due to its high transformation efficiency.
J. Trae Hampton; Wenshe Ray Liu Chem. Rev., 2024, 124(9), 6051-6077 https://doi.org/10.1021/acs.chemrev.4c00004AbstractSitting on the interface between biologics and small molecules, peptides represent an emerging class of therapeutics. Numerous techniques have been developed in the past 30 years to take advantage of biological methods to generate and screen peptide libraries for the identification of therapeutic compounds, with phage display being one of the most accessible techniques. Although traditional phage display can generate billions of peptides simultaneously, it is limited to expression of canonical amino acids. Recently, several groups have successfully undergone efforts to apply genetic code expansion to introduce noncanonical amino acids (ncAAs) with novel reactivities and chemistries into phage-displayed peptide libraries. In addition to biological methods, several different chemical approaches have also been used to install noncanonical motifs into phage libraries. This review focuses on these recent advances that have taken advantage of both biological and chemical means for diversification of phage libraries with ncAAs.
Paige Dickson ACS Chem. Biol., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acschembio.3c00803AbstractThe identification of novel covalent ligands for therapeutic purposes has long depended on serendipity, with dedicated hit finding techniques emerging only in the early 2000s. Advances in chemoproteomics have enabled robust characterization of putative drugs to derisk the unique liabilities associated with covalent hit molecules, leading to a renewed interest in this targeting modality. DNA-encoded library (DEL) technology has similarly emerged over the past two decades as a highly efficient method to identify new chemical equity toward protein targets of interest. A number of commercial and academic groups have reported methods in covalent DEL synthesis and hit identification; however, it is evident that there is still much to be done to fully realize the power of this technology for covalent ligand discovery. This perspective will explore the current approaches in covalent DEL technology and reflect on the next steps to advance this field.
Soobin Lee; Hyunchul Kwon; Eun-Kyoung Jee; Jaelim Kim; Kang Ju Lee; Jungyeon Kim; Nakeun Ko; Eunsung Lee; Hyun-Suk Lim Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c04387AbstractThe first synthesis of macrocyclic α-ABpeptoids with varying lengths is described. X-ray crystal structures reveal that cyclic trimer displays a chair-like conformation with a cct amide sequence and cyclic tetramer has a saddle-like structure with an uncommon cccc amide arrangement. The creation of a DNA-encoded combinatorial library of macrocyclic α-ABpeptoids is described.
Juan Zhang; Jinlu Liu; Gong Zhang; Xun He; Feng Xiong; Xiaohong Fan; Yangfeng Li; Yizhou Li Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c04374AbstractUtilizing already existing DNA-encoded libraries (DELs) for the generation of a distinct DEL represents an expedited strategy for expanding the chemical space. Herein, we leverage the unique photoreactivity of tetrazoles to synthesize diacylhydrazines on DNA. Widely available carboxylic acids serving as building blocks were employed under the mild photomediated reaction conditions, affording diverse DNA-conjugated diacylhydrazines. This methodology also demonstrates robustness in DEL-compatible synthesis and facilitates the preparation of oligonucleotide-based chemical probes.
Louise Plais; Louis Trachsel; Jörg Scheuermann Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.3c00559AbstractWhile dual-display DNA-encoded chemical libraries (DELs) are increasingly employed for ligand discovery, some of their fundamental properties have not yet been studied in-depth. Aided with fluorescence polarization experiments, we demonstrate that dual-display DELs are intrinsically asymmetrical entities, and we deduce practical guidelines to perform better-informed on-DNA hit validation from these libraries.
Ashley L. Ramos; Eric R. Goedken; Kristine E. Frank; Maria A. Argiriadi; Sana Bazzaz; Zhiguo Bian; Jesse T. C. Brown; Paolo A. Centrella; Hui-Ju Chen; Jeremy S. Disch; Pamela L. Donner; David B. Duignan; Diana Gikunju; Stephen N. Greszler; Marie-Aude Guié; Sevan Habeshian; Hajnalka E. Hartl; Christopher D. Hein; Charles W. Hutchins; Rachael Jetson; Anthony D. Keefe; Hasan Khan; Huan-Qiu Li; Allison Olszewski; Benjamin J. Ortiz Cardona; Augustine Osuma; Sanjay C. Panchal; Ryan Phelan; Wei Qiu; J. Brad Shotwell; Anurupa Shrestha; Myron Srikumaran; Zhi Su; Chaohong Sun; Anup K. Upadhyay; Michael D. Wood; Haihong Wu; Ruijie Zhang; Ying Zhang; Gang Zhao; Haizhong Zhu; Matthew P. Webster J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.3c02397AbstractDysregulation of IL17A drives numerous inflammatory and autoimmune disorders with inhibition of IL17A using antibodies proven as an effective treatment. Oral anti-IL17 therapies are an attractive alternative option, and several preclinical small molecule IL17 inhibitors have previously been described. Herein, we report the discovery of a novel class of small molecule IL17A inhibitors, identified via a DNA-encoded chemical library screen, and their subsequent optimization to provide in vivo efficacious inhibitors. These new protein–protein interaction (PPI) inhibitors bind in a previously undescribed mode in the IL17A protein with two copies binding symmetrically to the central cavities of the IL17A homodimer.
Rebecca Stevens; James D. F. Thompson; Julie C. L. Fournier; Glenn A. Burley; David J. Battersby; Afjal H. Miah Chem. Soc. Rev., 2024 https://doi.org/10.1039/D3CS01127EAbstractTargeted protein degraders such as PROTACs and molecular glues are a rapidly emerging therapeutic modality within industry and academia. Degraders possess unique mechanisms of action that lead to the removal of specific proteins by co-opting the cell's natural degradation mechanisms via induced proximity. Their optimisation thus far has often been largely empirical, requiring the synthesis and screening of a large number of analogues. In addition, the synthesis and development of degraders is often challenging, leading to lengthy optimisation campaigns to deliver candidate-quality compounds. This review highlights how the synthesis of degraders has evolved in recent years, in particular focusing on means of applying high-throughput chemistry and screening approaches to expedite these timelines, which we anticipate to be valuable in shaping the future of degrader optimisation campaigns.
Kristina Edfeldt; Aled M. Edwards; Ola Engkvist; Judith Günther; Matthew Hartley; David G. Hulcoop; Andrew R. Leach; Brian D. Marsden; Amelie Menge; Leonie Misquitta; Susanne Müller; Dafydd R. Owen; Kristof T. Schütt; Nicholas Skelton; Andreas Steffen; Alexander Tropsha; Erik Vernet; Yanli Wang; James Wellnitz; Timothy M. Willson; Djork-Arné Clevert; Benjamin Haibe-Kains; Lovisa Holmberg Schiavone; Matthieu Schapira Nat. Commun., 2024, 15(1), 5640 https://doi.org/10.1038/s41467-024-49777-xAbstractThe Structural Genomics Consortium is an international open science research organization with a focus on accelerating early-stage drug discovery, namely hit discovery and optimization. We, as many others, believe that artificial intelligence (AI) is poised to be a main accelerator in the field. The question is then how to best benefit from recent advances in AI and how to generate, format and disseminate data to enable future breakthroughs in AI-guided drug discovery. We present here the recommendations of a working group composed of experts from both the public and private sectors. Robust data management requires precise ontologies and standardized vocabulary while a centralized database architecture across laboratories facilitates data integration into high-value datasets. Lab automation and opening electronic lab notebooks to data mining push the boundaries of data sharing and data modeling. Important considerations for building robust machine-learning models include transparent and reproducible data processing, choosing the most relevant data representation, defining the right training and test sets, and estimating prediction uncertainty. Beyond data-sharing, cloud-based computing can be harnessed to build and disseminate machine-learning models. Important vectors of acceleration for hit and chemical probe discovery will be (1) the real-time integration of experimental data generation and modeling workflows within design-make-test-analyze (DMTA) cycles openly, and at scale and (2) the adoption of a mindset where data scientists and experimentalists work as a unified team, and where data science is incorporated into the experimental design.
Laura Lucaroni; Sebastian Oehler; Tony Georgiev; Marco Müller; Matilde Bocci; Roberto De Luca; Nicholas Favalli; Dario Neri; Samuele Cazzamalli; Luca Prati Chem. Sci., 2024 https://doi.org/10.1039/D3SC06668AAbstractProstate-Specific Membrane Antigen (PSMA) is a tumor-associated protein which has been successfully targeted with small organic ligands and with monoclonal antibodies. PluvictoTM is a PSMA-targeted Radio Ligand Therapeutic (RLT) recently approved by FDA for the treatment of metastatic castration-resistant prostate cancer (2022 FDA marketing authorization). Although a large Phase III clinical trial (VISION trial) demonstrated a clinical benefit in patients treated with PluvictoTM, the therapeutic window of the drug is narrowed by its undesired accumulation in healthy organs. Glutamate Carboxy Peptidase III (GCPIII), an enzyme sharing 70% identity with PSMA, may be responsible for the off-target accumulation of PSMA-RLTs in salivary glands and kidney. In this work, we designed and synthesized Affinity and Selectivity Maturation DNA-Encoded chemical Libraries (ASM-DELs) comprising 18’284’658 compounds, that were screened in parallel against PSMA and GCPIII with the aim to identify potent and selective PSMA ligands for tumor-targeting applications. Compound A70-B104 was isolated as the most potent and selective ligand (KD of 900 pM for PSMA, KD of 40 nM for GCPIII). 177Lu-A70-B104-DOTA, a radiolabeled derivative of compound A70-B104, presented a selective accumulation in PSMA-positive cancer lesions (i.e., 7.4% ID/g, 2 hours time point) after systemic administration in tumor-bearing mice. Results of autoradiography experiments showed that 177Lu-A70-B104-DOTA selectively binds to PSMA-positive cancer tissues, while negligible binding on human salivary glands was observed.
Wei Hou; Yiyuan Zhang; Fuchao Huang; Wanting Chen; Yuang Gu; Yan Wang; Jiacheng Pang; Hewei Dong; Kangyin Pan; Shuning Zhang; Peixiang Ma; Hongtao Xu Angew Chem. Int. Ed. Engl., 2024, e202318534 https://doi.org/10.1002/anie.202318534AbstractClick chemistry is a powerful molecular assembly strategy for rapid functional discovery. The development of click reaction with new connecting linkage is of great importance for expanding the click chemistry toolbox. We report the first Selenium-Nitrogen Exchange (SeNEx) click chemistry between benzoselenazolones and terminal alkynes (Se-N to Se-C), which is inspired by the biochemical SeNEx between Ebselen and cysteine (Cys) residue (Se-N to Se-S). The formed selenoalkyne connection is readily elaborated, thus endowing this chemistry with multidimensional molecular diversity. Besides, this reaction is characterized by modular, predictable, high-yielding, fast kinetics (k2 ≥ 14.43 M-1s-1), excellent functional group compatibility, and working well at miniaturization (nanomole-scale), opening up many interesting opportunities for organo-Se synthesis and bioconjugation, as exemplified by sequential click chemistry (coupled with ruthenium-catalyzed azide-alkyne cycloaddition (RuAAC) and sulfur-fluoride exchange (SuFEx), seleno-macrocycle synthesis, nanomole-scale synthesis of Se-containing natural product library and DNA-encoded library (DEL), late-stage peptide modification and ligation, and multiple functionalization of proteins. These results indicated that SeNEx is a useful strategy for new click chemistry development, and the established SeNEx chemistry will serve as a transformative platform in multidisciplinary fields such as synthetic chemistry, material science, chemical biology, medical chemistry, and drug discovery.
Christoph Dohmen; Heiko Ihmels ChemPhotoChem, 2024, e202300318 https://doi.org/10.1002/cptc.202300318AbstractIn the current field of photopharmacology, molecular photoswitches are applied whose interactions with DNA can be triggered or controlled by light. And although several photochromic reactions have been shown to serve this purpose well, the reversible photocycloaddition and photocycloreversion reactions have been largely neglected. This absence of research is surprising because especially the photodimerization of a DNA ligand leads to products with significant change of the size and shape which, in turn, leads to strongly diminished or even suppressed DNA association. Therefore, photocycloaddition-cycloreversion sequences have a huge potential for the photoinduced, reversible deactivation and activation of ligand-DNA interactions, as will be shown with selected examples in this Concept. Specifically, heterostyryl and -stilbene derivatives are presented whose DNA-binding properties are efficiently switched in reversible [2+2] photocycloaddition reactions. In addition, the photocontrolled DNA-binding of anthracene derivatives and their heterocyclic benzo[b]quinolizinium analogues in a [4+4] photocycloaddition, as well as the use of this reaction as part of dual-mode switches in combination with redox-active functionalities, are highlighted. Furthermore, examples of conjugates are provided, in which the photochromic unit is bound covalently to nucleic acids or proteins, such that the photocycloaddition reaction can be used for reversible photoinduced crosslinking, ligation, or inhibition of gene expression.
Chenhua Zhang; Yu-Chih Liu; Depu Wang; Yili Wang Biochemical and Biophysical Research Communications https://doi.org/10.1016/j.bbrc.2024.149537AbstractNeurodegeneration disorders, such as Alzheimer's disease (AD), have garnered significant attention due to their impact on individuals and society as a whole. Understanding the mechanisms behind these disorders and developing effective therapy strategies is of utmost importance. One potential therapeutic target that has emerged is Rho-associated coiled-coil containing protein kinase 2 (ROCK2), as its accumulation and activity have been closely linked to memory loss. In this report, we present the findings of a recent discovery involving a new molecule that has the ability to competitively inhibit ROCK2 activity. This molecule was identified through the utilization of a DNA-encoded library (DEL) screening platform. Following selection against ROCK2, an off-DNA compound was synthesized and examined to ascertain its inhibitory properties, selectivity, mechanism of action, and binding mode analysis. From the screening, compound CH-2 has demonstrated an IC50 value of 28 nM against ROCK2, while exhibiting a 5-fold selectivity over ROCK1. Further analysis through molecular docking has provided insights into the specific binding modes of this compound. Our findings suggest that DEL selection offers a rapid method for identifying new inhibitors. Among these, the CH-2 compound shows promise as a potential ROCK2 inhibitor and warrants further investigation.
Jason D. Shields; Rachel Howells; Gillian Lamont; Yin Leilei; Andrew Madin; Christopher Reimann; Hadi Rezaei; Tristan Reuillon; Bryony Smith; Clare Thomson; Yuting Zheng; Robert E. Ziegler RSC Med. Chem., 2024 https://doi.org/10.1039/D3MD00651DAbstractAstraZeneca chemists have been using the AI retrosynthesis tool AiZynth for three years. In this article, we present seven examples of how medicinal chemists using AiZynth positively impacted their drug discovery programs. These programs run the gamut from early-stage hit confirmation to late-stage route optimisation efforts. We also discuss the different use cases for which AI retrosynthesis tools are best suited.
Dean G. Brown J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.3c02165AbstractExpedited development and approval pathways at the Food and Drug Administration (FDA) such as Priority review, Fast Track Designation, Breakthrough Designation, and Accelerated Approval are programs available to drug sponsors that aim to incentivize and expedite the delivery of drugs to patients in need. In addition, other incentive programs such as Orphan Drug Designation (ODD), Qualified Infectious Disease Product Designation (QIDP), and Rare Pediatric Disease Designation (RPDD) are available to drug sponsors to help motivate development of drugs that may have lower economic incentive for commercialization. These programs have been largely effective, and many new innovative drugs since 2010 have accessed these programs. This Perspective highlights how these programs have been used in recent FDA drug approvals and discusses future ways sponsors and regulatory agencies may further enable development of these innovative drugs in the most expeditious fashion.
George Hartman; Paul Humphries; Robert Hughes; Andrew Ho; Rusty Montgomery; Aditi Deshpande; Maitriyee Mahanta; Sarah Tronnes; Samantha Cowdin; Xu He; Fangchao Liu; Lifang Zhang; Chuan Liu; Dengfeng Dou; Jin Li; Aleksander Spasic; Rebecca Coll; Michael Marleaux; Inga V. Hochheiser; Matthias Geyer; Paul Rubin; Kristen Fortney; Kevin Wilhelmsen Bioorg. Med. Chem. Lett., 2024, 102, 129675 https://doi.org/10.1016/j.bmcl.2024.129675AbstractNLRP3 is an intracellular sensor protein that detects a broad range of danger signals and environmental insults. Its activation results in a protective pro-inflammatory response designed to impair pathogens and repair tissue damage via the formation of the NLRP3 inflammasome. Assembly of the NLRP3 inflammasome leads to caspase 1-dependent secretory release of the pro-inflammatory cytokines IL-1β and IL-18 as well as to gasdermin d-mediated pyroptotic cell death. Herein, we describe the discovery of a novel indazole series of high affinity, reversible inhibitors of NLRP3 activation through screening of DNA-encoded libraries and the potent lead compound 3 (BAL-0028, IC50 = 25 nM) that was identified directly from the screen. SPR studies showed that compound 3 binds tightly (KD range 104–123 nM) to the NACHT domain of NLRP3. A CADD analysis of the interaction of compound 3 with the NLRP3 NACHT domain proposes a binding site that is distinct from those of ADP and MCC950 and includes specific site interactions. We anticipate that compound 3 (BAL-0028) and other members of this novel indazole class of neutral inhibitors will demonstrate significantly different physical, biochemical, and biological properties compared to NLRP3 inhibitors previously identified.
Xiaohong Fan; Xianfeng Li; Yangfeng Li; Yizhou Li Eur. J. Org. Chem., 2024, e202301172 https://doi.org/10.1002/ejoc.202301172AbstractAryl diazonium intermediates have emerged as highly versatile intermediates in modern organic synthesis and chemical biology, demonstrating remarkable reactivity under mild reaction conditions. Their significance has extended to DNA-encoded libraries (DELs), where they serve as valuable alternatives to traditional aryl halide-based cross-coupling reactions. Aryl diazonium intermediates as N2-annulation synthons can efficiently transform into benzotriazinone core on DNA besides C–C coupling. This concept paper illuminates recent advances in this field, emphasizing their applications and potential across various facets of DELs, off-DNA chemistry, DELs selection, and DNA-based probes. It is anticipated that these developments will propel the broader utilization of DNA-conjugated aryl diazonium salt intermediates in DELs, protein labeling, hit discovery, and other interdisciplinary fields.
W. Felix Zhu; Claire Empel; Sveva Pelliccia; Rene M. Koenigs; Ewgenij Proschak; Victor Hernandez-Olmos J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.3c02109AbstractPhotochemistry has emerged as a transformative force in organic chemistry, significantly expanding the chemical space accessible for medicinal chemistry. Light-induced reactions enable the efficient synthesis of intricate organic structures and have found applications throughout the different stages of the drug discovery and development processes. Moreover, photochemical techniques provide innovative solutions in chemical biology, allowing precise spatiotemporal drug activation and targeted delivery. In this Perspective, we highlight the already numerous remarkable applications and the even more promising future of photochemistry in medicinal chemistry and chemical biology.
Christoph Nitsche Synlett, 2023 https://doi.org/10.1055/a-2214-7612AbstractBioorthogonal reactions hold significant promise for applications in chemical biology. Despite their potential, nitriles have often been overlooked as reactive functional groups for selective bioconjugation. The condensation reaction between cyanopyridines and 1,2-aminothiols stands out as a particularly favorable nitrile modification strategy that proceeds under biocompatible conditions. Cyanopyridines can be seamlessly incorporated into peptides and proteins through both chemical and biotechnological approaches. Similarly, the selective integration of 1,2-aminothiols into peptides and proteins is achievable, leveraging the uniquely reactive N-terminal cysteine functional group.
Simon C. C. Lucas; J. Henry Blackwell; Sarah H. Hewitt; Hannah Semple; Benjamin C. Whitehurst; Hua Xu SLAS Discov., 2024 https://doi.org/10.1016/j.slasd.2024.01.003AbstractCovalent hits for drug discovery campaigns are neither fantastic beasts nor mythical creatures, they can be routinely identified through electrophile-first screening campaigns using a suite of different techniques. These include biophysical and biochemical methods, cellular approaches, and DNA-encoded libraries. Employing best practice, however, is critical to success. The purpose of this review is to look at state of the art covalent hit identification, how to identify hits from a covalent library and how to select compounds for medicinal chemistry programmes.
Inna V. Krieger; Subbarao Yalamanchili; Paige Dickson; Curtis A. Engelhart; Matthew D Zimmerman; Jeremy Wood; Ethan Clary; Jasmine Nguyen; Natalie Thornton; Paolo A. Centrella; Betty Chan; John W Cuozzo; Martin Gengenbacher; Marie-Aude Guié; John P Guilinger; Corey Bienstock; Hajnalka Hartl; Christopher D. Hupp; Rachael Jetson; Takashi Satoh; John T. S. Yeoman; Ying Zhang; Veronique Dartois; Dirk Schnappinger; Anthony D. Keefe; James C. Sacchettini ACS Infect. Dis., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsinfecdis.3c00592AbstractDNA-encoded chemical library (DEL) technology provides a time- and cost-efficient method to simultaneously screen billions of compounds for their affinity to a protein target of interest. Here we report its use to identify a novel chemical series of inhibitors of the thioesterase activity of polyketide synthase 13 (Pks13) from Mycobacterium tuberculosis (Mtb). We present three chemically distinct series of inhibitors along with their enzymatic and Mtb whole cell potency, the measure of on-target activity in cells, and the crystal structures of inhibitor-enzyme complexes illuminating their interactions with the active site of the enzyme. One of these inhibitors showed a favorable pharmacokinetic profile and demonstrated efficacy in an acute mouse model of tuberculosis (TB) infection. These findings and assay developments will aid in the advancement of TB drug discovery.
Andrea Testa Nat. Chem. Biol., 2023 https://doi.org/10.1038/s41589-023-01473-5AbstractDNA-encoded libraries are a powerful tool to identify novel chemical inducers of proximity such as targeted protein degraders, even without a known binder for the target protein.
András Gy. Németh; Levente Kollár; Krisztina Németh; Gitta Schlosser; Annamária Minus; György M. Keserű Org. Lett., 2023, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c03602AbstractThe increasing role of the DNA-encoded library technology in early phase drug discovery represents a significant demand for DNA-compatible synthetic methods for therapeutically relevant heterocycles. Herein, we report the first on-DNA synthesis of multisubstituted indoles via a cascade reaction of Sonogashira coupling and intramolecular ring closure. Further functionalization by Suzuki coupling at the third position exploits a diverse chemical space. The high fidelity of the method also enabled the construction of an indole-based mock library.
Soo Bok Kim; Dong Hyeon Kim; Han Yong Bae Nat. Commun., 2024, 15, 3876 https://doi.org/10.1038/s41467-024-47861-wAbstractSulfur(VI) fluoride exchange (SuFEx) has emerged as an innovative click chemistry to harness the pivotal connectivity of sulfonyl fluorides. Synthesizing such alkylated S(VI) molecules through a straightforward process is of paramount importance, and their water-compatibility opens the door to a plethora of applications in biorelevant and materials chemistry. Prior aquatic endeavors have primarily focused on delivering catalysts involving ionic mechanisms, studies regarding visible-light photocatalytic transformation are unprecedented. Herein we report an on-water accelerated dearomative aquaphotocatalysis for heterocyclic alkyl SuFEx hubs. Notably, water exerts a pronounced accelerating effect on the [2 + 2] cycloaddition between (hetero)arylated ethenesulfonyl fluorides and inert heteroaromatics. This phenomenon is likely due to the high-pressure-like reactivity amplification at the water-oil interface. Conventional solvents proved totally ineffective, leading to the isomerization of the starting material.
Panhong Gou; Wenchao Zhang Biomed. Pharmacother., 2024, 171, 116130 https://doi.org/10.1016/j.biopha.2024.116130AbstractCBP and p300 are homologous proteins exhibiting remarkable structural and functional similarity. Both proteins function as acetyltransferase and coactivator, underscoring their significant roles in cellular processes. The function of histone acetyltransferases is to facilitate the release of DNA from nucleosomes and act as transcriptional co-activators to promote gene transcription. Transcription factors recruit CBP/p300 by co-condensation and induce transcriptional bursting. Disruption of CBP or p300 functions is associated with different diseases, especially cancer, which can result from either loss of function or gain of function. CBP and p300 are multidomain proteins containing HAT (histone acetyltransferase) and BRD (bromodomain) domains, which perform acetyltransferase activity and maintenance of HAT signaling, respectively. Inhibitors targeting HAT and BRD have been explored for decades, and some BRD inhibitors have been evaluated in clinical trials for treating hematologic malignancies or advanced solid tumors. Here, we review the development and application of CBP/p300 inhibitors. Several inhibitors have been evaluated in vivo, exhibiting notable potency but limited selectivity. Exploring these inhibitors emphasizes the promise of targeting CBP and p300 with small molecules in cancer therapy.
Hongguang Deng; Yu Han; Liang Liu; Hong Zhang; Dan Liu; Jiachen Wen; Min Huang; Linxiang Zhao J. Med. Chem. 2024, 67, 8, 5963–5998 https://doi.org/10.1021/acs.jmedchem.3c01998AbstractAs a tripartite cell death switch, B-cell lymphoma protein 2 (Bcl-2) family members precisely regulate the endogenous apoptosis pathway in response to various cell signal stresses through protein–protein interactions. Myeloid leukemia-1 (Mcl-1), a key anti-apoptotic Bcl-2 family member, is positioned downstream in the endogenous apoptotic pathway and plays a central role in regulating mitochondrial function. Mcl-1 is highly expressed in a variety of hematological malignancies and solid tumors, contributing to tumorigenesis, poor prognosis, and chemoresistance, making it an attractive target for cancer treatment. This Perspective aims to discuss the mechanism by which Mcl-1 regulates apoptosis and non-apoptotic functions in cancer cells and to outline the discovery and optimization process of potent Mcl-1 modulators. In addition, we summarize the structural characteristics of potent inhibitors that bind to Mcl-1 through multiple co-crystal structures and analyze the cardiotoxicity caused by current Mcl-1 inhibitors, providing prospects for rational targeting of Mcl-1.
Juan Zhang; Jinlu Liu; Xianfeng Li; Yunzhu Ju; Yangfeng Li; Gong Zhang; Yizhou Li J. Am. Chem. Soc., 2024, 146(3), 2122–2131 https://doi.org/10.1021/jacs.3c11574AbstractBioconjugation chemistry has emerged as a powerful tool for the modification of diverse biomolecules under mild conditions. Tetrazole, initially proposed as a bioorthogonal photoclick handle for 1,3-dipolar cyclization with alkenes, was later demonstrated to possess broader photoreactivity with carboxylic acids, serving as a versatile bioconjugation and photoaffinity labeling probe. In this study, we unexpectedly discovered and validated the photoreactivity between tetrazole and primary amine to afford a new 1,2,4-triazole cyclization product. Given the significance of functionalized N-heterocycles in medicinal chemistry, we successfully harnessed the serendipitously discovered reaction to synthesize both pharmacologically relevant DNA-encoded chemical libraries (DELs) and small molecule compounds bearing 1,2,4-triazole scaffolds. Furthermore, the mild reaction conditions and stable 1,2,4-triazole linkage found broad application in photoinduced bioconjugation scenarios, spanning from intramolecular peptide macrocyclization and templated DNA reaction cross-linking to intermolecular photoaffinity labeling of proteins. Triazole cross-linking products on lysine side chains were identified in tetrazole-labeled proteins, refining the comprehensive understanding of the photo-cross-linking profiles of tetrazole-based probes. Altogether, this tetrazole-amine bioconjugation expands the current bioconjugation toolbox and creates new possibilities at the interface of medicinal chemistry and chemical biology.
Daniel Vik; David Pii; Chirag Mudaliar; Mads Nørregaard-Madsen; Aleksejs Kontijevskis Sci. Rep., 2024, 14, 8733 https://doi.org/10.1038/s41598-024-59620-4AbstractThis study explores how machine-learning can be used to predict chromatographic retention times (RT) for the analysis of small molecules, with the objective of identifying a machine-learning framework with the robustness required to support a chemical synthesis production platform. We used internally generated data from high-throughput parallel synthesis in context of pharmaceutical drug discovery projects. We tested machine-learning models from the following frameworks: XGBoost, ChemProp, and DeepChem, using a dataset of 7552 small molecules. Our findings show that two specific models, AttentiveFP and ChemProp, performed better than XGBoost and a regular neural network in predicting RT accurately. We also assessed how well these models performed over time and found that molecular graph neural networks consistently gave accurate predictions for new chemical series. In addition, when we applied ChemProp on the publicly available METLIN SMRT dataset, it performed impressively with an average error of 38.70 s. These results highlight the efficacy of molecular graph neural networks, especially ChemProp, in diverse RT prediction scenarios, thereby enhancing the efficiency of chromatographic analysis.
Michael Oelgemöller; Lijing Zhang; Fang Zhao; Yuanhai Su Front. Chem., 2023, 11, 1322556 https://doi.org/10.3389/fchem.2023.1322556AbstractDue to the development of novel light-sources, methodologies and technologies, photochemistry has seen a remarkable renaissance in academia and industry (Baumann et al., 2014; Bonfield et al., 2020; Cohen et al., 2023). Many photochemical investigations are now routinely performed under continuous-flow conditions in purpose-designed reactors (Loubière et al., 2016; Buglioni et al., 2022). Successful examples of pre-industrial applications have subsequently been developed and realized (Basso and Capurro, 2021; Donnelly and Baumann, 2021; Zhang and Roth, 2023). Likewise, photocalytic materials can be easily incorporated into reactor channels, thus further advancing the potential of flow-photochemistry (Franchi and Amara, 2020; Thomson et al., 2020; Zuliani and Cova, 2021). This Research Topic comprises of four submissions and highlights recent achievements in photochemical research. Li et al. developed a novel Fe3+-TiO2@CGS three-dimensional photoelectric system and applied it to the degradation of methylene blue. Under optimal operation conditions, the device reached a degradation yield of 99.98% after 60 min of photoelectrical treatment, clearly demonstrating the potential of this technology for the removal of organic contaminants. The constructed photoelectrical degradation reactor was equipped with inlet and outlet points, thus permitting (circulating) flow operation in future studies. Dinter et al. reported on the development of a flexible and affordable microfluidic photochemical flow reactor for rapid prototyping. The fabricated module was first utilized to optimize a photopinacolization reaction and was subsequently transferred to an application with DNA-tagged substrates. The study demonstrated the suitability of the developed modular flow photoreactor as a DNA-encoded library technology (DELT). Meinhardová et al. investigated the role of the lamp type for photocatalytic hydrogen production under batch and flow conditions. The authors initially established the efficiency of six commercial lamps in a batch reactor using a methanol-water solution and a NiO-TiO2 photocatalyst. Using a circulating microphotoreactors system incorporating TiO2 immobilized on borosilicate glass, continuous and reproducible hydrogen generation of 333.7 ± 21.1 µmol H2 or 252.8 ± 16.0 mmol·m−2 was achieved over a period of 168 h. Guo et al. summarized recent advances in catalyst development for the photocatalytic hydrogenation of nitrobenzene to aniline. In contrast to thermal methods, photocatalysis enables the sustainable production of the important platform chemical aniline at room temperature and low hydrogen pressures. Photocatalysts were divided into semiconductors, plasmonic metal-based catalysts and dyes, and the challenges, opportunities and future development prospects of these materials were described. Subsequent immobilization of these photocatalytic materials into flow devices may enable a continuous future production of aniline. All contributions unambiguously demonstrate the potential and importance of flow-photochemistry and photocatalysis as sustainable and energy-efficient technologies.
Yingbo Zhou; Zhangtao Cai; Yijia Zhai; Jintao Yu; Qiujing He; Yuan He; Siriporn Jitkaew; Zhenyu Cai Apoptosis, 2023 https://doi.org/10.1007/s10495-023-01905-6AbstractNecroptosis is a type of programmed cell death that is morphologically similar to necrosis. This type of cell death is involved in various pathophysiological disorders, including inflammatory, neurodegenerative, infectious, and malignant diseases. Receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like protein (MLKL) pseudokinase constitute the core components of the necroptosis signaling pathway and are considered the most promising targets for therapeutic intervention. The discovery and characterization of necroptosis inhibitors not only accelerate our understanding of the necroptosis signaling pathway but also provide important drug candidates for the treatment of necroptosis-related diseases. Here, we will review recent research progress on necroptosis inhibitors, mechanisms of action and their potential applications for disease treatment.
Xianfu Fang; Tianyang Zhang; Wei Fang; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett. 2023, 25(46), 8326–8331 https://doi.org/10.1021/acs.orglett.3c03404AbstractTriazoles are privileged structural motifs that are embedded in a number of molecules with interesting biological activities. In this work, we developed a practical and general synthetic strategy to construct a medicinally important 5-amino-1,2,3-triazole moiety on DNA by coupling DNA-conjugated azides and monosubstituted acetonitriles via azide-acetonitrile “click” reaction. Under mild reaction conditions, this reaction displayed a broad substrate scope. Most substrates gave moderate-to-excellent conversions. Thus, this DNA-compatible reaction could be employed in practical DNA-encoded library (DEL) construction and potentially expand the chemical space of DNA-encoded libraries.
Mark A. Reed; Carla E. Brown Synfacts, 2023, 19(11), 1076 https://doi.org/10.1055/s-0042-1752036AbstractN/A
Ramón Rama-Garda; María Dolores Martin-Ortega; Anthony de Jesús Sánchez; Julián Priego; Jesús de Blas; Alicia Torrado; Eduardo Domínguez; Rubén Haro; Alfonso Rivera-Sagredo; José Pablo Román; María José Lorite; Hans E Johansson; María Isabel Loza; Jorge Amigo; Beatriz Sobrino; María José Lallena; Miguel Ángel Toledo Bioorg. Med. Chem., 2024, 117596 https://doi.org/10.1016/j.bmc.2024.117596AbstractCodification of DNA Encoded Libraries (DELs) is critical for successful ligand identification of molecules that bind a protein of interest (POI). There are different encoding strategies that permit, for instance, the customization of a DEL for testing single or dual pharmacophores (single strand DNA) or for producing and screening large diversity libraries of small molecules (double strand DNA). Both approaches challenges, either from the synthetic and encoding point of view, or from the selection methodology to be utilized for the screening. The Head-Piece contains the DNA sequence that is attached to a chemical compound, allowing the encoding of each molecule with a unique DNA tag. Designing the Head-Piece for a DNA-encoded library involves careful consideration of several key aspects including DNA barcode identity, sequence length and attachment chemistry. Here we describe a double stranded DNA versatile Head-Piece that can be used for the generation of single or dual pharmacophore libraries, but also shows other advanced DEL functionalities, stability and enlarged encoding capacity.
Millicent Dockerill; Daniel J. Ford; Simona Angerani; Imala Alwis; Luke J. Dowman; Jorge Ripoll-Rozada; Rhyll E. Smythe; Joanna S. T. Liu; Pedro José Barbosa Pereira; Shaun P. Jackson; Richard J. Payne; Nicolas Winssinge Nat. Biotech., 2024 https://doi.org/10.1038/s41587-024-02209-zAbstractDrugs are administered at a dosing schedule set by their therapeutic index, and termination of action is achieved by clearance and metabolism of the drug. In some cases, such as anticoagulant drugs or immunotherapeutics, it is important to be able to quickly reverse the drug’s action. Here, we report a general strategy to achieve on-demand reversibility by designing a supramolecular drug (a noncovalent assembly of two cooperatively interacting drug fragments held together by transient hybridization of peptide nucleic acid (PNA)) that can be reversed with a PNA antidote that outcompetes the hybridization between the fragments. We demonstrate the approach with thrombin-inhibiting anticoagulants, creating very potent and reversible bivalent direct thrombin inhibitors (Ki = 74 pM). The supramolecular inhibitor effectively inhibited thrombus formation in mice in a needle injury thrombosis model, and this activity could be reversed by administration of the PNA antidote. This design is applicable to therapeutic targets where two binding sites can be identified.
Lijun Fan; Yang Yu; Charles Jayne; John R. Frost; Jack D. Scott Org. Lett., 2023, 25(44), 8038–8042 https://doi.org/10.1021/acs.orglett.3c03284AbstractDNA-encoded library (DEL) technology holds exciting potential for discovering novel therapeutic macrocyclic peptides (MPs). Herein, we describe the development of a DEL-compatible peptide macrocyclization method that proceeds via intramolecular click-condensation between 3-(2-cyano-4-pyridyl)-l-alanine (Cpa) and an N-terminal cysteine. Cyclization takes place spontaneously in a buffered aqueous solution and affords the cyclized products in excellent yields. The reaction exhibits a broad substrate scope and can be employed to generate MPs of variable ring size and amino acid composition.
Ann M. Rowley; Gang Yao; Logan Andrews; Aaron Bedermann; Ross Biddulph; Ryan Bingham; Jennifer J. Brady; Rachel Buxton; Ted Cecconie; Rona Cooper; Adam Csakai; Enoch N. Gao; Melissa C. Grenier-Davies; Meghan Lawler; Yiqian Lian; Justyna Macina; Colin Macphee; Lisa Marcaurelle; John Martin; Patricia McCormick; Rekha Pindoria; Martin Rauch; Warren Rocque; Yingnian Shen; Lisa M. Shewchuk; Michael Squire; Will Stebbeds; Westley Tear; Xin Wang; Paris Ward; Shouhua Xiao J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.3c01562AbstractHuman genetic evidence shows that PDE3B is associated with metabolic and dyslipidemia phenotypes. A number of PDE3 family selective inhibitors have been approved by the FDA for various indications; however, given the undesirable proarrhythmic effects in the heart, selectivity for PDE3B inhibition over closely related family members (such as PDE3A; 48% identity) is a critical consideration for development of PDE3B therapeutics. Selectivity for PDE3B over PDE3A may be achieved in a variety of ways, including properties intrinsic to the compound or tissue-selective targeting. The high (>95%) active site homology between PDE3A and B represents a massive obstacle for obtaining selectivity at the active site; however, utilization of libraries with high molecular diversity in high throughput screens may uncover selective chemical matter. Herein, we employed a DNA-encoded library screen to identify PDE3B-selective inhibitors and identified potent and selective boronic acid compounds bound at the active site.
Gavin W. Collie; Matthew A. Clark; Anthony D. Keefe; Andrew Madin; Jon A. Read; Emma L. Rivers; Ying Zhang J. Med. Chem., 2024, 67(2), 864–884 https://doi.org/10.1021/acs.jmedchem.3c01861AbstractThe DNA-encoded library (DEL) discovery platform has emerged as a powerful technology for hit identification in recent years. It has become one of the major parallel workstreams for small molecule drug discovery along with other strategies such as HTS and data mining. For many researchers working in the DEL field, it has become increasingly evident that many hits and leads discovered via DEL screening bind to target proteins with unique and unprecedented binding modes. This Perspective is our attempt to analyze reports of DEL screening with the purpose of providing a rigorous and useful account of the binding modes observed for DEL-derived ligands with a focus on binding mode novelty.
Kazuki Yamamoto Pharmacia, 2023, 59(10), 943 https://doi.org/10.14894/faruawpsj.59.10_943AbstractUnlike conventional inhibitors, which bind to target proteins and reduce their function, proteolysis inducers are attracting attention as new drug discovery modalities because they chemically knock down target proteins. Proteolysis targeting chimera (PROTAC) has an E3 ligase and a binding site for the target molecule in its molecule, and induces degradation by forcibly ubiquitinating the target protein. One of the advantages of PROTAC is that it acts catalytically, and lower doses can be expected to reduce toxicity. However, in the development of PROTACs, there are no guidelines for optimizing the modification position of the target binder or the length and shape of the linker, and it is necessary to customize it depending on the target, so efficiency is required. This time, we will introduce an approach to PROTAC optimization using Chen et al.'s DNA encoded library (DEL).
Suhyeorn Park; Jiayi Fan; Srinivas Chamakuri; Murugesan Palaniappan; Kiran Sharma; Xuan Qin; Jian Wang; Zhi Tan; Allison Judge; Liya Hu; Banumathi Sankaran; Feng Li; B. V. Venkataram Prasad; Martin M. Matzuk; Timothy Palzkill J. Med. Chem., 2024, 67(1), 620–642 https://doi.org/10.1021/acs.jmedchem.3c01834Abstractβ-Lactamase enzymes hydrolyze and thereby provide bacterial resistance to the important β-lactam class of antibiotics. The OXA-48 and NDM-1 β-lactamases cause resistance to the last-resort β-lactams, carbapenems, leading to a serious public health threat. Here, we utilized DNA-encoded chemical library (DECL) technology to discover novel β-lactamase inhibitors. We exploited the β-lactamase enzyme–substrate binding interactions and created a DECL targeting the carboxylate-binding pocket present in all β-lactamases. A library of 106 compounds, each containing a carboxylic acid or a tetrazole as an enzyme recognition element, was designed, constructed, and used to identify OXA-48 and NDM-1 inhibitors with micromolar to nanomolar potency. Further optimization led to NDM-1 inhibitors with increased potencies and biological activities. This work demonstrates that the carboxylate-binding pocket-targeting DECL, designed based on substrate binding information, aids in inhibitor identification and led to the discovery of novel non-β-lactam pharmacophores for the development of β-lactamase inhibitors for enzymes of different structural and mechanistic classes.
Animesh Kairi; Tapas Bhadra Eur. Chem. Bull., 2023, 12(10), 3597-3609 https://doi.org/10.48047/ecb/2023.12.si10.00413AbstractDNA cryptography is a particularly promising area of research in this field. The advantage of DNA cryptography is its potential to store vast amounts of data in a tiny volume. This high data density makes it well-suited for applications like secure archival storage, particularly for sensitive or classified information. Moreover, DNA cryptography's parallel processing capabilities, driven by DNA's multitasking nature can significantly accelerate encryption and decryption processes. The idea of utilizing DNA cryptography to encrypt and decode data is proposed in this article. The proposed approach performs well in computing, storage, and transmission and is especially resistant to different assaults, according to theoretical research and real world implementations. This offers a ground-breaking DNA cryptography technique based on the decimal bond algorithm. In this article, we develop the fundamental concepts underlying DNA cryptography, including the encoding of binary information into DNA sequences, encryption algorithms, and decoding procedures. We discuss various encoding strategies, their strengths, and their limitations, highlighting the trade-offs between data security and efficiency. Furthermore, we examine the challenges associated with DNA cryptography, such as the need for specialized laboratory equipment and the ongoing advancements required in biotechnology. This study also proposes a novel key generation scheme and an innovative cipher text generation approach. This article concludes with a comparison, demonstrating the effectiveness of the proposed approach against existing DNA cryptography methods.
Silin Zhang; Haiman Zhang; Xiawen Liu; Ping Qi; Tingting Tan; Shengdong Wang; Hui Gao; Hongtao Xu; Zhi Zhou; Wei Yi Adv.Sci., 2023, 2307049 https://doi.org/10.1002/advs.202307049AbstractAn ideal DNA-encoded library (DEL) selection requires the library to consist of diverse core skeletons and cover chemical space as much as possible. However, the lack of efficient on-DNA synthetic approaches toward core skeletons has greatly restricted the diversity of DEL. To mitigate this issue, this work disclosed a “Mask & Release” strategy to streamline the challenging on-DNA core skeleton synthesis. N-phenoxyacetamide is used as a masked phenol and versatile directing group to mediate diversified DNA-compatible C-H functionalization, introducing the 1st-dimensional diversity at a defined site, and simultaneously releasing the phenol functionality, which can facilitate the introduction of the 2nd diversity. This work not only provides a set of efficient syntheses toward DNA-conjugated drug-like core skeletons such as ortho-alkenyl/sulfiliminyl/cyclopropyl phenol, benzofuran, dihydrobenzofuran but also provides a paradigm for on-DNA core skeleton synthetic method development.
Nicolas Duchemin; Zhaomei Sun; Yunjin Hu Eur. Chem. Bull., 2023, 12(9), 3267-3278 https://doi.org/10.48047/ecb/2023.12.Si9.2952023.21/09/2023AbstractDNA-encoded libraries are collections of small molecules covalently bound to single or doublestranded oligonucleotides, which sequences individually carry encoded information about the identity of the molecule. These oligonucleotide conjugates are assembled using building blocks or cores, and their corresponding encoding DNA tags. Libraries, once synthesized, can be screened towards biological targets of interest, and binders, being physically separated from non-binders, can be identified simple decoding of the unique DNA tags, using PCR amplification and DNA sequencing. Ever since the conceptual inception of DNA-encoded library technology in 1992, [1] and following applicative studies, the field has been rapidly emerging as a powerful pathway to discover valuable chemical matter for drug discovery and probe molecules. Thanks to the efforts of many academics and industrial groups, this technology has been continuously evolving and benefited from the multidimensional advancements: new on-DNA chemical reactions, improved selection methods, increasingly affordable nucleotide sequencing, and ameliorated data analysis approach to reveal the enriched binders. [4, 5] The significance and strong impact of DELT in unveiling relevant hits in an efficient and economical manner was recently witnessed by the expedient access to SARS-CoV-2 M pro inhibitors, and naturally became increasingly significant in drug discovery. These efforts notably contributed to the discovery of two phase 2 clinical candidates GSK2256294 and GSK2982772 from GSK and recently FDA-approved X-165 for Phase I clinical trials.
Priyanka Raghavan; Brittany C. Haas; Madeline E. Ruos; Jules Schleinitz; Abigail G. Doyle; Sarah E. Reisman; Matthew S. Sigman; Connor W. Coley ACS Cent. Sci. 2023, 9(12), 2196-2204 https://doi.org/10.1021/acscentsci.3c01163AbstractModels can codify our understanding of chemical reactivity and serve a useful purpose in the development of new synthetic processes via, for example, evaluating hypothetical reaction conditions or in silico substrate tolerance. Perhaps the most determining factor is the composition of the training data and whether it is sufficient to train a model that can make accurate predictions over the full domain of interest. Here, we discuss the design of reaction datasets in ways that are conducive to data-driven modeling, emphasizing the idea that training set diversity and model generalizability rely on the choice of molecular or reaction representation. We additionally discuss the experimental constraints associated with generating common types of chemistry datasets and how these considerations should influence dataset design and model building.
Suzanne Willems; Elena Detta; Lorenzo Baldini; Deniz Tietz; Andrea Trabocchi; Andreas Brunschweiger ACS Omega, 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsomega.3c07136AbstractIn DNA-encoded library synthesis, amine-substituted building blocks are prevalent. We explored isocyanide multicomponent reactions to diversify DNA-tagged amines and reported the Ugi-azide reaction with high yields and a good substrate scope. In addition, the Ugi-aza-Wittig reaction and the Ugi-4-center-3-component reaction, which used bifunctional carboxylic acids to provide lactams, were explored. Five-, six-, and seven-membered lactams were synthesized from solid support-coupled DNA-tagged amines and bifunctional building blocks, providing access to structurally diverse scaffolds.
The Atomwise AIMS Program Sci. Rep., 2024, 14, 7526 https://doi.org/10.1038/s41598-024-54655-zAbstractHigh throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery.
Dirk Trauner; Joseph A. Flores Synfacts, 2023, 19(10), 1036 https://doi.org/10.1055/s-0042-1752021AbstractDNA-encoded libraries (DELs) represent powerful platforms for identifying small molecule hits in early-stage drug discovery. For hits identified from enrichment data generated with purified protein, methods for triaging ligands based on live cell occupancy would accelerate hit-to-lead prioritization. Here, the authors demonstrate a generalizable workflow to convert DEL hit ligands into bioluminescence resonance energy transfer (BRET) probes for target engagement studies in live cells.
Evan S. O’Brien; Vipin Ashok Rangari; Amal El Daibani; Shainnel O. Eans; Betsy White; Haoqing Wang; Yuki Shiimura; Kaavya Krishna Kumar; Kevin Appourchaux; Weijiao Huang; Chensong Zhang; Jesper M. Mathiesen; Tao Che; Jay P. McLaughlin; Susruta Majumdar; Brian K. Kobilka bioRxiv, 2023 https://doi.org/10.1101/2023.09.08.556921AbstractThe µ-opioid receptor (µOR) is a well-established target for analgesia, yet conventional opioid receptor agonists cause serious adverse effects, notably addiction and respiratory depression, which have led to the present opioid overdose epidemic. µOR negative allosteric modulators (NAMs) may serve as powerful tools in preventing opioid overdose deaths, but promising chemical scaffolds remain elusive. We screened a large DNA-encoded chemical library against inactive µOR, counter-screening with active, G-protein and agonist bound receptor to “steer” selections toward functional negative allosteric modulators. We discovered a NAM compound with high and selective enrichment to inactive µOR; the molecule potently blocks the activity of orthosteric agonists and enhances the affinity of the key opioid overdose reversal molecule, naloxone. It accomplishes this by binding to a site on the extracellular vestibule proximal to naloxone, stabilizing a unique inactive conformation of the extracellular portions of the second and seventh transmembrane helices. The NAM perturbs orthosteric ligand kinetics in therapeutically desirable ways and works cooperatively with low doses of naloxone in vivo to inhibit morphine-induced antinociception, respiratory depression and conditioned place preference while minimizing withdrawal behaviors. Our results provide detailed structural insights into the mechanism of a negative allosteric modulator for the µOR and demonstrate how it can be exploited in vivo.
Hangke Ma; Zhaomei Sun; Lijun Xue; Xue Zhao; Jie Zhang; Huanqing Zhang; Kexin Yang; Yunjin Hu ACS Omega 2023, 8(50), 48050-48055 https://doi.org/10.1021/acsomega.3c06768AbstractOver the past three decades, DNA-encoded library (DEL) technologies have become one of the most relevant strategies for hit-finding. Recent advances in synthetic methodologies for DNA-encoded libraries rendered the increased chemical space available, but it is unknown how every variety of chemistry affects DNA’s integrity. Available assays to quantify DNA damage are restricted to electrophoresis, ligation efficiency, and mostly qPCR quantification and sequencing, which may contain predisposition and inconsistency. We developed an external standard method through LC-MS analysis to accurately quantify DNA damage throughout the chemical transformations. An assessment was conducted on on-DNA chemical reactions that are frequently employed in DEL synthesis, and these results were compared to traditional qPCR measurements. Our study provides a simple, practicable, and accurate measurement for DNA degradation during DEL synthesis. Our finding reveals substantial disagreement among the usual DNA-damaging assessment methods, which have been largely neglected so far.
Vijay Kumar Siripuram; Yashoda Krishna Sunkari; Fei Ma; Thu-Lan Nguyen; Marc Flajolet ACS Omega, 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsomega.3c06755AbstractThe present work describes a complete and reversible transformation of DNA’s properties allowing solubilization in organic solvents and subsequent chemical modifications that are otherwise not possible in an aqueous medium. Organo-soluble DNA (osDNA) moieties are generated by covalently linking a dsDNA fragment to a polyether moiety with a built-in mechanism, rendering the process perfectly reversible and fully controllable. The precise removal of the polyether moiety frees up the initial DNA fragment, unaltered, both in sequence and nature. The solubility of osDNA was confirmed in six organic solvents of decreasing polarity and six types of osDNAs. As a proof of concept, in the context of DNA-encoded library (DEL) technology, an amidation reaction was successfully performed on osDNA in 100% DMSO. The development of osDNA opens up entirely new avenues for any DNA applications that could benefit from working in nonaqueous solutions, including chemical transformations.
Inna Zilberleyb; Christine Kugel; Purvit Patel; Christine Tam; Peter L Hsu; Yvonne Franke; Kanika Bajaj Pahuja Curr. Protoc., 2023, 3(9), e872 https://doi.org/10.1002/cpz1.872AbstractThe drug discovery landscape is ever-evolving and constantly demands revolutionary technology advancements in protein expression and production laboratories. We have built a higher-throughput mid-scale semi-automated protein expression and screening platform to accelerate drug discovery research. The workflow described here enables comprehensive expression and purification screening assessment of challenging or difficult-to-express recombinant proteins in a fast and efficient manner by delivering small but sufficient amounts of high-quality proteins. The platform has been implemented for a wide range of applications that include identification of optimal constructs and chaperones for poorly expressing proteins, assessment of co-expression partners for expressing stable multiprotein complexes, and suitable buffer/additive screening for insoluble or aggregation-prone proteins. The approach allows parallel expression, purification, and characterization of 24 different samples using co-infection or a polycistronic approach in insect cells and enables parallel testing of multiple parameters to improve protein yields. The strategy has been successfully adopted for screening intracellular and secreted proteins in Escherichia coli, mammalian transient expression, and baculovirus expression vector systems. Proteins purified from this platform are used for several structural and functional screens, such as negative staining, biochemical activity assays, mass spectrometry, surface plasmon resonance, and DNA-encoded chemical library screens. In this article, for simplicity, we have focused on detailed expression and purification screening of intracellular protein complexes from insect cells. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Baculovirus generation via homologous recombination Support Protocol 1: Anti-glycoprotein 64 antibody assay Basic Protocol 2: Generation of insect cell biomass expressing target protein(s) Basic Protocol 3: Mid-scale affinity purification Support Protocol 2: Automated method for affinity purification on Hamilton STAR Basic Protocol 4: Size exclusion chromatography Support Protocol 3: Chromeleon 7 operation on Vanquish Duo.
Miles McGibbon; Steven Shave; Jie Dong; Yumiao Gao; Douglas R Houston; Jiancong Xie; Yuedong Yang; Philippe Schwaller; Vincent Blay Author Notes Brief. Bioinform., 2024, 25(1), 1–13 https://doi.org/10.1093/bib/bbad422AbstractWithin drug discovery, the goal of AI scientists and cheminformaticians is to help identify molecular starting points that will develop into safe and efficacious drugs while reducing costs, time and failure rates. To achieve this goal, it is crucial to represent molecules in a digital format that makes them machine-readable and facilitates the accurate prediction of properties that drive decision-making. Over the years, molecular representations have evolved from intuitive and human-readable formats to bespoke numerical descriptors and fingerprints, and now to learned representations that capture patterns and salient features across vast chemical spaces. Among these, sequence-based and graph-based representations of small molecules have become highly popular. However, each approach has strengths and weaknesses across dimensions such as generality, computational cost, inversibility for generative applications and interpretability, which can be critical in informing practitioners’ decisions. As the drug discovery landscape evolves, opportunities for innovation continue to emerge. These include the creation of molecular representations for high-value, low-data regimes, the distillation of broader biological and chemical knowledge into novel learned representations and the modeling of up-and-coming therapeutic modalities.
Anna Pasieka; Eleonora Diamanti; Elisa Uliassi; Maria Laura Bolognesi ChemMedChem, 2023, e202300422 https://doi.org/10.1002/cmdc.202300422AbstractClick chemistry is universally recognized as a powerful strategy for the fast and precise assembly of diverse building blocks. Targeted Protein Degradation (TPD) is a new therapeutic modality based on heterobifunctional small-molecule degraders that provides new opportunities to medicinal chemists dealing with undruggable targets and incurable diseases. Here, we highlight how very recently the TPD field and that of click chemistry have merged, opening up the possibility for fine-tuning the properties of a degrader, chemically assembled through a “click” synthesis. By reviewing concrete examples, we want to provide the reader with the insight that the application of click and bioorthogonal chemistry in the TDP field may be a winning combination.
Geoffrey A. Holdgate; Catherine Bardelle; Sophia K. Berry; Alice Lanne; Maria Emanuela Cuomo SLAS Discov., 2023 https://doi.org/10.1016/j.slasd.2023.12.008AbstractMolecular glues are small molecules, typically smaller than PROTACs, and usually with improved physicochemical properties that aim to stabilise the interaction between two proteins. Most often this approach is used to improve or induce an interaction between the target and an E3 ligase, but other interactions which stabilise interactions to increase activity or to inhibit binding to a natural effector have also been demonstrated. This review will describe the effects of induced proximity, discuss current methods used to identify molecular glues and introduce approaches that could be adapted for molecular glue screening.
Priscila M. S. Castanha; Patrick J. McEnaney; Yongseok Park; Anthea Bouwer; Elton J. F. Chaves; Roberto D. Lins; Nicholas G. Paciaroni; Paige Dickson; Graham Carlson; Marli T. Cordeiro; Tereza Magalhaes; Jodi Craigo; Ernesto T. A. Marques; Thomas Kodadek; Donald S. Burke Proc. Natl. Acad. Sci. USA, 2024, 121(21), e2312755121 https://doi.org/10.1073/pnas.2312755121AbstractSignificance Zika virus (ZIKV) remains a global health threat of high priority. ZIKV is closely related to the dengue viruses, and its spread in dengue-endemic areas poses significant challenges to the development of virus-specific diagnostic tools and effective vaccines. We screened combinatorial libraries of small synthetic molecules to efficiently identify a nonbiological molecule “CZV1-1” that binds specifically to IgG present in serum from Zika-immune persons, but not in serum from dengue-immune persons. CZV1-1 mimics a major Zika-neutralizing envelope epitope and can serve as a biomarker for evidence of prior Zika infection in flavivirus-endemic areas. This approach can be used to find small-molecule mimics of important epitopes for a wide range of other infectious and noninfectious diseases. Abstract Antigenic similarities between Zika virus (ZIKV) and other flaviviruses pose challenges to the development of virus-specific diagnostic tools and effective vaccines. Starting with a DNA-encoded one-bead-one-compound combinatorial library of 508,032 synthetic, non-natural oligomers, we selected and characterized small molecules that mimic ZIKV epitopes. High-throughput fluorescence-activated cell sorter-based bead screening was used to select molecules that bound IgG from ZIKV-immune but not from dengue-immune sera. Deep sequencing of the DNA from the “Zika-only” beads identified 40 candidate molecular structures. A lead candidate small molecule “CZV1-1” was selected that correctly identifies serum specimens from Zika-experienced patients with good sensitivity and specificity (85.3% and 98.4%, respectively). Binding competition studies of purified anti-CZV1-1 IgG against known ZIKV-specific monoclonal antibodies (mAbs) showed that CZV1-1 mimics a nonlinear, neutralizing conformational epitope in the domain III of the ZIKV envelope. Purified anti-CZV1-1 IgG neutralized infection of ZIKV in cell cultures with potencies comparable to highly specific ZIKV-neutralizing mAbs. This study demonstrates an innovative approach for identification of synthetic non-natural molecular mimics of conformational virus epitopes. Such molecular mimics may have value in the development of accurate diagnostic assays for Zika, as well as for other viruses.
Fangzhou Zhao; Martin Frandsen; Sabrina Capodaglio; Hanadi F. Sleiman J. Am. Chem. Soc., 2024, 146(3), 1946–1956 https://doi.org/10.1021/jacs.3c08984AbstractThe design of new protein structures is challenging due to their vast sequence space and the complexity of protein folding. Here, we report a new modular DNA-templated strategy to construct protein mimics. We achieve the spatial control of multiple peptide units by conjugation with DNA and hybridization to a branched DNA trimer template followed by covalent stapling of the preorganized peptides into a single unit. A library of protein mimics with different lengths, sequences, and heptad registers has been efficiently constructed. DNA-templated protein mimics show an α-helix or coiled-coil motif formation even when they are constructed from weakly interacting peptide units. Their attached DNA handles can be used to exert dynamic control over the protein mimics’ secondary and tertiary structures. This modular strategy will facilitate the development of DNA-encoded protein libraries for the rapid discovery of new therapeutics, enzymes, and antibody mimics.
Shaozhao Qin; Lijian Feng; Qingyi Zhao; Ziqin Yan; Xilin Lyu; Kaige Li; Baiyang Mu; Yujie Chen; Weiwei Lu; Chao Wang; Yanrui Suo; Jinfeng Yue; Mengqing Cui; Yingjie Li; Yujun Zhao; Zhiqiang Duan; Jidong Zhu; Xiaojie Lu J. Med. Chem., 2024, 67(2), 1079–1092 https://doi.org/10.1021/acs.jmedchem.3c01463AbstractThe DNA-encoded library (DEL) is a powerful hit generation tool for chemical biology and drug discovery; however, the optimization of DEL hits remained a daunting challenge for the medicinal chemistry community. In this study, hit compounds targeting the WIN binding domain of WDR5 were discovered by the initial three-cycle linear DEL selection, and their potency was further enhanced by a cascade DEL selection from the focused DEL designed based on the original first run DEL hits. As expected, these new compounds from the second run of focused DEL were more potent WDR5 inhibitors in the protein binding assay confirmed by the off-DNA synthesis. Interestingly, selected inhibitors exhibited good antiproliferative activity in two human acute leukemia cell lines. Taken together, this new cascade DEL selection strategy may have tremendous potential for finding high-affinity leads against WDR5 and provide opportunities to explore and optimize inhibitors for other targets.
Xiaocan Su; Wenxia Shi; Yue Zhang; Wenjie Xiao; Jiaying Xue; Xinhai Zhu; Ming Yan; Xuejing Zhang Adv. Synth. Catal., 2023 https://doi.org/10.1002/adsc.202300844AbstractAn operationally simple anhydrous carbene insertion reaction to construct DNA-encoded libraries (DEL) is reported. The developed reaction employs visible light photolysis of diazo compounds and Reversible Adsorption to Solid Support (RASS) strategy. From readily available amines, alcohols, phenols, and carboxylic acids, a variety of high-value molecules, including useful unnatural amino acids, ethers, and esters, promising clinical drug derivatives, and bioactive molecules have been constructed.
Olena S. Tokareva; Kunhua Li; Tara L. Travaline; Ty M. Thomson; Jean-Marie Swiecicki; Mahmoud Moussa; Jessica D. Ramirez; Sean Litchman; Gregory L. Verdine; John H. McGee Nat. Commun., 2023, 14, 6992 https://doi.org/10.1038/s41467-023-42395-zAbstractMolecules that induce novel interactions between proteins hold great promise for the study of biological systems and the development of therapeutics, but their discovery has been limited by the complexities of rationally designing interactions between three components, and because known binders to each protein are typically required to inform initial designs. Here, we report a general and rapid method for discovering α-helically constrained (Helicon) polypeptides that cooperatively induce the interaction between two target proteins without relying on previously known binders or an intrinsic affinity between the proteins. We show that Helicons are capable of binding every major class of E3 ubiquitin ligases, which are of great biological and therapeutic interest but remain largely intractable to targeting by small molecules. We then describe a phage-based screening method for discovering “trimerizer” Helicons, and apply it to reprogram E3s to cooperatively bind an enzyme (PPIA), a transcription factor (TEAD4), and a transcriptional coactivator (β-catenin).
Lixin Zhou; Jiamei Yang; Kuojun Zhang; Tianyu Wang; Sheng Jiang; Xiangyu Zhang J. Med. Chem. 2024, 67(2), 816–837 https://doi.org/10.1021/acs.jmedchem.3c01361AbstractCasitas B cell lymphoma-b (Cbl-b) is a vital negative regulator of TCR and BCR signaling pathways, playing a significant role in setting an appropriate threshold for the activation of T cells and controlling the tolerance of peripheral T cells via a variety of mechanisms. Overexpression of Cbl-b leads to immune hyporesponsiveness of T cells. Conversely, the deficiency of Cbl-b in T cells results in markedly increased production of IL-2, even in the lack of CD28 costimulation in vitro. And Cbl-b–/– mice spontaneously reject multifarious cancers. Therefore, Cbl-b may be associated with immune-mediated diseases, and blocking Cbl-b could be considered as a new antitumor immunotherapy strategy. In this review, the possible regulatory mechanisms and biological potential of Cbl-b for antitumor immunotherapy are summarized. Besides, the potential roles of Cbl-b in immune-mediated diseases are comprehensively discussed, with emphasis on Cbl-b immune-oncology agents in the preclinical stage and clinical trials.
Haimei Wei; Tianyang Zhang; Yangfeng Li; Gong Zhang; Yizhou Li Chem. Asian J., 2023, e202300652 https://doi.org/10.1002/asia.202300652AbstractCovalent crosslinking probes have arisen as efficient toolkits to capture and elucidate biomolecular interaction networks. Exploiting the potential of crosslinking in DNA-encoded chemical library (DEL) selection methods significantly boosted bioactive ligand discovery in complex physiological contexts. Herein, we incorporated o-nitrobenzyl alcohol (o-NBA) as a photo-activated lysine-selective crosslinker into divergent DEL formats and achieved covalent capture of ligand-target interactions featuring improved crosslinking efficiency and site specificity. In addition, covalent DEL selection was realized with the modularly designed o-NBA-functionalized mock libraries.
Kenneth Lopez Perez; Edgar Lopez Lopez; Jose Luis Medina Franco; Ramon Miranda-Quintana ChemRxiv, 2023 https://doi.org/10.26434/chemrxiv-2023-rlxjkAbstractVisualization of the chemical space is useful in many aspects of chemistry including compound library design, diversity analysis, and exploring structure-property relationships, to name a few. Examples of notable research areas where visualization of chemical space has strong applications are drug discovery and natural product research. However, the sheer volume of even comparatively small sub-sections of chemical space implies that we need to use approximations at the time of navigating through chemical space. ChemMaps is a visualization methodology that approximates the distribution of compounds in large datasets based on the selection of satellite compounds that yield a similar mapping of the whole dataset when principal component analysis on similarity matrix was performed. Here, we show how the recently proposed extended similarity indices can help to find regions that are relevant to sample satellites and reduce the amount of high dimensional data needed to describe a library’s chemical space.
Jocelyn Wang; Kohki M. Nakafuku; Jeannie Ziff; Christine F. Gelin; Hadi Gholami; Aaron A. Thompson; Nathan K. Karpowich; Luis Limon; Heather R. Coate; Kelly L. Damm-Ganamet; Amy Y. Shih; Joanna C. Grant; Marjorie Côte; Puiying A. Mak; Heather A. Pascual; Marie-Laure Rives; James P. Edwards; Jennifer D. Venable; Hariharan Venkatesan; Zhicai Shi; Samantha J. Allen; Sujata Sharma; Pei-Pei Kung; Brock T. Shireman Bioorg. Med. Chem. Lett., 2023, 129492 https://doi.org/10.1016/j.bmcl.2023.129492AbstractNatural killer group 2D (NKG2D) is a homodimeric activating immunoreceptor whose function is to detect and eliminate compromised cells upon binding to the NKG2D ligands (NKG2DL) major histocompatibility complex (MHC) molecules class I-related chain A (MICA) and B (MICB) and UL16 binding proteins (ULBP1-6). While typically present at low levels in healthy cells and tissue, NKG2DL expression can be induced by viral infection, cellular stress or transformation. Aberrant activity along the NKG2D/NKG2DL axis has been associated with autoimmune diseases due to the increased expression of NKG2D ligands in human disease tissue, making NKG2D inhibitors an attractive target for immunomodulation. Herein we describe the discovery and optimization of small molecule PPI (protein–protein interaction) inhibitors of NKG2D/NKG2DL. Rapid SAR was guided by structure-based drug design and accomplished by iterative singleton and parallel medicinal chemistry synthesis. These efforts resulted in the identification of several potent analogs (14, 21, 30, 45) with functional activity and improved LLE.
Mohammed Sharique; Bianca Matsuo; Albert Granados; Saegun Kim; Mahwish Arshad; Hyunjung Oh; Victoria E. Wu; Minxue Huang; Adam Csakai; Lisa A. Marcaurelle; Gary A. Molander Chem. Sci., 2023, 14(48), 14193-14199 https://doi.org/10.1039/D3SC03731BAbstractThe emergence of DNA-encoded library (DEL) technology has provided a considerable advantage to the pharmaceutical industry in the pursuit of discovering novel therapeutic candidates for their drug development initiatives. This combinatorial technique not only offers a more economical, spatially efficient, and time-saving alternative to the existing ligand discovery methods, but also enables the exploration of additional chemical space by utilizing novel DNA-compatible synthetic transformations to leverage multifunctional building blocks from readily available substructures. In this report, a decarboxylative-based hydroalkylation of DNA-conjugated N-vinyl heterocycles enabled by single-electron transfer (SET) and subsequent hydrogen atom transfer through electron-donor/electron-acceptor (EDA) complex activation is detailed. The simplicity and robustness of this method permits inclusion of a broad array of alkyl radical precursors and DNA-tethered nitrogenous heterocyles to generate medicinally relevant substituted heterocycles with pendant functional groups. Moreover, a successful telescoped route provides the opportunity to access a broad range of intricate structural scaffolds by employing basic carboxylic acid feedstocks.
Bianca Matsuo; Saegun Kim; Scott T. Shreiber; Guillaume Levitre; Longbo Li; Erika A. Crane; Edward J. McClain; Eric A. Voight; Gary A. Molander Chem. Commun., 2023, 59(73), 10964-10967 https://doi.org/10.1039/D3CC03421FAbstractAzaspiro[3.3]heptanes are valuable synthetic targets for drug discovery programs. The challenges associated with the preparation and diversification of this moiety as compared to other small, saturated rings have led to limited applications of compounds containing this spirocycle. In this regard, important advances in the field of synthetic photochemistry have exploited the biradical nature of the triplet excited state of 2-isoxazoline-3-carboxylates, engaging these species in intermolecular coupling reactions under visible light irradiation. As a continuation of our program preparing F(sp3)-rich, structurally complex molecules for DNA-encoded library technology (DELT) applications via photocatalysis, we disclose herein the incorporation of unique and densely functionalized 2-oxa-1-azabicyclo[3.2.0]heptanes via [2+2] cycloaddition energy transfer sensitization, providing access to an unexplored library of azaspiro compounds, many of which include additional synthetic handles important for further functionalization of the DNA-conjugated products and for library production.
Hyemi Lee; Jiho Nam; Hahyeong Jang; Young-Sik Park; Min-Hee Son; In-Hyun Lee; Seong-il Eyun; Jimin Jeon; Siyoung Yang Biomed. Pharmacother., 2023, 166, 115426 https://doi.org/10.1016/j.biopha.2023.115426AbstractOsteoarthritis (OA) is induced by matrix degradation and inflammation mediated by bromo-domain-containing protein 4 (BRD4)-dependent catabolic factors. BRD4 acts as both a transcriptional regulator and an epigenetic reader. BBC0901 was identified as an inhibitor of BRD4 using a DNA-encoded library screening system. We aimed to demonstrate the effects of BBC0901 on OA pathogenesis by in vitro, ex vivo, and in vivo analyses. BBC0901 inhibited the expression of catabolic factors that degrade cartilage without significantly affecting the viability of mouse articular chondrocytes. Additionally, ex vivo experiments under conditions mimicking OA showed that BBC0901 suppressed extracellular matrix degradation. RNA sequencing analysis of gene expression patterns showed that BBC0901 inhibited the expression of catabolic factors, such as matrix metalloproteinases (MMPs) and cyclooxygenase (COX)2, along with reactive oxygen species (ROS) production. Furthermore, intra-articular (IA) injection of BBC0901 into the knee joint blocked osteoarthritic cartilage destruction by inhibition of MMP3, MMP13, COX2, interleukin (IL)6, and ROS production, thereby obstructing the nuclear factor kappa-light-chain-enhancer of activated B cell and mitogen activated protein kinase signaling. In conclusion, BBC0901-mediated BRD4 inhibition prevented OA development by attenuating catabolic signaling and hence, can be considered a promising IA therapeutic for OA.
Yajun Zhang; Li Lin; Sai Qiao; Xue Zhao; Ting Li; Qi Liang Int. J. Biol. Macromol., 2024, 256(1), 128206 https://doi.org/10.1016/j.ijbiomac.2023.128206AbstractThe enormous growing demand for drug candidates binding to endothelin receptor A (ETA) has made it necessary to continuously pursue new strategies for ligand screening and early evaluation. This work achieved the one-step immobilization of ETA based on the bioorthogonal chemistry between the epidermal growth factor receptor tag (EGFR-tag) and ibrutinib. Comprehensive characterizations including Western blot analysis are performed to realize the morphology, antibody/ligand recognition activity, and specificity of the immobilized ETA. Taking macitentan, ambrisentan, and bosentan as an example, we utilized the immobilized ETA to construct a thermodynamic model for the evaluation of the specific ligands binding to ETA. Using this model, we screened the potential compound NP845 from a DNA-encoded library with 10,686 members derived from natural products and calculated the association constant as (2.24 ± 0.15) × 105 M−1 at 37 °C, thereby demonstrating the good pharmacological activity of NP845. The entropy change (), enthalpy change (), and Gibbs free energy () were 1.75 J/mol·K, −31.1 kJ/mol, and −31.6 kJ/mol at 37 °C, whereby we recognized the electrostatic force was the driving force of the interaction between NP845 and ETA. In vitro cell tests proved that NP845 can downregulate the expression level of PKA, B-Raf, MEK, and ERK1 in VSMC. Our results indicated that NP845 was a potential lead compound for fighting the ailments mediated by ETA.
Evan S. O'Brien; Vipin Ashok Rangari; Amal El Daibani; Shainnel O. Eans; Haylee R. Hammond; Elizabeth White; Haoqing Wang; Yuki Shiimura; Kaavya Krishna Kumar; Qianru Jiang; Kevin Appourchaux; Weijiao Huang; Chensong Zhang; Brandon J. Kennedy; Jesper M. Mathiesen; Tao Che; Jay P. McLaughlin; Susruta Majumdar; Brian K. Kobilka Nature, 2024 https://doi.org/10.1038/s41586-024-07587-7AbstractThe µ-opioid receptor (µOR) is a well-established target for analgesia1, yet conventional opioid receptor agonists cause serious adverse effects, notably addiction and respiratory depression. These factors have contributed to the current opioid overdose epidemic driven by fentanyl2, a highly potent synthetic opioid. µOR negative allosteric modulators (NAMs) may serve as useful tools in preventing opioid overdose deaths, but promising chemical scaffolds remain elusive. Here we screened a large DNA-encoded chemical library against inactive µOR, counter-screening with active, G-protein and agonist-bound receptor to 'steer' hits towards conformationally selective modulators. We discovered a NAM compound with high and selective enrichment to inactive µOR that enhances the affinity of the key opioid overdose reversal molecule, naloxone. The NAM works cooperatively with naloxone to potently block opioid agonist signalling. Using cryogenic electron microscopy, we demonstrate that the NAM accomplishes this effect by binding a site on the extracellular vestibule in direct contact with naloxone while stabilizing a distinct inactive conformation of the extracellular portions of the second and seventh transmembrane helices. The NAM alters orthosteric ligand kinetics in therapeutically desirable ways and works cooperatively with low doses of naloxone to effectively inhibit various morphine-induced and fentanyl-induced behavioural effects in vivo while minimizing withdrawal behaviours. Our results provide detailed structural insights into the mechanism of negative allosteric modulation of the µOR and demonstrate how this can be exploited in vivo.
Pradeep Reddy Cingaram Mol. Ther. Nucleic Acids, 2023, 33, 142-143 https://doi.org/10.1016/j.omtn.2023.06.014AbstractEpigenetic dysregulation plays a crucial role in the onset and progression of cancer, positioning them as compelling targets for therapeutic intervention. In a recent issue of Molecular Therapy – Nucleic Acids, Jeong et al. describe the identification of a novel selective bromodomain and extra-terminal motif (BET) inhibitor, BBC1115, using DNA-encoded library (DEL) screening.1 The study highlights the successful application of DEL-based small-molecule compound screening coupled with intensive biological characterization to discover new chemotypes with selectivity, efficacy, and safety profiles for targeting proteins involved in epigenetic regulation in human malignancies.
William K. Weigel III; Alba L. Montoya; Raphael M. Franzini J. Chem. Inf. Model., 2023, 63(15), 4641-4653 https://doi.org/10.1021/acs.jcim.3c01008AbstractDNA-encoded libraries (DELs) are widely used in the discovery of drug candidates, and understanding their design principles is critical for accessing better libraries. Most DELs are combinatorial in nature and are synthesized by assembling sets of building blocks in specific topologies. In this study, different aspects of library topology were explored and their effect on DEL properties and chemical diversity was analyzed. We introduce a descriptor for DEL topological assignment (DELTA) and use it to examine the landscape of possible DEL topologies and their coverage in the literature. A generative topographic mapping analysis revealed that the impact of library topology on chemical space coverage is secondary to building block selection. Furthermore, it became apparent that the descriptor used to analyze chemical space dictates how structures cluster, with the effects of topology being apparent when using three-dimensional descriptors but not with common two-dimensional descriptors. This outcome points to potential challenges of attempts to predict DEL productivity based on chemical space analyses alone. While topology is rather inconsequential for defining the chemical space of encoded compounds, it greatly affects possible interactions with target proteins as illustrated in docking studies using NAD/NADP binding proteins as model receptors.
Qi Miao; Vilas D. Kadam; Ayan Mukherjee, Zhi Tan; Mingxing Teng J. Med. Chem., 2023, 66(19), 13369-13383 https://doi.org/10.1021/acs.jmedchem.3c01209AbstractChemically induced proximity-based targeted protein degradation (TPD) has become a prominent paradigm in drug discovery. With the clinical benefit demonstrated by certain small-molecule protein degraders that target the cullin-RING E3 ubiquitin ligases (CRLs), the field has proactively strategized to tackle anticipated drug resistance by harnessing additional E3 ubiquitin ligases to enrich the arsenal of this therapeutic approach. Here, we endeavor to explore the collaborative efforts involved in unlocking a broad range of CRL4DCAF for degrader drug development. Throughout the discussion, we also highlight how both conventional and innovative approaches in drug discovery can be taken to realize this objective. Moving ahead, we expect a greater allocation of resources in TPD to pursue these high-hanging fruits.
Peixiang Ma; Shuning Zhang; Qianping Huang; Yuang Gu; Zhi Zhou; Wei Hou; Wei Yi; Hongtao Xu Acta Pharmaceutica Sinica B, 2023 https://doi.org/10.1016/j.apsb.2023.10.001AbstractDNA-encoded chemical library (DEL) links the power of amplifiable genetics and the non-self-replicating chemical phenotypes, generating a diverse chemical world. In analogy with the biological world, the DEL world can evolve by using a chemical central dogma, wherein DNA replicates using the PCR reactions to amplify the genetic codes, DNA sequencing transcripts the genetic information, and DNA-compatible synthesis translates into chemical phenotypes. Importantly, DNA-compatible synthesis is the key to expanding the DEL chemical space. Besides, the evolution-driven selection system pushes the chemicals to evolve under the selective pressure, i.e., desired selection strategies. In this perspective, we summarized recent advances in expanding DEL synthetic toolbox and panning strategies, which will shed light on the drug discovery harnessing in vitro evolution of chemicals via DEL.
Andreas Varkaris; Ferran Fece de la Cruz; Elizabeth E. Martin; Bryanna L. Norden; Nicholas Chevalier; Allison M. Kehlmann; Ignaty Leshchiner; Haley Barnes; Sara Ehnstrom; Anastasia-Maria Stavridi; Xin Yuan; Janice S. Kim; Haley Ellis; Alkistis Papatheodoridi; Hakan Gunaydin; Brian P. Danysh; Laxmi Parida; Ioannis Sanidas; Yongli Ji; Kayao Lau; Gerburg M. Wulf; Aditya Bardia; Laura M. Spring; Steven J. Isakoff; Jochen K. Lennerz; Kathryn Del Vecchio; Levi Pierce; Ermira Pazolli; Gad Getz; Ryan B. Corcoran; Dejan Juric Cancer Discov., 2024, 14(2), 227-239 https://doi.org/10.1158/2159-8290.CD-23-0704AbstractPIK3CA mutations occur in ∼8% of cancers, including ∼40% of HR-positive breast cancers, where the PI3K-alpha (PI3Kα)-selective inhibitor alpelisib is FDA approved in combination with fulvestrant. Although prior studies have identified resistance mechanisms, such as PTEN loss, clinically acquired resistance to PI3Kα inhibitors remains poorly understood. Through serial liquid biopsies and rapid autopsies in 39 patients with advanced breast cancer developing acquired resistance to PI3Kα inhibitors, we observe that 50% of patients acquire genomic alterations within the PI3K pathway, including PTEN loss and activating AKT1 mutations. Notably, although secondary PIK3CA mutations were previously reported to increase sensitivity to PI3Kα inhibitors, we identified emergent secondary resistance mutations in PIK3CA that alter the inhibitor binding pocket. Some mutations had differential effects on PI3Kα-selective versus pan-PI3K inhibitors, but resistance induced by all mutations could be overcome by the novel allosteric pan-mutant-selective PI3Kα-inhibitor RLY-2608. Together, these findings provide insights to guide strategies to overcome resistance in PIK3CA-mutated cancers.In one of the largest patient cohorts analyzed to date, this study defines the clinical landscape of acquired resistance to PI3Kα inhibitors. Genomic alterations within the PI3K pathway represent a major mode of resistance and identify a novel class of secondary PIK3CA resistance mutations that can be overcome by an allosteric PI3Kα inhibitor.See related commentary by Gong and Vanhaesebroeck, p. 204.See related article by Varkaris et al., p. 240.This article is featured in Selected Articles from This Issue, p. 201
Atsushi Mikami; Shohei Mori; Takashi Osawa; Satoshi Obika Chem. Eur. J., 2023, e202301928 https://doi.org/10.1002/chem.202301928AbstractRecently, we reported that the alkynyl modification of nucleobases mitigates the toxicity of antisense oligonucleotides (ASO) while maintaining the efficacy. However, the general effect of alkynyl modifications on the duplex-forming ability of oligonucleotides (ONs) is unclear. In this study, post-synthetic nucleobase modification by Sonogashira coupling in aqueous medium was carried out to efficiently evaluate the physiological properties of various ONs with alkynyl-modified nucleobases. Although several undesired reactions, including nucleobase cyclization, were observed, various types of alkynyl-modified ONs were successfully obtained via Sonogashira coupling of ONs containing iodinated nucleobases. Evaluation of the stability of the duplex formed by the synthesized alkynyl-modified ONs showed that the alkynyl modification of pyrimidine was less tolerated than that of purine, although both the modifications occurred in the major groove of the duplex. These results can be attributed to the bond angle of the alkyne on the pyrimidine and the close proximity of the alkynyl substituents to the phosphodiester backbone. The synthetic method developed in this study may contribute to the screening of the optimal chemical modification of ASO because various alkynyl-modified ONs that are effective in reducing the toxicity of ASO can be easily synthesized by this method.
Zhaobing Ding; Yizhou Wu; Liu Liu; Bing Qi; Zuozhong Peng Org. Lett., 2023, 25(29), 5515-5519 https://doi.org/10.1021/acs.orglett.3c01986AbstractHerein we report a DNA-compatible Biginelli reaction to construct isocytosine scaffolds. This reaction utilizes a one-pot reaction of DNA-conjugated guanidines with aldehydes and methyl cyanoacetates to give isocytosine derivatives, and the method is well compatible with different types of substrates. This is the first report on the synthesis of an isocytosine backbone in the field of DNA-compatible organic synthesis. The successful development of this reaction can widen the chemical space of DELs.
Alfonso T. García-Sosa Expert Opin. Drug Discov., 2023 https://doi.org/10.1080/17460441.2023.2277342AbstractABSTRACT Introduction Modern drug discovery incorporates various tools and data, heralding the beginning of the data-driven drug design (DD) era. The distributions of chemical and physical data used for Artificial Intelligence (AI)/Machine Learning (ML) and to drive DD have thus become highly important to be understood and used effectively. Areas covered The authors perform a comprehensive exploration of the statistical distributions driving the data-intensive era of drug discovery, including Benford’s Law in AI/ML-based DD. Expert opinion As the relevance of data-driven discovery escalates, we anticipate meticulous scrutiny of datasets utilizing principles like Benford’s Law to enhance data integrity and guide efficient resource allocation and experimental planning. In this data-driven era of the pharmaceutical and medical industries, addressing critical aspects such as bias mitigation, algorithm effectiveness, data stewardship, effects, and fraud prevention are essential. Harnessing Benford’s Law and other distributions and statistical tests in DD provides a potent strategy to detect data anomalies, fill data gaps, and enhance dataset quality. Benford’s Law is a fast method for data integrity and quality of datasets, the backbone of AI/ML and other modeling approaches, proving very useful in the design process.
Noémi Csorba; Péter Ábrányi-Balogh; György M. Keserű Trends Pharmacol. Sci., 2023, 44(11), 802-816 https://doi.org/10.1016/j.tips.2023.08.014AbstractCovalent fragment approaches combine advantages of covalent binders and fragment-based drug discovery (FBDD) for target identification and validation. Although early applications focused mostly on cysteine labeling, the chemistries of available warheads that target other orthosteric and allosteric protein nucleophiles has recently been extended. The range of different warheads and labeling chemistries provide unique opportunities for screening and optimizing warheads necessary for targeting non-cysteine residues. In this review, we discuss these recently developed amino-acid-specific and promiscuous warheads, as well as emerging labeling chemistries, which includes novel transition metal catalyzed, photoactive, electroactive, and noncatalytic methodologies. We also highlight recent applications of covalent fragments for the development of molecular glues and proteolysis-targeting chimeras (PROTACs), and their utility in chemical proteomics-based target identification and validation.
Justice L. Merrifield; Edward B. Pimentel; Trenton M. Peters-Clarke; Daniel J. Nesbitt; Joshua J. Coon; Jeffrey D. Martell Bioconjugate Chem., 2023, 34(8), 1380-1386 https://doi.org/10.1021/acs.bioconjchem.3c00254AbstractAldehydes are important synthons for DNA-encoded library (DEL) construction, but the development of a DNA-compatible method for the oxidation of alcohols to aldehydes remains a significant challenge in the field of DEL chemistry. We report that a copper/TEMPO catalyst system enables the solution-phase DNA-compatible oxidation of DNA-linked primary activated alcohols to aldehydes. The semiaqueous, room-temperature reaction conditions afford oxidation of benzylic, heterobenzylic, and allylic alcohols in high yield, with DNA compatibility verified by mass spectrometry, qPCR, Sanger sequencing, and ligation assays. Subsequent transformations of the resulting aldehydes demonstrate the potential of this method for robust library diversification.
Yandan Bao; Minyan Xing; Naylor Matthew, Xiaohua Chen; Xuan Wang; Xiaojie Lu Org. Lett., 2023, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c01817AbstractWhile DNA-encoded macrocyclic libraries have gained substantial attention and several hit compounds have been identified from DNA-encoded library technology, efficient on-DNA macrocyclic methods are also required to construct DNA-linked libraries with a high degree of cyclization and DNA integrity. In this paper, we reported a set of on-DNA methodologies, including the use of an OPA-mediated three-component cyclization with native handles of amino acids and photoredox chemistries. These chemistries proceed smoothly under mild conditions in good to excellent conversions, successfully generating novel isoindole, isoindoline, indazolone, and bicyclic scaffolds.
Qingyi Zhao; Yandan Bao; Dawen Niu; Xuan Wang; Xiaojie Lu Org. Lett., 2023, 25(26), 4940-4944 https://doi.org/10.1021/acs.orglett.3c01799AbstractIn this study, we revealed two distinct S-glycosyl transformations in a DNA-encoded library (DEL)-compatible environment. The first approach involves 2-chloro-1,3-dimethylimidazolidinium chloride (DMC)-mediated S-glycosylation, which is facilitated by the coupling of unprotected sugar units with the thiol residue of the DNA-linked compounds. However, this methodology falls short of the requirement for DEL construction due to its limited substrate scope. We further investigated a photoinduced DNA-compatible S-glycosyl transformation through a radical process. In this alternative approach, allyl sugar sulfones serve as sugar donors and are conjugated to DNA-linked compounds upon irradiation with green light. Encouragingly, this on-DNA glycosyl chemistry demonstrated excellent compatibility with functional groups presented in both sugar units and peptides, affording the desired DNA-linked glycosyl derivatives with good to excellent conversions. This pioneering DNA-compatible S-glycosyl transformation represents a valuable tool, facilitating the preparation of glycosyl DELs and offering avenues for the exploration of sugar-incorporated delivery systems.
Xianfeng Li; Changyang Liu; Yuting Gao; Gong Zhang; Yangfeng Li; Yizhou Li Chem. Commun., 2023, 59, 9489-9492 https://doi.org/10.1039/D3CC02593DAbstract2-Thiobenzazole is among the privileged heterocyclic scaffolds in medicinal chemistry. Constructing such structural components in DNA-encoded libraries (DELs) may promote related bioactive hit discovery in a high-throughput fashion. Herein, we reported a DNA-compatible mild-condition synthetic methodology to efficiently forge functionalized 2-thiobenzazole scaffolds, realizing on-DNA sulfhydryl incorporation with broad substrate scope, thereby expanding the scope of 2-thiobenzazole-focused DNA-encoded chemical libraries.
Shuang Liu; Bingqi Tong; Jeremy W. Mason; Jonathan M. Ostrem; Antonin Tutter; Bruce K. Hua; Sunny A. Tang; Simone Bonazzi; Karin Briner; Frédéric Berst; Frédéric J. Zécri; Stuart L. Schreiber bioRxiv, 2023 https://doi.org/10.1101/2023.05.22.541439AbstractThe hallmark of a molecular glue is its ability to induce cooperative protein–protein interactions, leading to the formation of a ternary complex, despite weaker binding towards one or both individual proteins. Notably, the extent of cooperativity distinguishes molecular glues from bifunctional compounds, a second class of inducers of protein–protein interactions. However, apart from serendipitous discovery, there have been limited rational screening strategies for the high cooperativity exhibited by molecular glues. Here, we propose a binding-based screen of DNA-barcoded compounds on a target protein in the presence and absence of a presenter protein, using the ″presenter ratio″, the ratio of ternary enrichment to binary enrichment, as a predictive measure of cooperativity. Through this approach, we identified a range of cooperative, noncooperative, and uncooperative compounds in a single DNA-encoded library screen with bromodomain (BRD)9 and the VHL–elongin C–elongin B (VCB) complex. Our most cooperative hit compound, 13-7, exhibits micromolar binding affinity to BRD9 but nanomolar affinity for the ternary complex with BRD9 and VCB, with cooperativity comparable to classical molecular glues. This approach may enable the discovery of molecular glues for pre-selected proteins and thus facilitate the transition to a new paradigm of molecular therapeutics.
Guzel Minibaeva; Aleksandra Ivanova; Pavel Polishchuk J. Cheminform., 2023, 15(1), 102 https://doi.org/10.1186/s13321-023-00772-2AbstractDocking of large compound collections becomes an important procedure to discover new chemical entities. Screening of large sets of compounds may also occur in de novo design projects guided by molecular docking. To facilitate these processes, there is a need for automated tools capable of efficiently docking a large number of molecules using multiple computational nodes within a reasonable timeframe. These tools should also allow for easy integration of new docking programs and provide a user-friendly program interface to support the development of further approaches utilizing docking as a foundation. Currently available tools have certain limitations, such as lacking a convenient program interface or lacking support for distributed computations. In response to these limitations, we have developed a module called EasyDock. It can be deployed over a network of computational nodes using the Dask library, without requiring a specific cluster scheduler. Furthermore, we have proposed and implemented a simple model that predicts the runtime of docking experiments and applied it to minimize overall docking time. The current version of EasyDock supports popular docking programs, namely Autodock Vina, gnina, and smina. Additionally, we implemented a supplementary feature to enable docking of boron-containing compounds, which are not inherently supported by Vina and smina, and demonstrated its applicability on a set of 55 PDB protein-ligand complexes.
Ayun Luo; Nicolas Duchemin; Xiuming Wang; Hongxia Zhou; Fanming Zeng; Xue Zhao; Weina Yu; Kexin Yang; Yunjin Hu Chem. Asian J. 2023, e202300458 https://doi.org/10.1002/asia.202300458AbstractThiophene and its substituted derivatives are a highly important class of heterocyclic compounds, with noteworthy applications in pharmaceutical ingredients. In this study, we leverage the unique reactivity of alkynes to generate thiophenes on-DNA, using a cascade iodination, Cadiot-Chodkiewicz coupling and heterocyclization. This approach, tackling on-DNA thiophene synthesis for the first time, generates diverse, and unprecedented structural and chemical features, which could be significant motifs in DEL screening as molecular recognition agents for drug discovery.
Regina Pikalyova; Yuliana Zabolotna; Dragos Horvath; Gilles Marcou; Alexandre Varnek J. Chem. Inf. Model., 2023, 63(17), 5571-5582 https://doi.org/10.1021/acs.jcim.3c00719AbstractIn chemical library analysis, it may be useful to describe libraries as individual items, rather than as collections of compounds. This is particularly true for ultra-large non-cherry pickable compound mixtures, such as DNA-Encoded Libraries (DELs). In this sense, the Chemical Library Space (CLS) is useful for the management of a portfolio of libraries, just like Chemical Space (CS) helps managing a portfolio of molecules. Several possible CLSs were previously defined using vectorial library representations obtained from Generative Topographic Mapping (GTM). Given the steadily growing number of DEL designs, the CLS becomes “crowded”, and requires analysis tools beyond pairwise library comparison. Therefore, herein we investigate the cartography of CLS on meta-(µ)GTMs – “meta” to remind that these are maps of the CLS, itself based on responsibility vectors issued by regular CS GTMs. 2,5K DELs and ChEMBL (reference) were projected on the µGTM, producing landscapes of library-specific properties. These describe both inter-library similarity and intrinsic library characteristics in the same view, herewith facilitating the selection of the best project-specific libraries.
J. Miguel Mata; Edith van der Nol; Sebastian J. Pomplun J. Am. Chem. Soc., 2023, 145(34), 19129-19139 https://doi.org/10.1021/jacs.3c04899AbstractDiscovering new bioactive molecules is crucial for drug development. Finding a hit compound for a new drug target usually requires screening of millions of molecules. Affinity selection based technologies have revolutionized early hit discovery by enabling the rapid screening of libraries with millions or billions of compounds in short timeframes. In this Perspective, we describe recent technology breakthroughs that enable the screening of ultralarge synthetic peptidomimetic libraries with a barcode-free tandem mass spectrometry decoding strategy. A combination of combinatorial synthesis, affinity selection, automated de novo peptide sequencing algorithms, and advances in mass spectrometry instrumentation now enables hit discovery from synthetic libraries with over 100 million members. We provide a perspective on this powerful technology and showcase success stories featuring the discovery of high affinity binders for a number of drug targets including proteins, nucleic acids, and specific cell types. Further, we show the usage of the technology to discover synthetic peptidomimetics with specific functions and reactivity. We predict that affinity selection coupled with tandem mass spectrometry and automated de novo decoding will rapidly evolve further and become a broadly used drug discovery technology.
Vikrant Kumar; Puneeth Kumar Chunchagatta Lakshman; Thazhe Kootteri Prasad; Kavyashree Manjunath; Sneha Bairy; Akshaya S. Vasu; B. Ganavi; Subbarao Jasti; Neelagandan Kamariah Heliyon, 2024, 10(1), e23864 https://doi.org/10.1016/j.heliyon.2023.e23864AbstractTarget-based discovery of first-in-class therapeutics demands an in-depth understanding of the molecular mechanisms underlying human diseases. Precise measurements of cellular and biochemical activities are critical to gain mechanistic knowledge of biomolecules and their altered function in disease conditions. Such measurements enable the development of intervention strategies for preventing or treating diseases by modulation of desired molecular processes. Fluorescence-based techniques are routinely employed for accurate and robust measurements of in-vitro activity of molecular targets and for discovering novel chemical molecules that modulate the activity of molecular targets. In the current review, the authors focus on the applications of fluorescence-based high throughput screening (HTS) and fragment-based ligand discovery (FBLD) techniques such as fluorescence polarization (FP), Förster resonance energy transfer (FRET), fluorescence thermal shift assay (FTSA) and microscale thermophoresis (MST) for the discovery of chemical probe to exploring target’s role in disease biology and ultimately, serve as a foundation for drug discovery. Some recent advancements in these techniques for compound library screening against important classes of drug targets, such as G-protein-coupled receptors (GPCRs) and GTPases, as well as phosphorylation- and acetylation-mediated protein-protein interactions, are discussed. Overall, this review presents a landscape of how these techniques paved the way for the discovery of small-molecule modulators and biologics against these targets for therapeutic benefits.
Robin Dinter; Katharina Götte; Franziska Gronke; Leon Justen; Andreas Brunschweiger; Norbert Kockmann J. Flow Chem., 2023, https://doi.org/10.1007/s41981-023-00282-0AbstractAn automated flow chemistry platform for DNA-encoded library (DEL) technologies requires the integration of a purification process for DNA-tagged substrates. It facilitates the development of further DEL reactions, building block rehearsal, and library synthesis. Therefore, a recently developed, manual affinity-based batch purification process for DNA-tagged substrates based on dispersive solid-phase extraction (DSPE) was transferred to automated flow chemistry using tailored 3D-printed microfluidic devices and open-source lab automation equipment. The immobilization and purification steps use Watson–Crick base pairing for a compound-encoding single-stranded DNA, which allows for the thorough removal of impurities and contaminations by washing steps and operationally simple recovery of the purified DNA-encoded compounds. This work optimized the annealing step for flow incubation and DNA purification was accomplished by flow DSPE washing/elution steps. The manually performed batch affinity-based purification process was compared with the microfluidic process by determining qualitative and quantitative DNA recovery parameters. It aimed at comparing batch and flow purification processes with regard to DNA recovery and purity to benefit from the high potential for automation, precise process control, and higher information density of the microfluidic purification process for DNA-tagged substrates. Manual operations were minimized by applying an automation strategy to demonstrate the potential for integrating the microfluidic affinity-based purification process for DNA-tagged substrates into an automated DNA-encoded flow chemistry platform.
Lulu Winer; Leila Motiei; David Margulies Bioconjugate Chem., 2023, 34(9), 1509-1522 https://doi.org/10.1021/acs.bioconjchem.3c00203AbstractThe unfathomable role that fluorescence detection plays in the life sciences has prompted the development of countless fluorescent labels, sensors, and analytical techniques that can be used to detect and image proteins or investigate their properties. Motivated by the demand for simple-to-produce, modular, and versatile fluorescent tools to study proteins, many research groups have harnessed the advantages of oligodeoxynucleotides (ODNs) for scaffolding such probes. Tight control over the valency and position of protein binders and fluorescent dyes decorating the polynucleotide chain and the ability to predict molecular architectures through self-assembly, inherent solubility, and stability are, in a nutshell, the important properties of DNA probes. This paper reviews the progress in developing DNA-based, fluorescent sensors or labels that navigate toward their protein targets through small-molecule (SM) or peptide ligands. By describing the design, operating principles, and applications of such systems, we aim to highlight the versatility and modularity of this approach and the ability to use ODN-SM or ODN-peptide conjugates for various applications such as protein modification, labeling, and imaging, as well as for biomarker detection, protein surface characterization, and the investigation of multivalency.
Oleksandr V. Oksiuta; Alexander E. Pashenko; Radomyr V. Smalii; Dmitry M. Volochnyuk; Serhii V. Ryabukhin J. Org. Pharm. Chem., 2023, 21, 3-19 https://doi.org/10.24959/ophcj.23.275133AbstractAim. DNA-encoded libraries technologies (DELT) are gradually becoming an important part of standard drug discovery toolbox. DELT is looking to find its place between classic low-molecular-weight drug candidates on the one hand, and high-molecular-weight antibodies and peptides on the other hand. On its natural path to overcoming the “childhood diseases” typical for every novel technology, DELT has reached a point where the chemical diversity of DNA-encoded libraries (DELs) becomes an important factor to look out for. In this paper, we aim to take a closer look at the chemical diversity of DELs in their present state and find the ways to improve it. Results and discussion. We have identified the DEL-viable building blocks from the Enamine Ltd. stock collection, as well as from Chemspace Ltd. virtual collection, using the SMARTS set, which takes into account all the necessary structural restrictions. Using modern cheminformatics tools, such as Synt-On, we have analyzed the scaffold diversity of both stock and virtual core bi- and tri-functional building blocks (BBs) suitable for DNA-tolerant reactions. The identification of scaffolds from the most recently published on-DNA heterocyclization reactions and analysis of their inclusion into the existing BBs space have shown that novel DNA-tolerant heterocyclizations are extremely useful for expanding chemical diversity in DEL technologies. Conclusions. The analysis performed allowed us to recognize which functional groups should be prioritized as the most impactful when the new BBs are designed. It is also made clear that the development of new DNA-tolerant reactions, including heterocyclizations, have a significant potential to further expand DEL molecular diversity.
R. Madison Green; Paul R. Thompson Curr. Opin. Chem. Biol., 2023, 75, 102313 https://doi.org/10.1016/j.cbpa.2023.102313AbstractProtein citrullination is a post-translational modification of arginine that controls a diverse array of cellular processes, including gene regulation, protein stability, and neutrophil extracellular trap (NET) formation. Histone citrullination promotes chromatin decondensation and NET formation, a pro-inflammatory form of cell death that is aberrantly increased in numerous immune disorders. This review will provide insights into NETosis and how this novel form of cell death contributes to inflammatory diseases, with a particular emphasis on its role in thrombosis. We will also discuss recent efforts to develop PAD-specific inhibitors.
Alexander A. Peterson; David R. Liu Nat. Rev. Drug Discov., 2023, 22(9), 699-722 https://doi.org/10.1038/s41573-023-00713-6AbstractThe development of bioactive small molecules as probes or drug candidates requires discovery platforms that enable access to chemical diversity and can quickly reveal new ligands for a target of interest. Within the past 15 years, DNA-encoded library (DEL) technology has matured into a widely used platform for small-molecule discovery, yielding a wide variety of bioactive ligands for many therapeutically relevant targets. DELs offer many advantages compared with traditional screening methods, including efficiency of screening, easily multiplexed targets and library selections, minimized resources needed to evaluate an entire DEL and large library sizes. This Review provides accounts of recently described small molecules discovered from DELs, including their initial identification, optimization and validation of biological properties including suitability for clinical applications.
Yiting Wang; Xianfu Fang; Huilin Liao; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett., 2023, 25(24), 4473-4477 https://doi.org/10.1021/acs.orglett.3c01482AbstractThiazolidione, conferring drug-like properties, is an important heterocycle that widely exists in medicinally relevant molecules. In this work, by efficiently assembling various DNA-tagged primary amines, abundant aryl isothiocyanates, and ethyl bromoacetate, we present a DNA-compatible three-component annulation to generate a 2-iminothiazolidin-4-one scaffold, which was further decorated via Knoevenagel condensation by employing (hetero)aryl and alkyl aldehydes. These thiazolidione derivatives should find broad use in focused DNA-encoded library construction.
Michael Mortensen; Marco Bertolini; Jacqueline Mock; Jörg Scheuermann; Sebastian Oehler Bioconjugate Chem., 2023, 34(8), 1374-1379 https://doi.org/10.1021/acs.bioconjchem.3c00194AbstractThe targeted delivery of bioactive proteins, such as cytokines, for cancer immunotherapy approaches mostly relies on antibodies or antibody fragments. However, fusion proteins may display low tissue penetration due to a large molecular size. Small molecule ligands with high affinity toward tumor-associated antigens provide a promising alternative for the selective delivery of cytokines to tumor lesions. We developed a one-pot procedure for the site-specific thiazolidine formation between an aldehyde bearing small molecule and the in situ generated N-terminal cysteine of a bioactive protein. Thereby, neoleukin-2/15 (Neo-2/15), a computationally engineered interleukin-2 and -15 mimic, was chemically conjugated to acetazolamide plus, a potent carbonic anhydrase IX (CAIX) ligand. The conjugate retained the biological activity of Neo-2/15 and revealed its ability to accumulate in renal cell carcinoma (SK-RC-52) xenografts upon systemic intravenous administration. The results highlight the potential of small molecule targeting moieties to drive the accumulation of a protein cargo to the respective disease site while conserving the small construct size.
Xueyi Yang; Jessica L. Childs-Disney; Brian M. Paegel; Matthew D. Disney Isr. J. Chem., 2023, e202300073 https://doi.org/10.1002/ijch.202300073AbstractThe functional roles of structured RNAs in the regulation of biological processes, and hence RNA's potential as an effective therapeutic target, have only recently been appreciated. Robust and high-throughput methods that identify potent RNA ligands are critical to the development of chemical probes and therapeutics. DNA-encoded libraries (DEL) technology has emerged as a powerful tool for protein ligand discovery, and its ability to generate large, custom-tailored, and novel chemical space offers unprecedented opportunities to discover the rules of RNA ligand design. In this review, we discuss the basic principles of DEL selection, current progress on the application of DEL to RNA targets, and the outlook of targeting RNA by DEL.
Takashi Osawa; Satoshi Obika Chem. Pharm. Bull., 2024, 72, 143-148 https://doi.org/10.1248/cpb.c23-00295AbstractDNA-encoded libraries (DELs) are attracting attention as a screening tool in the early stages of drug discovery. In the development of DELs, drug candidate compounds are chemically synthesized on barcode DNA. Therefore, it is important to perform the synthesis under mild conditions so as to not damage the DNA. On the other hand, coumarins are gaining increasing research focus not only because they possess excellent fluorescence properties, but also because many medicines contain a coumarin skeleton. Among the various reactions developed for the synthesis of coumarins thus far, Knoevenagel condensation followed by intramolecular cyclization under mild conditions can yield coumarins. In this study, we developed a new synthetic method for preparing a coumarin-conjugated oligonucleotide library via Knoevenagel condensation. The results showed that coumarins substituted at the 5-, 6-, 7-, or 8-positions could be constructed on DNA to afford a total of 26 coumarin-conjugated DNAs. Moreover, this method was compatible with enzymatic ligation, demonstrating its utility in DEL synthesis. The developed strategy for the construction of coumarin scaffolds based on Knoevenagel condensation may contribute to the use of DELs in drug discovery and medicinal chemistry.
Denise B. Catacutan; Jeremie Alexander; Autumn Arnold; Jonathan M. Stokes Nat. Chem. Biol., 2024, 20, 960-973 https://doi.org/10.1038/s41589-024-01679-1AbstractDrug-discovery and drug-development endeavors are laborious, costly and time consuming. These programs can take upward of 12 years and cost US $2.5 billion, with a failure rate of more than 90%. Machine learning (ML) presents an opportunity to improve the drug-discovery process. Indeed, with the growing abundance of public and private large-scale biological and chemical datasets, ML techniques are becoming well positioned as useful tools that can augment the traditional drug-development process. In this Perspective, we discuss the integration of algorithmic methods throughout the preclinical phases of drug discovery. Specifically, we highlight an array of ML-based efforts, across diverse disease areas, to accelerate initial hit discovery, mechanism-of-action (MOA) elucidation and chemical property optimization. With advances in the application of ML across diverse therapeutic areas, we posit that fully ML-integrated drug-discovery pipelines will define the future of drug-development programs.
Patrick R. Fitzgerald; Wesley G. Cochrane; Brian M. Paegel ACS Med. Chem. Lett., 2023, 14(9), 1295-1303 https://doi.org/10.1021/acsmedchemlett.3c00159AbstractDose–response, or “conforming” behavior, increases confidence in a screening hit’s authenticity. Here, we demonstrate dose–response solid-phase DNA-encoded library (DEL) screening. Compound dose in microfluidic droplets is modulated via the UV intensity of photocleavage from DEL beads. A 55,296-member DEL was screened at different UV intensities against model enzyme drug targets factor Xa (FXa) and autotaxin (ATX). Both screens yielded photochemical dose-dependent hit rates (FXa hit rates of 0.08/0.05% at 100/30% UV exposure; ATX hit rates of 0.24/0.08% at 100/20% UV exposure). FXa hits contained structures reflective of FXa inhibitors and four hits inhibited FXa (IC50 = 4.2 ± 0.1, 7.4 ± 0.3, 9.0 ± 0.3, and 19 ± 2 μM.) The top ATX hits (two dihydrobenzamidazolones and a tetrahydroisoquinoline) were validated as inhibitors (IC50 = 7 ± 2, 13 ± 2, and 1 ± 0.3 μM). Photochemical dose–response DEL screening data prioritized hits for synthesis, the rate-limiting step in DEL lead identification.
Yujin An; Juyeon Lee; Hyewon Seo; Seri Bae; Jihee Kang; Jieon Lee; Jinwoo Kim; Myung Hee Nam; Minsoo Song; Gil Tae Hwang Org. Lett., 2023, 25(24), 4445-4450 https://doi.org/10.1021/acs.orglett.3c01366AbstractThis study presents a DNA-compatible synthesis of diverse 5-arylimidazo[1,2-a]pyridin-3-amine derivatives using the Suzuki–Miyaura reaction, followed by a Groebke–Blackburn–Bienaymé (GBB) reaction. The GBB reaction demonstrates a wide substrate scope, mild one-pot reaction conditions, and compatibility with subsequent enzymatic ligation, highlighting its potential in DNA-encoded library technology.
Chris Zhang; Mary Pitman; Anjali Dixit; Sumudu Leelananda; Henri Palacci; Meghan Lawler; Svetlana Belyanskaya; LaShadric Grady; Joe Franklin; Nicolas Tilmans; David L. Mobley J. Chem. Inf. Model., 2023, 63(16), 5120-5132 https://doi.org/10.1021/acs.jcim.3c00588AbstractDNA-encoded libraries (DELs) provide the means to make and screen millions of diverse compounds against a target of interest in a single experiment. However, despite producing large volumes of binding data at a relatively low cost, the DEL selection process is susceptible to noise, necessitating computational follow-up to increase signal-to-noise ratios. In this work, we present a set of informatics tools to employ data from prior DEL screen(s) to gain information about which building blocks are most likely to be productive when designing new DELs for the same target. We demonstrate that similar building blocks have similar probabilities of forming compounds that bind. We then build a model from the inference that the combined behavior of individual building blocks is predictive of whether an overall compound binds. We illustrate our approach on a set of three-cycle OpenDEL libraries screened against soluble epoxide hydrolase (sEH) and report performance of more than an order of magnitude greater than random guessing on a holdout set, demonstrating that our model can serve as a baseline for comparison against other machine learning models on DEL data. Lastly, we provide a discussion on how we believe this informatics workflow could be applied to benefit researchers in their specific DEL campaigns.
Yasser Tabana; Dinesh Babu; Richard Fahlman; Arno G. Siraki; Khaled Barakat BMC Biotechnol., 2023, 23, 44 https://doi.org/10.1186/s12896-023-00815-4AbstractTarget identification is an essential part of the drug discovery and development process, and its efficacy plays a crucial role in the success of any given therapy. Although protein target identification research can be challenging, two main approaches can help researchers make significant discoveries: affinity-based pull-down and label-free methods. Affinity-based pull-down methods use small molecules conjugated with tags to selectively isolate target proteins, while label-free methods utilize small molecules in their natural state to identify targets. Target identification strategy selection is essential to the success of any drug discovery process and must be carefully considered when determining how to best pursue a specific project. This paper provides an overview of the current target identification approaches in drug discovery related to experimental biological assays, focusing primarily on affinity-based pull-down and label-free approaches, and discusses their main limitations and advantages.
Adam D. Moorhouse; Joshua A. Homer; John E. Moses Chem, 2023, 9(8), 2063-2077 https://doi.org/10.1016/j.chempr.2023.03.017AbstractThe bigger picture Challenges and opportunities: • Click chemistry drives unrivaled opportunities for creating functional molecules. We look to an exciting future where discovery and process chemistry merge; that is, where click reactions and associated functional molecules find mass-market deployment. However, this will largely depend on the availability and accessibility of suitable screens, the development of new assays, and further innovation to ensure that the desirable properties from the myriad of candidate molecules generated by click chemistry are realized. • Numerous developments, such as click-activated drugs, inverse drug discovery, and “sleeping beauty” probes, have the potential to form platform technologies. No doubt, other yet-to-be-discovered methods for drug discovery and chemical biology will change the direction of these endeavors. • As the application of artificial intelligence in chemistry makes gains, we advocate a case for click chemistry principles to expedite the realization of in silico discoveries. The impact of click chemistry was recently recognized with the 2022 Nobel Prize in Chemistry. The breadth of areas where click chemistry has accelerated discovery is extensive. In one of the most written-about subjects in Chemistry over recent years, this short perspective zones in on a small fragment of what we consider to be some of the most critical developments in synthetic chemistry, which have expanded access to the click chemistry toolbox. In addition, we touch upon areas within medicinal chemistry and novel approaches to drug discovery enabled by click chemistry, where we believe there is untapped potential for biological functions to be found and exploited.
Gergely Takács; Dávid Havasi; Márk Sándor; Zsolt Dohánics; György T. Balogh; Róbert Kiss ACS Med. Chem. Lett., 2023, 14(9), 1188-1197 https://doi.org/10.1021/acsmedchemlett.3c00146AbstractThe advancement of in silico technologies such as library enumeration and synthetic feasibility prediction has made drug discovery pipelines rely more and more on virtual libraries, which provide a significantly larger pool of compounds than in-stock supplier catalogs. Virtual libraries from external sources, however, may be associated with long delivery time and high cost. In this study, we present a Do-It-Yourself (DIY) combinatorial chemistry library containing over 14 million almost completely novel products built from 1000 low-cost building blocks based on robust reactions frequently applied at medicinal chemistry laboratories. The applicability of the DIY library for various drug discovery approaches is demonstrated by extensive physicochemical property, structural diversity profiling, and the generation of focused libraries. We found that internally built DIY chemical libraries present a viable alternative of external virtual catalogs by providing access to a large number of low-cost and quickly accessible potential chemical starting points for drug discovery.
Asher Mullard Nat. Rev. Drug Discov., 2023, 22, 347-348 https://doi.org/10.1038/d41573-023-00064-2AbstractBob Lefkowitz has four quotes taped to the walls of his office at Duke University. “Life is filled with golden opportunities cleverly disguised as insoluble proteins,” reads the biochemist, before correcting the last words to “insolvable problems”.
Yuchen Tang; Xingxing Zheng; Tingjuan Gao Research, 2023, 6, 0136 https://doi.org/10.34133/research.0136AbstractHigh-throughput assays play an important role in the fields of drug discovery, genetic analysis and clinical diagnostics. Although super-capacity coding strategies may facilitate labeling and detecting large numbers of targets in a single assay, practically the constructed large-capacity codes have to be decoded with complicated procedures or are lack of survivability in the required reaction conditions. This challenge results in either inaccurate or insufficient decoding outputs. Here, we identified chemical-resistant Raman compounds to build a 10^3 combinatorial coding system for the high-throughput screening of cell-targeting ligands from a focused 8-mer cyclic peptide library. The accurate in-situ decoding results proved the signal, synthetic and functional orthogonality for this Raman coding strategy. The orthogonal Raman codes allowed for a rapid identification of 63 positive hits at one time, evidencing a high-throughput-out capability in the screening process. We anticipate this orthogonal Raman coding strategy being generalized to enable efficient high-throughput-out screening of more useful ligands for cell targeting and drug discovery.
Kun Zhou; Daquan Chen ChemBioChem, 2023, e202300301 https://doi.org/10.1002/cbic.202300301AbstractThe COVID-19 pandemic has brought a widespread influence on the world, especially in the face of sudden coronavirus infections, and there is still an urgent need for specific small molecule therapies to cope with possible future pandemics. The pathogen responsible for this pandemic is SARS-CoV-2, and understanding its structure and lifecycle is beneficial for designing specific drugs of treatment for COVID-19. The main protease (Mpro) which has conservative and specific advantages is essential for viral replication and transcription. It is regarded as one of the most potential targets for anti-SARS-CoV-2 drug development. This review introduces the popular knowledge of SARS-CoV-2 in drug development and lists a series of design principles and relevant activities of advanced Mpro inhibitors.
Navid J. Ayon Metabolites, 2023, 13(5), 625 https://doi.org/10.3390/metabo13050625AbstractDue to the continued emergence of resistance and a lack of new and promising antibiotics, bacterial infection has become a major public threat. High-throughput screening (HTS) allows rapid screening of a large collection of molecules for bioactivity testing and holds promise in antibacterial drug discovery. More than 50% of the antibiotics that are currently available on the market are derived from natural products. However, with the easily discoverable antibiotics being found, finding new antibiotics from natural sources has seen limited success. Finding new natural sources for antibacterial activity testing has also proven to be challenging. In addition to exploring new sources of natural products and synthetic biology, omics technology helped to study the biosynthetic machinery of existing natural sources enabling the construction of unnatural synthesizers of bioactive molecules and the identification of molecular targets of antibacterial agents. On the other hand, newer and smarter strategies have been continuously pursued to screen synthetic molecule libraries for new antibiotics and new druggable targets. Biomimetic conditions are explored to mimic the real infection model to better study the ligand–target interaction to enable the designing of more effective antibacterial drugs. This narrative review describes various traditional and contemporaneous approaches of high-throughput screening of natural products and synthetic molecule libraries for antibacterial drug discovery. It further discusses critical factors for HTS assay design, makes a general recommendation, and discusses possible alternatives to traditional HTS of natural products and synthetic molecule libraries for antibacterial drug discovery.
Jing Chai; Christopher C. Arico-Muendel; Yun Ding; Michael P. Pollastri; Sarah Scott; Mark A. Mantell; Gang Yao Bioconjugate Chem. 2023, 34(6), 988-993 https://doi.org/10.1021/acs.bioconjchem.3c00159AbstractMacrocycles occupy chemical space “beyond the rule of five”. They bridge traditional bioactive small molecule drugs and macromolecules and have the potential to modulate challenging targets such as PPI or proteases. Here we report an on-DNA macrocyclization reaction utilizing intramolecular benzimidazole formation. A 129-million-member macrocyclic library composed of a privileged benzimidazole core, a dipeptide sequence (natural or non-natural), and linkers of varying length and flexibility was designed and synthesized.
Regina Pikalyova; Yuliana Zabolotna; Dragos Horvath; Gilles Marcou; Alexandre Varnek J. Chem. Inf. Model., 2023, 63(13), 4042-4055 https://doi.org/10.1021/acs.jcim.3c00520AbstractThe development of DNA-encoded library (DEL) technology introduced new challenges for the analysis of chemical libraries. It is often useful to consider a chemical library as a stand-alone chemoinformatic object─represented both as a collection of independent molecules, and yet an individual entity─in particular, when they are inseparable mixtures, like DELs. Herein, we introduce the concept of chemical library space (CLS), in which resident items are individual chemical libraries. We define and compare four vectorial library representations obtained using generative topographic mapping. These allow for an effective comparison of libraries, with the ability to tune and chemically interpret the similarity relationships. In particular, property-tuned CLS encodings enable to simultaneously compare libraries with respect to both property and chemotype distributions. We apply the various CLS encodings for the selection problem of DELs that optimally “match” a reference collection (here ChEMBL28), showing how the choice of the CLS descriptors may help to fine-tune the “matching” (overlap) criteria. Hence, the proposed CLS may represent a new efficient way for polyvalent analysis of thousands of chemical libraries. Selection of an easily accessible compound collection for drug discovery, as a substitute for a difficult to produce reference library, can be tuned for either primary or target-focused screening, also considering property distributions of compounds. Alternatively, selection of libraries covering novel regions of the chemical space with respect to a reference compound subspace may serve for library portfolio enrichment.
Malte Korn; Christiane Ehrt; Fiorella Ruggiu; Marcus Gastreich; Matthias Rarey Curr. Opin. Struct. Biol., 2023, 102578 https://doi.org/10.1016/j.sbi.2023.102578AbstractThe size of actionable chemical spaces is surging, owing to a variety of novel techniques, both computational and experimental. As a consequence, novel molecular matter is now at our fingertips that cannot and should not be neglected in early-phase drug discovery. Huge, combinatorial, make-on-demand chemical spaces with high probability of synthetic success rise exponentially in content, generative machine learning models go hand in hand with synthesis prediction, and DNA-encoded libraries offer new ways of hit structure discovery. These technologies enable to search for new chemical matter in a much broader and deeper manner with less effort and fewer financial resources. These transformational developments require new cheminformatics approaches to make huge chemical spaces searchable and analyzable with low resources, and with as little energy consumption as possible. Substantial progress has been made in the past years with respect to computation as well as organic synthesis. First examples of bioactive compounds resulting from the successful use of these novel technologies demonstrate their power to contribute to tomorrow's drug discovery programs. This article gives a compact overview of the state-of-the-art.
Morgane Rivoal; Laurent Dubuquoy; Régis Millet; Natascha Leleu-Chavain J. Med. Chem., 2023, 66(21), 14391-14410 https://doi.org/10.1021/acs.jmedchem.3c00593AbstractReceptor interacting serine/threonine protein kinase 2 (RIPK2) is a downstream signaling molecule essential for the activation of several innate immune receptors, including the NOD-like receptors (NOD1 and NOD2). Recognition of pathogen-associated molecular pattern proteins by NOD1/2 leads to their interaction with RIPK2, which induces release of pro-inflammatory cytokines through the activation of NF-κB and MAPK pathways, among others. Thus, RIPK2 has emerged as a key mediator of intracellular signal transduction and represents a new potential therapeutic target for the treatment of various conditions, including inflammatory diseases and cancer. In this Perspective, first, an overview of the mechanisms that underlie RIPK2 function will be presented along with its role in several diseases. Then, the existing inhibitors that target RIPK2 and different therapeutic strategies will be reviewed, followed by a discussion on current challenges and outlook.
Mayar Tarek Ibrahim; Gennady M. Verkhivker; Jyoti Misra; Peng Tao Int. J. Mol. Sci., 2023, 24(10), 9009 https://doi.org/10.3390/ijms24109009AbstractThe Hippo pathway is an evolutionary conserved signaling network involved in several cellular regulatory processes. Dephosphorylation and overexpression of Yes-associated proteins (YAPs) in the Hippo-off state are common in several types of solid tumors. YAP overexpression results in its nuclear translocation and interaction with transcriptional enhanced associate domain 1-4 (TEAD1-4) transcription factors. Covalent and non-covalent inhibitors have been developed to target several interaction sites between TEAD and YAP. The most targeted and effective site for these developed inhibitors is the palmitate-binding pocket in the TEAD1-4 proteins. Screening of a DNA-encoded library against the TEAD central pocket was performed experimentally to identify six new allosteric inhibitors. Inspired by the structure of the TED-347 inhibitor, chemical modification was performed on the original inhibitors by replacing secondary methyl amide with a chloromethyl ketone moiety. Various computational tools, including molecular dynamics, free energy perturbation, and Markov state model analysis, were employed to study the effect of ligand binding on the protein conformational space. Four of the six modified ligands were associated with enhanced allosteric communication between the TEAD4 and YAP1 domains indicated by the relative free energy perturbation to original molecules. Phe229, Thr332, Ile374, and Ile395 residues were revealed to be essential for the effective binding of the inhibitors.
Xin Wen; Minmin Zhang; Zhiqiang Duan; Yanrui Suo; Weiwei Lu; Rui Jin; Baiyang Mu; Kaige Li; Xu Zhang; Linghua Meng; Yu Hong; Xingyu Wang; Hangchen Hu; Jian Zhu; Weixiao Song; Aijun Shen; Xiaojie Lu J. Med. Chem., 2023, 66(16), 11118-11132 https://doi.org/10.1021/acs.jmedchem.2c02129AbstractThe DNA-encoded library (DEL) is a powerful hit-generation tool in drug discovery. This study describes a new DEL with a privileged scaffold quinazolin-4(3H)-one developed by a robust DNA-compatible multicomponent reaction and a series of novel glutathione S-transferase (GST) inhibitors that were identified through affinity-mediated DEL selection. A novel inhibitor 16 was subsequently verified with an inhibitory potency value of 1.55 ± 0.02 μM against SjGST and 2.02 ± 0.20 μM against hGSTM2. Further optimization was carried out via various structure–activity relationship studies. And especially, the co-crystal structure of the compound 16 with the SjGST was unveiled, which clearly demonstrated its binding mode was quite different from the known GSH-like compounds. This new type of probe is likely to play a different role compared with the GSH, which may provide new opportunities to discover more potent GST inhibitors.
Lydia Borgelt; Peng Wu ACS Chem. Biol., 2023, 18(10), 2101-2113 https://doi.org/10.1021/acschembio.3c00191AbstractRibonucleases (RNases) cleave and process RNAs, thereby regulating the biogenesis, metabolism, and degradation of coding and noncoding RNAs. Thus, small molecules targeting RNases have the potential to perturb RNA biology, and RNases have been studied as therapeutic targets of antibiotics, antivirals, and agents for autoimmune diseases and cancers. Additionally, the recent advances in chemically induced proximity approaches have led to the discovery of bifunctional molecules that target RNases to achieve RNA degradation or inhibit RNA processing. Here, we summarize the efforts that have been made to discover small-molecule inhibitors and activators targeting bacterial, viral, and human RNases. We also highlight the emerging examples of RNase-targeting bifunctional molecules and discuss the trends in developing such molecules for both biological and therapeutic applications.
Anthony P. Silvestri; Qi Zhang; Yan Ping; Erik W. Muir; Jingsi Zhao; Sai Kumar Chakka; Gaonan Wang; Walter M. Bray; Wenhua Chen; Jennifer L. Fribourgh; Sarvind Tripathi; Yunyun He; Seth M. Rubin; Alexander L. Satz; Cameron R. Pye; Letian Kuai; Wenji Su; Joshua A. Schwochert ACS Med. Chem. Lett., 2023, 14(6), 820-826 https://doi.org/10.1021/acsmedchemlett.3c00117AbstractSynthetic macrocyclic peptides are an emerging molecular class for both targeting intracellular protein–protein interactions (PPIs) and providing an oral modality for drug targets typically addressed by biologics. Display technologies, such as mRNA and phage display, often yield peptides that are too large and too polar to achieve passive permeability or oral bioavailability without substantial off-platform medicinal chemistry. Herein, we use DNA-encoded cyclic peptide libraries to discover a neutral nonapeptide, UNP-6457, that inhibits MDM2–p53 interaction with an IC50 of 8.9 nM. X-ray structural analysis of the MDM2–UNP-6457 complex revealed mutual binding interactions and identified key ligand modification points which may be tuned to enhance its pharmacokinetic profile. These studies showcase how tailored DEL libraries can directly yield macrocyclic peptides benefiting from low MW, TPSA, and HBD/HBA counts that are capable of potently inhibiting therapeutically relevant protein–protein interactions.
Hongxin Zeng; Yanjing Li; Yang Zhang; Shiyang Xu; Yingwei Wang; Dawen Niu ChemRxiv, 2023 https://doi.org/10.26434/chemrxiv-2023-z0mqc-v2AbstractHerein, we report a method that enables the synthesis of carbohydrate-DNA conjugates by radical addition, in which the gen-eration of glycosyl radicals from readily available and bench-stable unprotected glycosyl sulfinates is the key. These reactions were carried out under mild conditions and tolerate a broad substrate scope.
Siyu He; Linjie Li; Rui Jin; Xiaojie Lu J. Med. Chem., 2023, 66(14), 9251-9277 https://doi.org/10.1021/acs.jmedchem.3c00390AbstractPeptidyl-prolyl cis/trans isomerase family (PPIase) is structurally divided into three subfamilies, cyclophilins (Cyps), FK506-binding proteins (FKBPs), and parvulins. Pin1 belongs to the parvulin family and is the only enzyme capable of isomerizing the phosphorylated Ser/Thr-Pro motif (p-Ser/Thr-Pro) in its interacting proteins. Due to its multibiological functions in vivo, including folding, intracellular signaling, transcription, cell cycle progression, and apoptosis, Pin1 is extensively studied as a promising drug target for various human diseases, especially cancer. In this Perspective, we summarized the literature covering diverse classes of Pin1 inhibitors and the inhibition mechanism, aiming to provide insights for the design of potent Pin1 inhibitors and suggest alternative strategies for developing potent Pin1 inhibitors.
Michael James Waring; Catherine LA Salvini; Benoit Darlot; Jack Davison; Mathew Martin; Susan Tudhope; Shannon Turberville; Akane Kawamura; Martin EM Noble; Stephen Wedge; James John Crawford Chem. Sci., 2023, 14, 8288-8294 https://doi.org/10.1039/D3SC01171BAbstractOptimisation of the affinity of lead compounds is a critical challenge in the identification of drug candidates and chemical probes and is a process that takes many years. Fragment-based drug discovery has become established as one of the methods of choice for drug discovery starting with small, low affinity compounds. Due to their low affinity, evolution of fragments to desirable levels of affinity is often a key challenge. The accepted best method for increasing the potency of fragments is by iterative fragment growing, which can be very time consuming and complex. Here, we introduce a paradigm for fragment hit optimisation using poised DNA-encoded chemical libraries (DELs). The synthesis of a poised DEL, a partially constructed library that retains a reactive handle, allows the coupling of any active fragment for a specific target protein, allowing rapid discovery of potent ligands. This is illustrated for bromodomain-containing protein 4 (BRD4), in which a weakly binding fragment was coupled to a 42-member poised DEL via Suzuki-Miyaura cross coupling resulting in the identification of an inhibitor with nanomolar affinity in a single step, representing an increase in potency of 6 orders of magnitude from an original fragment. The potency of the compound was shown to arise from the synergistic combination of substructures, which would have been very difficult to discover by any other method and was rationalised by X-ray crystallography. The compound showed attractive lead-like properties suitable for further optimisation and demonstrated BRD4-dependent cellular pharmacology. This work demonstrates the power of poised DELs to rapidly optimise fragments, representing an attractive generic approach to drug discovery.
Simon L. Rössler; Nathalie M. Grob; Stephen L. Buchwald; Bradley L. Pentelute Science, 2023, 379(6635), 939-945 https://doi.org/10.1126/science.adf1354AbstractStructured Abstract INTRODUCTION The discovery of therapeutics and biochemical probes hinges on the ability to identify molecules that interact with biological targets of interest. Technologies such as DNA-encoded libraries have transformed the process of drug discovery by enabling the rapid synthesis of vast collections of molecules, each encoded with a unique appendant DNA tag, and their subsequent screening in affinity selection experiments. However, the use of DNA to encode information confers synthetic limitations owing to the incompatibility of oligonucleotides with various chemical reaction conditions that result in the loss of stored information. As a consequence, chemical transformations developed for DNA-encoded library synthesis have to be optimized for oligonucleotide compatibility rather than reaction efficiency and scope. Given the vast potential of encoded library technologies in drug discovery, complementary platforms addressing the limitations of DNA encoding by leveraging carriers of information with higher stability and versatility are desirable. RATIONALE Molecular encoding can in principle be achieved in any polymer with at least two distinguishable monomers. Peptides constitute a biopolymer that is routinely sequenced for elucidation of protein identity through the use of tandem mass spectrometry. Accordingly, synthetic peptides can serve as carriers of information decoded through determination of their amino acid sequence. We hypothesized that the excellent chemical stability of peptides and their compatibility with a broad range of reaction conditions would render them particularly suited for the encoding of small-molecule synthesis. Peptides, which can be efficiently prepared by solid-phase synthesis, offer potential as carriers of information for a next-generation discovery platform with combinatorial libraries of small molecules. RESULTS We describe the design of abiotic peptides as carriers of information for the encoding of small-molecule synthesis. Therein, the identity of a small molecule was stored in an appendant peptide that is elongated in accordance with the synthetic elaboration of the small molecule. The encoding peptide featured a hexadecimal encoding alphabet of nonisobaric amino acids, resulting in high information density and chemical stability. The sequence of the encoding peptide was optimized through the systematic inclusion of selected amino acids to fine-tune polarity and ease of sequencing, resulting in high-fidelity decoding by tandem mass spectrometry. The chemical stability of the peptide tag enabled synthetic versatility for small-molecule transformations, including acidic conditions or transition metal catalysis with reported incompatibility with DNA tags. This broad compatibility allowed the implementation of palladium-mediated cross-coupling reactions characterized by a diverse scope and high reaction efficiency. The encoding of small molecule synthesis in peptides was leveraged for the split-and-pool synthesis of combinatorial libraries called peptide-encoded libraries (PELs) characterized by high purity. PELs featuring tens of thousands of drug-like small molecules resulting from optimized palladium-mediated C–C and C–N cross-coupling reactions were used in affinity selections against oncogenic proteins. The peptide sequences of enriched conjugates were decoded, and the corresponding small molecules were rapidly prepared by solid-phase synthesis and subsequently confirmed to exhibit affinity for their target protein. The PEL discovery platform is characterized by high efficiency and has afforded diverse, previously unknown small-molecule binders for the target proteins. CONCLUSION The results demonstrate that abiotic peptides can be used to encode and decode information to discover small molecules with affinity to proteins of interest. The PEL discovery platform establishes a starting point for the next generation of encoded library technology with broad implications for therapeutics discovery and biochemical research.
Amjad Ali; Marcus Bauser; Sophie Bertrand; Wesley Blackaby; Christoph Boss; Martin Bossart; Adrian Hall; Hayley Binch; Werngard Czechtizky; Harrie Gijsen; Helmut Haning; Ingo V. Hartung; Paul Kilburn; Gilbert Lassalle; Ulrich Lücking; Jürgen Mack; Martin Missbach; Leena Otsomaa; Antoni Torrens; Michael Wagner; Magnus Walter; Harald Weinstabl; Luc van Hijfte; Franz von Nussbaum ChemMedChem, 2023, e202300127 https://doi.org/10.1002/cmdc.202300127AbstractThe status of industrial Medicinal Chemistry was discussed with European Medicinal Chemistry Leaders from large to mid-sized pharma and CRO companies as well as biotechs. The chemical modality space has expanded recently from small molecules to address new challenging targets. Besides the classical SAR/SPR optimization of drug molecules also their ‘greenness’ has increasing importance. The entire pharma discovery ecosystem has developed significantly. Beyond pharma and academia new key players such as Biotech and integrated CROs as well as Digital companies have appeared and are now to a large extend fueled by VC money. Digitalization is happening everywhere but surprisingly did not change speed and success rates of projects so far. Future Medicinal Chemists will still have to be excellent synthetic chemists but in addition they must be knowledgeable in new computational areas such as data sciences. Their ability to collaborate and to work in teams is key.
Salma Haj-Yahia; Arijit Nandi; Raphael I. Benhamou Expert Opin. Drug Discov., 2023, 18(8), 929-942 https://doi.org/10.1080/17460441.2023.2224960AbstractRNA structural motifs can serve as recognition sites for proteins or regulatory elements. Notably, these specific RNA shapes are directly related to many diseases. Targeting specific RNA motifs using small molecules is an emerging domain of study within the area of drug discovery. Targeted degradation strategies are a relatively modern technology in drug discovery, with important clinical and therapeutic outcomes. These approaches involve using small molecules to selectively degrade specific biomacromolecules associated with a disease.”Ribonuclease-Targeting Chimeras” (RiboTaCs) represent a promising type of targeted degradation strategy because of their ability to selectively degrade structured RNA targets.
Jessica Graham; Harriet Stanway-Gordon; Michael James Waring Chem. Eur. J., 2023, e202300603 https://doi.org/10.1002/chem.202300603AbstractDNA-Encoded Libraries (DELs) are becoming widely established as a hit identification strategy for drug discovery campaigns. Their successful application relies on the availability and efficiency of the reactions that can be carried out on DNA. These reactions should proceed with high conversion to the desired product and have a broad substrate scope to synthesise chemically diverse and drug-like DELs. The Sonogashira coupling provides a unique means of coupling an sp-hybridized carbon centre to an aryl halide and methods to achieve this reaction on DNA are highly desirable. We report the application of our micellar technology for on-DNA chemistry to the Sonogashira reaction. This method gives highly efficient conversions for the coupling of (hetero)aromatic and aliphatic alkynes to (hetero)aryl iodides and bromides allowing the preparation of highly diverse DELs.
Yu Zhou; Wenyin Shen; Ying Gao; Jianzhao Peng; Qingrong Li; Xueying Wei; Shihao Liu; Fong Sang Lam; Joan Mayol-Llinàs; GuixianZhao; Gang Li; Yizhou Li; Hongzhe Sun; Yan Cao; Xiaoyu Li Nat. Chem., 2024 https://doi.org/10.1038/s41557-024-01442-yAbstractDNA-encoded chemical libraries (DELs) have become a powerful technology platform in drug discovery. Dual-pharmacophore DELs display two sets of small molecules at the termini of DNA duplexes, thereby enabling the identification of synergistic binders against biological targets, and have been successfully applied in fragment-based ligand discovery and affinity maturation of known ligands. However, dual-pharmacophore DELs identify separate binders that require subsequent linking to obtain the full ligands, which is often challenging. Here we report a protein-templated DEL selection approach that can identify full ligand/inhibitor structures from DNA-encoded dynamic libraries (DEDLs) without the need for subsequent fragment linking. Our approach is based on dynamic DNA hybridization and target-templated in situ ligand synthesis, and it incorporates and encodes the linker structures in the library, along with the building blocks, to be sampled by the target protein. To demonstrate the performance of this method, 4.35-million- and 3.00-million-member DEDLs with different library architectures were prepared, and hit selection was achieved against four therapeutically relevant target proteins.
Feiyan Zhan; Jingjie Zhu; Shaowen Xie; Jinyi Xu; Shengtao Xu Eur. J. Med. Chem., 2023, 253, 115338 https://doi.org/10.1016/j.ejmech.2023.115338AbstractCurrently, bioorthogonal coupling reactions have garnered considerable interest due to their high substrate selectivity and less restrictive reaction conditions. During recent decades, bioorthogonal coupling reactions have emerged as powerful tools in drug development. This review describes the current applications of bioorthogonal coupling reactions in compound library building mediated by the copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction and in situ click chemistry or conjunction with other techniques; druggability optimization with 1,2,3-triazole groups; and intracellular self-assembly platforms with ring tension reactions, which are presented from the viewpoint of drug development. There is a reasonable prospect that bioorthogonal coupling reactions will accelerate the screening of lead compounds, the designing strategies of small molecules and expand the variety of designed compounds, which will be a new trend in drug development in the future.
Regina Pikalyova; Yuliana Zabolotna; Dragos Horvath; Gilles Marcou; Alexandre Varnek ChemRxiv, 2023 https://doi.org/10.26434/chemrxiv-2023-w7x9tAbstractThe development of DNA-Encoded Library (DEL) technology introduced new challenges for the analysis of chemical libraries. Unlike classical HTS libraries, the DEL composition cannot be modified once synthesized and, therefore, it must be considered as a stand-alone chemoinformatic object represented both as a collection of independent molecules, and yet an individual entity. For the analysis of such collections, the concept of the Chemical Library Space (CLS) where entire libraries become objects is indispensable. In this article, we introduce, analyze and compare four vectorial library representations obtained using Generative Topographic Mapping (GTM), formally defining CLS. Such representations allow effective comparison of libraries, with the ability to tune and chemically interpret the similarity relationships. In particular, property-tuned CLS encodings enable to simultaneously compare libraries with respect to both property and chemotype distribution. We apply the various CLS encodings for the selection problem of DELs that optimally match a reference collection (here ChEMBL28), showing how the choice of the CLS descriptors may help to fine-tune the matching (overlap) criteria. The proposed CLS encodings are a new efficient way for polyvalent analysis of the space of thousands of chemical libraries.
Harriet A. Stanway-Gordon; Jake A. Odger; Michael J. Waring Bioconjugate Chem., 2023, 34(4), 756-763 https://doi.org/10.1021/acs.bioconjchem.3c00051AbstractThe capability of DNA encoded libraries (DELs) as a method of small molecule hit identification is becoming widely established in drug discovery. While their selection method offers advantages over more traditional means, DELs are limited by the chemistry that can be utilized to construct them. Significant advances in DNA compatible chemistry have been made over the past five years; however such procedures are still often burdened by substrate specificity and/or incomplete conversions, reducing the fidelity of the resulting libraries. One such reaction is the Heck coupling, for which current DNA-compatible protocols are somewhat unreliable. Utilizing micellar technology, we have developed a highly efficient DNA-compatible Heck reaction that proceeds on average to 95% conversion to product across a broad variety of structurally significant building blocks and multiple DNA conjugates. This work continues the application of micellar catalysis to the development of widely applicable, effective DNA-compatible reactions for use in DELs.
Nari N. Talaty; Robert W. Johnson; James Sawicki; Omprakash Nacham; Stevan W. Djuric ACS Med. Chem. Lett., 2023, 14(6), 711-718 https://doi.org/10.1021/acsmedchemlett.3c00040AbstractThe complexity of new therapeutics continues to increase and the timeline for the discovery of these therapeutics continues to shrink. This creates demand for new analytical techniques to facilitate quicker discovery and development of novel drugs. Mass spectrometry is one of the most prolific analytical techniques that has been applied across the entire drug discovery pipeline. New mass spectrometers and the associated methods for sampling have been introduced at a rate that keeps pace with new chemistries, therapeutic types, and screening practices used by modern drug hunters. This microperspective covers application and implementation of new mass spectrometry workflows that enable current and future efforts in screening and synthesis for drug discovery.
Liu Y; Sundah Nr; Ho Nry; Shen Wx; Xu Y; Natalia A; Yu Z; Seet Je; Chan Cw; Loh Tp; Lim By; Shao H Nat. Biomed. Eng., 2024 https://doi.org/10.1038/s41551-024-01225-3AbstractCapturing the full complexity of the diverse hierarchical interactions in the protein interactome is challenging. Here we report a DNA-barcoding method for the multiplexed mapping of pairwise and higher-order protein interactions and their dynamics within cells. The method leverages antibodies conju …
Rahul Sanawar; Vinodh J. Sahayasheela; Praseetha Sarath; Vipin Mohan Dan Mini. Rev. Med. Chem., 2023, 23(15), 1507-1513 https://dx.doi.org/10.2174/1389557523666230125114921AbstractDiscoidin domain receptor (DDR) 1, a collagen binding receptor kinase, is an intensively researched therapeutic target for cancer, fibrosis and other diseases. The majority of early known DDR1 inhibitors targeted the ATP binding pocket of this enzyme that shares structural similarities with other kinase pockets across the biological system. This structural similarity of DDR1 kinase with other protein kinases often leads to “off target “toxicity issues. Understanding of uniqueness in DDR:ATP–phosphate-binding loop (P-loop), DNA encoded library screen, structure-guided optimization studies, and machine learning drug design platforms that come under the umbrella of artificial intelligence has led to the discovery of a new array of inhibitors that are highly selective for DDR1 over DDR2 and other similar kinases. Most of the drug discovery platforms concentrated on the ATP binding region of DDR1 kinase and never looked beyond this region for novel therapeutic options. Recent findings have disclosed the kinase-independent functions of DDR1 in immune exclusion, which resides in the extracellular collagen-binding domain, thus opening avenues for the development of inhibitors that veer away from targeting ATP binding pockets. This recent understanding of the functional modalities of DDR1 opens the complexity of targeting this transmembrane protein as per its functional prominence in the respective disease and thus demands the development of specific novel therapeutics. The perspective gives a short overview of recent developments of DDR1 inhibitors with the aid of the latest technologies, future directions for therapeutic development, and possibility of combinational therapeutic treatments to completely disengage functions of DDR1.
Sarah E. Lindley; Yiyang Lu; Diwakar Shukla ACS Appl. Bio Mater., 2023, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsabm.3c00054AbstractInitially part of the field of artificial intelligence, machine learning (ML) has become a booming research area since branching out into its own field in the 1990s. After three decades of refinement, ML algorithms have accelerated scientific developments across a variety of research topics. The field of small molecule design is no exception, and an increasing number of researchers are applying ML techniques in their pursuit of discovering, generating, and optimizing small molecule compounds. The goal of this review is to provide simple, yet descriptive, explanations of some of the most commonly utilized ML algorithms in the field of small molecule design along with those that are highly applicable to an experimentally focused audience. The algorithms discussed here span across three ML paradigms: supervised learning, unsupervised learning, and ensemble methods. Examples from the published literature will be provided for each algorithm. Some common pitfalls of applying ML to biological and chemical data sets will also be explained, alongside a brief summary of a few more advanced paradigms, including reinforcement learning and semi-supervised learning.
Zhaomei Sun; Lingqian Xiao; Yahui Chen; Jiangying Wang; Fanming Zeng; Huanqing Zhang; Jie Zhang; Kexin Yang; Yunjin Hu ACS Med. Chem. Lett., 2023, 14(4), 473-478 https://doi.org/10.1021/acsmedchemlett.3c00022AbstractOrganophosphonic compounds are distinctive among natural products in terms of stability and mimicry. Numerous synthetic organophosphonic compounds, including pamidronic acid, fosmidromycin, and zoledronic acid, are approved drugs. DNA encoded library technology (DELT) is a well-established platform for identifying small molecule recognition to target protein of interest (POI). Therefore, it is imperative to create an efficient procedure for the on-DNA synthesis of α-hydroxy phosphonates for DEL builds.
Jia Fu; Wei Qin; Luqi Cao; Zhesheng Chen; Huiling Cao Drug Discov. Today, 2023, 103576 https://doi.org/10.1016/j.drudis.2023.103576AbstractReceptor chromatography involves high-throughput separation and accurate drug screening based on specific drug–receptor recognition and affinity, which has been widely used to screen active compounds in complex samples. This review summarizes the immobilization methods for receptors from three aspects: random covalent immobilization methods, site-specific covalent immobilization methods and dual-target receptor chromatography. Meanwhile, it focuses on its applications from three angles: screening active compounds in natural products, in natural-product-derived DNA-encoded compound libraries and drug–receptor interactions. This review provides new insights for the design and application of receptor chromatography, high-throughput and accurate drug screening, drug–receptor interactions and more. Teaser: This review summarizes the immobilization methods of receptors and the application of receptor chromatography, which will provide new insights for the design and application of receptor chromatography, rapid drug screening, drug–receptor interactions and more.
Longbo Li, Bianca Taeko Matsuo, Guillaume Levitre, Edward J. McClain, Eric A. Voight; Erika A. Crane; Gary A. Molander Chem. Sci., 2023, 14, 2713-2720 https://doi.org/10.1039/D3SC00144JAbstractDNA-encoded library (DEL) screens have significantly impacted new lead compound identification efforts within drug discovery. An advantage of DELs compared to traditional screening methods is that an exponentially broader chemical space can be effectively screened using only nmol quantities of billions of DNA-tagged, drug-like molecules. The synthesis of DELs containing diverse, sp3-rich spirocycles, an important class of molecules in drug discovery, has not been previously reported. Herein, we demonstrate the synthesis of complex and novel spirocyclic cores via an on-DNA, visible light-mediated intermolecular [2+2] cycloaddition of olefins with heterocycles, including indoles, azaindoles, benzofurans, and coumarins. The DNA-tagged exo-methylenecyclobutane substrates were prepared from easily accessible alkyl iodides and styrene derivatives. Broad reactivity with many other DNA-conjugated alkene substrates was observed, including unactivated and activated alkenes, and the process is tolerant of various heterocycles. The cycloaddition was successfully scaled from 10 to 100 nmols without diminished yield, indicative of this reaction’s suitability for DNA-encoded library production. Evaluation of DNA compatibility with the developed reaction in a mock-library format showed that the DNA barcode was maintained with high fidelity, with <1% mutated sequences and >99% amplifiable DNA from quantitative polymerase chain reaction (PCR) and next generation sequencing (NGS).
Zhi Zhou; Yuang Gu; Liexin Wu; Yan Wang; Huiying Xu; Lei Ma; Zhaoyong Zhang; Jincun Zhao; Wei Zhang; Wei Peng; Guang Yang; Xiyong Yu; Hongtao Xu; Wei Yi Chem, 2023, 9(11), 3335-3346 https://doi.org/10.1016/j.chempr.2023.07.022AbstractOrgano-Se species represent a type of increasingly important core motif widely found in endogenous proteins and bioactive small molecules. Consequently, exploring their structural diversity to entrust their powerful potential has emerged as an important endeavor in basic research, but it remains extremely limited and highly challenging. Here, we realize either racemic or asymmetric synthesis toward a class of organo-Se species, namely benzothiaselenazole-1-oxides (BTSAs), and reveal its diversified applications. For example, we identified that BTSAs can be recognized as the click selenylation reagents, which not only enabled on-plate modular construction of parallel (1,066 examples) and DNA-encoded (261 examples) indole-C3-selenide libraries but also provided the site-specific modification of sulfhydryl-containing chemicals for application in anti-COVID-19 drug discovery and bioorthogonal labeling-based HER2 fluorescence imaging analysis. This work greatly expands the space of organo-Se chemistry, and given the potent performance of BTSAs in click selenylation, it should find robust application in chemistry-related research areas.
Jean Quancard; Anna Vulpetti; Anders Bach; Brian Cox; Stéphanie M. Guéret; Ingo V. Hartung; Hannes F. Koolman; Stefan Laufer; Josef Messinger; Gianluca Sbardella; Russell Craft ChemMedChem, 2023, e202300002 https://doi.org/10.1002/cmdc.202300002AbstractHit generation is a crucial step in drug discovery that will determine the speed and chance of success of identifying drug candidates. Many strategies are now available to identify chemical starting points, or hits, and each biological target warrants a tailored approach. In this set of best practices, we detail the essential approaches for target centric hit generation and the opportunities and challenges they come with. We then provide guidance on how to validate hits to ensure medicinal chemistry is only performed on compounds and scaffolds that engage the target of interest and have the desired mode of action. Finally, we discuss the design of integrated hit generation strategies that combine several approaches to maximize the chance of identifying high quality starting points to ensure a successful drug discovery campaign.
An T.H. Le; Svetlana M. Krylova; Sergey N. Krylov TrAC, Trends Anal. Chem., 2023, 117061 https://doi.org/10.1016/j.trac.2023.117061AbstractIdentifying protein binders is the first step in drug discovery. The combinatorial approach, in which a library of compounds is subjected to affinity screening against a target protein, is a major way for identifying protein binders. Oligonucleotide libraries constitute the largest source of material for such affinity screening. Selecting protein binders from such libraries requires a highly efficient method for separation of protein-oligonucleotide complexes from the excess of unbound oligonucleotides. Kinetic Capillary electrophoresis (KCE) is a fast-developing trend in affinity applications. It has the highest reported efficiency of partitioning, but screening oligonucleotide libraries by KCE has many challenges which must be addressed before KCE can compete with conventional surface-based screening. Here we provide the critical analysis of advantages and limitations of KCE in screening oligonucleotide libraries. We identify potential ways of overcoming the limitations in an attempt to direct researchers towards the most important and urgent tasks in this area.
Charlotte Franck; Karishma Patel; Louise J. Walport; Mary Christie; Alexander Norman; Toby Passioura; Hiroaki Suga; Richard J. Payne; Joel P. Mackay Structure, 2023, 31(8), 912-923 https://doi.org/10.1016/j.str.2023.05.009AbstractDNA encoded cyclic peptide libraries offer unique opportunities to discover high-potency, high-specificity ligands directed against a target protein. We set out to explore the potential for such libraries to provide ligands that can distinguish between bromodomains from the closely related paralogues of the Bromodomain and ExtraTerminal domain (BET) family of epigenetic regulators. Analysis of peptides isolated from a screen against the C-terminal bromodomain of family member BRD2, together with new peptides discovered in previous screens against the corresponding domain from BRD3 and BRD4, reveals peptides with nanomolar and subnanomolar affinities. X-ray crystal structures of several of these bromodomain-peptide complexes reveal diverse structures and binding modes, which nevertheless display several conserved binding features. A subset of the peptides demonstrates significant paralogue-level specificity, though structural analysis does not reveal clear physicochemical explanations for this specificity. Our data demonstrate the power of cyclic peptides to discriminate between highly similar proteins with high potency and hint that differences in conformational dynamics between BET-family bromodomains might modulate binding affinities amongst family members for particular ligands.
Yaqiong Yan; Lei Wang; Hao Wang Chem. Res. Chin. Univ., 2023, 39, 83–91 https://doi.org/10.1007/s40242-023-2356-2AbstractCombinatorial chemistry provides a cost-effective method for the rapid discovery of new functional peptides. One-bead one-compound(OBOC) high-throughput screening technique offers a lot of structurally diverse peptides to be rapidly synthesized and screened for binding to a target of interest. The OBOC peptide library screening involves three main steps: library construction, positive beads separation, and peptide sequencing. This review mainly summarizes some special technique tips during functional peptide screening and potential future directions of the OBOC high-throughput screening technique.
Suzanne Ackloo; Albert A. Antolin; Jose Manuel Bartolome; Hartmut Beck; Alex Bullock; Ulrich A. K. Betz; Jark Böttcher; Peter J. Brown; Menorca Chaturvedi; Alisa Crisp; Danette Daniels; Jan Dreher; Kristina Edfeldt; Aled M. Edwards; Ursula Egner; Jon Elkins; Christian Fischer; Tine Glendorf; Steven Goldberg; Ingo V. Hartung; Alexander Hillisch; Evert Homan; Stefan Knapp; Markus Köster; Oliver Krämer; Josep Llaveria; Uta Lessel; Sven Lindemann; Lars Linderoth; Hisanori Matsui; Maurice Michel; Florian Montel; Anke Mueller-Fahrnow; Susanne Müller; Dafydd R. Owen; Kumar Singh Saikatendu; Vijayaratnam Santhakumar; Wendy Sanderson; Cora Scholten; Matthieu Schapira; Sujata Sharma; Brock Shireman; Michael Sundström; Matthew H. Todd; Claudia Tredup; Jennifer Venable; Timothy M. Willson; Cheryl H. Arrowsmith RSC Med. Chem., 2023, 14, 1002-1011 https://doi.org/10.1039/D2MD00441KAbstractTarget 2035, an international federation of biomedical scientists from the public and private sectors, is leveraging ‘open’ principles to develop a pharmacological tool for every human protein. These tools are important reagents for scientists studying human health and disease and will facilitate the development of new medicines. It is therefore not surprising that pharmaceutical companies are joining Target 2035, contributing both knowledge and reagents to study novel proteins. Here, we present a brief progress update on Target 2035 and highlight some of industry's contributions.
Kirill Shmilovich; Benson Chen; Theofanis Karaletsos; Mohammad M. Sultan J. Chem. Inf. Model., 2023, 63(9), 2719-2727 https://doi.org/10.1021/acs.jcim.2c01608AbstractDNA-encoded library (DEL) technology has enabled significant advances in hit identification by enabling efficient testing of combinatorially generated molecular libraries. DEL screens measure protein binding affinity though sequencing reads of molecules tagged with unique DNA barcodes that survive a series of selection experiments. Computational models have been deployed to learn the latent binding affinities that are correlated to the sequenced count data; however, this correlation is often obfuscated by various sources of noise introduced in its complicated data-generation process. In order to denoise DEL count data and screen for molecules with good binding affinity, computational models require the correct assumptions in their modeling structure to capture the correct signals underlying the data. Recent advances in DEL models have focused on probabilistic formulations of count data, but existing approaches have thus far been limited to only utilizing 2-D molecule-level representations. We introduce a new paradigm, DEL-Dock, that combines ligand-based descriptors with 3-D spatial information from docked protein–ligand complexes. 3-D spatial information allows our model to learn over the actual binding modality rather than using only structure-based information of the ligand. We show that our model is capable of effectively denoising DEL count data to predict molecule enrichment scores that are better correlated with experimental binding affinity measurements compared to prior works. Moreover, by learning over a collection of docked poses we demonstrate that our model, trained only on DEL data, implicitly learns to perform good docking pose selection without requiring external supervision from expensive-to-source protein crystal structures.
Xianfu Fang; Huilin Liao; Xiaohong Fan; Yiting Wang; Huihong Wang; Gong Zhang; Wei Fang; Yangfeng; Yizhou Li Org. Biomol. Chem., 2023, 21(10), 2162-2166 https://doi.org/10.1039/D2OB02278HAbstractViridicatin alkaloids as natural products have attracted great interest due to their unique core scaffold. To fully exploit their potential application in DNA-encoded chemical libraries that would facilitate drug discovery, we here describe an efficient on-DNA synthesis of viridicatin alkaloid-like scaffolds from isatins and DNA-tagged aldehydes. Promoted by benzenesulfonyl hydrazide, this reaction provided the corresponding DNA-conjugated viridicatin alkaloid-like products in moderate-to-excellent conversion yields, and DNA compatibility validated by enzymatic ligation and qPCR evaluation exhibited the feasible utility of this methodology in DEL synthesis. Cross substrate scope study, together with subsequent on-DNA chemical diversification, further showed the competence of this approach in focused natural product-like encoded library construction.
Komal Sharma; Krishna K. Sharma; Anku Sharma; Rahul Jain Drug Discov. Today, 2022, 103464 https://doi.org/10.1016/j.drudis.2022.103464AbstractThe progressive development of peptides from reaction vessels to life-saving drugs via rigorous preclinical and clinical assessments is fascinating. Peptide therapeutics have gained momentum with the evolution of techniques in peptide chemistry, such as microwave irradiation in solid- and solution-phase synthesis, ligation chemistry, recombinant synthesis, and amalgamation with synthetic tools, including metal catalysis. Diverse emerging technologies, such as DNA-encoded libraries (DELs) and display techniques, are changing the status quo in the discovery of peptide therapeutics. In this review, we analyzed US Food and Drug Administration (FDA)-approved peptide drugs and those in clinical trials, highlighting recent advances in peptide-based drug discovery.
Alex Hoose; Richard Vellacott; Marko Storch; Paul S. Freemont; Maxim G. Ryadnov Nat. Rev. Chem., 2023, 7, 144-161 https://doi.org/10.1038/s41570-022-00456-9AbstractSynthetic DNA is of increasing demand across many sectors of research and commercial activities. Engineering biology, therapy, data storage and nanotechnology are set for rapid developments if DNA can be provided at scale and low cost. Stimulated by successes in next generation sequencing and gene editing technologies, DNA synthesis is already a burgeoning industry. However, the synthesis of >200 bp sequences remains unaffordable. To overcome these limitations and start writing DNA as effectively as it is read, alternative technologies have been developed including molecular assembly and cloning methods, template-independent enzymatic synthesis, microarray and rolling circle amplification techniques. Here, we review the progress in developing and commercializing these technologies, which are exemplified by innovations from leading companies. We discuss pros and cons of each technology, the need for oversight and regulatory policies for DNA synthesis as a whole and give an overview of DNA synthesis business models.
Mark A. Mantell; Lisa Marcaurelle; Yun Ding Org. Lett., 2023, 25(8), 1241-1245 https://doi.org/10.1021/acs.orglett.2c04043AbstractDeveloping new on-DNA reactions is paramount to the development of new encoded libraries in the pursuit of novel pharmaceutical lead compounds. Lactam-containing molecules have been shown to be effective in a wide range of therapeutic areas and therefore represent a promising target for further investigation by DNA-encoded library screening. In pursuit of this motif, we report a novel method for the introduction of lactam-containing structures onto a DNA headpiece through the Ugi four-center three-component reaction (4C-3CR). This novel method is successful in three different approaches to give unique on-DNA lactam structures: on-DNA aldehyde coupled with isonitriles and amino acids; on-DNA isonitrile coupled with aldehydes and amino acids; and on-DNA isonitrile coupled with amines and acid aldehydes.
Jiayi Yin; Xintong Lu; Mengling Hou; Ting Cao; Zhen Tian Biochem. Pharmacol., 2023, 115543 https://doi.org/10.1016/j.bcp.2023.115543AbstractSirtuin1 (SIRT1) is a conserved nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase that plays key roles in a range of cellular events, including the maintenance of genome stability, gene regulation, cell proliferation, and apoptosis. P53 is one of the most studied tumor suppressors and the first identified non-histone target of SIRT1. SIRT1 deacetylates p53 in a NAD+-dependent manner and inhibits its transcriptional activity, thus exerting action on a series of pathways related to tissue homeostasis and various pathological states. The SIRT1-p53 axis is thought to play a central role in tumorigenesis. Although SIRT1 was initially identified as a tumor promoter, evidence now indicates that SIRT1 may also act as a tumor suppressor. This seemingly contradictory evidence indicates that the functionality of SIRT1 may be dictated by different cell types and intracellular localization patterns. In this review, we summarize recent evidence relating to the interactions between SIRT1 and p53 and discuss the relative roles of these two molecules with regards to cancer-associated cellular events. We also provide an overview of current knowledge of SIRT1-p53 signaling in tumorigenesis. Given the vital role of the SIRT1-p53 pathway, targeting this axis may provide promising strategies for the treatment of cancer.
Sven Stegemann; Chris Moreton; Sami Svanbäck; Karl Box; Geneviève Motte; Amrit Paudel Drug Discov. Today, 2022, 103344 https://doi.org/10.1016/j.drudis.2022.103344AbstractIn 1997, the ‘Rule of Five’ (Ro5) suggested physicochemical limitations for orally administered drugs, based on the analysis of chemical libraries from the early 1990s. In this review, we report on the trends in oral drug product development by analyzing products launched between 1994 and 1997 and between 2013 and 2019. Our analysis confirmed that most new oral drugs are within the Ro5 descriptors; however, the number of new drug products of drugs with molecular weight (MW) and calculated partition coefficient (clogP) beyond the Ro5 has slightly increased. Analysis revealed that there is no single scientific or technological reason for this trend, but that it likely results from incremental advances are being made in molecular biology, target diversity, drug design, medicinal chemistry, predictive modeling, drug metabolism and pharmacokinetics, and drug delivery.
Pratik R. Chheda; Nicholas Simmons; David P. Schuman; Zhicai Shi Org. Lett., 2022, 24(51), 9514-9519 https://doi.org/10.1021/acs.orglett.2c03994AbstractDNA-encoded library (DEL) screens have become a key technology to find small molecule binders to biological targets for drug discovery applications. The development of new DNA-compatible chemistries to expand the accessible DEL chemical space is imperative to enhance screen success across broad target classes and modalities. Additionally, reactions that use commonly available building blocks as well as those that enable the fsp3 of library members to be increased would have high impact for accessing diverse drug-like structures. Herein, we report a DNA-compatible Giese-type addition of nonstabilized C-centered radicals generated by the deoxygenation of preactivated alcohols into on-DNA olefins. Although alcohols have been historically underused as a building block class within DEL synthesis, their activation to a xanthate enables Csp3–Csp3 coupling to furnish sp3-rich products. This reaction is compatible with multiple classes of functional groups, does not damage the DNA tag, and is suitable for use in DEL productions.
He Zhu; Mona Sharafi; Wei Pin Teh; Ariana S. Bratt; Sara J. Buhrlage; Jarrod A. Marto Isr. J. Chem., 2023, 63, e202200113 https://doi.org/10.1002/ijch.202200113AbstractSince its introduction by Cravatt and colleagues in 1999, activity-based protein profiling (ABPP) has become widely utilized throughout academia, government, and industry laboratories to study enzymes spanning numerous gene families in a multitude of biological systems. As a variation of ABPP, competitive ABPP provides a powerful approach to characterize the binding behavior of small molecule probes and clinical drugs throughout the functional proteome. The power and flexibility of competitive ABPP are exemplified by a wide range of creative adaptions which increase assay throughput, enable diverse detection schemes, and support the implementation of this approach within a hybrid target-based screening platform. We review major developments in competitive ABPP through a compare-contrast format to provide a useful introduction to this enabling technology for scientists in chemical biology and drug discovery.
Bianca Matsuo; Albert Granados; Guillaume Levitre; Gary A. Molander Acc. Chem. Res., 2023, 56(3), 385-401 https://doi.org/10.1021/acs.accounts.2c00778AbstractDNA-encoded library technology (DELT) is a new screening modality that allows efficient, cost-effective, and rapid identification of small molecules with potential biological activity. This emerging technique represents an enormous advancement that, in combination with other technologies such as high-throughput screening (HTS), fragment-based lead generation, and structure-based drug design, has the potential to transform how drug discovery is carried out. DELT is a hybrid technique in which chemically synthesized compounds are linked to unique genetic tags (or “barcodes”) that contain readable information. In this way, millions to billions of building blocks (BBs) attached on-DNA via split-and-pool synthesis can be evaluated against a biological target in a single experiment. Polymerase chain reaction (PCR) amplification and next-generation sequencing (NGS) analysis of the unique sequence of oligonucleotides in the DNA tag are used to identify those ligands with high affinity for the target. This innovative fusion of genetic and chemical technologies was conceived in 1992 by Brenner and Lerner (Proc. Natl. Acad. Sci. 1992, 89, 5381–5383) and is under accelerated development with the implementation of new synthetic techniques and protocols that are compatible with DNA. In fact, reaction compatibility is a key parameter to increasing the chances of identification of a drug target ligand, and a central focus has been the development of new transformations and the transition to robust protocols for on-DNA synthesis. Because the sole use of the DNA tag is as an amplifiable identification barcode, its structural integrity during a new chemical process is mandatory. As such, the use of these sensitive, polyfunctional biological molecules as substrates typically requires aqueous solutions within defined pH and temperature ranges, which is considered a notable challenge in DEL synthesis. Using low-energy visible light as the driving force to promote chemical transformations represents an attractive alternative to classical synthetic methods, and it is an important and well-established synthetic tool for forging chemical bonds in a unique way via radical intermediates. Recent advances in the field of photocatalysis are extraordinary, and this powerful research arena is still under continuous development. Several applications taking advantage of the mild reaction conditions of photoinduced transformations have been directed toward DEL synthesis, allowing the expansion of chemical space available for the evaluation of new building blocks on-DNA. There are no doubts that visible-light-driven reactions have become one of the most powerful approaches for DELT, given the easy way they provide to construct new bonds and the challenges to achieve equal success via classical protocols. Key characteristics of photocatalytic synthesis include the short reaction times and efficiency, which translate into retention of DNA integrity. In this Account, we describe recent advances in the photoinduced diversification of building blocks prepared on-DNA, highlighting the amenability of the techniques employed for preserving the genetic structure of the molecules. We demonstrate with recent research from our group the applicability of photocatalysis to the field and include in the summary a table containing all the photoinduced methods reported to date for DELT, demonstrating their key aspects such as scope, applications, and DNA compatibilities. With this information, practitioners are provided with compelling reasons for developing/choosing photocatalytic methods for DELT applications.
Yves P. Auberson; Paola B. Arimondo; Maria Duca; Sebastian Essig; Uwe Grether; Arne C. Rufer; Gianluca Sbardella; Ulrich Schopfer; Antoni Torrens; Mario van der Stelt; Boris Vauzeilles; Olalla Vázquez; Andrew X. Zhang ChemBioChem, 2023, 24(7), e202200690 https://doi.org/10.1002/cbic.202200690AbstractGround-breaking research in disease biology and continuous efforts in method development have uncovered a range of potential new drug targets. Increasingly, the drug discovery process is informed by technologies involving chemical probes as tools. Applications for chemical probes comprise target identification and assessment, as well as the qualification of small molecules as chemical starting points and drug candidates. Progress in probe chemistry has opened the way to novel assay formats and pharmaceutical compound classes. The European Federation of Medicinal Chemistry and Chemical Biology (EFMC) has launched the Chemical Biology Initiative to advance science in the field of medicinal chemistry and chemical biology, while representing all members of this extended scientific community. This review provides an overview of the many important developments in the field of chemical biology that have happened at the lively interface of academic and industrial research.
Ram K. Modukuri; Diana Monsivais; Feng Li; Murugesan Palaniappan; Kurt M. Bohren; Zhi Tan; Angela F. Ku; Yong Wang; Chandrashekhar Madasu; Jian-Yuan Li; Suni Tang; Gabriella Miklossy; Stephen S. Palmer; Damian W. Young; Martin M. Matzuk J. Med. Chem., 2023, 66(3), 2143-2160 https://doi.org/10.1021/acs.jmedchem.2c01886AbstractThe discovery of monokinase-selective inhibitors for patients is challenging because the 500+ kinases encoded by the human genome share highly conserved catalytic domains. Until now, no selective inhibitors unique for a single transforming growth factor β (TGFβ) family transmembrane receptor kinase, including bone morphogenetic protein receptor type 2 (BMPR2), have been reported. This dearth of receptor-specific kinase inhibitors hinders therapeutic options for skeletal defects and cancer as a result of an overactivated BMP signaling pathway. By screening 4.17 billion “unbiased” and “kinase-biased” DNA-encoded chemical library molecules, we identified hits CDD-1115 and CDD-1431, respectively, that were low-nanomolar selective kinase inhibitors of BMPR2. Structure–activity relationship studies addressed metabolic lability and high-molecular-weight issues, resulting in potent and BMPR2-selective inhibitor analogs CDD-1281 (IC50 = 1.2 nM) and CDD-1653 (IC50 = 2.8 nM), respectively. Our work demonstrates that DNA-encoded chemistry technology (DEC-Tec) is reliable for identifying novel first-in-class, highly potent, and selective kinase inhibitors.
Ningke Hou; Lei Shuai; Lijing Zhang; Xuping Xie; Kaiming Tang; Yunkai Zhu; Yin Yu; Wenyi Zhang; Qiaozhu Tan; Gongxun Zhong; Zhiyuan Wen; Chong Wang; Xijun He; Hong Huo; Haishan Gao; You Xu; Jing Xue; Chen Peng; Jing Zou; Craig Schindewolf; Vineet Menachery; Wenji Su; Youlang Yuan; Zuyuan Shen; Rong Zhang; Shuofeng Yuan; Hongtao Yu; Peiyong Shi; Zhigao Bu; Jing Huang; Qi Hu ACS Cent. Sci., 2023, 9(2), 217-227 https://doi.org/10.1021/acscentsci.2c01359AbstractThe 3C-like protease (3CLpro) is an essential enzyme for the replication of SARS-CoV-2 and other coronaviruses and thus is a target for coronavirus drug discovery. Nearly all inhibitors of coronavirus 3CLpro reported so far are covalent inhibitors. Here, we report the development of specific, noncovalent inhibitors of 3CLpro. The most potent one, WU-04, effectively blocks SARS-CoV-2 replications in human cells with EC50 values in the 10-nM range. WU-04 also inhibits the 3CLpro of SARS-CoV and MERS-CoV with high potency, indicating that it is a pan-inhibitor of coronavirus 3CLpro. WU-04 showed anti-SARS-CoV-2 activity similar to that of PF-07321332 (Nirmatrelvir) in K18-hACE2 mice when the same dose was administered orally. Thus, WU-04 is a promising drug candidate for coronavirus treatment.
Xin Han; Yi Sun MedComm, 2023, 4(3), e290 https://doi.org/10.1002/mco2.290AbstractProteolysis targeting chimera (PROTAC) technology has become a powerful strategy in drug discovery, especially for undruggable targets/proteins. A typical PROTAC degrader consists of three components: a small molecule that binds to a target protein, an E3 ligase ligand (consisting of an E3 ligase and its small molecule recruiter), and a chemical linker that hooks first two components together. In the past 20 years, we have witnessed advancement of multiple PROTAC degraders into the clinical trials for anticancer therapies. However, one of the major challenges of PROTAC technology is that only very limited number of E3 ligase recruiters are currently available as E3 ligand for targeted protein degradation (TPD), although human genome encodes more than 600 E3 ligases. Thus, there is an urgent need to identify additional effective E3 ligase recruiters for TPD applications. In this review, we summarized the existing RING-type E3 ubiquitin ligase and their small molecule recruiters that act as effective E3 ligands of PROTAC degraders and their application in anticancer drug discovery. We believe that this review could serve as a reference in future development of efficient E3 ligands of PROTAC technology for cancer drug discovery and development.
Sameek Singh; Wenzhi Tian; Zachary C. Severance; Santosh K. Chaudhary; Viktoriya Anokhina; Basudeb Mondal; Rajaiah Pergu; Prashant Singh; Uttam Dhawa; Santanu Singha; Amit Choudhary Chem. Soc. Rev., 2023, 52, 5485-5515 https://doi.org/10.1039/D2CS00943AAbstractLiving systems use proximity to regulate biochemical processes. Inspired by this phenomenon, bifunctional modalities that induce proximity have been developed to redirect cellular processes. An emerging example of this class is molecules that induce ubiquitin-dependent proteasomal degradation of a protein of interest, and their initial development sparked a flurry of discovery for other bifunctional modalities. Recent advances in this area include modalities that can change protein phosphorylation, glycosylation, and acetylation states, modulate gene expression, and recruit components of the immune system. In this review, we highlight bifunctional modalities that perform functions other than degradation and have great potential to revolutionize disease treatment, while also serving as important tools in basic research to explore new aspects of biology.
Keqing Nian; Yifang Liu; Laura Brigandi; Sara H. Rouhanifard bioRxiv, 2022 https://doi.org/10.1101/2022.11.10.515969AbstractOf the various conjugation strategies for cellular biomolecules, Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) is the preferred click chemistry approach due to its fast reaction rate and the commercial availability of a wide range of conjugates. While extracellular labeling of biomolecules using CuAAC has been widely adopted, intracellular labeling in live cells has been challenging; the high copper concentration required for CuAAC reaction can be toxic to biological systems. As a critical first step towards achieving intracellular labeling with CuAAC, an ultrasensitive CuAAC ligand is needed to reduce the required copper concentration while maintaining fast reaction kinetics. Here, we develop a new DNA oligomer-conjugated CuAAC ligand for click reaction biomolecular labeling. The DNA oligo attachment serves several purposes: 1. Increases availability of local copper atoms in proximity to the ligand, which drives up reaction rates, 2. Enables the ligation of azide tags with up to 10-fold lower copper concentrations as compared to commercially available CuAAC ligands, 3. Allows nucleic acid template-driven proximity ligation through the choice of the attached DNA sequence and 4. Allows the CuAAC ligand and copper to traverse the cell and nuclear membrane. We demonstrate that this ligand enhances the intracellular 5-ethynyl uridine labeling of nascent RNAs using fluorogenic dyes. We also show that our DNA-enhanced CuAAC ligand enables the ligation of fluorogenic dyes to label both sialylated glycans on the surface on live cells as well as the live-cell intracellular labeling of nascent RNAs. This new ligand advances our efforts toward the final goal of applying CuAAC for live-cell applications.
Yiqun Chang; Bryson A. Hawkins; Jonathan J. Du; Paul W. Groundwater; David E. Hibbs; Felcia Lai Pharmaceutics. 2023, 15(1), 49 https://doi.org/10.3390/pharmaceutics15010049AbstractThe drug discovery process is a rocky path that is full of challenges, with the result that very few candidates progress from hit compound to a commercially available product, often due to factors, such as poor binding affinity, off-target effects, or physicochemical properties, such as solubility or stability. This process is further complicated by high research and development costs and time requirements. It is thus important to optimise every step of the process in order to maximise the chances of success. As a result of the recent advancements in computer power and technology, computer-aided drug design (CADD) has become an integral part of modern drug discovery to guide and accelerate the process. In this review, we present an overview of the important CADD methods and applications, such as in silico structure prediction, refinement, modelling and target validation, that are commonly used in this area.
Ayun Luo; Hongxia Zhou; Qini Hua; Yufang An; Hangke Ma; Xue Zhao; Kexin Yang; Yunjin Hu ACS Med. Chem. Lett., 2023, 14(3), 270-277 https://doi.org/10.1021/acsmedchemlett.2c00477AbstractAn efficient approach for aryl acetylene DNA-encoded library (DEL) synthesis was developed in this study by transition-metal-mediated inverse Sonogashira reaction of 1-iodoalkyne with boronic acid under ambient conditions, with moderate to excellent conversions and broad substrate adaptability for the first time. Compared to palladium-phosphine, copper iodide performed better in the on-DNA inverse Sonogashira reaction. Interestingly, substrate diversity can be enhanced by first interrogating coupling reagents under copper-promoted conditions, and then revalidating them under palladium-facilitated conditions for those reagents which failed under the former. This complementary validation strategy is particularly well-fitted to any DEL validation studies.
Changyang Liu; Xianfeng Li; Juan Zhang; Yangfeng Li; Gong Zhang; Yizhou Li J. Org. Chem., 2023, 88(11), 6565-6572 https://doi.org/10.1021/acs.joc.2c02686AbstractThe incorporation of N-containing heterocycles with potential bioactivity into DNA-encoded chemical libraries (DELs) represents an important approach to synthesizing medicinally useful compound collections for high-throughput screening. Herein, we reported a synthetic methodology to afford a benzotriazinone core as a drug-like scaffold in a DNA-compatible manner through aryl diazonium intermediates. Starting from DNA-conjugated amines, anthranilic acid or isatoic anhydride building blocks were coupled to form chemically diversified anthranilamides, which were subsequently transformed into 1,2,3-benzotriazin-4(3H)-one via tert-butyl nitrite-triggered cyclization. This methodology features DEL synthesis compatibility through a mild diazonium intermediate mechanism, allowing late-stage decoration of the bioactive benzotriazinone cap on DNA-conjugated amines. The broad substrate scope and high conversion render this methodology a promising approach to diversifying and decorating DNA-encoded combinatorial peptide-like libraries with medicinally relevant heterocyclic moieties.
Qigui Nie; Jie Sun; Xianfu Fang; Xun He; Feng Xiong; Gong Zhang; Yangfeng Li; Yizhou Li Chin. Chem. Lett., 2023, 33(5), 2559-2563 https://doi.org/10.1016/j.cclet.2023.108132AbstractDNA-encoded chemical libraries technology has become a novel approach to finding hit compounds in early drug discovery. The chemical space in a DEL would be expanded to realize its full potential, especially when integrating privileged scaffold dihydroquinazoline that has demonstrated a variety of diverse bioactivities. Driven by the requirement of parallel combinatorial synthesis, we here report a facile synthesis of on-DNA dihydroquinazolinone from aldehyde and anthranilamide. This DNA-compatible reaction was promoted by antimony trichloride, which has been proven to accelerate the reaction and improve conversions. Notably, the broad substrate scope of aldehydes and anthranilamides was explored under the mild reaction condition to achieve moderate-to-excellent conversion yields. We further applied the reaction into on-DNA macrocyclization, obtaining macrocycles embedded dihydroquinazolinone scaffold in synthetically useful conversion yields.
Yanrui Suo; Min Xu; Meimei Sun; Weiwei Lu; Xuan Wang; Xiaojie Lu Org. Lett., 2022, 24(49), 9092-9096 https://doi.org/10.1021/acs.orglett.2c03759AbstractIndane and isoindoline are attractive bicyclic systems in biologically active compounds but are rarely reported in DNA-encoded libraries. In this paper, we reported an efficient and versatile approach for assembling indane and isoindoline scaffolds via a ruthenium-catalyzed [2 + 2 + 2] cyclotrimerization reaction. This method exhibits a broad substrate scope and has been successfully applied to construct a 53K-membered DNA-encoded library (DEL). In order to test its application, we carried out a preliminary selection of this DEL against Aurora A protein and identified a hit compound with 9.3 μM inhibition activity.
Xiong Feng; Honggui Xu; Mingao Yu; Xingyu Chen; Zhenmin Zhong; Yuhan Guo; Meihong Chen; Huanfang Ou; Jiaqi Wu; Anhua Xie; Jiaqi Xiong; Linlin Xu; Lanmei Zhang; Qijian Zhong; Liye Huang; Zhenwei Li; Tianyuan Zhang; Feng Jin; Xun He Front. Pharmacol., 2022, 13, 1085665 https://doi.org/10.3389/fphar.2022.1085665AbstractMolecular generation (MG) via machine learning (ML) has speeded drug structural optimization, especially for targets with a large amount of reported bioactivity data. However, molecular generation for structural optimization is often powerless for new targets. DNA-Encoded Libraries (DEL) can generate systematic, target-specific activity data, including novel targets with few or unknown activity data. Therefore, this study aims to overcome the limitation of molecular generation in the structural optimization for the new target. Firstly, we generated molecules using the structure-affinity data (2.96 million samples) for 3C-like protease (3CLpro) from our own-built DEL platform to get rid of using public databases (e.g., CHEMBL and ZINC). Subsequently, to analyze the effect of transfer learning on the positive rate of the molecule generation model, molecular docking and affinity model based on DEL data was applied to explore the enhanced impact of transfer learning on molecule generation. In addition, the generated molecules are subjected to multiple filtering, including physicochemical properties, drug-like properties, and pharmacophore evaluation to determine the molecules for further study and verified by molecular dynamics simulation.
Weilin Lin; Shanil Gandhi; Prabesh Bhattarai; Keida Kasa; Caghan Kizil; Yixin Zhang E. J. Med. Chem., 2023, 115078 https://doi.org/10.1016/j.ejmech.2022.115078AbstractIdentifying new chemical structures against protein targets of interest represents one of the major challenges in drug discovery. As the major experimental method, high throughput screenings are performed with existing chemical libraries, thus restricting its capability to explore high molecular diversity. Herein, we report the use of high throughput array synthesis technology, in combination with growth algorithm, to discover binders for proinflammatory cytokine TNF-α. After 6 iterations of Library design – Array synthesis – Screening (i-LAS), one identified compound T17 has shown a kd value of 14.8 μM, and can rescue L929 cells from TNF-α mediated cytotoxicity. Further engineering T17 in various forms of oligomers have led to low nM binders. More interestingly, through tuning the multi-valent interaction with TNF-α, the high affinity oligomers can be switched from inhibitors to activators, leading to the hypothesis of an oligomerization-induced receptor activation mechanism. The i-LAS technology has allowed us to discover new binder structures, which can be further engineered into molecules with novel properties.
Yasaman Mahdavi-Amiri; Molly S. J. Hu; Nicole Frias; Matina Movahedi; Adam Csakai; Lisa A. Marcaurelle; Ryan Hili Org. Biomol. Chem., 2023, 21(7), 1463-1467 https://doi.org/10.1039/D2OB01956FAbstractAn efficient approach to the photoredox-catalysed hydroaminoalkylation between on-DNA secondary N-substituted (hetero)arylamines and vinylarenes has been developed and explored. The methodology was examined with a broad scope of vinylarenes and secondary arylamines to establish a preferred building block profile for the process. Compatible substrates furnished the desired derivitised amine products in modest to excellent conversions and with minimal or no detectable by-products.
Liam Hudson; Jeremy W. Mason; Matthias V. Westphal; Matthieu J. R. Richter; Jonathan R. Thielman; Bruce K. Hua; Christopher J. Gerry; Guoqin Xia; Heather L. Osswald; John M. Knapp; Zher Yin Tan; Praveen Kokkonda; Ben I. C. Tresco; Shuang Liu; Andrew G. Reidenbach; Katherine S. Lim; Jennifer Poirier; John Capece; Simone Bonazzi; Christian M. Gampe; Nichola J. Smith; James E. Bradner; Connor W. Coley; Paul A. Clemons; Bruno Melillo; C. Suk-Yee Hon; Johannes Ottl; Christoph E. Dumelin; Jonas V. Schaefer; Ann Marie E. Faust; Frédéric Berst; Stuart L. Schreiber; Frédéric J. Zécri; Karin Briner Nat. Commun. 2023, 14, 4930 https://doi.org/10.1038/s41467-023-40575-5AbstractDiversity-oriented synthesis (DOS)is a powerful strategy to prepare molecules with underrepresented features in commercial screening collections, resulting in the elucidation of novel biological mechanisms. In parallel to the development of DOS, DNA-encoded libraries (DELs) have emerged as an effective, efficient screening strategy to identify protein binders. Despite recent advancements in this field, most DEL syntheses are limited by the presence of sensitive DNA-based constructs. Here, we describe the design, synthesis, and validation experiments performed for a 3.7 million-member DEL, generated using diverse skeleton architectures with varying exit vectors, derived from DOS, to achieve structural diversity beyond what is possible by varying appendages alone. We will make this DEL available to the academic scientific community to increase access to novel structural features and accelerate early-phase drug discovery.
Millicent Dockerill, Nicolas Winssinger Angew. Chem. Int. Ed. Engl., 2023, 62(9), e202215542 https://doi.org/10.1002/anie.202215542AbstractDNA-encoded library (DEL) technologies are transforming the drug discovery process, enabling the identification of ligands at unprecedented speed and scale. DEL makes use of libraries that are orders of magnitude larger than traditional high throughput screens. While a DNA tag alludes to a genotype-phenotype connection that is exploitable for evolutionary cycles, most of the work in the field is performed with libraries where the tag serves as an amplifiable bar code but does not allow ‘translation’ into the synthetic product it is linked to. In this review, we cover technologies that enable the ‘translation’ of the genetic tag into synthetic molecules, both biochemically and chemically, and explore how it can be used to harness Darwinian evolutionary pressure.
Seoyeon Jeong; Hwa-Ryeon Kim; June-Ha Shin; Min-Hee Son; In-Hyun Lee; Jae-Seok Roe Mol. Ther. Nucleic Acids, 2023, 32, 637-649 https://doi.org/10.1016/j.omtn.2023.04.023AbstractTargeting aberrant epigenetic programs that drive tumorigenesis is a promising approach to cancer therapy. DNA-encoded small-molecule library (DEL) screening is a core platform technology increasingly used to identify drugs that bind to protein targets. Here, we use DEL screening against bromodomain and extra-terminal motif (BET) proteins to identify inhibitors with new chemotypes, and successfully identified BBC1115 as a selective BET inhibitor. While BBC1115 does not structurally resemble OTX-015, a clinically active pan-BET inhibitor, our intensive biological characterization revealed that BBC1115 binds to BET proteins, including BRD4, and suppresses aberrant cell fate programs. Phenotypically, BBC1115-mediated BET inhibition impaired proliferation in acute myeloid leukemia, pancreatic, colorectal, and ovarian cancer cells in vitro. Moreover, intravenous administration of BBC1115 inhibited subcutaneous tumor xenograft growth with minimal toxicity and favorable pharmacokinetic properties in vivo. Since epigenetic regulations are ubiquitously distributed across normal and malignant cells, it will be critical to evaluate if BBC1115 affects normal cell function. Nonetheless, our study shows integrating DEL-based small-molecule compound screening and multi-step biological validation represents a reliable strategy to discover new chemotypes with selectivity, efficacy, and safety profiles for targeting proteins involved in epigenetic regulation in human malignancies.
Qiuxia Chen; Chuan Liu; Wei Wang; Xiaoyun Meng; Xuemin Cheng; Xianyang Li; Longying Cai; Linfu Luo; Xu He; Huan Qu; Jing Luo; Hong Wei; Sen Gao; Guansai Liu; Jinqiao Wan; David I. Israel; Jin Li; Dengfeng Dou ACS Chem. Biol., 2023, 18(1), 25-33 https://doi.org/10.1021/acschembio.2c00797AbstractThe proteolysis targeting chimera (PROTAC) strategy results in the down-regulation of unwanted protein(s) for disease treatment. In the PROTAC process, a heterobifunctional degrader forms a ternary complex with a target protein of interest (POI) and an E3 ligase, which results in ubiquitination and proteasomal degradation of the POI. While ternary complex formation is a key attribute of PROTAC degraders, modification of the PROTAC molecule to optimize ternary complex formation and protein degradation can be a labor-intensive and tedious process. In this study, we take advantage of DNA-encoded library (DEL) technology to efficiently synthesize a vast number of possible PROTAC molecules and describe a parallel screening approach that utilizes DNA barcodes as reporters of ternary complex formation and cooperative binding. We use a designed PROTAC DEL against BRD4 and CRBN to describe a dual protein affinity selection method and the direct discovery of novel, potent BRD4 PROTACs that importantly demonstrate clear SAR. Such an approach evaluates all the potential PROTACs simultaneously, avoids the interference of PROTAC solubility and permeability, and uses POI and E3 ligase proteins in an efficient manner.
Christopher R. Gardner; Katherine A. Davies; Ying Zhang; Martin Brzozowski; Peter E. Czabotar; James M. Murphy; Guillaume Lessene J. Med. Chem., 2023, 66(4), 2361-2385 https://doi.org/10.1021/acs.jmedchem.2c01621AbstractNecroptosis is a regulated caspase-independent form of necrotic cell death that results in an inflammatory phenotype. This process contributes profoundly to the pathophysiology of numerous neurodegenerative, cardiovascular, infectious, malignant, and inflammatory diseases. Receptor-interacting protein kinase 1 (RIPK1), RIPK3, and the mixed lineage kinase domain-like protein (MLKL) pseudokinase have been identified as the key components of necroptosis signaling and are the most promising targets for therapeutic intervention. Here, we review recent developments in the field of small-molecule inhibitors of necroptosis signaling, provide guidelines for their use as chemical probes to study necroptosis, and assess the therapeutic challenges and opportunities of such inhibitors in the treatment of a range of clinical indications.
Devan J. Shell; Justin M. Rectenwald; Peter H. Buttery; Rebecca L. Johnson; Caroline A. Foley; Shiva K. R. Guduru; Mélanie Uguen; Juanita Sanchez Rubiano; Xindi Zhang; Fengling Li; Jacqueline L. Norris-Drouin; Matthew Axtman; P. Brian Hardy; Masoud Vedadi; Stephen V. Frye; Lindsey I. James; Kenneth H. Pearce SLAS Discov., 2022, 27(8), 428-439 https://doi.org/10.1016/j.slasd.2022.10.003AbstractMethyl-lysine (Kme) reader domains are prevalent in chromatin regulatory proteins which bind post-translational modification sites to recruit repressive and activating factors; therefore, these proteins play crucial roles in cellular signaling and epigenetic regulation. Proteins that contain Kme domains are implicated in various diseases, including cancer, making them attractive therapeutic targets for drug and chemical probe discovery. Herein, we report on expanding the utility of a previously reported, Kme-focused DNA-encoded library (DEL), UNCDEL003, as a screening tool for hit discovery through the specific targeting of Kme reader proteins. As an efficient method for library generation, focused DELs are designed based on structural and functional features of a specific class of proteins with the intent of novel hit discovery. To broadly assess the applicability of our library, UNCDEL003 was screened against five diverse Kme reader protein domains (53BP1 TTD, KDM7B JmjC-PHD, CDYL2 CD, CBX2 CD, and LEDGF PWWP) with varying structures and functions. From these screening efforts, we identified hit compounds which contain unique chemical scaffolds distinct from previously reported ligands. The selected hit compounds were synthesized off-DNA and confirmed using primary and secondary assays and assessed for binding selectivity. Hit compounds from these efforts can serve as starting points for additional development and optimization into chemical probes to aid in further understanding the functionality of these therapeutically relevant proteins.
Alba L. Montoya; Marta Glavatskikh; Brayden J. Halverson; Lik Hang Yuen; Herwig Schüler; Dmitri Kireev; Raphael M. Franzini Eur. J. Med. Chem., 2023, 246, 114980 https://doi.org/10.1016/j.ejmech.2022.114980AbstractDNA-encoded chemical libraries (DECLs) interrogate the interactions of a target of interest with vast numbers of molecules. DECLs hence provide abundant information about the chemical ligand space for therapeutic targets, and there is considerable interest in methods for exploiting DECL screening data to predict novel ligands. Here we introduce one such approach and demonstrate its feasibility using the cancer-related poly-(ADP-ribose)transferase tankyrase 1 (TNKS1) as a model target. First, DECL affinity selections resulted in structurally diverse TNKS1 inhibitors with high potency including compound 2 with an IC50 value of 0.8 nM. Additionally, TNKS1 hits from four DECLs were translated into pharmacophore models, which were exploited in combination with docking-based screening to identify TNKS1 ligand candidates in databases of commercially available compounds. This computational strategy afforded TNKS1 inhibitors that are outside the chemical space covered by the DECLs and yielded a drug-like lead compound 12 with an IC50 value of 22 nM. The study further provided insights in the reliability of screening data and the effect of library design on hit compounds. In particular, the study revealed that while in general DECL screening data are in good agreement with off-DNA ligand binding, unpredictable interactions of the linker scar with the target protein contribute to the noise in the affinity selection data.
Jeremy W. Mason; Liam Hudson; Matthias V. Westphal; Antonin Tutter; Gregory Michaud; Wei Shu; Xiaolei Ma; Connor W. Coley; Paul A Clemons; Simone Bonazzi; Frédéric Berst; Frédéric J. Zécri; Karin Briner; Stuart L. Schreiber bioRxiv, 2022 https://doi.org/10.1101/2022.10.13.512184AbstractMolecular glues and bifunctional compounds that induce protein-protein associations provide a powerful and general mechanism to modulate cell circuitry. We sought to develop a platform for the direct discovery of compounds able to induce association of any two pre-selected proteins, using the first bromodomain of BRD4 and the VHL-elongin C-elongin B (VCB) complex as a test system. Leveraging the screening power of DNA-encoded libraries (DELs), we synthesized ~one million DNA-encoded compounds that possess a VHL-targeting fragment, a variety of connectors, and a diversity element generated by split-and-pool combinatorial chemistry. By screening our DEL against BRD4 in the presence and absence of VCB, we could identify VHL-bound molecules that simultaneously bind BRD4. For highly barcode-enriched library members, ternary complex formation leading to BRD4 degradation was confirmed in cells. Furthermore, a ternary complex crystal structure was obtained for the most enriched library member. Our work provides a foundation for adapting DEL screening to the discovery of proximity-inducing small molecules.
Yue Zhang; Shengdi Xia; Wenxia Shi; Bizhen Lin; Xiaocan Su; Weiwei Lu; Xinyuan Wu; Xuan Wang; Xiaojie Lu; Ming Yan; Xuejing Zhang Org. Lett., 2022, 24(43), 7961-7966 https://doi.org/10.1021/acs.orglett.2c03077AbstractThe photocatalyst-free electron donor–acceptor (EDA) complex photochemistry was deemed to expand the potential of photodriven radical chemistry. Here, we report a cross-coupling reaction of thianthrenium salt functionalized arenes and sodium sulfinates via a photopromoted single electron transfer (SET) process of an EDA complex. A series of biarylsulfones were obtained with high site-selectivity and reactivity. This mild and practical radical reaction could be applied on the bioactive and DNA-encoded molecules.
Vincent Blay; Xiaoyu Li; Jacob Gerlach; Fabio Urbina; Sean Ekins Drug Discov. Today, 2022, 103351 https://doi.org/10.1016/j.drudis.2022.103351AbstractDNA-encoded libraries (DELs) allow starting chemical matter to be identified in drug discovery. The volume of experimental data generated also makes DELs an attractive resource for machine learning (ML). ML allows modeling complex relationships between compounds and numerical endpoints, such as the binding to a target measured by DELs. DELs could also empower other areas of drug discovery. Here, we propose that DELs and ML could be combined to model binding to off-targets, enabling better predictive toxicology. With enough data, ML models can make accurate predictions across a vast chemical space, and they can be reused and expanded across projects. Although there are limitations, more general toxicology models could be applied earlier during drug discovery, illuminating safety liabilities at a lower cost.
Sebastian Oehler; Laura Lucaroni; Francesca Migliorini; Abdullah Elsayed; Luca Prati; Sara Puglioli; Mattia Matasci; Kristina Schira; Jörg Scheuermann; Denis Yudin; Min Jia; Nenad Ban; Dave Bushnell; Roger Kornberg; Samuele Cazzamalli; Dario Neri; Nicholas Favalli; Gabriele Bassi Nat. Chem., 2023, 15(10), 1431-1443 https://doi.org/10.1038/s41557-023-01257-3AbstractDNA-encoded chemical libraries (DELs) consist of large chemical compound collections individually linked to DNA barcodes, facilitating pooled construction and screening. However, screening campaigns often fail if the molecular arrangement of the building blocks is not conducive to an efficient interaction with a protein target. Here we postulated that the use of rigid, compact and stereo-defined central scaffolds for DEL synthesis may facilitate the discovery of very specific ligands capable of discriminating between closely related protein targets. We synthesized a DEL comprising 3,735,936 members, featuring the four stereoisomers of 4-aminopyrrolidine-2-carboxylic acid as central scaffolds. The library was screened in comparative selections against pharmaceutically relevant targets and their closely related protein isoforms. Hit validation results revealed a strong impact of stereochemistry, with large affinity differences between stereoisomers. We identified potent isozyme-selective ligands against multiple protein targets. Some of these hits, specific to tumour-associated antigens, demonstrated tumour-selective targeting in vitro and in vivo. Collectively, constructing DELs with stereo-defined elements contributed to high library productivity and ligand selectivity.
Divya Vemula; Perka Jayasurya; Varthiya Sushmitha; Yethirajula Naveen Kumar; Vasundhra Bhandari Eur. J. Pharm. Sci., 2023, 181, 106324 https://doi.org/10.1016/j.ejps.2022.106324AbstractComputer-aided drug design (CADD) is an emerging field that has drawn a lot of interest because of its potential to expedite and lower the cost of the drug development process. Drug discovery research is expensive and time-consuming, and it frequently took 10-15 years for a drug to be commercially available. CADD has significantly impacted this area of research. Further, the combination of CADD with Artificial Intelligence (AI), Machine Learning (ML), and Deep Learning (DL) technologies to handle enormous amounts of biological data has reduced the time and cost associated with the drug development process. This review will discuss how CADD, AI, ML, and DL approaches help identify drug candidates and various other steps of the drug discovery process. It will also provide a detailed overview of the different in silico tools used and how these approaches interact.
Quentin M. R. Gibaut; Yoshihiro Akahori; Jessica A. Bush; Amirhossein Taghavi; Toru Tanaka; Haruo Aikawa; Lucas S. Ryan; Brian M. Paegel; Matthew D. Disney J. Am. Chem. Soc., 2022, 144(48), 21972-21979 https://doi.org/10.1021/jacs.2c08883AbstractA solid-phase DNA-encoded library (DEL) was studied for binding the RNA repeat expansion r(CUG)exp, the causative agent of the most common form of adult-onset muscular dystrophy, myotonic dystrophy type 1 (DM1). A variety of uncharged and novel RNA binders were identified to selectively bind r(CUG)exp by using a two-color flow cytometry screen. The cellular activity of one binder was augmented by attaching it with a module that directly cleaves r(CUG)exp. In DM1 patient-derived muscle cells, the compound specifically bound r(CUG)exp and allele-specifically eliminated r(CUG)exp, improving disease-associated defects. The approaches herein can be used to identify and optimize ligands and bind RNA that can be further augmented for functionality including degradation.
Yuting Gao; Yang Sun; Guixian Zhao; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett., 2022, 24, 36, 6664–6669 https://doi.org/10.1021/acs.orglett.2c02714AbstractThe functionalized 4H-pyran scaffold has aroused synthetic attention because it is widely found in many interesting pharmacologically relevant compounds. We here disclose its incorporation into DNA-encoded chemical libraries, combining this scaffold with the merits of scaffold architecture in drug design. Under the optimized DNA-compatible conditions, functionalized 4H-pyrans were efficiently formed with a broad substrate scope. Among the 4H-pyrans formed, the axial structure features rotational restriction, and the spirocyclic structure provides rigidity and three-dimensionality. These efforts open the door for the construction of DNA-encoded chemical libraries with more consideration for this structural architecture.
Xianfeng Li; Juan Zhang; Changyang Liu; Jie Sun; Yangfeng Li; Gong Zhang; Yizhou Li Chem. Sci., 2022, 13, 13100-13109 https://doi.org/10.1039/D2SC04482JAbstractForging carbon-carbon (C-C) linkage in DNA-encoded combinatorial library synthesis represents a fundamental task for drug discovery, especially with broad substrate scope and exquisite functional group tolerance. Here we reported the palladium-catalyzed Suzuki-Miyaura , Heck and Hiyama type cross-coupling via DNA-conjugated aryl diazonium intermediates for DNA-encoded chemical library (DEL) synthesis. Starting from commodity arylamines, this synthetic route facilely delivers vast chemical diversity under mild temperature and pH, thus circumventing damage to fragile functional groups. Given its orthogonality with traditional aryl halide-based cross-coupling, the aryl diazonium-centered strategy expands the compatible synthesis of complex C-C bond-connected scaffolds. In addition, DNA-tethered pharmaceutical compounds (e.g., HDAC inhibitor) are constructed without decomposition of susceptible bioactive warheads (e.g., hydroxamic acid), emphasizing the superiority of the aryl diazonium-based approach. Together with the convenient transformation to aryl azide photo-crosslinker, aryl diazonium’s DNA-compatible diversification synergistically demonstrated its competence to create medicinally relevant combinatorial libraries and investigate protein-ligand interactions in pharmaceutical research.
Corina M. Borza; Gema Bolas; Ambra Pozzi Front. Pharmacol., 2022, 13, 1001122 https://doi.org/10.3389/fphar.2022.1001122AbstractFollowing injury the kidney undergoes a repair process, which results in replacement of the injured tissue with little evidence of damage. However, repetitive injuries or inability of the kidney to stop the repair process result in abnormal deposition of extracellular matrix (ECM) components leading to fibrosis and organ dysfunction. The synthesis/degradation of ECM components is finely regulated by several factors, including discoidin domain receptors (DDRs). These are receptor tyrosine kinases that are activated by collagens. Upon activation, DDRs control several cell functions that, when exacerbated, contribute to kidney injury and fibrosis. DDRs are undetectable in healthy kidney, but become rapidly upregulated in several kidney fibrotic conditions, thus making them attractive anti-fibrotic targets. DDRs contribute to kidney injury and fibrosis by promoting apoptosis of injured kidney cells, stimulating the production of pro-inflammatory cytokines, and regulating the production of ECM components. They achieve these effects by activating canonical intracellular molecules or by directly interacting with nuclear chromatin and promoting the transcription of pro-fibrotic genes. The goal of this review is to highlight canonical and non-canonical mechanisms whereby DDRs contribute to kidney injury/fibrosis. This review will summarize key findings obtained using cells and mice lacking DDRs and it will discuss the discovery and development of targeted DDR small molecule- and antisense-based inhibitors. Understanding the molecular mechanisms whereby DDRs control kidney injury and fibrosis might enable us to not only develop more selective and potent inhibitors, but to also determine when DDR inhibition needs to be achieved to prevent and/or halt the development of kidney fibrosis.
Doretta Cuffaro; Lidia Ciccone; Armando Rossello; Elisa Nuti; Salvatore Santamaria J. Med. Chem., 2022, 65(20), 13505-13532 https://doi.org/10.1021/acs.jmedchem.2c01177AbstractOsteoarthritis (OA) is the most common degenerative joint disease. In 1999, two members of the A Disintegrin and Metalloproteinase with Thrombospondin Motifs (ADAMTS) family of metalloproteinases, ADAMTS4 and ADAMTS5, or aggrecanases, were identified as the enzymes responsible for aggrecan degradation in cartilage. The first aggrecanase inhibitors targeted the active site by chelation of the catalytic zinc ion. Due to the generally disappointing performance of zinc-chelating inhibitors in preclinical and clinical studies, inhibition strategies tried to move away from the active-site zinc in order to improve selectivity. Exosite inhibitors bind to proteoglycan-binding residues present on the aggrecanase ancillary domains (called exosites). While exosite inhibitors are generally more selective than zinc-chelating inhibitors, they are still far from fulfilling their potential, partly due to a lack of structural and functional data on aggrecanase exosites. Filling this gap will inform the design of novel potent, selective aggrecanase inhibitors.
Lisa M. Baker; James B. Murray; Roderick E. Hubbard PDBJ - 8AHI https://www.rcsb.org/structure/8AHIAbstractPAC-FragmentDEL: Photoactivated covalent capture of DNA encoded fragments for hit discovery PDB DOI: 10.2210/pdb8AHI/pdb Classification: TRANSFERASE Organism(s): Homo sapiens Expression System: Escherichia coli BL21(DE3) Mutation(s): Yes Deposited: 2022-07-21 Released: 2022-09-14 Deposition Author(s): Baker, L.M., Murray, J.B., Hubbard, R.E. Funding Organization(s): Not funded Experimental Data Snapshot Method: X-RAY DIFFRACTION Resolution: 2.69 Å R-Value Free: 0.249 R-Value Work: 0.155
Zhaomei Sun; Jie Zhang; Huanqing Zhang; Hongli Cao; Lingqian Xiao; Kexin Yang; Yunjin Hu Bioconjugate Chem., 2022, 33, 9, 1585-1594 https://doi.org/10.1021/acs.bioconjchem.2c00340AbstractThrough a modified Kinugasa reaction, a novel method of amidation on terminal oligo alkyne conjugates by copper-promoted oxidation with nitrones has been developed. Unprotected bifunctional carboxylic acid–amine reagents can be transformed directly to the respective amide products under these edited Kinugasa reaction conditions. 3-Cycle DNA-encoded libraries (DELs) can be built in three steps of chemical conversion.
Meiying Cui; MIchelle Patino Gaillez; Dzung Nguyen; Stephan Heiden; Weilin Lin; Michael Thompson; Francesco Reddavide; Qinchang Chen; Yixin Zhang Nat. Commun., 2023, 14, 1481 https://doi.org/10.1038/s41467-023-37071-1AbstractThe split-and-pool method has been widely used to synthesize chemical libraries of a large size, albeit without the possibility of meaningful quality control. In contrast, a self-assembled DNA-encoded chemical library (DEL) allows us to construct an m x n-member library by mixing an m-member and an n-member pre-purified sub-libraries. Herein, we report a trio-pharmacophore DEL (T-DEL) of m x l x n members through assembling three pre-purified and validated sub-libraries. The middle sub-library is synthesized using DNA-templated synthesis with different reaction mechanisms and designed as a linkage connecting the fragments displayed on the flanking two sub-libraries. In spite of assembling three fragments, the resulting compounds do not exceed the up-to-date standard of molecular weight regarding drug-likeness. The utility of T-DEL has been demonstrated in linker optimization for known binding fragments against trypsin and carbonic anhydrase II and de novo selections against matrix metalloprotease-2 and − 9.
Gokul Gowri; Kuanwei Sheng; Peng Yin Nat. Comput. Sci., 2024 https://doi.org/10.1038/s43588-024-00646-zAbstractOrthogonal DNA barcode library design is an essential task in bioengineering. Here we present seqwalk, an efficient method for designing barcode libraries that satisfy a sequence symmetry minimization (SSM) heuristic for orthogonality, with theoretical guarantees of maximal or near-maximal library size under certain design constraints. Seqwalk encodes SSM constraints in a de Bruijn graph representation of sequence space, enabling the application of recent advances in discrete mathematics1 to the problem of orthogonal sequence design. We demonstrate the scalability of seqwalk by designing a library of >106 SSM-satisfying barcode sequences in less than 20 s on a standard laptop.
Ayun Luo; Ziqi Zhang; Fanming Zeng; Xiuming Wang; Xue Zhao; Kexin Yang; Yun Jin Hu Org. Lett., 2022, 24, 31, 5756–5761 https://doi.org/10.1021/acs.orglett.2c02237Abstractβ-Lactam antibiotics are one of the most important antibacterial drug classes worldwide. This work will present the first prototype on-DNA β-lactam combinatorial library with novel structures and chemical space properties that would be significant for phenotypic screening to identify the next generation of antibiotics to combat the pervasive problem of bacterial resistance.
Lu Chen; Xiaoqin Zhang; Yaqing Ou; Maoyu Liu; Dongke Yu; Zhiheng Song; Lihong Niu; Lijuan Zhang; Jianyou Shi Front. Pharmacol., 2022, 13, 976435 https://doi.org/10.3389/fphar.2022.976435AbstractProgrammed necrosis is a new modulated cell death mode with necrotizing morphological characteristics. Receptor interacting protein 1 (RIPK1) is a critical mediator of the programmed necrosis pathway that is involved in stroke, myocardial infarction, fatal systemic inflammatory response syndrome, Alzheimer’s disease, and malignancy. At present, the reported inhibitors are divided into four categories. The first category is the type I ATP-competitive kinase inhibitors that targets the area occupied by the ATP adenylate ring; The second category is type Ⅱ ATP competitive kinase inhibitors targeting the DLG-out conformation of RIPK1; The third category is type Ⅲ kinase inhibitors that compete for binding to allosteric sites near ATP pockets; The last category is others. This paper reviews the structure, biological function, and recent research progress of receptor interaction protein-1 kinase inhibitors.
Pratik R. Chheda; Nicholas Simmons; David P. Schuman; Zhicai Shi Org. Lett., 2022, 24(28), 5214-5219 https://doi.org/10.1021/acs.orglett.2c02113AbstractDeveloping new DNA-compatible reactions is key to expanding the accessible chemical space of DNA-encoded library (DEL) technology. Here we disclose the first report of a DNA-compatible carbonylative Suzuki coupling of DNA-conjugated (hetero)aryl iodides with (hetero)aryl boronic acids to access di(hetero)aryl ketones, a valuable structural motif present within several approved or clinically advanced small molecules. The reported DNA-compatible, Pd(OAc)2-mediated system is mild, uses a robust protocol, has a wide substrate scope for both coupling partners, is suitable for large-scale DEL productions, and provides a source of previously unexplored chemical matter for DEL screens.
Jessica L. Childs-Disney; Xueyi Yang; Quentin M. R. Gibaut; Yuquan Tong; Robert T. Batey; Matthew D. Disney Nat. Rev. Drug Discov., 2022, 21(10), 736-762 https://doi.org/10.1038/s41573-022-00521-4AbstractRNA adopts 3D structures that confer varied functional roles in human biology and dysfunction in disease. Approaches to therapeutically target RNA structures with small molecules are being actively pursued, aided by key advances in the field including the development of computational tools that predict evolutionarily conserved RNA structures, as well as strategies that expand mode of action and facilitate interactions with cellular machinery. Existing RNA-targeted small molecules use a range of mechanisms including directing splicing — by acting as molecular glues with cellular proteins (such as branaplam and the FDA-approved risdiplam), inhibition of translation of undruggable proteins and deactivation of functional structures in noncoding RNAs. Here, we describe strategies to identify, validate and optimize small molecules that target the functional transcriptome, laying out a roadmap to advance these agents into the next decade.
Gavin W. Collie; Louise Barlind; Sana Bazzaz; Ulf Börjesson; Ian L. Dale; Jeremy S. Disch; Sevan Habeshian; Rachael Jetson; Puneet Khurana; Andrew Madin; Iacovos N. Michaelides; Ling Peng; Arjan Snijder; Christopher J. Stubbs Bioorg. Med. Chem. Lett., 2022, 128948 https://doi.org/10.1016/j.bmcl.2022.128948AbstractThe c-MET receptor tyrosine kinase has received considerable attention as a cancer drug target yet there remains a need for inhibitors which are selective for c-MET and able to target emerging drug-resistant mutants. We report here the discovery, by screening a DNA-encoded chemical library, of a highly selective c-MET inhibitor which was shown by X-ray crystallography to bind to the kinase in an unprecedented manner. These results represent a novel mode of inhibiting c-MET with a small molecule and may provide a route to targeting drug-resistant forms of the kinase whilst avoiding potential toxicity issues associated with broad kinome inhibition.
Matic Proj; Krištof Bozovičar; Martina Hrast; Rok Frlan; Stanislav Gobec Bioorg. Med. Chem. Lett., 2022, 128915 https://doi.org/10.1016/j.bmcl.2022.128915AbstractScreening of DNA-encoded libraries is an emerging technology for discovering hits against protein targets. With the recent launch of the DELopen platform, a facile screening of 4.4 billion compounds is available to accelerate the drug discovery process. Here we report an affinity-based screening of the DELopen library for the first time. Screening was performed against two bacterial enzymes of the peptidoglycan biosynthetic pathway, N-acetylglucosamine-enolpyruvyl transferase (MurA) and D-alanine:D-alanine ligase (DdlB). Several binders were obtained and selected for off-DNA synthesis. Hits with confirmed inhibitory potency were deconstructed into smaller fragments. In this way, two new MurA inhibitors with antibacterial activity were obtained and are available for further optimization.
Yuting Gao; Yang Sun; Xianfu Fang; Guixian Zhao; Xufeng Li; Gong Zhang; Yangfeng Li; Yizhou Li Org. Chem. Front., 2022, 9, 4542-4548 https://doi.org/10.1039/D2QO00881EAbstractThe DNA-encoded chemical libraries (DELs) technology is a novel drug discovery tool, benefiting from its large number of compound collections with chemical space coverage. Vinyl sulfone is a privileged functional group, eliciting diverse pharmacological activities as the covalent warhead. In this report, we incorporated vinyl sulfone into the DEL chemistry field by developing a mild and efficient DNA-compatible method starting from simple sodium sulfinates and DNA-conjugated alkenes, affording a broad substrate scope with moderate-to-excellent conversions. Further chemical transformations including thiol-Michael addition reaction, together with enzymatic ligation, were also successfully implemented to demonstrate the feasibility of this DNA-compatible synthesis in DEL construction.
Ke Li; Craig M. Crews Chem. Soc. Rev., 2022, 51(12), 5214-5236 https://doi.org/10.1039/D2CS00193DAbstractProteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules consisting of one ligand that binds to a protein of interest (POI) and another that can recruit an E3 ubiquitin ligase. The chemically-induced proximity between the POI and E3 ligase results in ubiquitination and subsequent degradation of the POI by the ubiquitin-proteasome system (UPS). The event-driven mechanism of action (MOA) of PROTACs offers several advantages compared to traditional occupancy-driven small molecule inhibitors, such as a catalytic nature, reduced dosing and dosing frequency, a more potent and longer-lasting effect, an added layer of selectivity to reduce potential toxicity, efficacy in the face of drug-resistance mechanisms, targeting nonenzymatic functions, and expanded target space. Here, we highlight important milestones and briefly discuss lessons learned about targeted protein degradation (TPD) in recent years and conjecture on the efforts still needed to expand the toolbox for PROTAC discovery to ultimately provide promising therapeutics.
Aapo Aho; Tommi Österlund; Jani Rahkila; Pasi Virta Eur. J. Org. Chem., 2022, e202200583 https://doi.org/10.1002/ejoc.202200583AbstractDNA-templated formation and N,O-transacetalization of N-methoxyoxazolidines have been studied. Compared to the reaction without a DNA-catalyst, the hybridization-driven N-methoxyoxazolidine formation shows a marked rate acceleration, whereas the rate of corresponding N,O-transacetalization is limited by the rate of decay to aldehyde intermediates. In both cases, the equilibrium yield increases markedly on the DNA template. Different hairpin architectures have been studied to evaluate the role and limits of the template effect. Furthermore, an attention has been paid to stereochemical integrity (R/S) of the N-methoxyoxazolidine linkage. The N-methoxyoxazolidine formation represents a dynamic pH-responsive DNA-templated ligation that occurs readily in slightly acidic conditions (pH 5).
Katharina Götte; Robin Dinter; Leon Justen; Norbert Kockmann; Andreas Brunschweiger ACS Omega, 2022, 7, 32, 28369–28377 https://doi.org/10.1021/acsomega.2c02906AbstractDNA-encoded library technologies require high-throughput, compatible, and well automatable platforms for chemistry development, building block rehearsal, and library synthesis. An affinity-based process using Watson–Crick interactions was developed that enables purification of DNA-tagged compounds from complex reaction mixtures. The purification relies on a single-stranded DNA-oligonucleotide, called capture strand, which was covalently coupled to an agarose matrix and to which a DNA-compound conjugate from a DNA-encoded library (DEL) reaction can be reversibly annealed to. The thus-formed DNA duplex tolerated surprisingly stringent washing conditions with multiple solvents to remove excess reactants and reagents. The tolerated solvents included aqueous buffers, aqueous EDTA solutions to remove metal ions, aqueous mixtures of organic solvents, and even pure organic solvents. The purified DNA-conjugate was eluted with aqueous ammonia and could be used for reaction analysis or for instance in DNA-encoded library synthesis. The lab equipment for purification was tailored for automation with open-source hardware and constructed by 3D printing.
Jie Sun; Qigui Nie; Xianfu Fang; Zhiwei He; Gong Zhang; Yangfeng Li; Yizhou Li Org. Biomol. Chem., 2022, 20(25), 5045-5049 https://doi.org/10.1039/D2OB00862AAbstractInspired by diversity-oriented synthesis, we have developed a series of DNA-compatible transformations utilizing on-DNA vinyl azide as a synthon to forge divergent N-heterocyclic scaffolds. Polysubstituted imidazoles and isoquinolines were efficiently obtained with moderate-to-excellent conversions. Besides, the “one-pot” strategy to prepare in-house on-DNA vinyl azides afforded synthons readily. Results from substrate scope exploration and enzymatic ligation further demonstrate the feasibility of these N-heterocycle syntheses in DNA-encoded chemical library construction.
Silvia Chines; Christiane Ehrt; Marco Potowski; Felix Biesenkamp; Lars Grützbach; Susanne Brunner; Frederik van den Broek; Shilpa Bali; Katja Ickstadt; Andreas Brunschweiger Chem. Sci., 2022, 13(37), 11221-11231 https://doi.org/10.1039/D2SC02474HAbstractDatabases contain millions of reactions for compound synthesis, rendering selection of reactions for forward synthetic design of small molecule screening libraries, such as DNA-encoded libraries (DELs), a big data challenge. To support reaction space navigation, we developed the computational workflow Reaction Navigator. Reaction files from a large chemistry database were processed using the open-source KNIME Analytics Platform. Initial processing steps included a customizable filtering cascade that removed reactions with a high probability to be incompatible with DEL, as they would e.g. damage the genetic barcode, to arrive at a comprehensive list of transformations for DEL design with applicability potential. These reactions were displayed and clustered by user-defined molecular reaction descriptors which are independent of reaction core substitution patterns. Thanks to clustering, these can be searched to identify reactions for DEL synthesis according to desired reaction criteria, such as ring formation or sp3 content. The workflow was initially applied for mapping chemical reaction space for aromatic aldehydes as an exemplary functional group often used in DEL synthesis. Exemplary reactions have been successfully translated to DNA-tagged substrates and can be applied to library synthesis. The versatility of the Reaction Navigator was then shown by mapping reaction space for different reaction conditions, for amines, and for data from a second database.
Sara Puglioli; Eleonore Schmidt; Christian Pellegrino; Luca Prati; Sebastian Oehler; Roberto De Luca; Andrea Galbiati; Claudia Comacchio; Lisa Nadal; Jörg Scheuermann; Markus G. Manz; Dario Neri; Samuele Cazzamalli; Gabriele Bassi; Nicholas Favalli Chem, 2023, 9, 411-429 https://doi.org/10.1016/j.chempr.2022.10.006AbstractThe Bigger Picture The discovery of tumor-targeting ligands with exquisite accumulation at the site of disease represents an innovative avenue for the delivery of bioactive payloads which include radionuclides, drugs, and immunomodulators. This approach promises to overcome limitations associated with cancer therapy based on the use of external beam radiation and untargeted cytotoxic drugs. Fibroblast activation protein (FAP) is an antigen that is abundantly expressed on the surface of activated fibroblasts in the stroma of most of solid malignancies. Through DNA-encoded chemical library technology, we have isolated OncoFAP-11, a novel and highly potent ligand which overcomes the limitations of previously described FAP binders. The ligand presented in this article shows high accumulation and prolonged residence time in tumors. A bivalent version of the molecule further improves tumor uptake and represents a promising clinical candidate for radioligand therapy and other targeted therapeutic modalities. Summary The availability of ultra-high-affinity small organic ligands, which are specific to accessible cancer antigens, is crucially important to enable efficient and selective tumor targeting applications. In this article, we describe the isolation of highly potent inhibitors of fibroblast activation protein (FAP), an enzyme that is abundantly and selectively expressed in the stroma of most of aggressive human solid malignancies. Affinity-maturation DNA-encoded chemical libraries, based on three series of 50,730 propargylglycine derivatives, enabled the identification of picomolar FAP inhibitors. A 177Lu-DOTAGA conjugate of the most potent novel ligand (named “OncoFAP-11”) localized to tumors implanted in mice, with tumor-to-blood ratios of ∼220:1 17 h after intravenous administration. A dimeric derivative (named “BiOncoFAP-11”) further enhanced tumor residence time with a low uptake in healthy organs.
Ziyi Xu; Wenwen Huo; Thomas Ireland; Lihang Huang; Tonatiuh Ocampo; Arturo J. Vegas Bioconjugate Chem., 2022, 33(7), 1279-1285 https://doi.org/10.1021/acs.bioconjchem.2c00195AbstractThe indiscriminate biodistribution of therapeutics can be a key barrier to their safety and efficacy. Localization of compounds into non-diseased tissues often leads to both toxic and dose-limiting effects. To overcome this barrier, nanomedicine implements targeting agents to localize or selectively uptake drugs at disease sites. However, to date there are only a small number of targeting agents with limited scope for targeting tissues. Small-molecule ligands are particularly attractive as targeting agents due to their relatively low cost, tunability, and ease of conjugation. Currently, there are no systematic approaches to the discovery of new small-molecule targeting ligands. Here, we developed a quantitative metal-encoded conjugate platform to determine the biodistribution of multiple small molecules in vivo. By utilizing lanthanide metal complexes, this platform successfully distinguished known ligands with differential tissue targeting in vivo. This system will facilitate the discovery of small molecules as targeting ligands and can accelerate the identification of novel biological targets for tissue-targeted drug delivery.
Anastasiia V. Sadybekov; Vsevolod Katritch Nature, 2023, 616, 673–685 https://doi.org/10.1038/s41586-023-05905-zAbstractComputer-aided drug discovery has been around for decades, although the past few years have seen a tectonic shift towards embracing computational technologies in both academia and pharma. This shift is largely defined by the flood of data on ligand properties and binding to therapeutic targets and their 3D structures, abundant computing capacities and the advent of on-demand virtual libraries of drug-like small molecules in their billions. Taking full advantage of these resources requires fast computational methods for effective ligand screening. This includes structure-based virtual screening of gigascale chemical spaces, further facilitated by fast iterative screening approaches. Highly synergistic are developments in deep learning predictions of ligand properties and target activities in lieu of receptor structure. Here we review recent advances in ligand discovery technologies, their potential for reshaping the whole process of drug discovery and development, as well as the challenges they encounter. We also discuss how the rapid identification of highly diverse, potent, target-selective and drug-like ligands to protein targets can democratize the drug discovery process, presenting new opportunities for the cost-effective development of safer and more effective small-molecule treatments.
Michelle Keller; Kristina Schira; Jörg Scheuermann Chimia, 2022, 76, 388 https://doi.org/10.2533/chimia.2022.388AbstractDNA-Encoded Chemical Libraries (DELs) have gained momentum over the recent years for the discovery of small-molecule ligands and the technology has been integrated in most of the larger pharmaceutical companies. With this perspective we would like to summarize the development of DEL technology and present some representative DEL-derived hits which may soon enter the pharmaceutical market.
Yashoda Krishna Sunkari; Laurent Meijer; Marc Flajolet Front. Mol. Biosci., 2022, 9, 916232 https://doi.org/10.3389/fmolb.2022.916232AbstractProtein kinases play a vital role in biology and deregulation of kinases is implicated in numerous diseases ranging from cancer to neurodegenerative diseases, making them a major target class for the pharmaceutical industry. However, the high degree of conservation that exists between ATP-binding sites among kinases makes it difficult for current inhibitors to be highly specific. In the context of neurodegeneration, several groups including ours, have linked different kinases such as CK1 and Alzheimer’s disease for example. Strictly CK1-isoform specific regulators do not exist and known CK1 inhibitors are inhibiting the enzymatic activity, targeting the ATP-binding site. Here we review compounds known to target CK1, as well as other inhibitory types that could benefit CK1. We introduce the DNA-encoded library (DEL) technology that might represent an interesting approach to uncover allosteric modulators instead of ATP competitors. Such a strategy, taking into account known allosteric inhibitors and mechanisms, might help designing modulators that are more specific towards a specific kinase, and in the case of CK1, toward specific isoforms.
Sevan Habeshian; Manuel Leonardo Merz; Gontran Sangouard; Ganesh Kumar Mothukuri; Mischa Schüttel; Zsolt Bognár; Cristina Díaz-Perlas; Jonathan Vesin; Julien Bortoli Chapalay; Gerardo Turcatti; Laura Cendron; Alessandro Angelini; Christian Heinis Nat. Commun., 2022, 13, 3823 https://doi.org/10.1038/s41467-022-31428-8AbstractMacrocycles have excellent potential as therapeutics due to their ability to bind challenging targets. However, generating macrocycles against new targets is hindered by a lack of large macrocycle libraries for high-throughput screening. To overcome this, we herein established a combinatorial approach by tethering a myriad of chemical fragments to peripheral groups of structurally diverse macrocyclic scaffolds in a combinatorial fashion, all at a picomole scale in nanoliter volumes using acoustic droplet ejection technology. In a proof-of-concept, we generate a target-tailored library of 19,968 macrocycles by conjugating 104 carboxylic-acid fragments to 192 macrocyclic scaffolds. The high reaction efficiency and small number of side products of the acylation reactions allowed direct assay without purification and thus a large throughput. In screens, we identify nanomolar inhibitors against thrombin (Ki = 44 ± 1 nM) and the MDM2:p53 protein-protein interaction (Kd MDM2 = 43 ± 18 nM). The increased efficiency of macrocycle synthesis and screening and general applicability of this approach unlocks possibilities for generating leads against any protein target.
Xianfu Fang; Yiting Wang; Pengyang He; Huilin Liao; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett., 2022, 24(17), 3291-3296 https://doi.org/10.1021/acs.orglett.2c01187AbstractBenzoheterocyclics have been widely adopted as drug-like core scaffolds that can be incorporated into DNA-encoded chemical library technology for high-throughput hit discovery. Here, we present a visible light-promoted divergent synthesis of on-DNA benzoheterocycles from aldehydes. Four types of DNA-conjugated benzoheterocyclics were obtained under mild conditions with a broad substrate scope. A cross substrate scope study, together with enzymatic ligation and subsequent chemical diversifications, were conducted, demonstrating the feasibility of this approach in DNA-encoded chemical library construction.
Chetan K. Chana; Pierre Maisonneuve; Ganna Posternak; Nicolas G.A. Grinberg; Juline Poirson; Samara M. Ona; Derek F. Ceccarelli; Pavel Mader; Daniel J. St-Cyr; Victor Pau; Igor Kurinov; Xiaojing Tang; Dongjing Deng; Weiren Cui; Wenji Su; Letian Kuai; Richard Soll; Mike Tyers; Hannes L. Röst; Robert A. Batey; Mikko Taipale; Anne-Claude Gingras; Frank Sicheri J. Med. Chem., 2022, 65(19), 12725–12746 https://doi.org/10.1021/acs.jmedchem.2c00509AbstractTargeted protein degradation (TPD) strategies exploit bivalent small molecules to bridge substrate proteins to an E3 ubiquitin ligase to induce substrate degradation. Few E3s have been explored as degradation effectors due to a dearth of E3-binding small molecules. We show that genetically induced recruitment to the GID4 subunit of the CTLH E3 complex induces protein degradation. An NMR-based fragment screen followed by structure-guided analog elaboration identified two binders of GID4, 16 and 67, with Kd values of 110 and 17 μM in vitro. A parallel DNA-encoded library (DEL) screen identified five binders of GID4, the best of which, 88, had a Kd of 5.6 μM in vitro and an EC50 of 558 nM in cells with strong selectivity for GID4. X-ray co-structure determination revealed the basis for GID4–small molecule interactions. These results position GID4-CTLH as an E3 for TPD and provide candidate scaffolds for high-affinity moieties that bind GID4.
Pratik R. Chheda; Nicholas Simmons; David P. Schuman; Zhicai Shi Org. Lett., 2022, 24(18), 3401-3406 https://doi.org/10.1021/acs.orglett.2c01175AbstractDNA-encoded chemical library (DEL) screens are a powerful hit generation tool in drug discovery, but the diversity of DEL chemical matter is dependent on developing robust reaction conditions that may be used on hundreds to millions of substrate combinations and that are compatible with the platform. Here, we disclose the first report of a general, aqueous, DNA-compatible C–N coupling condition that can now couple aliphatic amines, in addition to (hetero)aromatic amines, with a variety of (hetero)aryl iodides, bromides, and chlorides. The reported BippyPhos-Pd(OAc)2 catalyst system has a wide substrate scope for both coupling partners, is operationally feasible for large scale DEL productions, uses common DEL building block solution stocks, and enables an expansion of DEL-accessible, drug-like chemical space.
Stephany M. Ramos De Dios; Virendra K. Tiwari; Christopher D. McCune; Ranjeet A. Dhokale; David B. Berkowitz Chem. Rev., 2022, 122, 16, 13800–13880 https://doi.org/10.1021/acs.chemrev.2c00213AbstractReaction discovery and catalyst screening lie at the heart of synthetic organic chemistry. While there are efforts at de novo catalyst design using computation/artificial intelligence, at its core, synthetic chemistry is an experimental science. This review overviews biomacromolecule-assisted screening methods and the follow-on elaboration of chemistry so discovered. All three types of biomacromolecules discussed─enzymes, antibodies, and nucleic acids─have been used as “sensors” to provide a readout on product chirality exploiting their native chirality. Enzymatic sensing methods yield both UV-spectrophotometric and visible, colorimetric readouts. Antibody sensors provide direct fluorescent readout upon analyte binding in some cases or provide for cat-ELISA (Enzyme-Linked ImmunoSorbent Assay)-type readouts. DNA biomacromolecule-assisted screening allows for templation to facilitate reaction discovery, driving bimolecular reactions into a pseudo-unimolecular format. In addition, the ability to use DNA-encoded libraries permits the barcoding of reactants. All three types of biomacromolecule-based screens afford high sensitivity and selectivity. Among the chemical transformations discovered by enzymatic screening methods are the first Ni(0)-mediated asymmetric allylic amination and a new thiocyanopalladation/carbocyclization transformation in which both C–SCN and C–C bonds are fashioned sequentially. Cat-ELISA screening has identified new classes of sydnone-alkyne cycloadditions, and DNA-encoded screening has been exploited to uncover interesting oxidative Pd-mediated amido-alkyne/alkene coupling reactions.
Teresa I. Ng; Ivan Correia; Jane Seagal; David A. DeGoey; Michael R. Schrimpf; David J. Hardee; Elizabeth L. Noey; Warren M. Kati Viruses, 2022, 14(5), 961 https://doi.org/10.3390/v14050961AbstractThe coronavirus disease 2019 (COVID-19) pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a recently emerged human coronavirus. COVID-19 vaccines have proven to be successful in protecting the vaccinated from infection, reducing the severity of disease, and deterring the transmission of infection. However, COVID-19 vaccination faces many challenges, such as the decline in vaccine-induced immunity over time, and the decrease in potency against some SARS-CoV-2 variants including the recently emerged Omicron variant, resulting in breakthrough infections. The challenges that COVID-19 vaccination is facing highlight the importance of the discovery of antivirals to serve as another means to tackle the pandemic. To date, neutralizing antibodies that block viral entry by targeting the viral spike protein make up the largest class of antivirals that has received US FDA emergency use authorization (EUA) for COVID-19 treatment. In addition to the spike protein, other key targets for the discovery of direct-acting antivirals include viral enzymes that are essential for SARS-CoV-2 replication, such as RNA-dependent RNA polymerase and proteases, as judged by US FDA approval for remdesivir, and EUA for Paxlovid (nirmatrelvir + ritonavir) for treating COVID-19 infections. This review presents an overview of the current status and future direction of antiviral drug discovery for treating SARS-CoV-2 infections, covering important antiviral targets such as the viral spike protein, non-structural protein (nsp) 3 papain-like protease, nsp5 main protease, and the nsp12/nsp7/nsp8 RNA-dependent RNA polymerase complex.
Dino Gluhacevic von Krüchten; Magdalena Roth; Oliver Seitz J. Am. Chem. Soc., 2022, 144(24), 10700-10704 https://doi.org/10.1021/jacs.2c03188AbstractNucleic-acid-templated chemical reactions are currently explored for applications in DNA-encoded drug discovery, nucleic acid diagnostics, and theranostics. Of particular interest are reactions enabling the template to gain catalytic activity, so that enzymatic amplification of low copy targets would no longer be necessary. Herein, we introduce a new reaction design relying on the template-controlled cleavage of PNA-spermine conjugates. With turnover frequencies in the range of 3–10 min–1 and a kcat/KM = 1.3 × 106 M–1 s–1, the loss of affinity upon reaction provides a catalytic efficiency equal to most enzymatic conversions and superior to nucleic-acid-templated reactions reported to date.
Árpád Furka Drug Discov. Today, 2022, 27(10), 103308 https://doi.org/10.1016/j.drudis.2022.06.008AbstractCombinatorial technology has been facilitating the synthesis and screening of large molecular libraries containing millions of organic compounds ever since its introduction 40 years ago. It has changed the paradigms of pharmaceutical research from focusing on single compounds to focusing on immense collections of compounds. It inspired the development of dynamic combinatorial libraries, fragment-based drug discovery and virtual library screening. Combinatorial technology was revitalized by the development of DNA encoding. Amplification of DNA oligomers plus next-generation sequencing has made it possible to successfully screen billions of compounds in a single process.
Natalia Milosevich; Katrina H. Vizely; Roko P. A. Nikolic; Jacob F. McCallum; Lee M. Treanor; Sarah S. Khan; Ratmir Derda; Fraser Hof ChemRxiv, 2022 https://doi.org/10.26434/chemrxiv-2022-1lkkxAbstractWe report a synthetic methodology for the installation of methyllysine mimics on cysteine-containing peptides and bacteriophage peptide libraries. Strategies that allow for diversity and high throughput screening of PTM-containing peptides are critical for successfully targeting the many methyllysine reader proteins that are misregulated in cancer and disease. We have de- veloped conditions for alkylation of cysteine containing peptides with (2-haloethyl) amines, providing products that closely mimic methyllysine residues. Extensive optimization on C7C peptide phage constructs allowed for the successful installation of Kme3 mimics in 60–70% yields to create post-translational ε-Lys-N-methylated peptide phage libraries. Optimized reaction conditions between 2-bromo-N,N,N-trimethylethaninium bromide and commercially available PhD C7C library produce >2 × 1011 phage parti- cles and libraries of ~2 × 108 diversity in which each peptide sequence contains the Kme3 mimic. This process adds a new fragment into readily available genetically encoded libraries and opens new avenues for high throughput screening that may give rise to new ligands for a variety of methyllysine reader proteins.
Louis Bellmann; Raphael Klein; Matthias Rarey J. Chem. Inf. Model., 2022, 62(11), 2800-2810 https://doi.org/10.1021/acs.jcim.2c00334AbstractThe distributions of physicochemical property values, like the octanol–water partition coefficient, are routinely calculated to describe and compare virtual chemical libraries. Traditionally, these distributions are derived by processing each member of a library individually and summarizing all values in a distribution. This process becomes impractical when operating on chemical spaces which surpass billions of compounds in size. In this work, we present a novel algorithmic method called SpaceProp for the property distribution calculation of large nonenumerable combinatorial fragment spaces. The novel method follows a combinatorial approach and is able to calculate physicochemical property distributions of prominent spaces like Enamine’s REAL Space, WuXi’s GalaXi Space, and OTAVA’s CHEMriya Space for the first time. Furthermore, we present a first approach of optimizing property distributions directly in combinatorial fragment spaces.
Sai Rohini Narayanan Kolusu; Manuel Nappi Chem. Sci., 2022, 13, 6982-6989 https://doi.org/10.1039/D2SC01621DAbstractAlcohols are among the most widely occurring functional groups found in naturally abundant, biologically-relevant organic compounds, which in many cases are considered feedstock chemicals. Herein, we report a metal-free method for the deoxygenative coupling of alcohol-derived benzoates and pyridines promoted by visible light. Given the practical, mild and water-compatible conditions, small molecules and DNA headpieces can be successfully functionalized with a range of primary, secondary and tertiary alcohols. This protocol is distinguished by its wide substrate scope and broad applicability, especially in the context of late-stage functionalization and DNA-drug coupling.
Qi Liang; Bowen Shi; Qingqing Yao; Taotao Wang; Xu Ji; Yajun Zhang; Jing Wang; Xinfeng Zhao Bioorg. Med. Chem., 2022, 68, 116864 https://doi.org/10.1016/j.bmc.2022.116864AbstractEarly potential evaluation of lead compounds is critical to decrease downstream lead-optimization cycle times and clinical attrition rates for drug development. This increasingly necessitates the methodologies for accurately evaluating the potential compounds. This work immobilized β2-adrenoceptor (β2-AR) onto microspheres through Halo-tag mediated reaction. Characterizing the resulting microspheres by elemental and functional analysis, we utilized the immobilized receptor to determine the thermodynamics of terbutaline, tulobuterol, clorprenaline, salbutamol, and methoxyphenamine. The association constants correlated to their capacity factors on the column containing the immobilized β2-AR, thus providing a possibility for early potential evaluation of lead compounds from complex matrices like a DNA-encoded library. By this model, the lead compound (XC267) was predicted to have an association constant higher than terbutaline, salbutamol, and methoxyphenamine, but lower than tulobuterol and clorprenaline. The binding interaction between XC267 and β2-AR is a spontaneous endothermic process with an association constant of (6.62 ± 0.13) × 104 M−1 at 37 °C. The change of Gibbs free energy(ΔGθ), enthalpy change (ΔHθ), and entropy change (ΔSθ) was −28.49 kJ/mol, −10.58 kJ/mol, and 57.79 J/moL·K at 37 °C. By the semi-empirical rule of Ross, the driving force of the interaction between XC267 and β2-AR was electrostatic interaction. Such binding force was also achieved by molecular docking. These results suggested that XC267 is a candidate to treat asthma by specific binding to β2-AR. We reasoned that receptor chromatography is able to the early potential evaluation of lead compounds from complex matrices.
Expédite Yen-Pon; Longbo Li; Guillaume Levitre; Jadab Majhi; Edward J. McClain; Eric A. Voight; Erika A. Crane; Gary A. Molander J. Am. Chem. Soc., 2022, 144(27), 12184-12191 https://doi.org/10.1021/jacs.2c03025AbstractDNA-encoded libraries have proven their tremendous value in the identification of new lead compounds for drug discovery. To access libraries in new chemical space, many methods have emerged to transpose traditional mol-scale reactivity to nmol-scale, on-DNA chemistry. However, procedures to access libraries with a greater fraction of C(sp3) content are still limited, and the need to “escape from flatland” more readily on-DNA remains. Herein, we report a Giese addition to install highly functionalized bicyclo[1.1.1]pentanes (BCPs) using tricyclo[1.1.1.01,3]pentane (TCP) as a radical linchpin, as well as other diverse alkyl groups, on-DNA from the corresponding organohalides as non-stabilized radical precursors. Telescoped procedures allow extension of the substrate pool by at least an order of magnitude to ubiquitous alcohols and carboxylic acids, allowing us to “upcycle” these abundant feedstocks to afford non-traditional libraries with different physicochemical properties for the small-molecule products (i.e., non-peptide libraries with acids). This approach is amenable to library production, as a DNA damage assessment revealed good PCR amplifiability and only 6% mutated sequences for a full-length DNA tag.
Bidong Zhang; Alexander Dömling Expert. Opin. Ther. Pat., 2022, 1-13 https://doi.org/10.1080/13543776.2022.2143264AbstractInterleukin-17A (IL-17A) is a well-established pro-inflammatory cytokine, which plays a pivotal role in immune and autoimmune diseases including psoriasis, asthma, psoriatic arthritis, and rheumatoid arthritis. Three currently approved monoclonal antibodies (mAbs) are in clinical practice for the treatment of multiple immune diseases. However, the disadvantages of the mAbs, such as non-oral administration, poor tissue penetration, lacking blood-brain barrier penetration, often long half-life times, narrow its application. Thus, intensive research is performed to discover potent small molecules, peptides, and macrocycles targeting the IL-17A/IL-17 RA protein-protein interaction (PPI) to modulate immune responses as an attractive approach for immunotherapy.
Linjie Li; Mingbo Su; Weiwei Lu; Hongzhi Song; Jiaxiang Liu; Xin Wen; Yanrui Suo; Jingjing Qi; Xiaomin Luo; Yubo Zhou; Xinhua Liao; Jia Li; Xiaojie Lu ACS Med. Chem. Lett., 2022, 13(10), 1574–1581 https://doi.org/10.1021/acsmedchemlett.2c00127AbstractSince ibrutinib was approved by the FDA as an effective monotherapy for chronic lymphocytic leukemia (CLL) and multilymphoma, more and more FDA-approved covalent drugs are coming back into the market. On this occasion, the resurgence of interest in covalent drugs calls for more hit discovery techniques. However, the limited numbers of covalent libraries prevent the development of this area. Herein, we report the design of covalent DNA-encoded library (DEL) and its selection method for the discovery of covalent inhibitors for target proteins. These triazine-based covalent DELs yielded potent compounds after covalent selection against target proteins, including Bruton’s Tyrosine Kinase (BTK), Janus kinase 3 (JAK3), and peptidyl-prolyl cis/trans isomerase NIMA-interacting-1 (Pin1).
Rui Tian; Jiatai Yin; Qingqing Yao; Taotao Wang; Jiahuan Chen; Qi Liang; Qian Li; Xinfeng Zhao Anal. Chem., 2022, 94(25), 9048-9057 https://doi.org/10.1021/acs.analchem.2c01210AbstractAllosteric ligands are promising drugs owing to their remote regulations of the orthosteric ligand signaling pathway. There are few allosteric ligands due to the lack of handy and efficacious method for the screening. Herein, we developed an affinity chromatographic method for allosteric ligand screening by immobilizing purified beta2 adrenoceptor (β2-AR) onto macroporous silica gel by a two-point tethering method. The method relies on the occupation of the orthosteric site by an antagonist and the chelation of N-terminal His-tag of the receptor and Ni2+ coated on the gel. The immobilized β2-AR demonstrated the greatest allosteric responsive feature when Cmpd-15 (0.25 μM) was included in the mobile phase. Under the same conditions, the association constants of three agonists (salbutamol, terbutaline, and tulobuterol) reduced to 47%, 19%, and 27% compared with the data without the inclusion of Cmpd-15 in the mobile phase. APF was screened as a potential allosteric modulator of β2-AR by applying the immobilized receptor in a natural product-derived DNA-encoded chemical library (DEL). Relying on these results, we reasoned that the current method has potential in screening allosteric ligands of the receptor. We expect that it is applicable for the discovery of new allosteric binding sites of a target protein and screening allosteric modulators of the other receptors from complex samples.
Yasuhiro Uozumi; Aya Tazawa Synfacts, 2022, 18(04), 0417 https://doi.org/10.1055/s-0041-1737383AbstractTransfer Hydrogenolyses and hydrogenations of DNA-conjugated substrates were effected using the micelle-forming surfactant TPGS-750-M. DNA-conjugated N-Cbz and benzyl ethers were deprotected with 10% Pd/C and HCO2NH4 in aqueous TPGS-750-M to give the corresponding amines and alcohols in up to 100% conversion. Transfer hydrogenations of DNA-conjugated nitro-benzenes, aryl halides, alkenes, alkynes, nitriles, and aldehydes were also preformed under similar conditions to afford the corresponding hydrogenated products in up to 100% conversion.
Marco Potowski; Verena B. K. Kunig; Lukas Eberlein; Mateja Klika Škopić; Alexandros Vakalopoulos; Stefan M. Kast; Andreas Brunschweiger Front. Chem., 2022, 10, 894563 https://doi.org/10.3389/fchem.2022.894563AbstractDNA-encoded libraries are a prime technology for target-based small molecule screening. Native DNA used as genetic compound barcode is chemically vulnerable under many reaction conditions. DNA barcodes that are composed of pyrimidine nucleobases, 7-deazaadenine, and 7-deaza-8-azaguanine have been investigated for their suitability for encoded chemistry both experimentally and computationally. These four-letter barcodes were readily ligated by T4 ligation, amplifiable by Taq polymerase, and the resultant amplicons were correctly sequenced. Chemical stability profiling showed a superior chemical stability compared to native DNA, though higher susceptibility to depurination than a three-letter code based on pyrimidine DNA and 7-deazaadenine.
Ana Domostegui; Luis Nieto-Barrado; Carles Perez-Lopez; Cristina Mayor-Ruiz Chem. Soc. Rev., 2022, 51(13), 5498-5517 https://doi.org/10.1039/D2CS00197GAbstractProtein–protein interactions (PPIs) govern all biological processes. Some small molecules modulate PPIs through induced protein proximity. In particular, molecular glue degraders are monovalent compounds that orchestrate interactions between a target protein and an E3 ubiquitin ligase, prompting the proteasomal degradation of the former. This and other pharmacological strategies of targeted protein degradation (e.g. proteolysis-targeting chimeras – PROTACs) overcome some limitations of traditional occupancy-based therapeutics. Here, we provide an overview of the “molecular glue” concept, with a special focus on natural and synthetic inducers of proximity to E3s. We then briefly highlight the serendipitous discoveries of some clinical and preclinical molecular glue degraders, and discuss the first examples of intentional discoveries. Specifically, we outline the different screening strategies reported in this rapidly evolving arena and our thoughts on future perspectives. By mastering the ability to influence PPIs, molecular glue degraders can induce the degradation of unligandable proteins, thus providing an exciting path forward to broaden the targetable proteome.
Vijay Kumar Siripuram; Yashoda Krishna Sunkari; Thu-Lan Nguyen; Marc Flajolet Front. Chem., 2022, 10, 894603 https://doi.org/10.3389/fchem.2022.894603AbstractAn efficient method for the C-C bond formation via water soluble Na2PdCl4/sSPhos mediated Suzuki-Miyaura cross-coupling reaction of DNA-conjugated aryl iodide with (het)aryl boronic acids has been developed. This reaction proceeds at 37°C in water and acetonitrile (4:1) system. We also demonstrated that numerous aromatic and heteroaromatic boronic acids of different electronic natures, and harboring various functional groups, were highly compatible providing the desired coupling products in good to excellent yields. This DNA-compatible Suzuki-Miyaura cross-coupling reaction has strong potential to construct DNA-Encoded Libraries (DELs) in the context of drug discovery.
Alexey V. Stepanov; Jia Xie; Qiaoqiao Zhu; Zuyuan Shen; Wenji Su; Letian Kuai; Richard Soll; Christoph Rader; Geramie Shaver; Lacey Douthit; Ding Zhang; Roman Kalinin; Xiang Fu; Yingying Zhao; Tian Qin; Phil S. Baran; Alexander G. Gabibov; David Bushnell; Dario Neri; Roger D. Kornberg; Richard A. Lerner Nat. Biomed. Eng., 2023 https://doi.org/10.1038/s41551-023-01102-5AbstractOn-target off-tumour toxicity limits the anticancer applicability of chimaeric antigen receptor (CAR) T cells. Here we show that the tumour-targeting specificity and activity of T cells with a CAR consisting of an antibody with a lysine residue that catalytically forms a reversible covalent bond with a 1,3-diketone hapten can be regulated by the concentration of a small-molecule adapter. This adapter selectively binds to the hapten and to a chosen tumour antigen via a small-molecule binder identified via a DNA-encoded library. The adapter therefore controls the formation of a covalent bond between the catalytic antibody and the hapten, as well as the tethering of the CAR T cells to the tumour cells, and hence the cytotoxicity and specificity of the cytotoxic T cells, as we show in vitro and in mice with prostate cancer xenografts. Such small-molecule switches of T-cell cytotoxicity and specificity via an antigen-independent ‘universal’ CAR may enhance the control and safety profile of CAR-based cellular immunotherapies.
Shuyue Wang; Xiaojie Shi; Jie Li; Qianping Huang; Qun Ji; Ying Yao; Tao Wang; Lili Liu; Min Ye; Yun Deng; Peixiang Ma; Hongtao Xu; Guang Yang Adv. Sci., 2022, e2201258 https://doi.org/10.1002/advs.202201258AbstractTumor necrosis factor α (TNF-α) inhibitors have shown great success in the treatment of autoimmune diseases. However, to date, approved drugs targeting TNF-α are restricted to biological macromolecules, largely due to the difficulties in using small molecules for pharmaceutical intervention of protein–protein interactions. Herein the power of a natural product-enriched DNA-encoded library (nDEL) is exploited to identify small molecules that interfere with the protein–protein interaction between TNF-α and the cognate receptor. Initially, to select molecules capable of binding to TNF-α , “late-stage” DNA modification method is applied to construct an nDEL library consisted of 400 sterically diverse natural products and pharmaceutically active chemicals. Several natural products, including kaempferol, identified not only show direct interaction with TNF-α, but also lead to the blockage of TNF-α/TNFR1 interaction. Significantly, kaempferol attenuates the TNF-α signaling in cells and reduces the 12-O-tetradecanoylphorbol-13-acetateinduced ear inflammation in mice. Structure-activity-relationship analyses demonstrate the importance of substitution groups at C-3, C-7, and C-4' of kaempferol. The nDEL hit, kaempferol, represents a novel chemical scaffold capable of specifically recognizing TNF-α and blocking its signal transduction, a promising starting point for the development of a small molecule TNF-α inhibitor for use in the clinical setting.
Hongtao Xu; Yan Wang; Hewei Dong; Yiyuan Zhang; Yuang Gu; Shuning Zhang; Yu Meng; Jie Li; XiaoJie Shi; Qun Ji; Lili Liu; Peixiang Ma; Fei Ma; Guang Yang; Wei Hou Angew. Chem. Int. Ed. Engl., 2022, e202206516 https://doi.org/10.1002/ange.202206516AbstractClick chemistry is a concept wherein modular synthesis is used for rapid functional discovery. To this end, continuous discovery of clickable chemical transformations is the pillar to support the development of this field. This report details the development of a clickable C3-H selenylation of indole that is suitable for on-plate parallel and DNA-encoded library ( SeDEL) synthesis via bioinspired LUMO activation strategy. This reaction is modular, robust and highly site-selective, and it features simple and mild reaction system (catalyzed by nonmetallic B(C6F5)3 at room temperature), high yields and excellent functional group compatibility. Using this method, a library of 1350 indole-selenides was parallel synthesized in an efficient and practical manner, enabling the rapid identification of 3ai as a promising compound with nanomolar antiproliferative activity in cancer cells via in situ phenotypic screening. These results indicate the great potential of this new clickable selenylation reaction in highthroughput medicinal chemistry and chemical biology.
Xuexuan Wu; Qiuyue Zhang; Yuqi Guo; Hengheng Zhang; Xiaoke Guo; Qidong You; Lei Wang Antioxidants, 2022, 11(4), 619 https://doi.org/10.3390/antiox11040619AbstractThe oxidative stress response pathway is one of the hotspots of current pharmaceutical research. Many proteins involved in these pathways work through protein–protein interactions (PPIs). Hence, targeting PPI to develop drugs for an oxidative stress response is a promising strategy. In recent years, small molecules targeting protein–protein interactions (PPIs), which provide efficient methods for drug discovery, are being investigated by an increasing number of studies. However, unlike the enzyme–ligand binding mode, PPIs usually exhibit large and dynamic binding interfaces, which raise additional challenges for the discovery and optimization of small molecules and for the biochemical techniques used to screen compounds and study structure–activity relationships (SARs). Currently, multiple types of PPIs have been clustered into different classes, which make it difficult to design stationary methods for small molecules. Deficient experimental methods are plaguing medicinal chemists and are becoming a major challenge in the discovery of PPI inhibitors. In this review, we present current methods that are specifically used in the discovery and identification of small molecules that target oxidative stress-related PPIs, including proximity-based, affinity-based, competition-based, structure-guided, and function-based methods. Our aim is to introduce feasible methods and their characteristics that are implemented in the discovery of small molecules for different types of PPIs. For each of these methods, we highlight successful examples of PPI inhibitors associated with oxidative stress to illustrate the strategies and provide insights for further design.
Andi Guo; Kenian Yan; Hao Hu; Linhui Zhai; Tengfei Hu; Haixia Su; Yijia Chi; Jinyin Zha; Yechun Xu; Dongxin Zhao; Xiaojie Lu; Yongjiang Xu; Jian Zhang; Minjia Tan; Xiaohua Chen Nat. Chem., 2023, 15, 803-814 https://doi.org/10.1038/s41557-023-01196-zAbstractPrecise dissection of DNA–protein interactions is essential for elucidating the recognition basis, dynamics and gene regulation mechanism. However, global profiling of weak and dynamic DNA–protein interactions remains a long-standing challenge. Here, we establish the light-induced lysine (K) enabled crosslinking (LIKE-XL) strategy for spatiotemporal and global profiling of DNA–protein interactions. Harnessing unique abilities to capture weak and transient DNA–protein interactions, we demonstrate that LIKE-XL enables the discovery of low-affinity transcription-factor/DNA interactions via sequence-specific DNA baits, determining the binding sites for transcription factors that have been previously unknown. More importantly, we successfully decipher the dynamics of the transcription factor subproteome in response to drug treatment in a time-resolved manner, and find downstream target transcription factors from drug perturbations, providing insight into their dynamic transcriptional networks. The LIKE-XL strategy offers a complementary method to expand the DNA–protein profiling toolbox and map accurate DNA–protein interactomes that were previously inaccessible via non-covalent strategies, for better understanding of protein function in health and disease.
Brian Goldman; Steven Kearnes; Trevor Kramer; Patrick Riley; W. Patrick Walters J. Med. Chem., 2022, 65(10), 7073-7087 https://doi.org/10.1021/acs.jmedchem.2c00334AbstractOne application area of computational methods in drug discovery is the automated design of small molecules. Despite the large number of publications describing methods and their application in both retrospective and prospective studies, there is a lack of agreement on terminology and key attributes to distinguish these various systems. We introduce Automated Chemical Design (ACD) Levels to clearly define the level of autonomy along the axes of ideation and decision making. To fully illustrate this framework, we provide literature exemplars and place some notable methods and applications into the levels. The ACD framework provides a common language for describing automated small molecule design systems and enables medicinal chemists to better understand and evaluate such systems.
John V. L. Nguyen; Ahlem Meziadi; Christina Nassif; Dillon Da Fonte; Lidija Malic; Maryam Tabrizian ACS Appl. Bio Mater., 2025, 8(1), 844–853 https://doi.org/10.1021/acsabm.4c01681AbstractSynthetic ssDNA oligonucleotides hold great potential for various applications, including DNA aptamers, DNA digital data storage, DNA origami, and synthetic genomes. In these contexts, precise control over the synthesis of the ssDNA strands is essential for generating combinatorial sequences with user-defined parameters. Desired features for creating synthetic DNA oligonucleotides include easy manipulation of DNA strands, effective detection of unique DNA sequences, and a straightforward mechanism for strand elongation and termination. In this study, we present a split-and-pool method for generating synthetic DNA oligonucleotides on nanoparticles, enabling the creation of scalable combinatorial libraries. Our approach involves coupling DNA to nanoparticles, ligating double-digested fragments for orientation-specific synthesis, and attaching a final single-digested fragment to ensure strand termination. We assess the quality of our method by characterizing both the DNA and the nanoparticles used as solid supports, confirming that our method produces scalable, combinatorial nanoparticle-bound ssDNA libraries with controllable strand lengths.
Elijah T. Marris; Daniel S. Rampon; Jennifer M. Schomaker Acc. Chem. Res., 2025, 58, 2, 231-249 https://doi.org/10.1021/acs.accounts.4c00670AbstractConspectusThe manipulation of strained rings is a powerful strategy for accessing the valuable chemical frameworks present in natural products and active pharmaceutical ingredients. Aziridines, the smallest N-containing heterocycles, have long served as building blocks for constructing more complex amine-containing scaffolds. Traditionally, the reactivity of typical aziridines has been focused on ring-opening by nucleophiles or the formation of 1,3-dipoles. However, over the past decade, our group has pioneered highly chemo-, site-, and stereoselective Ag- and Rh-catalyzed nitrene transfer (NT) reactions of allenes and alkenes to furnish unusual, or "anomalous", aziridines. The unique features of these aziridines, coupled with our ability to control the fate of strained intermediates resulting from diverse reactions of these precursors, allow for their transformation to densely substituted, stereochemically complex N-heterocyclic structures that would otherwise be difficult to access using conventional strategies. Our research is driven by a keen interest in versatile synthetic approaches that explore high Fsp3 (the fraction of sp3 carbons relative to the total number of carbons in a molecule) amine chemical space, which holds promise for uncovering novel bioactivity toward challenging protein targets. We begin by outlining the design and synthesis of selected anomalous aziridines and highlighting the key features that are pertinent to their versatility as synthetic intermediates. We detail chemo-, site-, and stereoselective Rh-catalyzed NT of homoallenic sulfamates leading to the key (E)-methyleneaziridines (MAs) and the development of new Ag catalysts to achieve chemo- and enantioselective aziridinations of homoallenic and homoallylic carbamates/carbamimidates to yield bicyclic (methylene)aziridines. The chemoselective Ag-catalyzed NT of carbamimidates to accomplish intramolecular aza-Büchner reactions via polycyclic aziridine intermediates is also highlighted. Next, we focus on unlocking several modes of reactivity of our anomalous aziridines. These include regioselective ring-opening with nucleophiles and subsequent functionalization to afford amine stereotriads, the stereocontrolled formation and reaction of 2-amidoallyl cations, and alkene oxidation of endo- and exocyclic MAs. Additionally, due to the unusual molecular geometry of bicyclic (methylene)aziridines, the nitrogen lone pair can react with carbenes to generate aziridinium ylides that can be diverted along multiple pathways. From only a handful of anomalous aziridines, our chemistry is capable of delivering amine stereotriads, azetidines, azetidinones, piperidines, aminated cyclopentanes and cycloheptanes, azepines, and azepanes. Finally, we discuss our efforts to leverage this chemistry to explore the complex amine chemical space relevant to the natural products jogyamycin and methyl detoxinine. Ultimately, our goal is to use these methods to generate DNA-encoded libraries of high Fsp3 compounds with potential activity against difficult-to-target proteins.
Ioana Oprea; Terry K. Smith ACS Chem. Biol., 2025, 20(1), 19–32 https://doi.org/10.1021/acschembio.4c00608AbstractClick chemistry is an immensely powerful technique for the synthesis of reliable and efficient covalent linkages. When undertaken in living cells, the concept is thereby coined bioorthogonal chemistry. Used in conjunction with the photo-cross-linking methodology, it serves as a sound strategy in the exploration of biological processes and beyond. Its broad scope has led to widespread use in many disciplines; however, this Review focuses on the use of click and bioorthogonal chemistry within medicinal chemistry, specifically with regards to drug development applications, namely, the use of DNA-encoded libraries as a novel technique for lead compound discovery, as well as the synthesis of antisense oligonucleotides and protein-drug conjugates. This Review aims to provide a critical perspective and a future outlook of this methodology, such as potential widespread use in cancer therapy and personalized medicine.
Qigui Nie; Tingting Xu; Xianfu Fang; Yanrong Dan; Gong Zhang; Yangfeng Li; Jianbo Li; Yizhou Li Org. Lett., 2025, 27(1), 498–503 https://doi.org/10.1021/acs.orglett.4c04505AbstractWe here report an efficient DNA-compatible furan-thiol-amine reaction for macrocyclization and late-stage amine transformation. This reaction, conducted under mild conditions, enables the facile cyclization of DNA-conjugated linear peptides into thiopyrrole-grafted macrocycles regardless of ring size or side-chain modification with good to excellent conversion yields. Additionally, this strategy was employed for the late-stage transformation of terminal amines, serving as critical intermediates in the construction of DNA-encoded peptide libraries. Diverse amines were successfully converted into their corresponding thiopyrrole scaffolds, thereby expanding the structural diversity that can be achieved within DNA-encoded libraries.
Mélanie Uguen; Tongkun Liu; Lindsey I. James; Stephen V. Frye ACS Chem. Biol., 2025, 20(1), 33–47 https://doi.org/10.1021/acschembio.4c00661AbstractTudor domains are histone readers that can recognize various methylation marks on lysine and arginine. This recognition event plays a key role in the recruitment of other epigenetic effectors and the control of gene accessibility. The Tudor-containing protein family contains 42 members, many of which are involved in the development and progression of various diseases, especially cancer. The development of chemical tools for this family will not only lead to a deeper understanding of the biological functions of Tudor domains but also lay the foundation for therapeutic discoveries. In this review, we discuss the role of several Tudor domain-containing proteins in a range of relevant diseases and progress toward the development of chemical tools such as peptides, peptidomimetics, or small-molecules that bind Tudor domains. Overall, we highlight how Tudor domains are promising targets for therapeutic development and would benefit from the development of novel chemical tools.
Fangyuan Chen; Qingmei Liu; Lei Ma; Cuishi Yan; Haiman Zhang; Zhi Zhou; Wei Yi J. Med. Chem., 2025, 68(1), 819–831 https://doi.org/10.1021/acs.jmedchem.4c02803AbstractRecent studies have identified selective peroxisome proliferator-activated receptor γ (PPARγ) modulators, which synergistically engage in the inhibition mechanism of PPARγ-Ser273 phosphorylation, as a promising approach for developing safer and more effective antidiabetic drugs. Herein, we present the design, synthesis, and evaluation of a new class of organo-Se compounds, namely, benzothiaselenazole-1-oxides (BTSAs), acting as potent, reversible, and selective PPARγ covalent modulators. Notably, 2n, especially (R)-2n, displayed a high binding affinity and superior antidiabetic effects with diminished side effects. This is mainly because it can reversibly form a unique covalent bond with the Cys285 residue in PPARγ-LBD. Further mechanistic investigations revealed that it manifested such desired pharmacological profiles primarily by effectively suppressing PPARγ-Ser273 phosphorylation, enhancing glucose metabolism, and selectively upregulating the expression of insulin-sensitive genes. Collectively, our results suggest that (R)-2n holds promise as a lead compound for treating T2DM and also provides an innovative reversible covalent warhead reference for future covalent drug design.
Philipp N. Sander; Jared T. Gillen Miller; Luke L. Lairson RSC Chem. Biol., 2025,6, 273-280 https://doi.org/10.1039/d4cb00137kAbstractBased on their ability to canvas vast genetic or chemical space at low cost and high speed, DNA-encoded libraries (DEL) have served to enable both genomic and small molecule discovery. Current DEL chemical library screening approaches focus primarily on in vitro target-based affinity or activity. Here we describe an approach to record the phenotype-based activity of DNA-encoded small molecules on their cognate barcode in living cells. We transfected chloroalkane-derivatized DNA barcodes carrying photoreleasable small molecules into cells. Following photorelease, bioactive compounds induced expression of a reporter gene cassette containing self-labeling HaloTag protein that becomes covalently modified by encoding barcodes. We demonstrate that we can recover activity information from cells that received active compound following immunoprecipitation-based enrichment. This generalizable approach should enable future strategies that facilitate phenotype-based screens of DNA-encoded chemical libraries in complex cellular or organism level systems.
Niemeier, Felix; Servos, Lisa-Marie; Papadopoulos, Zisis; Montesdeoca, Nicolás; Ni, Kaixin; Heinrich, Sascha; Karges, Johannes J. Med. Chem., 2025, 68(2), 1316–1327 https://doi.org/10.1021/acs.jmedchem.4c01875AbstractCancer remains one of the deadliest diseases worldwide, with some tumors proving difficult to treat and increasingly resistant to current therapies. Capitalizing on this, there is a need for new therapeutic agents with novel mechanisms of action. Among promising candidates, Fe(III) complexes have gained significant attention as potential chemotherapeutic agents. However, research on these compounds has been limited to a small number, leading to inefficiencies in drug discovery. This study addresses these limitations by developing a combinatorial library of 495 new Fe(III) complexes synthesized from aminophenol, hydroxybenzaldehyde, and pyridine derivatives. The compounds were screened for cytotoxicity against human breast adenocarcinoma and noncancerous fibroblasts, identifying a novel class of Fe(III) complexes with modest cancer cell selectivity. The lead compound effectively eradicated breast cancer tumor spheroids at low micromolar concentrations, highlighting the potential of this approach for rapid drug discovery.
Chunjie Hu; Qingya Wen; Qiuyang Lai; Ziyi Xie; Kaiyue Zhang; Lu Zhou; Zhi-bei Qu Chem. Commun., 2025, 61, 881-884 https://doi.org/10.1039/D4CC05109BAbstractA headpiece was introduced in the construction of a DNA-based data storage platform. It was demonstrated that the involvement of the headpiece could largely improve the stability, recovery, resistance to DNA contamination, and accuracy in sequencing and data retrieval.
Yi Gan; Yumei Zeng; Haojie Guan; Alex Shaginian; Jin Li; Guanyu Yang; Guansai Liu Org. Lett., 2024, 26, 49, 10640–10644 https://doi.org/10.1021/acs.orglett.4c04210AbstractDNA-encoded library (DEL) technology has been developed to serve as a practical platform for the discovery of biologically active macrocyclic peptide compounds. However, the cyclization of linear peptides has been widely regarded as the challenging step in the production of macrocyclic peptide DELs. Herein, we describe a novel DNA-compatible macrocyclization strategy, which enables the construction of ring systems via visible-light-mediated desulfurative C–C bond formation. The macrocyclization proceeds smoothly under mild conditions and in a good yield. Moreover, the reaction is compatible with a variety of linear substrates and can thus be employed to generate structurally diverse DNA-encoded macrocycles with various ring sizes.
Baoli Ding; Li Lu; Jiawen Hu; Rongtian Zhang; Feifan Wang; Zhesheng Zhou; Yushen Lin; Chenghao Pan; Yihui Zhou; Bo Yang; Cheng-Liang Zhu; Chun Zhou; Ji Cao Bioorg. Chem., 2025, 154, 107948 https://doi.org/10.1016/j.bioorg.2024.107948AbstractWD repeat-containing protein 5 (WDR5) is a scaffolding protein involved in critical protein-protein interactions and a promising target for therapeutic development. Novel small-molecule ligands targeting WDR5 were identified using the DELopen platform, a free-access DNA-encoded chemical library (DEL) for academic research. Through off-DNA structure-activity relationship studies and photoaffinity labeling, two promising initial leads, DBL-6-13 and DBL-6-33, were identified as new binders of WDR5. These compounds exhibited moderate to good binding affinities and were confirmed to bind the WIN-site through co-crystal structure analysis. Our findings demonstrate the utility of DEL technology in identifying ligands for challenging targets like WDR5, particularly within an academic research setting using the DELopen platform. The identified WDR5 ligands offer a foundation for further optimization and exploration as chemical probes for WDR5 research.
Francesca Migliorini; Andrea Ciamarone; Dario Neri; Samuele Cazzamalli; Nicholas Favalli Asian J.Org. Chem. 2025, e202400556 https://doi.org/10.1002/ajoc.202400556AbstractIsocyanates, isothiocyanates, and sulfonyl chlorides can be found in valuable building blocks for the construction of DNA-Encoded Chemical Libraries (DELs), leading to the generation of ureas, thioureas, and sulfonamides. We present the development of reaction conditions for the addition of isocyanates, isothiocyanates, and sulfonyl chlorides to single-stranded oligonucleotides, with a focus on maximizing on-DNA conversion while preserving DNA integrity. We explored and optimized reaction parameters to achieve efficient coupling and minimize DNA degradation and by-product formation. The protocols reported in this paper broaden the scope of DNA-compatible reactions and may be applicable to the construction of novel DELs.
Pengyong Li; Kaihao Zhang; Tianxiao Liu; Ruiqiang Lu; Yangyang Chen; Xiaojun Yao; Lin Gao; Xiangxiang Zeng Nat. Comput. Sci., 2024, 4, 851-864 https://doi.org/10.1038/s43588-024-00718-0AbstractDeep generative models are gaining attention in the field of de novo drug design. However, the rational design of ligand molecules for novel targets remains challenging, particularly in controlling the properties of the generated molecules. Here, inspired by the DNA-encoded compound library technique, we introduce DeepBlock, a deep learning approach for block-based ligand generation tailored to target protein sequences while enabling precise property control. DeepBlock neatly divides the generation process into two steps: building blocks generation and molecule reconstruction, accomplished by a neural network and a rule-based reconstruction algorithm we proposed, respectively. Furthermore, DeepBlock synergizes the optimization algorithm and deep learning to regulate the properties of the generated molecules. Experiments show that DeepBlock outperforms existing methods in generating ligands with affinity, synthetic accessibility and drug likeness. Moreover, when integrated with simulated annealing or Bayesian optimization using toxicity as the optimization objective, DeepBlock successfully generates ligands with low toxicity while preserving affinity with the target.
Moreno Wichert; Laura Guasch; Raphael M. Franzini Chem. Rev., 2024, 124(22), 12551–12572 https://doi.org/10.1021/acs.chemrev.4c00284AbstractDNA-encoded library (DEL) technology is a powerful platform for the efficient identification of novel chemical matter in the early drug discovery process enabled by parallel screening of vast libraries of encoded small molecules through affinity selection and deep sequencing. While DEL selections provide rich data sets for computational drug discovery, the underlying technical factors influencing DEL data remain incompletely understood. This review systematically examines the key parameters affecting the chemical information in DEL data and their impact on hit triaging and machine learning integration. The need for rigorous data handling and interpretation is emphasized, with standardized methods being critical for the success of DEL-based approaches. Major challenges include the relationship between sequence counts and binding affinities, frequent hitters, and the influence of factors such as inhomogeneous library composition, DNA damage, and linkers on binding modes. Experimental artifacts, such as those caused by protein immobilization and screening matrix effects, further complicate data interpretation. Recent advancements in using machine learning to denoise DEL data and predict drug candidates are highlighted. This review offers practical guidance on adopting best practices for integrating robust methodologies, comprehensive data analysis, and computational tools to improve the accuracy and efficacy of DEL-driven hit discovery.
Suzanne Ackloo; Fengling Li; Magda Szewczyk; Almagul Seitova; Peter Loppnau; Hong Zeng; Jin Xu; Shabbir Ahmad; Yelena A. Arnautova; A. J. Baghaie; Serap Beldar; Albina Bolotokova; Paolo A. Centrella; Irene Chau; Matthew A. Clark; John W. Cuozzo; Saba Dehghani-Tafti; Jeremy S. Disch; Aiping Dong; Antoine Dumas; Jianwen A. Feng; Pegah Ghiabi; Elisa Gibson; Justin Gilmer; Brian Goldman; Stuart R. Green; Marie-Aude Guié; John P. Guilinger; Nathan Harms; Oleksandra Herasymenko; Scott Houliston; Ashley Hutchinson; Steven Kearnes; Anthony D. Keefe; Serah W. Kimani; Trevor Kramer; Maria Kutera; Haejin A. Kwak; Cristina Lento; Yanjun Li; Jenny Liu; Joachim Loup; Raquel A. C. Machado; Christopher J. Mulhern; Sumera Perveen; Germanna L. Righetto; Patrick Riley; Suman Shrestha; Eric A. Sigel; Madhushika Silva; Michael D. Sintchak; Belinda L. Slakman; Rhys D. Taylor; James Thompson; Wen Torng; Carl Underkoffler; Moritz von Rechenberg; Ryan T. Walsh; Ian Watson; Derek J. Wilson; Esther Wolf; Manisha Yadav; Aliakbar K. Yazdi; Junyi Zhang; Ying Zhang; Vijayaratnam Santhakumar; Aled M. Edwards; Dalia Barsyte-Lovejoy; Matthieu Schapira; Peter J. Brown; Levon Halabelian; Cheryl H. Arrowsmith J. Med. Chem., 2025, 68(2), 1092–1112 https://doi.org/10.1021/acs.jmedchem.4c02010AbstractTarget class-focused drug discovery has a strong track record in pharmaceutical research, yet public domain data indicate that many members of protein families remain unliganded. Here we present a systematic approach to scale up the discovery and characterization of small molecule ligands for the WD40 repeat (WDR) protein family. We developed a comprehensive suite of protocols for protein production, crystallography, and biophysical, biochemical, and cellular assays. A pilot hit-finding campaign using DNA-encoded chemical library selection followed by machine learning (DEL-ML) to predict ligands from virtual libraries yielded first-in-class, drug-like ligands for 7 of the 16 WDR domains screened, thus demonstrating the broader ligandability of WDRs. This study establishes a template for evaluation of protein family wide ligandability and provides an extensive resource of WDR protein biochemical and chemical tools, knowledge, and protocols to discover potential therapeutics for this highly disease-relevant, but underexplored target class.
Kilian Colas; Daniel Bindl; Hiroaki Suga Chem. Rev., 2024, 124, 21, 12213–12241 https://doi.org/10.1021/acs.chemrev.4c00422AbstractTechnological advances and breakthrough developments in the pharmaceutical field are knocking at the door of the “undruggable” fortress with increasing insistence. Notably, the 21st century has seen the emergence of macrocyclic compounds, among which cyclic peptides are of particular interest. This new class of potential drug candidates occupies the vast chemical space between classic small-molecule drugs and larger protein-based therapeutics, such as antibodies. As research advances toward clinical targets that have long been considered inaccessible, macrocyclic peptides are well-suited to tackle these challenges in a post-rule of 5 pharmaceutical landscape. Facilitating their discovery is an arsenal of high-throughput screening methods that exploit massive randomized libraries of genetically encoded compounds. These techniques benefit from the incorporation of non-natural moieties, such as non- proteinogenic amino acids or stabilizing hydrocarbon staples. Exploiting these features for the strategic architectural design of macrocyclic peptides has the potential to tackle challenging targets such as protein–protein interactions, which have long resisted research efforts. This Review summarizes the basic principles and recent developments of the main high-throughput techniques for the discovery of macrocyclic peptides and focuses on their specific deployment for targeting undruggable space. A particular focus is placed on the development of new design guidelines and principles for the cyclization and structural stabilization of cyclic peptides and the resulting success stories achieved against well-known inaccessible drug targets.
Hanqun Cao; Mutian He; Ning Ma; Chang-yu Hsieh; Chunbin Gu; Pheng-Ann Heng arXiv, 2024 https://doi.org/10.48550/arXiv.2410.14946AbstractDNA-encoded library (DEL) screening has revolutionized the detection of protein-ligand interactions through read counts, enabling rapid exploration of vast chemical spaces. However, noise in read counts, stemming from nonspecific interactions, can mislead this exploration process. We present DEL-Ranking, a novel distribution-correction denoising framework that addresses these challenges. Our approach introduces two key innovations: (1) a novel ranking loss that rectifies relative magnitude relationships between read counts, enabling the learning of causal features determining activity levels, and (2) an iterative algorithm employing self-training and consistency loss to establish model coherence between activity label and read count predictions. Furthermore, we contribute three new DEL screening datasets, the first to comprehensively include multi-dimensional molecular representations, protein-ligand enrichment values, and their activity labels. These datasets mitigate data scarcity issues in AI-driven DEL screening research. Rigorous evaluation on diverse DEL datasets demonstrates DEL-Ranking's superior performance across multiple correlation metrics, with significant improvements in binding affinity prediction accuracy. Our model exhibits zero-shot generalization ability across different protein targets and successfully identifies potential motifs determining compound binding affinity. This work advances DEL screening analysis and provides valuable resources for future research in this area.
Markus Orsi; Jean-Louis Reymond ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-bqd8cAbstractHerein we report a virtual library of 1E+60 members, a common estimate for the total size of the drug-like chemical space. The library is obtained from 100 commercially available peptide and peptoid building blocks assembled into linear or cyclic oligomers of up to 30 units, forming molecules within the size range of peptide drugs and accessible by solid-phase synthesis. We demonstrate ligand-based virtual screening (LBVS) using the peptide design genetic algorithm (PDGA), which evolves a population of 50 members to resemble a given target molecule using molecular fingerprint similarity as fitness function. Target molecules are reached in less than 10,000 generations. Like in many journeys, the value of the chemical space journey using PDGA lies not in reaching the target but in the journey itself, here by encountering molecules otherwise difficult to design. We also show that PDGA can be used to generate median molecules and analogs of non-peptide target molecules.
Markus Orsi; Jean-Louis Reymond Mol. Inform., 2025, 44, e202400186 https://doi.org/10.1002/minf.202400186AbstractHerein we report a virtual library of 1E+60 members, a common estimate for the total size of the drug-like chemical space. The library is obtained from 100 commercially available peptide and peptoid building blocks assembled into linear or cyclic oligomers of up to 30 units, forming molecules within the size range of peptide drugs and potentially accessible by solid-phase synthesis. We demonstrate ligand-based virtual screening (LBVS) using the peptide design genetic algorithm (PDGA), which evolves a population of 50 members to resemble a given target molecule using molecular fingerprint similarity as fitness function. Target molecules are reached in less than 10,000 generations. Like in many journeys, the value of the chemical space journey using PDGA lies not in reaching the target but in the journey itself, here by encountering non-obvious analogs. We also show that PDGA can be used to generate median molecules and analogs of non-peptide target molecules.
Jiayin Zhan; Xiang Li; Zhe Feng; Zheng Liu; Zhiyuan Feng; Jun-Jie Zhu; Jingjing Zhang Angew. Chem. Int. Ed., 2025, 64, e202415323 https://doi.org/10.1002/anie.202415323AbstractProteolysis-targeting chimeras (PROTACs) have accelerated drug development; however, some challenges still exist owing to their lack of tumor selectivity and on-demand protein degradation. Here, we developed a miRNA-initiated assembled pre-PROTAC (miRiaTAC) platform that enables the on-demand activation and termination of target degradation in a cell type-specific manner. Using miRNA-21 as a model, we engineered DNA hairpins labeled with JQ-1 and pomalidomide and facilitated the modular assembly of DNA-encoded pre-PROTACs through a hybridization chain reaction. This configuration promoted the selective polyubiquitination and degradation of BRD4 upon miR-21 initiation, highlighting significant tumor selectivity and minimal systemic toxicity. Furthermore, the platform incorporates photolabile groups, enabling the precise optical control of pre-PROTACs during DNA assembly/disassembly, mitigating the risk of excessive protein degradation. Additionally, by introducing a secondary ligand targeting CDK6, these pre-PROTACs were used as a modular scaffold for the programmable assembly of active miRiaTACs containing two different warheads in exact stoichiometry, enabling orthogonal multitarget degradation. The integration of near-infrared light-mediated photodynamic therapy through an upconversion nanosystem further enhanced the efficacy of the platform with potent in vivo anticancer activity. We anticipate that miRiaTAC represents a significant intersection between dynamic DNA nanotechnology and PROTAC, potentially expanding the versatility of PROTAC toolkit for cancer therapy.
Laura J. Byrnes, Won Young Choi, Paul Balbo, Mary Ellen Banker, Jeanne Chang, Shi Chen, Xuemin Cheng, Yang Cong, Jeff Culp, Hongxia Di, Matt Griffor, Justin Hall, Xiaoyun Meng, Barry Morgan, James J. Mousseau, Jennifer Nicki, Thomas O'Connell, Simeon Ramsey, Alex Shaginian, Suman Shanker, John Trujillo, Jinqiao Wan, Fabien Vincent, Stephen W. Wright, Felix Vajdos ACS Chemical Biology (2024). https://doi.org/10.1021/acschembio.4c00397 Abstract Peptidyl arginine deiminases (PADs) are enzymes implicated in various inflammatory and autoimmune diseases. Developing isoform-specific inhibitors has remained a challenge. This study reports the discovery of a potent, noncovalent, and selective PAD2 inhibitor, PF-3166, identified through DNA-encoded library (DEL) screening. Biochemical and biophysical assays revealed that PF-3166 inhibits PAD2 via a novel Ca²⁺-competitive allosteric mechanism. X-ray crystallography confirmed that PF-3166 induces conformational changes that eject Ca²⁺ from the high-affinity CA1 site and prevent Ca²⁺ binding at the CA2 site, both essential for enzyme activation. PF-3166 demonstrated selective cellular activity in human neutrophils, offering a valuable tool for elucidating PAD2 function in disease contexts. Summary A large-scale DEL screen targeting PAD2 yielded three compound series. Among them, PF-3166 stood out as a potent and selective PAD2 inhibitor (IC₅₀ = 150–600 nM), while the other two (PF-2635 and PF-8522) bound PAD2 but lacked inhibitory activity. Structural studies revealed that all three compounds bind at the PAD2 dimer interface. However, only PF-3166 induced significant conformational changes that disrupted Ca²⁺ binding at both CA1 and CA2 sites, leading to enzyme inhibition. This novel allosteric mechanism was further validated in cellular assays, where PF-3166 selectively inhibited PAD2-dependent citrullination in human neutrophils. Highlights 1. First report of a noncovalent, selective PAD2 inhibitor (PF-3166) from DEL screening. 2. PF-3166 inhibits PAD2 via a novel allosteric mechanism involving Ca²⁺ displacement from CA1 and CA2 sites. 3. X-ray structures reveal compound-induced conformational changes that prevent Ca²⁺ binding and enzyme activation. 4. PF-3166 is selective over PAD3 and PAD4 and shows cellular activity in human neutrophils. 5. Represents a new chemical tool for studying PAD2 biology and therapeutic potential. Conclusion PF-3166 is a first-in-class, cell-active, and selective PAD2 inhibitor that acts through a novel allosteric mechanism by disrupting Ca²⁺ binding at two critical sites (CA1 and CA2), essential for PAD2 activation. Its discovery underscores the power of DEL screening in uncovering unique inhibitory mechanisms. Despite liabilities such as high molecular weight and low permeability, PF-3166 provides a strong foundation for further optimization and a valuable tool for dissecting PAD2’s role in inflammation, autoimmunity, and cancer.
Levente M. Mihalovits; Tibor V. Szalai; Dávid Bajusz; György M. Keserű J. Chem. Inf. Model., 2024, 64(23), 8963–8979 https://doi.org/10.1021/acs.jcim.4c00803AbstractThe concept of DNA-encoded libraries (DELs) enables the experimental screening of billions of compounds simultaneously, offering an unprecedented boost in the coverage of chemical space. In parallel, however, dramatically increased access to supercomputers and a number of ultrahigh throughput virtual screening (uHTVS) tools have made screening of billion-membered virtual libraries available. Here, we investigate whether current, brute-force, or AI-enabled uHTVS approaches might constitute a computational alternative to DEL screening. While it is tempting to look at uHTVS as a computational analogue of DEL screening, we found specific advantages and limitations of both methodologies that suggest them being complementary rather than competitive.
Yoshitsugu Akiyama Nano Biomed., 2024, 16(1), 1-17 https://doi.org/10.11344/nano.16.1AbstractNucleic acids, serving as both a repository for genetic information and the basis for biological activity, can be regarded as highly ordered functional molecules. Oligonucleotides (ONs) are oligomers of nucleic acids that can now be obtained by chemical synthesis. The ONs thus obtained have been approved as nucleic acid therapeutics for the treatment of various diseases. Importantly, chemically modified non-natural ONs with augmented functionality are useful when the functions of natural ONs fall short of achieving therapeutic efficacy. Chemical modification of ONs can also facilitate their complexation with other functional materials and further enhance the functionality of the resulting hybrid materials. Examples of such modifications include the combination of ONs with small molecular compounds, peptides, proteins, or highly ordered nanostructures with enhanced biological stability as building blocks for self-assembly. Thus, strategies for the precise chemical synthesis of ONs will play an important role in the sustained progress of biomedical materials.
Zhongyao Ma; Bin Zou; Jiannan Zhao; Rui Zhang; Qiaoqiao Zhu; Xiaofeng Wang; Linan Xu; Xiang Gao; Xinyue Hu; Wei Feng; Wen Luo; Min Wang; Yunyun He; Zhifeng Yu; Weiren Cui; Qi Zhang; Letian Kuai; Wenji Su SLAS Discov., 2025 https://doi.org/10.1016/j.slasd.2024.100204 Abstract To date, RNA-targeted chemical matter is under explored due to a lack of robust screening assays. In this study, we present a novel RNA-targeted small molecule screening approach using a specialized DNA-encoded library (DEL). Our findings reveal that the specialized DEL library, called "DEL Zipper", can significantly reduce single-stranded DNA-RNA region interaction signals during various kinds of RNA selection. By performing the selection against both G-quadruplex, we have identified novel hits that interact with RNA targets and the results are validated through binding. This study demonstrates that the "DEL Zipper" method is a robust screening assay that has potential for discovering small molecule ligands for diverse RNA targets.
Kenneth López-Pérez; Juan F. Avellaneda-Tamayo; Lexin Chen; Edgar López-López; K. Eurídice Juárez-Mercado; José L. Medina-Franco; Ramón Alain Miranda-Quintana ChemRxiv, 2023 https://doi.org/10.26434/chemrxiv-2023-cs3wbAbstractMolecular similarity pervades much of our understanding and rationalization of chemistry. This has become particularly evident in the current data-intensive era of chemical research, with similarity measures serving as the backbone of many Machine Learning (ML) supervised and unsupervised procedures. Here, we present a discussion on the role of molecular similarity in drug design, chemical space exploration, chemical “art” generation, molecular representations, and many more. We also discuss more recent topics in molecular similarity, like the ability to efficiently compare large molecular libraries.
Ke Li; Mackenzie W. Krone; Arseniy Butrin; Michael J. Bond; Brian M. Linhares; Craig M. Crews J. Am. Chem. Soc., 2024, 146(36), 24884–24891 https://doi.org/10.1021/jacs.4c05393AbstractType I melanoma antigen (MAGE) family members are detected in numerous tumor types, and expression is correlated with poor prognosis, high tumor grade, and increased metastasis. Type I MAGE proteins are typically restricted to reproductive tissues, but expression can recur during tumorigenesis. Several biochemical functions have been elucidated for them, and notably, MAGEs regulate proteostasis by serving as substrate recognition modules for E3 ligase complexes. The repertoire of E3 ligase complexes that can be hijacked for targeted protein degradation continues to expand, and MAGE-E3 complexes are an especially attractive platform given their cancer-selective expression. Additionally, type I MAGE-derived peptides are presented on cancer cell surfaces, so targeted MAGE degradation may increase antigen presentation and improve immunotherapy outcomes. Motivated by these applications, we developed novel, small-molecule ligands for MAGE-A3, a type I MAGE that is widely expressed in tumors and associates with TRIM28, a RING E3 ligase. Chemical matter was identified through DNA-encoded library (DEL) screening, and hit compounds were validated for in vitro binding to MAGE-A3. We obtained a cocrystal structure with a DEL analog and hypothesize that the small molecule binds at a dimer interface. We utilized this ligand to develop PROTAC molecules that induce MAGE-A3 degradation through VHL recruitment and inhibit the proliferation of MAGE-A3 positive cell lines. These ligands and degraders may serve as valuable probes for investigating MAGE-A3 biology and as foundations for the ongoing development of tumor-specific PROTACs.
Rui Han; Tongtong Zhu; Zhiwei Kong; Xin Zhang; Dongli Wang; Junfeng Liu Int. J. Biol. Macromol., 2024, 2783), 134821 https://doi.org/10.1016/j.ijbiomac.2024.134821AbstractNecrosis-inducing secreted protein 1 (NIS1) is a core effector of Ascomycota and Basidiomycota fungi. They inhibit the immune responses of host plants mainly through interaction with the multi-functional coreceptor BRI1-associated receptor kinase 1 (BAK1). However, the structural mechanism of the NIS1 family and how they are recognized by BAK1 are unknown. Herein, we report the first crystal structure of the NIS1 family protein, the Magnaporthe oryzae NIS1 (MoNIS1), analyze the recognition mechanism of NIS1s by BAK1, and explore regulation of the NIS1-BAK1 interaction by a chemical compound. MoNIS1 exists as a β barrel formed by eight β strands, a folding mode that has not been reported. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) assay suggested that β4-β5 loop and β5 strand of MoNIS1 participate in OsBAK1 interaction, which was supported by further single-point mutational assays. For OsBAK1, HDX-MS assay suggested four regions involved in MoNIS1 interaction. Additionally, we identified a compound that blocks MoNIS1-OsBAK1 interaction in vitro and inhibits the virulence of M. oryzae on rice. Collectively, we determined the first structure of NIS1 family effectors, presented the recognition mechanism of NIS1 by BAK1, and showed that blocking NIS1-BAK1 interaction could be a new target for fungicide development.
Matteo Gasparetto; Balázs Fődi; Gellért Sipos Beilstein J. Org. Chem., 2024, 20, 1922-1932 https://doi.org/10.3762/bjoc.20.168AbstractAmino acids are vital motifs in the domain of biochemistry, serving as the foundational unit for peptides and proteins, while also holding a crucial function in many biological processes. Due to their bifunctional character, they have been also used for combinatorial chemistry purposes, such as the preparation of DNA-encoded chemical libraries. We developed a practical synthesis for α-heteroaryl-α-amino acids starting from an array of small heteroaromatic halides. The reaction sequence utilizes a photochemically enhanced Negishi cross-coupling as a key step, followed by oximation and reduction. The prepared amino esters were validated for on-DNA reactivity via a reverse amidation-hydrolysis-reverse amidation protocol.
Ping Liu; Refeya Jannatul; Juan Chen; Lihua Hou; Mingjuan Gao; Pengjie Wang; Lulu Wang; Dekui Jin; Hao Chen; Rong Liu; Ran Wang; Yinhua Zhu; Fang Bing; Lirong Jia; Yanan Sun; Yixin Zhang; Fazheng Ren; Weilin Lin Angew. Chem. Int. Ed. 2024, e202409746 https://doi.org/10.1002/anie.202409746AbstractNon-natural building blocks (BBs) present a vast reservoir of chemical diversity for molecular recognition and drug discovery. However, leveraging evolutionary principles to efficiently generate bioactive molecules with a larger number of diverse BBs poses challenges within current laboratory evolution systems. Here, we introduce programmable chemical evolution (PCEvo) by integrating chemoinformatic classification and high-throughput array synthesis/screening. PCEvo initiates evolution by constructing a diversely combinatorial library to create ancestral molecules, streamlines the molecular evolution process and identifies high-affinity binders within 2-4 cycles. By employing PCEvo with 108 BBs and exploring >10^17 chemical space, we identify bicyclic peptidomimetic binders against targets SAR-CoV-2 RBD and Claudin18.2, achieving nanomolar affinity. Remarkably, Claudin18.2 binders selectively stain gastric adenocarcinoma cell lines and patient samples. PCEvo achieves expedited evolution in a few rounds, marking a significant advance in utilizing non-natural building blocks for rapid chemical evolution applicable to targets with or without prior structural information and ligand preference.
Ruth Nussinov; Hyunbum Jang Expert Opin. Drug Discov., 2024 https://doi.org/10.1080/17460441.2024.2384467AbstractAllosteric drugs are advantageous. However, they still face hurdles, including identification of allosteric sites that will effectively alter the active site. Current strategies largely focus on id...
Yi-Ru Bai; Dong-Jie Seng; Ying Xu; Yao-Dong Zhang; Wen-Juan Zhou; Yang-Yang Jia; Jian Song; Zhang-Xu He; Hong-Min Liu; Shuo Yuan Eur. J. Med. Chem., 2024, 116706 https://doi.org/10.1016/j.ejmech.2024.116706AbstractIn 2023, the U.S. Food and Drug Administration has approved 55 novel medications, consisting of 17 biologics license applications and 38 new molecular entities. Although the biologics license applications including antibody and enzyme replacement therapy set a historical record, the new molecular entities comprising small molecule drugs, diagnostic agent, RNA interference therapy and biomacromolecular peptide still account for over 50 % of the newly approved medications. The novel and privileged scaffolds derived from drugs, active molecules and natural products are consistently associated with the discovery of new mechanisms, the expansion of clinical indications and the reduction of side effects. Moreover, the structural modifications based on the promising scaffolds can provide the clinical candidates with the improved biological activities, bypass the patent protection and greatly shorten the period of new drug discovery. Therefore, conducting an appraisal of drug approval experience and related information will expedite the identification of more potent drug molecules. In this review, we comprehensively summarized the pertinent information encompassing the clinical application, mechanism, elegant design and development processes of 28 small molecule drugs, and expected to provide the promising structural basis and design inspiration for pharmaceutical chemists.
Chloe Puteaux; Isabelle Toubia; Lina Truong; Marie Hubert-Roux; Laetitia Bailly; Hassan Oulyadi; Pierre-Yves Renard; Cyrille Sabot Angew. Chem. Int. Ed. 2024, e202407888 https://doi.org/10.1002/anie.202407888AbstractWe showcase the successful combination of photochemistry and kinetic target-guided synthesis (KTGS) for rapidly pinpointing enzyme inhibitors. KTGS is a fragment-based drug discovery (FBDD) methodology in which the biological target (BT) orchestrates the construction of its own ligand from fragments featuring complementary reactive functionalities. Notably, fragments interacting with the protein binding sites leverage their spatial proximity, facilitating a preferential reaction. Consequently, the resulting bivalent ligand exhibits heightened affinity. Within the realm of KTGS strategies, in situ click chemistry stands out as the most widely used to identify potent protein binders. This approach requires significant protein contributions, such as binding interactions and appropriate orientations of fragments, to overcome high activation barriers. This leads to prolonged incubation times and the potential for generating false negatives, thereby limiting this strategy to proteins that are stable enough in buffer. We herein unveil the possibility to integrate photochemistry into the realm of KTGS, accelerating the ligation reaction between fragments to a time scale of minutes. This approach should significantly expand the narrow reactivity window of traditional KTGS reactions, paving the way for the exploration and development of novel photo-KTGS reactions.
Lingling Hu; Zhidan Zhang; Congyu Li; Minghao Han; Mengyao Hao; Xu Zhang; Nida Ahmed; Jianmei Luo; Xiaoyun Lu; Jibin Sun; Huifeng Jiang Anal. Chem., 2024, 96(29), 12040-12048 https://doi.org/10.1021/acs.analchem.4c02110AbstractOligonucleotides represent a class of shorter DNA or RNA nucleic acid polymers extensively applied in the biomedical field. Despite progress in detecting and analyzing oligonucleotides, high-throughput analysis of the samples remains challenging. In this work, a high-throughput analysis method for oligonucleotide analysis was developed based on acoustic droplet ejection-open port interface-mass spectrometry (ADE-OPI-MS) technology. This approach was applied to determine the enzymatic activity of terminal deoxynucleotide transferase (TdT) for DNA synthesis, with a rate of 3 s/sample, which enhanced single-sample analysis efficiency approximately 60-fold over the previous gel analysis. After testing approximately 10,000 TdT mutants, we obtained three new variants with higher catalytic activities. Finally, by integrating these mutants, the catalytic activity of TdT was improved about 4 times compared to the starting mutant. Our results successfully established a high-throughput screening method for oligonucleotide analysis, which not only provides a foundation to engineer highly efficient TdT for ab initio synthesis of DNA but also paves the way for the potential application of oligonucleotide analysis in biomedical fields.
Takumi Hosozawa; Masatoshi Niwa; Hisayuki Takeuchi; Takehiko Inohana; Kaori Okumura; Shin Itoh Bioorg. Med. Chem. Lett., 2024, 129859 https://doi.org/10.1016/j.bmcl.2024.129859AbstractIn this study, we report on the ability of DMTMM PF6 to improve the amidation reaction. The on-DNA amidation reaction using DMTMM PF6 demonstrates higher conversion rates than those using HATU or DMTMM Cl, particularly with challenging sterically hindered amines and carboxylic acids. The developed method enables the expansion of available building blocks and the efficient synthesis of high-purity DNA-encoded libraries.
Yaolin Guo; Wen shuai; Aiping Tong; Yuxi Wang TrAC Trends Anal. Chem., https://doi.org/10.1016/j.trac.2024.117833AbstractCovalent inhibitors, forming reversible or irreversible covalent bonds with nucleophilic groups in target protein active sites, effectively inhibit protein function for therapeutic purposes. They can also be used for target validation, biomarker identification, and as chemical probes. Boasting high bioefficiency, low drug resistance, and minimal off-target effects, covalent inhibitors possess significant application potential within the small molecule drug market. Their discovery usually involves rational drug design. Understanding screening and identification tools for covalent inhibitors aids in selecting suitable and efficient development methods. This review encompasses screening platforms, target-ligand covalent binding, and selective analysis tools used in covalent inhibitor development. It focuses on discovery strategies such as computer-aided drug design, library screening, and classic techniques for binding determination and selectivity. Examples highlight the advantages and limitations of screening platforms, offering insights for covalent inhibitor discovery and advancing drug development.
Jason Deng; Svetlana Belyanskaya; Ninad Prabhu; Christopher Arico-Muendel; Hongfeng Deng; Christopher B. Phelps; David I. Israel; Hongfang Yang; Joseph Boyer; G. Joseph Franklin; Jeremy L. Yap; Kenneth E. Lind; Ching-Hsuan Tsai; Christine Donahue; Jennifer D. Summerfield SLAS Discov., 2024 https://doi.org/10.1016/j.slasd.2024.100171AbstractDNA-encoded small molecule library technology has recently emerged as a new paradigm for identifying ligands against drug targets. To date, it has been used to identify ligands against targets that are soluble or overexpressed on cell surfaces. Here, we report applying cell-based selection methods to profile surfaces of mouse C2C12 myoblasts and myotube cells in an unbiased, target agnostic manner. A panel of on-DNA compounds were identified and confirmed for cell binding selectivity. We optimized the cell selection protocol and employed a novel data analysis method to identify cell selective ligands against a panel of human B and T lymphocytes. We discuss the generality of using this workflow for DNA encoded small molecule library selection and data analysis against different cell types, and the feasibility of applying this method to profile cell surfaces for biomarker and target identification.
Lijun Xue; Jiaqing Yu; Ying Zhong; Junyun Chen; Chao Li; Kexin Yang; Nicolas Duchemin; Yunjin Hu Chem. Commun., 2024, 60, 6885-6888 https://doi.org/10.1039/d4cc02193bAbstractWe here describe a visible-light photooxidation of sulfinate salts with common alkenes to yield β-hydroxy sulfones on DNA. This process demonstrates a broad substrate compatibility and achieves conversion rates ranging from moderate to excellent. Most importantly, it presents a straightforward, efficient, and metal-free approach for synthesizing Csp3-rich DNA-encoded libraries.
Ying Huang; Yixin Zhang; Chenchen Hu; Yiyun Chen Org. Chem. Front., 2024 https://doi.org/10.1039/D4QO00752BAbstractExpanding DNA functionality has significant implications in nucleic acid chemistry, biology, and beyond. Therefore, developing new chemical tools for site-specific post-synthetic modification of nucleic acids is urgently needed. Herein, we demonstrate the first site-specific DNA post-synthetic modification via visible-light-catalyzed decarboxylative allylation. Allyl sulfone groups were introduced into DNA, not only at the terminal sites via amide formation but also at internal and terminal positions during DNA solid-phase synthesis. This visible-light-catalyzed decarboxylative allylation proceeds rapidly on DNA bearing allyl sulfone groups under open-air conditions within minutes, exhibiting excellent chemoselectivity and compatibility with various functional groups while retaining DNA integrity. Specifically, introducing allyl sulfones into DNA via solid-phase synthesis enables site-specific modification on chemically synthesized single-stranded DNA (internal and terminal positions), hybridized double-stranded DNA, and enzymatically amplified long-chain DNA under visible light irradiation. The versatile reactivity of allyl sulfone scaffolds further enables diverse on-DNA photocatalytic transformations, promising to advance the chemical toolbox for DNA post-synthetic modification through diverse photochemical methods.
Antoine Douchez; Julien Poupart; Gaoqiang Yang; Louis Vaillancourt; Anne Marinier Bioconjugate Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.bioconjchem.4c00160AbstractDNA-encoded libraries (DELs) can be considered as one of the most powerful tools for the discovery of small molecules of biological interest. However, the ability to access large DELs is contingent upon having chemical transformations that work in aqueous phase and generate minimal DNA alterations and the availability of building blocks compatible with on-DNA chemistry. In addition, accessing scaffolds of interest to medicinal chemists can be challenging in a DEL setting because of inherent limitations of DNA-supported chemistry. In this context, a squaramide formation reaction was developed by using a two-step process. The mild and high-yielding reaction tolerates a wide array of functional groups and was shown to be safe for DNA, thereby making this methodology ideal for DELs.
Hui Zhao; Anne Ravn; Michael Haibach; Keary Engle; Carin Johansson Seechurn ChemRxiv, 2024 https://doi.org/10.26434/chemrxiv-2024-twzr2AbstractRecent global events have led to the cost of platinum group metals (PGMs) reaching unprecedented heights. Many chemical companies are therefore starting to seriously consider and evaluate if, and where, they can substitute PGMs for non-PGMs in their catalytic processes. This review covers recent large-scale applications of non-PGM catalysts in the modern pharmaceutical industry. By highlighting these selected successful examples of non-PGM-catalyzed processes from the literature, we hope to emphasize the enormous potential of non-PGM catalysis and inspire further development within this field to enable this technology to progress towards manufacturing processes. We also present some historical context and review the perceived advantages and challenges of implementing non-PGM catalysts in the pharmaceutical manufacturing environment.
Yuyu Xing; Huiya Zhang; Yanhui Wang; Zhaoyun Zong; Matthew Bogyo; Shiyu Chen RSC Chem. Biol., 2024, 5, 691-702 https://doi.org/10.1039/D4CB00097HAbstractCovalent protease inhibitors serve as valuable tools for modulating protease activity and are essential for investigating the functions of protease targets. These inhibitors typically consist of a recognition motif and a covalently reactive electrophile. Substrate peptides, featuring residues capable of fitting into the substrate pockets of proteases, undergo chemical modification at the carbonyl carbon of the P1 residue with an electrophile and have been widely applied in the development of covalent inhibitors. In this study, we utilized a DNA-encoded peptide library to replicate peptide binder sequences and introduced a vinyl sulfone warhead at the C-termini to construct the DNA-encoded peptide covalent inhibitor library (DEPCIL) for targeting cysteine proteases. Screening results toward 3CL protease demonstrated the efficacy of this library, not only in identifying protease inhibitors, but also in discovering amino acids that can conform to aligned protease pockets. The identified peptide sequences provide valuable insight into the amino acid preferences within substrate binding pockets, and our novel technology is indicative of the potential for similar strategies to discover covalent inhibitors and profile binding preferences of other proteases.
Shan-Shan Zhang; Lance Larrabee; Andrew H. Chang; Sapna Desai; Lisa Sloan; Xin Wang; Yixuan Wu; Nazia Parvez; Karen Amaratunga; Allison C. Hartman; Abby Whitnall; Joseph Mason; Nicholas P. Barton; Audrey Y. Chu; Jonathan M. Davitte; Adam J. Csakai; Caitlin Vestal Tibbetts; Audrey E. Tolbert; Heather O'Keefe; Jessie Polanco; Joseph Foley; Casey Kmett; Jonathan Kehler; Gabriela Kozejova; Feng Wang; Andrew P. Mayer; Patrick Koenig; Davide Foletti; Steven J. Pitts; Christine G. Schnackenberg Sci. Rep., 2024, 14, 13209 https://doi.org/10.1038/s41598-024-62804-7AbstractHypertension remains a leading cause of cardiovascular and kidney diseases. Failure to control blood pressure with ≥ 3 medications or control requiring ≥ 4 medications is classified as resistant hypertension (rHTN) and new therapies are needed to reduce the resulting increased risk of morbidity and mortality. Here, we report genetic evidence that relaxin family peptide receptor 2 (RXFP2) is associated with rHTN in men, but not in women. This study shows that adrenal gland gene expression of RXFP2 is increased in men with hypertension and the RXFP2 natural ligand, INSL3, increases adrenal steroidogenesis and corticosteroid secretion in human adrenal cells. To address the hypothesis that RXFP2 activation is an important mechanism in rHTN, we discovered and characterized small molecule and monoclonal antibody (mAb) blockers of RXFP2. The novel chemical entities and mAbs show potent, selective inhibition of RXFP2 and reduce aldosterone and cortisol synthesis and release. The RXFP2 mAbs have suitable rat pharmacokinetic profiles to evaluate the role of RXFP2 in the development and maintenance of rHTN. Overall, we identified RXFP2 activity as a potential new mechanism in rHTN and discovered RXFP2 antagonists for the future interrogation of RXFP2 in cardiovascular and renal diseases.
Liwei Zheng; Yujia Sun; Michael Eisenstein; Hyongsok Tom Soh bioRxiv, 2024 https://doi.org/10.1101/2024.05.31.596913AbstractScalable methods that can accurately sequence peptides at single-amino acid resolution could significantly advance proteomic studies. We present a protein sequencing method based on the “reverse translation” of peptide sequence information into DNA barcodes that document the identity, position, and the originating peptide of each amino acid. We employ a modified Edman degradation process that converts peptides into DNA-barcoded amino acids, which are subsequently detected by proximity extension assay, yielding multi-barcoded DNA outputs that can be PCR amplified and sequenced. Using our method, we sequenced multiple consecutive amino acids within a model peptide. This method also enables the differentiation of single amino acid substitutions, and the identification of post-translational modifications and their positions within multiple peptides simultaneously. With further development, we anticipate that this method will enable highly parallel de novo protein sequencing with single-molecule sensitivity.
Lucy L. Robbie; Tyler Beyett J. Pharmacol. Exp. Ther., 2024, 389(S3), 293 https://doi.org/10.1124/jpet.293.129469AbstractAbstract ID 129469Poster Board 293Thymocytes are involved in the adaptive immune system where they eliminate pathogens. Mature T cells develop from immature thymocytes through a regulated developmental process called selection. During this process, thymocytes are selected for their ability to recognize antigens. THymocyte-Expressed Molecule Involved In Selection (THEMIS) is a protein expressed in developing thymocytes that is required for maturation of T cells. THEMIS contains tandem Cystine-containing All Beta In THEMIS (CABIT) domains, which are only found in a limited number of human proteins. THEMIS interacts with and regulates tyrosine phosphatases SHP-1 and -2, though the precise molecular mechanism is unknown. THEMIS expression increases in CD4+/CD8+ cells, suggesting THEMIS may hold a pivotal role in regulating selection at the double-positive stage by modulating phosphotyrosine signaling downstream of the T cell receptor.Despite its significance to T cell development, it is unknown whether THEMIS represents a therapeutic target due to a lack of understanding of its function and an absence of chemical probes. To remedy this, we are conducting two screens to identify ligands that bind THEMIS in collaboration with Structural Genomics Consortium (SGC). First, we are using a DNA Encoded Library (DEL) screen with &gt;109 small molecule ligands. In this approach, THEMIS is used as a bait in a pulldown and molecules that bind are identified by sequencing their unique DNA tags. In parallel, we are using Affinity Selection-Mass Spectrometry (AS-MS) to identify ligands. AS-MS utilizes size-exclusion chromatography to enrich for small molecules from a mixture that co-elute with the protein of interest prior to identification by mass spectrometry.Prior to screening, THEMIS was recombinantly expressed and purified and mass spectrometry was utilized to confirm the sequence of the protein and identify any post-translational modifications. As ligands are identified from DEL and AS-MS screening, compounds are clustered based on structural similarities to streamline validation and analysis. We are utilizing a variety of biophysical approaches to assess the binding affinities and sites of select ligands on THEMIS. Further investigation into these probes can lead to the discovery of THEMIS modulators, which could be utilized to block or enhance protein-protein interactions between THEMIS and SHP1/2, manipulating downstream signaling. This may reveal unrealized therapeutic applications for THEMIS in T cell malignancies and immunotherapies.We acknoledge Emory University for startup funds and the Structural Genomics Consortium for financial support.
Samuele Cazzamalli; Tony Georgiev; Francesca Migliorini; Andrea Ciamarone; Marco Müller; Ilaria Biancofiore; Nicholas Favalli; Dario Neri; Sebastian Oehler J. Nucl. Med., 2024, 65(supplement 2), 241120 https://jnm.snmjournals.org/content/65/supplement_2/241120 Abstract 241120 Introduction: Lutetium Vipivotide Tetraxetan (PluvictoTM, a Novartis product), a PSMA-targeted Radio Ligand Therapeutic (RLT), has revolutionized the treatment of metastatic castration-resistant prostate cancer. Despite the clinical benefit observed in large pivotal trials, virtually all patients treated with the drug eventually relapse (VISION trial). Moreover, the product shows an on-target off tumor uptake in healthy tissues, such as salivary glands and kidneys, which is responsible for certain toxicities observed in the clinic. Prostatic Acid Phosphatase (ACP3) is a non-specific phosphomonoesterase synthesized by prostate epithelial cells (Babain et al., Seminars in Nuclear Medicine, 1989, 4:309). ACP3 expression levels are elevated in most prostate cancer lesions, while the antigen is virtually absent in other healthy organs, including salivary glands and kidneys (). Technetium-99m and Indium-111 labeled anti-ACP3 antibody fragments have been successfully used for the radioimmunodetection of metastatic prostate cancer lesions in patients (Vihko et al., Prostate, 1987, 11:51). Here, we present the isolation of first-in-class high-affinity ACP3 ligands from DNA-Encoded Chemical Libraries. Methods: ACP3 ligands were isolated from DNA-Encoded Chemical Libraries of ~5.8 million members. The most enriched compounds were synthesized as DOTAGA and fluorescein conjugates to measure their affinity against recombinant human ACP3 and binding on antigen-positive cancer cells. Lutetium-177-labeled hit compounds were injected in tumor-bearing mice to assess their in vivo biodistribution and anti-cancer activity. Results: ProX1, ProX2, and ProX3 were identified as highly potent small organic binders and inhibitors of human ACP3. Affinities in the picomolar range were measured for the three compounds by surface plasmon resonance. The ProX1 ligand (also named “OncoACP3”) resulted in the best tumor accumulation and residence time after intravenous dosing in mice bearing HT-1080.hACP3 or PC3.hACP3 subcutaneous cancer lesions (i.e., >35 %ID/g, 72 hours after administration) with low salivary gland and kidney uptake. Low doses of 177Lu-OncoACP3 (i.e., 5 and 20 MBq/mouse) induced durable cancer remissions in the HT-1080.hACP3 in vivo tumor model. Conclusions: Given its exceptional tumor-targeting performance and the lack of salivary glands and kidney uptake, OncoACP3 promises to be the next best-in-class targeting agent for prostate cancer for ligand-based pharmacodelivery applications.
Jun Liang; Michael J. Lambrecht; Teresita L. Arenzana; Samuel Aubert-Nicol; Linda Bao; Fabio Broccatelli; Jianping Cai; Celine Eidenschenk; Christine Everett; Thomas Garner; Felix Gruber; Pouyan Haghshenas; Malcolm P. Huestis; Peter L. Hsu; Ponien Kou; Araz Jakalian; Robin Larouche-Gauthier; Jean-Philippe Leclerc; Dennis H. Leung; Aaron Martin; Jeremy Murray; Madeleine Prangley; Sascha Rutz; Satoko Kakiuchi-Kiyota; Alexander Lee Satz; Nicholas J. Skelton; Micah Steffek; Daniel Stoffler; Jawahar Sudhamsu; Sophia Tan; Jian Wang; Shouliang Wang; Qiuyue Wang; Timothy J. Wendorff; Moreno Wichert; Arun Yadav; Christine Yu; Xiaojing Wang ACS Med. Chem. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acsmedchemlett.4c00068AbstractWe were attracted to the therapeutic potential of inhibiting Casitas B-lineage lymphoma proto-oncogene-b (Cbl-b), a RING E3 ligase that plays a critical role in regulating the activation of T cells. However, given that only protein–protein interactions were involved, it was unclear whether inhibition by a small molecule would be a viable approach. After screening an ∼6 billion member DNA-encoded library (DEL) using activated Cbl-b, we identified compound 1 as a hit for which the cis-isomer (2) was confirmed by biochemical and surface plasmon resonance (SPR) assays. Our hit optimization effort was greatly accelerated when we obtained a cocrystal structure of 2 with Cbl-b, which demonstrated induced binding at the substrate binding site, namely, the Src homology-2 (SH2) domain. This was quite noteworthy given that there are few reports of small molecule inhibitors that bind to SH2 domains and block protein–protein interactions. Structure- and property-guided optimization led to compound 27, which demonstrated measurable cell activity, albeit only at high concentrations.
Louise M. Sternicki; Sally-Ann Poulsen RSC Med. Chem., 2024 https://doi.org/10.1039/D4MD00273CAbstractNative mass spectrometry (nMS) is well established as a biophysical technique for characterising biomolecules and their interactions with endogenous or investigational small molecule ligands. The high sensitivity mass measurements make nMS particularly well suited for applications in fragment-based drug discovery (FBDD) screening campaigns where the detection of weakly binding ligands to a target biomolecule is crucial. We first reviewed the contributions of nMS to guiding FBDD hit identification in 2013, providing a comprehensive perspective on the early adoption of nMS for fragment screening. Here we update this initial progress with a focus on contributions of nMS that have guided FBDD for the period 2014 until end of 2023. We highlight the development of nMS adoption in FBDD in the context of other biophysical fragment screening techniques. We also discuss the roadmap for increased adoption of nMS for fragment screening beyond soluble proteins, including for guiding the discovery of fragments supporting advances in PROTAC discovery, RNA-binding small molecules and covalent therapeutic drug discovery.
Peiyuan Zhang; Xiyun Ye; John C. K. Wang; Hannah T. Baddock; Zena Jensvold; Ian T. Foe; Andrei Loas; Dan L. Eaton; Qi Hao; Aaron H. Nile; Bradley L. Pentelute J. Am. Chem. Soc., 2024, 146, 22, 15627–15639 https://doi.org/10.1021/jacs.4c05571AbstractCovalent peptide binders have found applications as activity-based probes and as irreversible therapeutic inhibitors. Currently, there is no rapid, label-free, and tunable affinity selection platform to enrich covalent reactive peptide binders from synthetic libraries. We address this challenge by developing a reversibly reactive affinity selection platform termed ReAct-ASMS enabled by tandem high-resolution mass spectrometry (MS/MS) to identify covalent peptide binders to native protein targets. It uses mixed disulfide-containing peptides to build reversible peptide–protein conjugates that can enrich for covalent variants, which can be sequenced by MS/MS after reduction. Using this platform, we identified covalent peptide binders against two oncoproteins, human papillomavirus 16 early protein 6 (HPV16 E6) and peptidyl-prolyl cis–trans isomerase NIMA-interacting 1 protein (Pin1). The resulting peptide binders efficiently and selectively cross-link Cys58 of E6 at 37 °C and Cys113 of Pin1 at room temperature, respectively. ReAct-ASMS enables the identification of highly selective covalent peptide binders for diverse molecular targets, introducing an applicable platform to assist preclinical therapeutic development pipelines.
Guixian Zhao; Mengping Zhu; Yangfeng Li; Gong Zhang; Yizhou Li Expert Opin. Drug Discov., 2024, 19(6), 725-740 https://doi.org/10.1080/17460441.2024.2354287AbstractINTRODUCTION: The effectiveness of Fragment-based drug design (FBDD) for targeting challenging therapeutic targets has been hindered by two factors: the small library size and the complexity of the fragment-to-hit optimization process. The DNA-encoded library (DEL) technology offers a compelling and robust high-throughput selection approach to potentially address these limitations. AREA COVERED: In this review, the authors propose the viewpoint that the DEL technology matches perfectly with the concept of FBDD to facilitate hit discovery. They begin by analyzing the technical limitations of FBDD from a medicinal chemistry perspective and explain why DEL may offer potential solutions to these limitations. Subsequently, they elaborate in detail on how the integration of DEL with FBDD works. In addition, they present case studies involving both de novo hit discovery and full ligand discovery, especially for challenging therapeutic targets harboring broad drug-target interfaces. EXPERT OPINION: The future of DEL-based fragment discovery may be promoted by both technical advances and application scopes. From the technical aspect, expanding the chemical diversity of DEL will be essential to achieve success in fragment-based drug discovery. From the application scope side, DEL-based fragment discovery holds promise for tackling a series of challenging targets.
Xu Cui; Yi Liu; Lih Feng Cheow bioRxiv, 2024 https://doi.org/10.1101/2024.05.07.593072AbstractScreening for peptide fragments that can be displayed on antigen-presenting cells is an essential step in vaccine development. The current approach for this process is slow and costly as it involves separately pulsing cells with chemically synthesized peptides. We present Microfluidic-Enabled production of DNA-barcoded APC Library (MEDAL), a high throughput microfluidic droplet platform for parallel production of DNA-barcoded APCs loaded with enzymatically synthesized peptides. Droplets containing peptides and their encoding DNA are produced from microfluidic PCR-IVTT reaction. APCs presenting both peptides and DNA barcodes are obtained by injecting cells into these droplets. Up to 9,000 different APCs can be produced and screened within a 10-hour workflow. This approach allowed us to identify peptide sequences that bind to APCs expressing H-2Kb MHC class 1 molecule with next-generation sequencing of DNA barcodes.
Anna Vulpetti; Jean-Michel Rondeau; Marie-Hélène Bellance; Jutta Blank; Ralf Boesch; Andreas Boettcher; Frédéric Bornancin; Sylvia Buhr; Lauren E. Connor; Christoph E. Dumelin; Esser, Oliver; Michael Hediger; Hintermann, Samuel; Ulrich Hommel; Elke Koch; Guillaume Lapointe; Lukas Leder; Sylvie Lehmann; Philipp Lehr; Peter Meier; Lionel Muller; Daniela Ostermeier; Paul Ramage; Sihame Schiebel-Haddad; Alexander Baxter Smith; Aleksandar Stojanovic; Juraj Velcicky; Rina Yamamoto; Konstanze Hurth J. Med. Chem., 2024, 67(10), 8141–8160 https://doi.org/10.1021/acs.jmedchem.4c00240AbstractHuman interleukin-1β (IL-1β) is a pro-inflammatory cytokine that plays a critical role in the regulation of the immune response and the development of various inflammatory diseases. In this publication, we disclose our efforts toward the discovery of IL-1β binders that interfere with IL-1β signaling. To this end, several technologies were used in parallel, including fragment-based screening (FBS), DNA-encoded library (DEL) technology, peptide discovery platform (PDP), and virtual screening. The utilization of distinct technologies resulted in the identification of new chemical entities exploiting three different sites on IL-1β, all of them also inhibiting the interaction with the IL-1R1 receptor. Moreover, we identified lysine 103 of IL-1β as a target residue suitable for the development of covalent, low-molecular-weight IL-1β antagonists.
Marco Müller; Laura Lucaroni; Nicholas Favalli; Gabriele Bassi; Dario Neri; Samuele Cazzamalli; Sebastian Oehler J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.4c00332AbstractProstate-specific membrane antigen (PSMA)-targeted radio ligand therapeutics (RLTs), such as [177Lu]Lu-PSMA-617 (Pluvicto), have been shown to accumulate in salivary glands and kidneys, potentially leading to undesired side effects. As unwanted accumulation in normal organs may derive from the cross-reactivity of PSMA ligands to glutamate carboxypeptidase III (GCPIII), it may be convenient to block this interaction with GCPIII-selective ligands. Parallel screening of a DNA-encoded chemical library (DEL) against GCPIII and PSMA allowed the identification of GCPIII binders. Structure–activity relationship (SAR) studies resulted in the identification of nanomolar GCPIII ligands with up to 1000-fold selectivity over PSMA. We studied the ability of GCPIII ligands to counteract the binding of [177Lu]Lu-PSMA-617 to human salivary glands by autoradiography and could demonstrate a partial radioprotection.
Chandrashekhar Madasu; Zian Liao; Sydney E. Parks; Kiran L. Sharma; Kurt M. Bohren; Qiuji Ye; Feng Li; Murugesan Palaniappan; Zhi Tan; Fei Yuan; Chad J. Creighton; Suni Tang; Ramya P. Masand; Xiaoming Guan; Damian W. Young; Diana Monsivais; Martin M. Matzuk Proc. Natl. Acad. Sci. USA, 2024, 121(19), e2322934121 https://doi.org/10.1073/pnas.2322934121AbstractEPH receptors (EPHs), the largest family of tyrosine kinases, phosphorylate downstream substrates upon binding of ephrin cell surface-associated ligands. In a large cohort of endometriotic lesions from individuals with endometriosis, we found that EPHA2 and EPHA4 expressions are increased in endometriotic lesions relative to normal eutopic endometrium. Because signaling through EPHs is associated with increased cell migration and invasion, we hypothesized that chemical inhibition of EPHA2/4 could have therapeutic value. We screened DNA-encoded chemical libraries (DECL) to rapidly identify EPHA2/4 kinase inhibitors. Hit compound, CDD-2693, exhibited picomolar/nanomolar kinase activity against EPHA2 (Ki: 4.0 nM) and EPHA4 (Ki: 0.81 nM). Kinome profiling revealed that CDD-2693 bound to most EPH family and SRC family kinases. Using NanoBRET target engagement assays, CDD-2693 had nanomolar activity versus EPHA2 (IC50: 461 nM) and EPHA4 (IC50: 40 nM) but was a micromolar inhibitor of SRC, YES, and FGR. Chemical optimization produced CDD-3167, having picomolar biochemical activity toward EPHA2 (Ki: 0.13 nM) and EPHA4 (Ki: 0.38 nM) with excellent cell-based potency EPHA2 (IC50: 8.0 nM) and EPHA4 (IC50: 2.3 nM). Moreover, CDD-3167 maintained superior off-target cellular selectivity. In 12Z endometriotic epithelial cells, CDD-2693 and CDD-3167 significantly decreased EFNA5 (ligand) induced phosphorylation of EPHA2/4, decreased 12Z cell viability, and decreased IL-1β-mediated expression of prostaglandin synthase 2 (PTGS2). CDD-2693 and CDD-3167 decreased expansion of primary endometrial epithelial organoids from patients with endometriosis and decreased Ewing's sarcoma viability. Thus, using DECL, we identified potent pan-EPH inhibitors that show specificity and activity in cellular models of endometriosis and cancer.
Giulia Alboreggia; Parima Udompholkul; Carlo Baggio; Kendall Muzzarelli; Zahra Assar; Maurizio Pellecchia J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.4c00277AbstractSeveral novel and effective cysteine targeting (Cys) covalent drugs are in clinical use. However, the target area containing a druggable Cys residue is limited. Therefore, methods for creating covalent drugs that target different residues are being looked for; examples of such ligands include those that target the residues lysine (Lys) and tyrosine (Tyr). Though the histidine (His) side chain is more frequently found in protein binding locations and has higher desirable nucleophilicity, surprisingly limited research has been done to specifically target this residue, and there are not many examples of His-targeting ligands that have been rationally designed. In the current work, we created novel stapled peptides that are intended to target hMcl-1 His 252 covalently. We describe the in vitro (biochemical, NMR, and X-ray) and cellular design and characterization of such agents. Our findings further suggest that the use of electrophiles to specifically target His residues is warranted.
Ramón Rama-Garda; Eduardo Domínguez; María Isabel Loza; María José Lallena; Jesús de Blas; Miguel Ángel Toledo; Rubén Haro ASSAY and Drug Development Technologies, 2024, 010https://doi.org/10.1089/adt.2024.010 Abstract DNA-encoded libraries (DELs) have demonstrated to be one of the most powerful technologies within the ligand identification toolbox, widely used either in academia or biotech and pharma companies. DEL methodology utilizes affinity selection (AS) as the approach to interrogate the protein of interest for the identification of binders. Here we present a high-throughput, fully automated AS platform developed to fulfill industrial standards and compatible with different assay formats to improve the reproducibility of the AS process for DEL binders identification. This platform is flexible enough to virtually set aside all kinds of DELs and AS methods and conditions using immobilized proteins. It bears the two main immobilization methods to support of the proteins of interest: magnetic beads or resin tip columns. A combination of a broad variety of protocol options with a wide range of different experimental conditions can be set up with a throughput of 96 samples at the same time. In addition, small modifications of the protocols provide the platform with the versatility to run not only the routine DEL screens, but also test covalent libraries, the successful immobilization of the proteins of interest, and many other experiments that may be required. This versatile AS platform for DEL can be a powerful instrument for direct application of the technology in academic and industry settings.
Hyemi Lee; Jiho Nam; Hahyeong Jang; Young-Sik Park; Min-Hee Son; In-Hyun Lee; Seong-Il Eyun; Jae-Hyun Yang; Jimin Jeon; Siyoung Yang Br. J. Pharmacol., 2024, 10.1111/bph.16359 https://doi.org/10.1111/bph.16359AbstractBACKGROUND AND PURPOSE: The discovery of new bromo- and extra-terminal inhibitors presents new drugs to treat osteoarthritis (OA). EXPERIMENTAL APPROACH: The new drug, BBC0403, was identified in the DNA-encoded library screening system by searching for compounds that target BRD (bromodomain-containing) proteins. The binding force with BRD proteins was evaluated using time-resolved fluorescence energy transfer (TR-FRET) and binding kinetics assays. Subsequently, in vitro and ex vivo analyses demonstrated the effects of the BRD2 inhibitor, BBC0403, on OA. For animal experiments, medial meniscus destabilization was performed to create a 12-week-old male C57BL/6 mouse model, and intra-articular (i.a.) injections were administered. Histological and immunohistochemical analyses were then performed. The underlying mechanism was confirmed by gene set enrichment analysis (GSEA) using RNA-seq. KEY RESULTS: TR-FRET and binding kinetics assays revealed that BBC0403 exhibited higher binding specificity for BRD2 compared to BRD3 and BRD4. The anti-OA effects of BBC0403 were tested at concentrations of 5, 10 and 20 μM (no cell toxicity in the range tested). The expression of catabolic factors, prostaglandin E2 (PGE2) production and extracellular matrix (ECM) degradation was reduced. Additionally, the i.a. injection of BBC0403 prevented OA cartilage degradation in mice. Finally, BBC0403 was demonstrated to suppress NF-κB and MAPK signalling pathways. CONCLUSION AND IMPLICATIONS: This study demonstrated that BBC0403 is a novel BRD2-specific inhibitor and a potential i.a.-injectable therapeutic agent to treat OA.
Yu Fan; Ruibing Feng; Xinya Zhang; Zhen-Liang Wang; Feng Xiong; Shuihua Zhang; Zhang-Feng Zhong; Hua Yu; Qing-Wen Zhang; Zhang Zhang; Yitao Wang; Guodong Li Acta Pharm. Sin. B, 2024, 14(8), 3362-3384 https://doi.org/10.1016/j.apsb.2024.04.006AbstractDrug discovery is a sophisticated process that incorporates scientific innovations and cutting-edge technologies. Compared to traditional bioactivity-based screening methods, encoding and display technologies for combinatorial libraries have recently advanced from proof-of-principle experiments to promising tools for pharmaceutical hit discovery due to their high screening efficiency, throughput, and resource minimization. This review systematically summarizes the development history, typology, and prospective applications of encoding and displayed technologies, including phage display, ribosomal display, mRNA display, yeast cell display, one-bead one-compound, DNA-encoded, peptide nucleic acid-encoded, and new peptide-encoded technologies, and examples of preclinical and clinical translation. We discuss the progress of novel targeted therapeutic agents, covering a spectrum from small-molecule inhibitors and nonpeptidic macrocycles to linear, monocyclic, and bicyclic peptides, in addition to antibodies. We also address the pending challenges and future prospects of drug discovery, including the size of screening libraries, advantages and disadvantages of the technology, clinical translational potential, and market space. This review is intended to establish a comprehensive high-throughput drug discovery strategy for scientific researchers and clinical drug developers.
Cyril Balsollier; Simon Bijkerk; Arjan de Smit; Kevin van Eekelen; Krištof Bozovičar; Dirk Husstege; Tihomir Tomašič; Marko Anderluh; Roland J. Pieters Bioorg. Chem., 2024, 107321 https://doi.org/10.1016/j.bioorg.2024.107321AbstractFinding potent inhibitors of O-GlcNAc transferase (OGT) has proven to be a challenge, especially because the diversity of published inhibitors is low. The large majority of available OGT inhibitors are uridine-based or uridine-like compounds that mimic the main interactions of glycosyl donor UDP-GlcNAc with the enzyme. Until recently, screening of DNA-encoded libraries for discovering hits against protein targets was dedicated to a few laboratories around the world, but has become accessible to wider public with the recent launch of the DELopen platform. Here we report the results and follow-up of a DNA-encoded library screening by using the DELopen platform. This led to the discovery of two new hits with structural features not resembling UDP. Small focused libraries bearing those two scaffolds were made, leading to low micromolar inhibition of OGT and elucidation of their structure–activity relationship.
Pengyang He; Guixian Zhao; Mengping Zhu; Yangfeng Li; Gong Zhang; Yizhou Li Org. Chem. Front., 2024 https://doi.org/10.1039/D4QO00203BAbstractFunctional group transformation represents a potent strategy for enriching the chemical diversity in DNA-encoded chemical library (DEL) synthesis. Within the realm of functional group transformation reactions, oxidation stands out as a predominant and facile approach to deliver altered chemical reactivity. However, reports on DNA-compatible oxidative reactions facilitating functional group transformations remain limited. Herein, we present a DNA-compatible K2RuO4-mediated oxidation methodology, which enables the in situ generation of carboxylic acids from alcohols and nitriles from primary amines, respectively. With several advantages, such as mild reaction conditions, high conversion rates, and broad substrate scopes, this oxidative transformation facilitates the potential utilization of carboxylic and nitrile groups as reactive handles in DEL synthesis. This work implicates how oxidation could be harnessed as a versatile approach to expand the chemical space of DEL for potential drug discovery purposes.
Yue Zhang; Jia-ying Xue; Xiao-can Su; Wen-jie Xiao; Jing-yi Lv; Wen-xia Shi; Yong Zou; Ming Yan; Xue-jing Zhang Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.4c00377AbstractIn this report, we present a photopromoted, metal-free transannulation of phenyl azides for the synthesis of DNA-encoded seven-membered rings. The transformation is efficiently achieved through a skeletal editing strategy targeting the benzene motif coupled with a Reversible Adsorption to Solid Support (RASS) strategy. A variety of valuable DNA-encoded seven-membered ring compounds, including DNA-encoded 3H-azepines, azepinones, and unnatural amino acids, are now accessible. Crucially, this DNA-compatible protocol can also be applied for the introduction of complex molecules, as exemplified by Lorcaserin and Betahistine. The selective conversion of readily available phenyl rings into high-value seven-membered rings offers a promising avenue for the construction of diversified and drug-like DNA-encoded library.
Dongliang Guan; Jiaxiang Liu; Feifei Chen; Jian Li; Xiaowen Wang; Weiwei Lu; Yanrui Suo; Feng Tang; Lefu Lan; Xiaojie Lu; Wei Huang J. Med. Chem., 2024, 67, 5, 3778–3794 https://doi.org/10.1021/acs.jmedchem.3c02197AbstractIt is an urgent need to tackle the global crisis of multidrug-resistant bacterial infections. We report here an innovative strategy for large-scale screening of new antibacterial agents using a whole bacteria-based DNA-encoded library (DEL) of vancomycin derivatives via peripheral modifications. A bacterial binding affinity assay was established to select the modification fragments in high-affinity compounds. The optimal resynthesized derivatives demonstrated excellently enhanced activity against various resistant bacterial strains and provided useful structures for vancomycin derivatization. This work presents the new concept in a natural product-templated DEL and in antibiotic discovery through bacterial affinity screening, which promotes the fight against drug-resistant bacteria.
Aliakbar Khalili Yazdi; Sumera Perveen; Xiaosheng Song; Aiping Dong; Magdalena M. Szewczyk; Matthew F. Calabrese; Agustin Casimiro-Garcia; Subramanyam Chakrapani; Matthew S. Dowling; Emel Ficici; Jisun Lee; Justin I. Montgomery; Thomas N. O’Connell; Grzegorz J. Skrzypek; Tuan P. Tran; Matthew D. Troutman; Feng Wang; Jennifer A. Young; Jinrong Min; Dalia Barsyte-Lovejoy; Peter J. Brown; Vijayaratnam Santhakumar; Cheryl H. Arrowsmith; Masoud Vedadi; Dafydd R. Owen bioRxiv, 2023 https://doi.org/10.1101/2023.11.13.566858AbstractWe have developed a novel chemical handle (PFI-E3H1) and a chemical probe (PFI-7) as ligands for the Gid4 subunit of the human E3 ligase CTLH degradation complex. Through an efficient initial hit-ID campaign, structure-based drug design (SBDD) and leveraging the sizeable Pfizer compound library, we identified a 500 nM ligand for this E3 ligase through file screening alone. Further exploration identified a vector that is tolerant to addition of a linker for future chimeric molecule design. The chemotype was subsequently optimized to sub-100 nM Gid4 binding affinity for a chemical probe. These novel tools, alongside the suitable negative control also identified, should enable the interrogation of this complex human E3 ligase macromolecular assembly.
Ayun Luo; Hongxia Zhou; Xiuming Wang; Fanming Zeng; Weina Yu; Kexin Yang; Nicolas Duchemin; Yun Jin Hu Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.4c00187AbstractUsing a novel homologation-heterocyclization cascade, the on-DNA synthesis of benzofurans from aldehydes has been developed. The methodology, based on an innovative use of the Seyferth-Gilbert homologation, followed by a high yielding Sonogashira coupling in situ intramolecular cyclization one-pot, two-step reaction, provides a powerful and unique pathway for DNA-encoded library (DEL) synthesis of a wide array of pharmaceutically relevant benzofuran-based scaffolds.
Lanmeng Cui; Wei Hou; Hongtao Xu Future Med. Chem., 2024, 16(6) https://doi.org/10.4155/fmc-2024-0041AbstractThese physicochemical properties are closely correlated to the diverse biological and pharmacological activities of Se when incorporated into small molecules or biomacromolecules.
Sandeep Pal; Zandile Nare; Vincenzo A. Rao; Brian O. Smith; Ian Morrison; Edward A. Fitzgerald; Andrew Scott; Matilda Jane Bingham; Thomas Pesnot ChemMedChem, 2024, e202300590 https://doi.org/10.1002/cmdc.202300590AbstractWe report the development of BioPhysical and Active Learning Screening (BioPALS); a rapid and versatile hit identification protocol combining AI-powered virtual screening with a GCI-driven biophysical confirmation workflow. Its application to the BRPF1b bromodomain afforded a range of novel micromolar binders with favorable ADMET parameters. In addition to the excellent in silico/in vitro confirmation rate demonstrated with BRPF1b, binding kinetics are determined, and binding topologies predicted for all hits. BioPALS is a lean, data-rich, and standardized approach to hit identification applicable to wide range of biological targets.
Gaonan Wang; Yu Tan; Hanzhi Zou; Xihang Sui; Zhanlong Wang; Alexander L. Satz; Letian Kuai; Wenji Su; Qi Zhang Org. Lett., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c04240AbstractDNA-encoded chemical library (DECL) technology is a commonly employed screening platform in both the pharmaceutical industry and academia. To expand the chemical space of DECLs, new and robust DNA-compatible reactions are sought after. In particular, DNA-compatible cyclization reactions are highly valued, as these reactions tend to be atom economical and thus may provide lead- and drug-like molecules. Herein, we report two new methodologies employing DNA-conjugated thiosemicarbazides as a common precursor, yielding highly substituted 1,3,4-oxadiazoles and 1,2,4-triazoles. These two novel DNA-compatible reactions feature a high conversion efficiency and broad substrate scope under mild conditions that do not observably degrade DNA.
Felix Gruber; Anthony W. McDonagh; Victoria Rose; James Hunter; Laura Guasch; Rainer E. Martin; Stefanie N. Geigle; Robert Britton Angew. Chem. Int. Ed. Engl., 2024, e202319836 https://doi.org/10.1002/anie.202319836AbstractDNA encoded library (DEL) synthesis represents a convenient means to produce, annotate and store large collections of compounds in a small volume. While DELs are well suited for drug discovery campaigns, the chemistry used in their production must be compatible with the DNA tag, which can limit compound class accessibility. As a result, most DELs are heavily populated with peptidomimetic and sp2 -rich molecules. Herein, we show that sp3 -rich mono- and bicyclic heterocycles can be made on DNA from ketochlorohydrin aldol products through a reductive amination and cyclization process. The resulting hydroxypyrrolidines possess structural features that are desirable for DELs and target a distinct region of pharmaceutically relevant chemical space.
Martin Vogt Expert Opin. Drug Discov., https://doi.org/10.1080/17460441.2024.2313475AbstractINTRODUCTION: Large chemical spaces (CSs) include traditional large compound collections, combinatorial libraries covering billions to trillions of molecules, DNA-encoded chemical libraries comprising complete combinatorial CSs in a single mixture, and virtual CSs explored by generative models. The diverse nature of these types of CSs require different chemoinformatic approaches for navigation. AREAS COVERED: An overview of different types of large CSs is provided. Molecular representations and similarity metrics suitable for large CS exploration are discussed. A summary of navigation of CSs in generative models is provided. Methods for characterizing and comparing CSs are discussed. EXPERT OPINION: The size of large CSs might restrict navigation to specialized algorithms and is limited to considering neighborhoods of structurally similar molecules. Efficient navigation of large CSs not only requires methods that scale with size but also requires smart approaches that focus on better but not necessarily larger molecule selections. Deep generative models aim to provide such approaches by implicitly learning features relevant for target biological properties. It is unclear whether these models can fulfill this ideal as validation is difficult as long as the covered CSs remain mainly virtual without experimental verification.
Koen F. W. Hekking; Sergio Maroto; Kees van Kekem; Frank S. Haasjes; Jack C. Slootweg; Patrick G. B. Oude Alink; Ron Dirks; Malvika Sardana; Marjon G. Bolster; Brian Kuijpers; Dennis Smith; Robin Doodeman; Marcel Scheepstra; Birgit Zech; Mark Mulvihill; Louis M. Renzetti; Lee Babiss; Paolo A. Centrella; Matthew A. Clark; John W. Cuozzo; Marie-Aude Guié; Eric Sigel; Sevan Habeshian; Christopher D. Hupp; Julie Liu; Heather A. Thomson; Ying Zhang; Anthony D. Keefe; Gerhard Müller; Stijn Gremmen J. Med. Chem., 2024, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.jmedchem.3c02206AbstractEvasion of apoptosis is critical for the development and growth of tumors. The pro-survival protein myeloid cell leukemia 1 (Mcl-1) is an antiapoptotic member of the Bcl-2 family, associated with tumor aggressiveness, poor survival, and drug resistance. Development of Mcl-1 inhibitors implies blocking of protein-protein interactions, generally requiring a lengthy optimization process of large, complex molecules. Herein, we describe the use of DNA-encoded chemical library synthesis and screening to directly generate complex, yet conformationally privileged macrocyclic hits that serve as Mcl-1 inhibitors. By applying a conceptual combination of conformational analysis and structure-based design in combination with a robust synthetic platform allowing rapid analoging, we optimized in vitro potency of a lead series into the low nanomolar regime. Additionally, we demonstrate fine-tuning of the physicochemical properties of the macrocyclic compounds, resulting in the identification of lead candidates 57/59 with a balanced profile, which are suitable for future development toward therapeutic use.
Haochun Jin; Dianxin Cui; Yu Fan; Guodong Li; Zhangfeng Zhong; Yitao Wang Drug Discov. Today, 2024, 103885 https://doi.org/10.1016/j.drudis.2024.103885AbstractBioaffinity drug screening strategies have gained popularity in preclinical and clinical drug discovery for natural products, small molecules and antibodies owing to their superior selectivity, the large number of compounds to be screened and their ability to reduce the time and cost of the drug discovery process. This paper provides a systematic summary of the principles of commonly used bioaffinity-based screening methods, elaborates on the success of bioaffinity in clinical drug development and summarizes the active compounds, preclinical drugs and marketed drugs obtained through affinity screening methods. Owing to the high demand for new drugs, bioaffinity-guided screening techniques will play a greater part in clinical drug development. Teaser: Here, we analyze the different affinity screening methods and their clinical outcomes, underscoring the significance of this technique in drug discovery.
Matthew Anderson; Thomas Carton; Catherine Salvini; James Crawford; Garry Pairaudeau; Michael James Waring Chem. Eur. J., 2024, e202400239 https://doi.org/10.1002/chem.202400239AbstractDNA-encoded libraries (DELs) have become a leading technology for hit identification in drug discovery projects as large, diverse libraries can be generated. DELs are commonly synthesised via split-and-pool methodology; thus, chemical transformations utilised must be highly efficient, proceeding with high conversions. Reactions performed in DEL synthesis also require a broad substrate scope to produce diverse, drug-like libraries. Many pharmaceutical compounds incorporate multiple C-N bonds, over a quarter of which are synthesised via reductive aminations. However, few on-DNA reductive amination procedures have been developed. Herein is reported the application of the micelle-forming surfactant, TPGS-750-M, to the on-DNA reductive amination of DNA-conjugated amines, yielding highly efficient conversions with a broad range of aldehydes, including medicinally relevant heterocyclic and aliphatic substrates. The procedure is compatible with DNA amplification and sequencing, demonstrating its applicability to DEL synthesis.
Pratik Worah arXiv, 2024, 2401.11562 https://doi.org/10.48550/arXiv.2401.11562AbstractGiven two labeled data-sets $\mathcal{S}$ and $\mathcal{T}$, we design a simple and efficient greedy algorithm to reweigh the loss function such that the limiting distribution of the neural network weights that result from training on $\mathcal{S}$ approaches the limiting distribution that would have resulted by training on $\mathcal{T}$. On the theoretical side, we prove that when the metric entropy of the input data-sets is bounded, our greedy algorithm outputs a close to optimal reweighing, i.e., the two invariant distributions of network weights will be provably close in total variation distance. Moreover, the algorithm is simple and scalable, and we prove bounds on the efficiency of the algorithm as well. Our algorithm can deliberately introduce distribution shift to perform (soft) multi-criteria optimization. As a motivating application, we train a neural net to recognize small molecule binders to MNK2 (a MAP Kinase, responsible for cell signaling) which are non-binders to MNK1 (a highly similar protein). We tune the algorithm's parameter so that overall change in holdout loss is negligible, but the selectivity, i.e., the fraction of top 100 MNK2 binders that are MNK1 non-binders, increases from 54\% to 95\%, as a result of our reweighing. Of the 43 distinct small molecules predicted to be most selective from the enamine catalog, 2 small molecules were experimentally verified to be selective, i.e., they reduced the enzyme activity of MNK2 below 50\% but not MNK1, at 10$\mu$M -- a 5\% success rate.
Erica Benedetti; Laurent Micouin Expert Opin. Drug Discov., 2024 https://doi.org/10.1080/17460441.2024.2305735AbstractSpirocycles are structures in which two or more rings are linked by one common atom. Although the use of such scaffolds has been known for more than 60 years in medicinal chemistry, the increasing interest in compounds with enhanced three-dimensionality and higher proportion of sp3-hybridized atoms has led to a growing number of examples of potential drug candidates incorporating spirocyclic elements [Citation1,Citation2]. Shifting from planar, aromatic structures to compounds with a higher fraction of saturated carbon (Fsp3) generally correlates with improved physicochemical properties as well as pharmacokinetic (PK) profiles. The modulation of these properties has been nicely illustrated by works on azaspirocycles, showing higher solubility, higher basicity, decreased liphophilicity and better metabolic stability than their six-membered ring piperazine, piperidines, morpholines or thiomorpholines counterparts [Citation3]. A similar trend can be observed with spirocyclic oxetanes [Citation4]. Furthermore, the rigidity of the spirocyclic system can lock the conformation of the molecule and optimize the orientation of binding elements in a controlled manner, leading to an improved efficacy and/or selectivity profile. In this editorial, we discuss how the advantages of using a spirocyclic scaffold have been exploited in drug design, with a focus on recent examples reported in this field.
Ruizhi Lai; Zhiqian Lin; Chunyan Yang; Li Hai; Zhongzhen Yang; Li Guo; Ruifang Nie; Yong Wu E. J. Med. Chem., 2024, 116116 https://doi.org/10.1016/j.ejmech.2023.116116AbstractAdenoviral E1A binding protein p300 (EP300 or p300) and its similar paralog, cyclic-AMP response element binding protein (CBP), are important histone acetyltransferases (HAT) and transcriptional co-activators in epigenetics, participating in numerous cellular pathways including proliferation, differentiation and apoptosis. The overexpression or dysregulation of p300/CBP is closely related to oncology-relevant disease. The inhibition of p300 HAT has been found to be a potential drug target. Berberine has been reported to show anticancer activity and synergistic effect in combination with some of the clinical anticancer drugs via modulation of various pathways. Here, the present study sought to discover more chemotypes of berberine derivatives as p300 HAT inhibitors and to examine the combination of these novel analogues with doxorubicin for the treatment of breast cancer. A series of novel berberine derivatives with modifications of A/B/D rings of berberine have been designed, synthesized and screened. Compound 7b was found to exhibit inhibitory potency against p300 HAT with IC50 values of 1.51 μM. Western blotting proved that 7b decreased H3K27Ac and interfered with the expression of oncology-relevant protein in MCF-7 cells. Further bioactive evaluation showed that combination of compound 7b with doxorubicin could significantly inhibit tumor growth and invasion in vitro and in vivo.
Joshua ML Casan; John F. Seymour Blood, 2023, 2023022993. https://doi.org/10.1182/blood.2023022993AbstractTargeted protein degradation (TPD) is a revolutionary approach to targeted therapy in hematological malignancies that potentially circumvents many constraints of existing small molecule inhibitors. Heterobifunctional proteolysis targets chimeras (PROTACs) are the leading TPD drug class, with numerous agents now in clinical trials for a range of blood cancers. PROTACs harness the cell intrinsic protein recycling infrastructure, the ubiquitin-proteasome system (UPS), to completely degrade target proteins. Distinct from targeted small molecule inhibitor therapies, PROTACs can eliminate critical but conventionally 'undruggable' targets, overcome resistance mechanisms to small molecule therapies and can improve tissue specificity and off-target toxicity. Orally bioavailable, PROTACs are not dependent on the occupancy driven pharmacology inherent to inhibitory therapeutics, facilitating sub-stoichiometric dosing that does not require an active or allosteric target binding site. Preliminary clinical data demonstrate promising therapeutic activity in heavily pre-treated populations and novel technology platforms are poised to exploit myriad permutations of PROTAC molecular design to enhance efficacy and targeting specificity. As the field rapidly progresses and various non-PROTAC TPD drug candidates emerge, this review explores the scientific and pre-clinical foundations of PROTACs and presents them within common clinical contexts. Additionally, we examine the latest findings from ongoing active PROTAC clinical trials.
Natalie Fuchs; Longfei Zhang; Laura Calvo-Barreiro; Katarzyna Kuncewicz; Moustafa Gabr J. Pers. Med., 2024, 14(1), 68 https://doi.org/10.3390/jpm14010068AbstractThe revolutionary progress in cancer immunotherapy, particularly the advent of immune checkpoint inhibitors, marks a significant milestone in the fight against malignancies. However, the majority of clinically employed immune checkpoint inhibitors are monoclonal antibodies (mAbs) with several limitations, such as poor oral bioavailability and immune-related adverse effects (irAEs). Another major limitation is the restriction of the efficacy of mAbs to a subset of cancer patients, which triggered extensive research efforts to identify alternative approaches in targeting immune checkpoints aiming to overcome the restricted efficacy of mAbs. This comprehensive review aims to explore the cutting-edge developments in targeting immune checkpoints, focusing on both small molecule- and peptide-based approaches. By delving into drug discovery platforms, we provide insights into the diverse strategies employed to identify and optimize small molecules and peptides as inhibitors of immune checkpoints. In addition, we discuss recent advances in nanomaterials as drug carriers, providing a basis for the development of small molecule- and peptide-based platforms for cancer immunotherapy. Ongoing research focused on the discovery of small molecules and peptide-inspired agents targeting immune checkpoints paves the way for developing orally bioavailable agents as the next-generation cancer immunotherapies.
Yinliang Bai; Yujun Qiao; Mingming Li; Wenzhen Yang; Haile Chen; Yanqing Wu; Honghua Zhang E. J. Med. Chem., 2024, 116123 https://doi.org/10.1016/j.ejmech.2024.116123AbstractWithin the field of medical science, there is a great deal of interest in investigating cell death pathways in the hopes of discovering new drugs. Over the past two decades, pharmacological research has focused on necroptosis, a cell death process that has just been discovered. Receptor-interacting protein kinase 1 (RIPK1), an essential regulator in the cell death receptor signalling pathway, has been shown to be involved in the regulation of important events, including necrosis, inflammation, and apoptosis. Therefore, researching necroptosis inhibitors offers novel ways to treat a variety of disorders that are not well-treated by the therapeutic medications now on the market. The research and medicinal potential of RIPK1 inhibitors, a promising class of drugs, are thoroughly examined in this study. The journey from the discovery of Necrostatin-1 (Nec-1) to the recent advancements in RIPK1 inhibitors is marked by significant progress, highlighting the integration of traditional medicinal chemistry approaches with modern technologies like high-throughput screening and DNA-encoded library technology. This review presents a thorough exploration of the development and therapeutic potential of RIPK1 inhibitors, a promising class of compounds. Simultaneously, this review highlights the complex roles of RIPK1 in various pathological conditions and discusses potential inhibitors discovered through diverse pathways, emphasizing their efficacy against multiple disease models, providing significant guidance for the expansion of knowledge about RIPK1 and its inhibitors to develop more selective, potent, and safe therapeutic agents.
Takashi Osawa YAKUGAKU ZASSHI, 2024, 144, 931-936 https://doi.org/10.1248/yakushi.24-00134AbstractOligonucleotides, including DNA and RNA, can be functionalized by chemical modification based on synthetic organic chemistry. For example, ligand–oligonucleotide conjugates have a wide variety of applications. Conjugates of functional ligands and oligonucleotides have attracted attention in recent years as a drug delivery system (DDS) for improving the efficacy of oligonucleotide therapeutics. In addition, oligonucleotide conjugates with drug candidate compounds as ligands have been applied to drug screening using DNA-encoded libraries (DELs). Against this background, we have focused on the development of practical synthetic methods for ligand–oligonucleotide conjugates. Recently, we have developed a new synthetic method to construct oligonucleotides conjugated with coumarins and dipeptides, which are expected to have bioactivity, for application to DDS research of oligonucleotide therapeutics and drug discovery research using DEL. In this review, we will discuss the details, including how to construct a coumarin scaffold on oligonucleotides based on Knoevenagel condensation.
Jie Zhang; Lu Wang; Qian Ji; Fei Liu Org. Lett., 2023, 25(37), 6931-6936 https://doi.org/10.1021/acs.orglett.3c02850AbstractA DNA-compatible reaction has been developed for the cyanomethylation of (hetero)aryl halides or triflates via a tandem process involving palladium-mediated Suzuki–Miyaura coupling and base-promoted isoxazole fragmentation. This one-pot protocol employs easily accessible starting materials, exhibits a wide substrate scope, and results in no significant DNA damage. Additionally, the resulting (hetero)arylacetonitriles can be converted into the corresponding carboxylic acids, which may be utilized for the synthesis of DNA-encoded chemical libraries.
Shabbir Ahmad; Jin Xu; Jianwen A Feng; Ashley Hutchinson; Hong Zeng; Pegah Ghiabi; Aiping Dong; Paolo A Centrella; Matthew A Clark; Marie-Aude Guie; John P Guilinger; Anthony D Keefe; Ying Zhang; Thomas Cerruti; John W Cuozzo; Moritz von Rechenberg; Albina Bolotokova; Yanjun Li; Peter Loppnau; Alma Seitova; Yen-Yen Li; Vijayaratnam Santhakumar; Peter Brown; Suzanne Ackloo; Levon Halabelian bioRxiv, 2023 https://doi.org/10.1101/2023.08.21.552681AbstractWD40 repeat-containing protein 91 regulates endosomal phosphatidylinositol 3-phosphate levels at the critical stage of endosome maturation and plays vital roles in endosome fusion, recycling, and transport by mediating protein-protein interactions. Due to its various roles in endocytic pathways, WDR91 has recently been identified as a potential host factor responsible for viral infection. We employed DNA-Encoded Chemical Library (DEL) selection against the WDR domain of WDR91, followed by machine learning to generate a model that was then used to predict ligands from the synthetically accessible Enamine REAL database. Screening of predicted compounds enabled us to identify the hit compound 1, which binds selectively to WDR91 with a KD of 6 ± 2 μM by surface plasmon resonance. The co-crystal structure confirmed the binding of 1 to the WDR91 side pocket, in proximity to cysteine 487. Machine learning-assisted structure activity relationship-by-catalog validated the chemotype of 1 and led to the discovery of covalent analogs 18 and 19. Intact mass LC-MS and differential scanning fluorimetry confirmed the formation of a covalent adduct, and thermal stabilization, respectively. The discovery of 1, 18, 19, accompanying SAR, and co-crystal structures will provide valuable insights for designing more potent and selective compounds against WDR91, thus accelerating the development of novel chemical tools to evaluate the therapeutic potential of WDR91 in disease.
Xinyin Wu; Pan Gao; Feng Chen Eur. J. Org. Chem., 2023, e202300864 https://doi.org/10.1002/ejoc.202300864AbstractThe utilization of aryl radicals as open-shelled intermediates has become an essential tool for both conventional and state-of-the-art synthetic chemistry. However, the current methods for generating aryl radicals are still inefficient, greatly impeding their practical applications. Encouragingly, sulfonium salts have emerged as appealing sources of aryl radicals for a wide range of transformations aimed at creating novel chemical bonds driven by their distinctive structural attributes and chemical tendencies. This review primarily focuses on the specific reaction mechanisms underlying the cleavage of C-S bonds in sulfonium salts, leading to the generation of corresponding aryl radicals within diverse reaction conditions.
Andreas Varkaris; Ermira Pazolli; Hakan Gunaydin; Qi Wang; Levi Pierce; Alessandro A. Boezio; Lucian DiPietro; Adam Frost; Fabrizio Giordanetto; Erika P. Hamilton; Katherine Harris; Michael Holliday; Tamieka L Hunter; Amanda Iskandar; Yongli Ji; Alexandre Larivée; Jonathan R. LaRochelle; André Lescarbeau; Fabien Llambi; Brenda Lormil; Mary M. Mader; Brenton G. Mar; Iain Martin; Thomas H. McLean; Klaus Michelsen; Yakov Pechersky; Erika Puente-Poushnejad; Ramin Samadani; Alison M. Schram; Kelley Shortsleeves; Sweta Swaminathan; Shahein Tajmir; Gege Tan; Yong Tang; Roberto Valverde; Bryan Wehrenberg; Jeremy Wilbur; Bret R. Williams; Hongtao Zeng; W. Patrick Walters; Beni B. Wolf; David E. Shaw; Donald A. Bergstrom; James Watters; James S. Fraser; Pascal D. Fortin; D. Randal Kipp Cancer Discov., 2023 https://doi.org/10.1158/2159-8290.CD-23-0944AbstractPIK3CA (PI3Ka) is a lipid kinase commonly mutated in cancer, including ~40% of hormone receptor-positive breast cancer. The most frequently observed mutants occur in the kinase and helical domains. Orthosteric PI3Ka inhibitors suffer from poor selectivity leading to undesirable side effects, most prominently hyperglycemia due to inhibition of wild-type (WT) PI3Ka. Here, we used molecular dynamics simulations and cryo-electron microscopy to identify an allosteric network that provides an explanation for how mutations favor PI3Ka activation. A DNA-encoded library screen leveraging electron microscopy-optimized constructs, differential enrichment, and an orthosteric-blocking compound led to the identification of RLY-2608, a first-in-class allosteric mutant-selective inhibitor of PI3Ka. RLY-2608 inhibited tumor growth in PIK3CA mutant xenograft models with minimal impact on insulin, a marker of dysregulated glucose homeostasis. RLY-2608 elicited objective tumor responses in two patients diagnosed with advanced hormone receptor-positive breast cancer with kinase or helical domain PIK3CA mutations, with no observed WT PI3Ka-related toxicities.
Lulu Jiang; Sixiu Liu; Xinglong Jia; Qinting Gong; Xin Wen; Weiwei Lu; Jintong Yang; Xinyuan Wu; Xuan Wang; Yanrui Suo; Yilin Li; Motonari Uesugi; Zhi-bei Qu; Minjia Tan; Xiaojie Lu; Lu Zhou J. Am. Chem. Soc., 2023, 145(46), 25283-25292 https://doi.org/10.1021/jacs.3c08852AbstractDNA-encoded chemical library (DEL) has been extensively used for lead compound discovery for decades in academia and industry. Incorporating an electrophile warhead into DNA-encoded compounds recently permitted the discovery of covalent ligands that selectively react with a particular cysteine residue. However, noncysteine residues remain underexplored as modification sites of covalent DELs. Herein, we report the design and utility of tyrosine-targeting DELs of 67 million compounds. Proteome-wide reactivity analysis of tyrosine-reactive sulfonyl fluoride (SF) covalent probes suggested three enzymes (phosphoglycerate mutase 1, glutathione s-transferase 1, and dipeptidyl peptidase 3) as models of tyrosine-targetable proteins. Enrichment with SF-functionalized DELs led to the identification of a series of tyrosine-targeting covalent inhibitors of the model enzymes. In-depth mechanistic investigation revealed their novel modes of action and reactive ligand-accessible hotspots of the enzymes. Our strategy of combining activity-based proteome profiling and covalent DEL enrichment (ABPP-CoDEL), which generated selective covalent binders against a variety of target proteins, illustrates the potential use of this methodology in further covalent drug discovery.
Shabbir Ahmad; Jin Xu; Jianwen A. Feng; Ashley Hutchinson; Hong Zeng; Pegah Ghiabi; Aiping Dong; Paolo A. Centrella; Matthew A. Clark; Marie-Aude Guié; John P. Guilinger; Anthony D. Keefe; Ying Zhang; Thomas Cerruti; John W. Cuozzo; Moritz von Rechenberg; Albina Bolotokova; Yanjun Li; Peter Loppnau; Alma Seitova; Yen-Yen Li; Vijayaratnam Santhakumar; Peter J. Brown; Suzanne Ackloo; Levon Halabelian J. Med. Chem., 2023, 66(23), 16051-16061 https://doi.org/10.1021/acs.jmedchem.3c01471AbstractWD40 repeat-containing protein 91 (WDR91) regulates early-to-late endosome conversion and plays vital roles in endosome fusion, recycling, and transport. WDR91 was recently identified as a potential host factor for viral infection. We employed DNA-encoded chemical library (DEL) selection against the WDR domain of WDR91, followed by machine learning to predict ligands from the synthetically accessible Enamine REAL database. Screening of predicted compounds identified a WDR91 selective compound 1, with a KD of 6 ± 2 μM by surface plasmon resonance. The co-crystal structure confirmed the binding of 1 to the WDR91 side pocket, in proximity to cysteine 487, which led to the discovery of covalent analogues 18 and 19. The covalent adduct formation for 18 and 19 was confirmed by intact mass liquid chromatography–mass spectrometry. The discovery of 1, 18, and 19, accompanying structure–activity relationship, and the co-crystal structures provide valuable insights for designing potent and selective chemical tools against WDR91 to evaluate its therapeutic potential.
Huicong Wang; Guixian Zhao; Tianyang Zhang; Yangfeng Li; Gong Zhang; Yizhou Li ACS Pharmacol. Transl. Sci., 2023, 2023, 6(11) 1724–1733 https://doi.org/10.1021/acsptsci.3c00181AbstractDNA-encoded libraries (DEL) have emerged as an important drug discovery technical platform for target-based compound library selection. The success rate of DEL depends on both the chemical diversity of combinatorial libraries and the accuracy of DNA barcoding. Therefore, it is critical that the chemistry applied to library construction should efficiently transform on a wide range of substrates while preserving the integrity of DNA tags. Although several analytical methods have been developed to measure DNA damage caused by DEL chemical reactions, efficient and cost-effective evaluation criteria for DNA damage detection are still demanding. Herein, we set standards for evaluating the DNA compatibility of chemistry development at the laboratory level. Based on four typical DNA damage models of three different DEL formats, we evaluated the detection capabilities of four analytical methods, including ultraperformance liquid chromatography (UPLC-MS), electrophoresis, quantitative polymerase chain reaction (qPCR), and Sanger sequencing. This work systematically revealed the scope and capability of different analytical methods in assessing DNA damages caused by chemical transformation. Based on the results, we recommended UPLC-MS and qPCR as efficient methods for DNA barcode integrity analysis in the early-stage development of DNA-compatible chemistry. Meanwhile, we identified that Sanger sequencing was unreliable to assess DNA damage in this application.
Yinkai Duan; Haofeng Wang; Zhenghong Yuan; Haitao Yang Curr. Opin. Struct. Biol., 2023, 102667 https://doi.org/10.1016/j.sbi.2023.102667AbstractSince its outbreak in late 2019, the COVID-19 pandemic has drawn enormous attention worldwide as a consequence of being the most disastrous infectious disease in the past century. As one of the most immediately druggable targets of SARS-CoV-2, the main protease (Mpro) has been studied thoroughly. In this review, we provide a comprehensive summary of recent advances in structural studies of Mpro, which provide new knowledge about Mpro in terms of its biological function, structural characteristics, substrate specificity, and autocleavage process. We examine the remarkable strides made in targeting Mpro for drug discovery during the pandemic. We summarize insights into the current understanding of the structural features of Mpro and the discovery of existing Mpro-targeting drugs, illuminating pathways for the future development of anti-SARS-CoV-2 therapeutics.
Daniel Riege; Sven Herschel; Teresa Fenkl; Dennis Schade ACS Pharmacol. Transl. Sci., 2023, 6(11), 1574–1599 https://doi.org/10.1021/acsptsci.3c00170AbstractThe bone morphogenetic protein (BMP) pathway is highly conserved and plays central roles in health and disease. The quality and quantity of its signaling outputs are regulated at multiple levels, offering pharmacological options for targeted modulation. Both target-centric and phenotypic drug discovery (PDD) approaches were applied to identify small-molecule BMP inhibitors and stimulators. In this Review, we accumulated and systematically classified the different reported chemotypes based on their targets as well as modes-of-action, and herein we illustrate the discovery history of selected candidates. A comprehensive summary of available biochemical, cellular, and in vivo activities is provided for the most relevant BMP modulators, along with recommendations on their preferred use as chemical probes to study BMP-related (patho)physiological processes. There are a number of high-quality probes used as BMP inhibitors that potently and selectively interrogate the kinase activities of distinct type I (16 chemotypes available) and type II receptors (3 chemotypes available). In contrast, only a few high-quality BMP stimulator modalities have been introduced to the field due to a lack of profound target knowledge. FK506-derived macrolides such as calcineurin-sparing FKBP12 inhibitors currently represent the best-characterized chemical tools for direct activation of BMP-SMAD signaling at the receptor level. However, several PDD campaigns succeeded in expanding the druggable space of BMP stimulators. Albeit the majority of them do not entirely fulfill the strict chemical probe criteria, many chemotypes exhibit unique and unrecognized mechanisms as pathway potentiators or synergizers, serving as valuable pharmacological tools for BMP perturbation.
Shuang Liu; Bingqi Tong; Jeremy W. Mason; Jonathan M. Ostrem; Antonin Tutter; Bruce K. Hua; Sunny A. Tang; Simone Bonazzi; Karin Briner; Frédéric Berst; Frédéric J. Zécri; Stuart L. Schreiber J. Am. Chem. Soc., 2023, 145(42), 23281-23291 https://doi.org/10.1021/jacs.3c08307AbstractThe hallmark of a molecular glue is its ability to induce cooperative protein–protein interactions, leading to the formation of a ternary complex, despite weaker binding toward one or both individual proteins. Notably, the extent of cooperativity distinguishes molecular glues from bifunctional compounds, which constitute a second class of inducers of protein–protein interactions. However, apart from serendipitous discovery, there have been limited rational screening strategies for the high cooperativity exhibited by molecular glues. Here, we propose a binding-based screen of DNA-barcoded compounds on a target protein in the presence or absence of a presenter protein, using the “presenter ratio”, the ratio of ternary enrichment to binary enrichment, as a predictive measure of cooperativity. Through this approach, we identified a range of cooperative, noncooperative, and uncooperative compounds in a single DNA-encoded library screen with bromodomain containing protein (BRD)9 and the VHL–elongin C–elongin B (VCB) complex. Our most cooperative hit compound, 13-7, exhibits micromolar binding affinity to BRD9 but nanomolar affinity for the ternary complex with BRD9 and VCB, with cooperativity comparable to classical molecular glues. This approach may enable the rational discovery of molecular glues for preselected proteins and thus facilitate the transition to a new paradigm of small-molecule therapeutics.
Ye Buehler; Jean-Louis Reymond J. Chem. Inf. Model., 2023, 63(20), 6239-6248 https://doi.org/10.1021/acs.jcim.3c01096AbstractIdentifying innovative fragments for drug design can help medicinal chemistry address new targets and overcome the limitations of the classical molecular series. By deconstructing molecules into ring fragments (RFs, consisting of ring atoms plus ring-adjacent atoms) and acyclic fragments (AFs, consisting of only acyclic atoms), we find that public databases of molecules (i.e., ZINC and PubChem) and natural products (i.e., COCONUT) contain mostly RFs and AFs of up to 13 atoms. We also find that many RFs and AFs are enriched in bioactive vs inactive compounds from ChEMBL. We then analyze the generated database GDB-13s, which enumerates 99 million possible molecules of up to 13 atoms, for RFs and AFs resembling ChEMBL bioactive RFs and AFs. This analysis reveals a large number of novel RFs and AFs that are structurally simple, have favorable synthetic accessibility scores, and represent opportunities for synthetic chemistry to contribute to drug innovation in the context of fragment-based drug discovery.
Nicholas Favalli; Sara Puglioli; Sebastian Oehler; Luca Prati; Jörg Scheuermann; Gabriele Bassi; Samuele Cazzamalli; Dario Neri Chem. Sci., 2023, 14(43), 12026-12033 https://doi.org/10.1039/D3SC03688JAbstractDNA-Encoded Chemical Libraries (DELs) are powerful drug discovery tools, enabling the parallel screening of millions of DNA-barcoded compounds. We investigated how the DEL input affects the hit discovery rate in DEL screenings. Evaluation of selection fingerprints revealed that the use of approximately 105 copies of each library member is required for the confident identification of nanomolar hits, using generally applicable methodologies.
Jan Legaard Andersson; Jesper Christensen; Daniela Kleine-Kohlbrecher; Itys Vacher Comet; Jonatan Fullerton Støier; Yasuko Antoku; Visnja Poljak; Loris Moretti; Johannes Dolberg; Tomas Jacso; Søren Jensby Nielsen; Mads Nørregård Madsen; Thomas Franch; Kristian Helin; Paul Andreas Compare Cloos ChemBioChem, 2023, e202300515 https://doi.org/10.1002/cbic.202300515AbstractNSD2 is a histone methyltransferase predominantly catalyzing di-methylation of histone H3 on lysine K36. Increased NSD2 activity due to mutations or fusion-events affecting the gene encoding NSD2 is considered an oncogenic event and a driver in various cancers, including multiple myelomas carrying t(4;14) chromosomal translocations and acute lymphoblastic leukemia's expressing the hyperactive NSD2 mutant E1099K. Using DNA-encoded libraries, we have identified small molecule ligands that selectively and potently bind to the PWWP1 domain of NSD2, inhibit NSD2 binding to H3K36me2-bearing nucleosomes, but do not inhibit the methyltransferase activity. The ligands were subsequently converted to selective VHL1-recruiting NSD2 degraders and by using one of the most efficacious degraders in cell lines, we show that its leads to NSD2 degradation, decrease in K3K36me2 levels and inhibition of cell proliferation.
Zhaobing Ding; Yizhou Wu; Feifei Li; Siyu Jia; Bing Qi; Zuozhong Peng Bioconjugate Chem., 2023, 34(9), 1523-1527 https://doi.org/10.1021/acs.bioconjchem.3c00291AbstractHere, we report a DNA-compatible reaction for the generation of cyclopropane derivatives using thiolides with α,β-unsaturated ketones in the absence of transition metal and N2 protection, which is convenient for DNA encoded library (DEL) construction. This approach allows the rapid and efficient production of a series of DEL libraries of potentially biologically active cyclopropanes and spirocyclopropyl oxindole derivatives.
Zhuxin She; Yinuo Yao; Conglong Wang; Yi Li; Xiaohui Xiong; Yuanyuan Liu J. Chem. Res., 2023 https://doi.org/10.1177/17475198231184799AbstractThe COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 is a global health emergency. The main protease is an important drug target in coronaviruses. It plays an important role in the processing of viral RNA-translated polyproteins and is highly conserved in the amino acid sequence and three-dimensional structure, making it a good drug target for which several small molecule inhibitors are available. This paper describes the various anti-severe acute respiratory syndrome coronavirus 2 inhibitor drugs targeting Mpro discovered since the severe acute respiratory syndrome coronavirus 2 outbreak at the end of 2019, with all these compounds inhibiting severe acute respiratory syndrome coronavirus 2 Mpro activity in vitro. This provides a reference for the development of severe acute respiratory syndrome coronavirus 2 Mpro-targeted inhibitors and the design of therapeutic approaches to address newly emerged severe acute respiratory syndrome coronavirus 2 mutant strains with immune evasion capabilities.
Devan J. Shell; Caroline A. Foley; Qinhong Wang; Chelsea M. Smith; Shiva K. R. Guduru; Hong Zeng; Aiping Dong; Jacqueline L. Norris-Drouin; Matthew Axtman; P. Brian Hardy; Gaorav Gupta; Levon Halabelian; Stephen V. Frye; Lindsey I. James; Kenneth H. Pearce J. Med. Chem., 2023, 66(20), 14133-14149 https://doi.org/10.1021/acs.jmedchem.3c01192AbstractMethyl-lysine reader p53 binding protein 1 (53BP1) is a central mediator of DNA break repair and is associated with various human diseases, including cancer. Thus, high-quality 53BP1 chemical probes can aid in further understanding the role of 53BP1 in genome repair pathways. Herein, we utilized focused DNA-encoded library screening to identify the novel hit compound UNC8531, which binds the 53BP1 tandem Tudor domain (TTD) with an IC50 of 0.47 ± 0.09 μM in a TR-FRET assay and Kd values of 0.85 ± 0.17 and 0.79 ± 0.52 μM in ITC and SPR, respectively. UNC8531 was cocrystallized with the 53BP1 TTD to guide further optimization efforts, leading to UNC9512. NanoBRET and 53BP1-dependent foci formation experiments confirmed cellular target engagement. These results show that UNC9512 is a best-in-class small molecule 53BP1 antagonist that can aid further studies investigating the role of 53BP1 in DNA repair, gene editing, and oncogenesis.
Xin Li; Yongcheng Song E. J. Med. Chem., 2023, 260, 115772 https://doi.org/10.1016/j.ejmech.2023.115772AbstractSevere acute respiratory syndrome-associated coronavirus (SARS-CoV) identified in 2003 infected ∼8000 people in 26 countries with 800 deaths, which was soon contained and eradicated by syndromic surveillance and enhanced quarantine. A closely related coronavirus SARS-CoV-2, the causative agent of COVID-19 identified in 2019, has been dramatically more contagious and catastrophic. It has infected and caused various flu-like symptoms of billions of people in >200 countries, including >6 million people died of or with the virus. Despite the availability of several vaccines and antiviral drugs against SARS-CoV-2, finding new therapeutics is needed because of viral evolution and a possible emerging coronavirus in the future. The main protease (Mpro) of these coronaviruses plays important roles in their life cycle and is essential for the viral replication. This article represents a comprehensive review of the function, structure and inhibition of SARS-CoV and -CoV-2 Mpro, including structure-activity relationships, protein-inhibitor interactions and clinical trial status.
N/A Nat. Synth., 2023, 2, 1018–1019 https://doi.org/10.1038/s44160-023-00352-0AbstractThe most popular reactions used by medicinal chemists are often incompatible with nanoscale ultrahigh-throughput experimentation (ultraHTE). Now, a set of ultraHTE-amenable reaction conditions is reported for four of the most important transformations in drug discovery, and their generality and scalability tested on a range of complex natural products and drug candidates.
Robin Dinter; Suzanne Willems; Thilo Nissalk; Oguz Hastürk; Andreas Brunschweiger; Norbert Kockmann Front. Chem., 2023, 11, 1244043 https://doi.org/10.3389/fchem.2023.1244043AbstractThe transfer from batch to flow chemistry is often based on commercial microfluidic equipment, such as costly complete reactor systems, which cannot be easily tailored to specific requirements of technologies such as DNA-encoded library technology (DELT), in particular for increasingly important photochemical reactions. Customized photoreactor concepts using rapid prototyping technology offer a modular, flexible, and affordable design that allows for adaptation to various applications. In order to validate the prototype reactors, a photochemical pinacol coupling reaction at 368 nm was conducted to demonstrate the transfer from batch to flow chemistry. The conversion rates were optimized by adapting the design parameters of the microfluidic flow photoreactor module. Subsequently, the photoreactor module has been extended to an application with DNA-tagged substrates by switching to LEDs with a wavelength of 454 nm. The successful recovery of DNA confirmed the feasibility of the modular-designed flow photo reactor. This collaborative approach holds enormous potential to drive the development of DELT and flow equipment design.
Huihong Wang; Xiaohong Fan; Teng Chen; Yangfeng Li; Gong Zhang; Wei Fang; Yizhou Li Org. Chem. Front., 2023, 10, 4105-4110 https://doi.org/10.1039/D3QO00901GAbstractConstruction of DNA-encoded chemical libraries which contain focused pharmacophores is of great value for high-throughput hit discovery. Enaminone serves as a privileged scaffold in a series of bioactive compounds, and additionally as a versatile intermediate due to its reactivity for abundant diversifications. Herein, we presented a DNA-compatible amination reaction of allenic ketones to access enaminones. This synthetic approach generates enaminones in a controllable manner, thus offering a wealth of opportunities for further transformations. Meanwhile, this method exhibits a wide substrate scope with various primary or secondary amines. Further annulation transforms enaminones into unprecedented polyfunctionalized benzenes in a DNA-encoded stepwise fusion fashion, demonstrating the potential of allelic ketone synthons in DNA-encoded library construction.
Xinjian Ji; Alexander L. Nielsen; Christian Heinis Angew. Chem. Int. Ed. Engl., 2023, e202308251 https://doi.org/10.1002/anie.202308251AbstractCyclic peptides are fascinating molecules abundantly found in nature and exploited as molecular format for drug development as well as other applications, ranging from research tools to food additives. Advances in peptide technologies made over many years through improved methods for synthesis and drug development have resulted in a steady stream of new drugs, with an average of around one cyclic peptide drug approved per year. Powerful technologies for screening random peptide libraries, and de novo generating ligands, have enabled the development of cyclic peptide drugs independent of naturally derived molecules and now offer virtually unlimited development opportunities. In this review, we feature therapeutically relevant cyclic peptides derived from nature and discuss the unique properties of cyclic peptides, the enormous technological advances in peptide ligand development in recent years, and current challenges and opportunities for developing cyclic peptides that address unmet medical needs.
Alberto G Borges Felipe; Camila B Miguel; Ana P Lazarin Bernardes; Rodrigo Perissinotto; Carlos A Trindade-da-Silva; Melissa C Martins-de-Abreu; Jamil Miguel-Neto; Wellington F Rodrigues; Carlo J Freire Oliveira Am. J. Transl. Res., 2023, 15(11), 6649–6659 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10703658/AbstractAn evaluation of the inflammatory enzymatic interactions related to pulmonary function can help identify biomarkers for interventions or prophylactic measures to improve patient prognosis. This study aimed to determine the effect of epoxide hydrolase inhibition by GSK2256294 in different pulmonary inflammation models. A secondary search was performed using Medline/PubMed, Web of Science, SciELO, Cochrane Library, Embase, Academic Google, and gray literature by two independent reviewers, who analyzed the methodological quality and consistency of the data. Different variables were compared using a meta-analysis. A total of 86 studies were found, 4 of which were selected from the gray literature. Based on the eligibility criteria, two clinical and one preclinical studies were evaluated. GSK2256294 inhibited the soluble epoxide hydrolase enzyme in both clinical and preclinical models, exhibiting greater effectiveness in clinical studies and contributing to the anti-inflammatory activity mediated by the eicosatrienoic pathway by reducing the levels of dihydroxyeicosatrienoic acids and leukotoxin-diol. Overall, GSK2256294 was identified as a promising drug for controlling the deleterious manifestations of lung inflammation. Further clinical and preclinical studies are required to ensure consistency among the evidence and identify other biological activities mediated by GSK2256294.
Lukas A. Schneider; Basilius Sauter; Koder Dagher; Dennis Gillingham J. Am. Chem. Soc., 2023, 145(38), 20874-20882 https://doi.org/10.1021/jacs.3c05961AbstractTerminal deoxynucleotidyl transferase (TdT) is an unusual DNA polymerase that adds untemplated dNTPs to 3′-ends of DNA. If a target protein is expressed as a TdT fusion and incubated with a DNA-encoded library (DEL) in the presence of dATP, the binders of the target induce proximity between TdT and the DNA, promoting the synthesis of a poly-adenine (polyA) tail. The polyA tail length is proportional to the binding affinity, effectively serving as a stable molecular record of binding events. The polyA tail is also a convenient handle to enrich binders with magnetic poly(dT)25 beads before sequencing. In a benchmarking system, we show that ligands spanning nanomolar to double-digit micromolar binding can be cleanly identified by TdT extension, whereas only the tightest binding ligands are identified by classical affinity selection. The method is simple to implement and can function on any DEL that bears a free 3′-end.
Sha Liu; Xiang Li; Xin Li; Xiang David Li Curr. Opin. Chem. Biol., 2023, 75, 102334 https://doi.org/10.1016/j.cbpa.2023.102334AbstractInhibitors for epigenetic readers of histone modifications are useful chemical probes to interrogate the functional roles played by their cognate targets in epigenetic regulation and can even serve as drugs for the treatment of diseases associated with the dysregulated targets. However, many epigenetic readers are intractable to small molecules, as the recognition of modified histone peptides commonly involves flat and extended protein surfaces. In contrast, the relatively large sizes and structural complexity of peptides help them achieve tight and specific binding to the target proteins. Increasing efforts have been made to target epigenetic readers using peptide-based inhibitors that can complement small molecules. In this review, we discuss the recent advances in the development of peptide-based inhibitors of lysine acetylation and methylation readers.
Kangyin Pan; Ying Yao; Yiyuan Zhang; Yuang Gu; Yan Wang; Peixiang Ma; Wei Hou; Guang Yang; Shuning Zhang; Hongtao Xu Bioconjugate Chem., 2023, 34(8), 1459-1466 https://doi.org/10.1021/acs.bioconjchem.3c00235AbstractThe DNA-encoded chemical library (DEL) is a powerful hit selection technique in either basic science or innovative drug discovery. With the aim to circumvent the issue concerning DNA barcode damage in a conventional on-DNA copper-catalyzed azide–alkyne cycloaddition reaction (CuAAC), we have successfully developed the first DNA-compatible enolate–azide [3 + 2] cycloaddition reaction. The merits of this DEL chemistry include metal-free reaction and high DNA fidelity, high conversions and easy operation, broad substrate scope, and ready access to the highly substituted 1,4,5-trisubstituted triazoles. Thus, it will not only further enrich the DEL chemistry toolbox but also will have great potential in practical DEL synthesis.
Mengnisa Seydimemet; Yixuan Yang; Yuhan Lv; Jiaxiang Liu; Ziqin Yan; Yujun Zhao; Xuan Wang; Xiaojie Lu ACS Med. Chem. Lett., 2023, 14(8), 1073-1078 https://doi.org/10.1021/acsmedchemlett.3c00205AbstractPyrimidine is a ubiquitous component in natural products and approved drugs, providing an ideal modular scaffold for generating libraries with drug-like properties. DNA-encoded library technology introduces a novel library modality where each small molecule is covalently linked to a unique oligo tag. This technology offers the advantages of rapidly generating and interrogating large-scale libraries containing billions of members, substantially reducing the entry barrier to their use in both academia and the pharmaceutical industry. In this Letter, we describe the synthesis of three DNA-encoded libraries based on different functionalized pyrimidine cores featuring diversified chemoselectivity and regioselectivity. Preliminary screening of these DNA-encoded libraries against BRD4 identified compounds with nanomolar inhibition activities.
Lijun Xue; Hangke Ma; Weijie Liu; Nicolas Duchemin; Fanming Zeng; Jie Zhang; Huanqing Zhang; Kexin Yang; Yunjin Hu ACS Omega, 2023, 8(26), 24072-24077 https://doi.org/10.1021/acsomega.3c03160AbstractA novel on-DNA oxidative disulfide formation method has been developed. Under ambient conditions, the methodology showcased wide applicability and swift implementation in routine DNA-encoded library synthesis to access pharmaceutically relevant motifs.
Phil Addis; Utsav Bali; Frank Baron; Adrian Campbell; Steven Harborne; Liz Jagger; Gavin Milne; Martin Pearce; Elizabeth M Rosethorne; Rupert Satchell; Denise Swift; Barbara Young; John F Unitt SLAS Discov., 2024, 29(1), 1-22 https://doi.org/10.1016/j.slasd.2023.08.007AbstractG-protein-coupled receptors (GPCRs) are the largest and most versatile cell surface receptor family with a broad repertoire of ligands and functions. We've learned an enormous amount about discovering drugs of this receptor class since the first GPCR was cloned and expressed in 1986, such that it's now well-recognized that GPCRs are the most successful target class for approved drugs. Here we take the reader through a GPCR drug discovery journey from target to the clinic, highlighting the key learnings, best practices, challenges, trends and insights on discovering drugs that ultimately modulate GPCR function therapeutically in patients. The future of GPCR drug discovery is inspiring, with more desirable drug mechanisms and new technologies enabling the delivery of better and more successful drugs.
Lijun Xue; Weijie Liu; Shu Li; Nicolas Duchemin; Mengjia Lou; Jingyu Yuan; Huanqing Zhang; Junyun Chen; Weina Yu; Kexin Yang; Yunjin Hu Bioconjugate Chem., 2023, 34(8), 1366-1373 https://doi.org/10.1021/acs.bioconjchem.3c00138AbstractWe herein present the first application of the on-DNA Morita-Baylis-Hillman (MBH) reaction for the creation of pharmaceutically relevant targeted covalent inhibitors (TCIs) with an α-hydroxyl Michael acceptor motif. Adapting a DNA-compatible organocatalytic process, this MBH reaction for covalent selection-capable DNA encoded library (DEL) synthesis grants access to densely functionalized and versatile precursors to explore novel chemical space for molecule recognition in drug discovery. Most importantly, this methodology sheds light on potentially unexpected reaction outcomes of the MBH reaction.
Ravikumar Jimmidi; Srinivas Chamakuri; Shuo Lu; Melek Nihan Ucisik; Peng-Jen Chen; Kurt M. Bohren; Seyed Arad Moghadasi; Leroy Versteeg; Christina Nnabuife; Jian-Yuan Li; Xuan Qin; Ying-Chu Chen; John C. Faver; Pranavanand Nyshadham; Kiran L. Sharma; Banumathi Sankaran; Allison Judge; Zhifeng Yu; Feng Li; Jeroen Pollet; Reuben S. Harris; Martin M. Matzuk; Timothy Palzkill; Damian W. Young Commun. Chem., 2023, 6, 164 https://doi.org/10.1038/s42004-023-00961-yAbstractThe development of SARS-CoV-2 main protease (Mpro) inhibitors for the treatment of COVID-19 has mostly benefitted from X-ray structures and preexisting knowledge of inhibitors; however, an efficient method to generate Mpro inhibitors, which circumvents such information would be advantageous. As an alternative approach, we show here that DNA-encoded chemistry technology (DEC-Tec) can be used to discover inhibitors of Mpro. An affinity selection of a 4-billion-membered DNA-encoded chemical library (DECL) using Mpro as bait produces novel non-covalent and non-peptide-based small molecule inhibitors of Mpro with low nanomolar Ki values. Furthermore, these compounds demonstrate efficacy against mutant forms of Mpro that have shown resistance to the standard-of-care drug nirmatrelvir. Overall, this work demonstrates that DEC-Tec can efficiently generate novel and potent inhibitors without preliminary chemical or structural information.
Dean G. Brown J. Med. Chem., 2023, 66(11), 7101-7139 https://doi.org/10.1021/acs.jmedchem.3c00521AbstractAn analysis of 156 published clinical candidates from the Journal of Medicinal Chemistry between 2018 and 2021 was conducted to identify lead generation strategies most frequently employed leading to drug candidates. As in a previous publication, the most frequent lead generation strategies resulting in clinical candidates were from known compounds (59%) followed by random screening approaches (21%). The remainder of the approaches included directed screening, fragment screening, DNA-encoded library screening (DEL), and virtual screening. An analysis of similarity was also conducted based on Tanimoto-MCS and revealed most clinical candidates were distant from their original hits; however, most shared a key pharmacophore that translated from hit-to-clinical candidate. An examination of frequency of oxygen, nitrogen, fluorine, chlorine, and sulfur incorporation in clinical candidates was also conducted. The three most similar and least similar hit-to-clinical pairs from random screening were examined to provide perspective on changes that occur that lead to successful clinical candidates.
Alice Lanne; Laura E. J. Usselmann; Poppy Llowarch; Lacovos N. Michaelides; Martin Fillmore; Geoffrey A. Holdgate Drug. Discov. Today, 2023, 103670 https://doi.org/10.1016/j.drudis.2023.103670AbstractRecently, there has been a change in the types of drug target entering early drug discovery portfolios. A significant increase in the number of challenging targets or which would have historically been classed as intractable has been observed. Such targets often have shallow or non-existent ligand-binding sites, can have disordered structures or domains or can be involved in protein–protein or protein–DNA interactions. The nature of the screens required to identify useful hits has, by necessity, also changed. The range of drug modalities explored has also increased and the chemistry required to design and optimise these molecules has adapted. In this review, we discuss this changing landscape and provide insights into the future requirements for small-molecule hit and lead generation.
Lewis D. Pennington; Philip N. Collier; Eamon Comer Med. Chem. Res., 2023, 32, 1278-1293 https://doi.org/10.1007/s00044-023-03073-3AbstractDespite 80 years of progress in modern small molecule drug discovery, medicinal chemists still struggle to minimize the number and duration of design cycles required to optimize hit and lead compounds into high-quality chemical probes or safe and efficacious clinical candidates. High-impact design elements are needed to make multiparameter optimization efforts more efficient and effective. The exchange of an aromatic methine group with a nitrogen atom is a minor bioisosteric structure modification that can lead to major improvements in a variety of pharmacological parameters. These improvements can enhance the efficiency of multiparameter optimization efforts and increase the probability of success in small molecule drug discovery. In this review, we will discuss general aspects of this structure modification and then highlight twelve case studies in which this design element played a pivotal role in multiparameter optimization efforts, solving key program issues and leading to key tool compounds, pre-clinical candidates, and clinical candidates.
Xujun Zhang; Odin Zhang; Chao Shen; Wanglin Qu; Shicheng Chen; Hanqun Cao; Yu Kang; Zhe Wang; Ercheng Wang; Jintu Zhang; Yafeng Deng; Furui Liu; Tianyue Wang; Hongyan Du; Langcheng Wang; Peichen Pan; Guangyong Chen; Chang-Yu Hsieh; Tingjun Hou Nat. Comput. Sci., 2023, 3, 789-804 https://doi.org/10.1038/s43588-023-00511-5AbstractLigand docking is one of the core technologies in structure-based virtual screening for drug discovery. However, conventional docking tools and existing deep learning tools may suffer from limited performance in terms of speed, pose quality and binding affinity accuracy. Here we propose KarmaDock, a deep learning approach for ligand docking that integrates the functions of docking acceleration, binding pose generation and correction, and binding strength estimation. The three-stage model consists of the following components: (1) encoders for the protein and ligand to learn the representations of intramolecular interactions; (2) E(n) equivariant graph neural networks with self-attention to update the ligand pose based on both protein–ligand and intramolecular interactions, followed by post-processing to ensure chemically plausible structures; (3) a mixture density network for scoring the binding strength. KarmaDock was validated on four benchmark datasets and tested in a real-world virtual screening project that successfully identified experiment-validated active inhibitors of leukocyte tyrosine kinase (LTK).
Hongxin Zeng; Yanjing Li; Rongfeng Wu; Daqi Liu; Yang Zhang; Shiyang Xu; Dawen Niu Org. Lett., 2023, XXXX, XXX, XXX-XXX https://doi.org/10.1021/acs.orglett.3c00833AbstractHerein, we report a method that enables the synthesis of carbohydrate–DNA conjugates by radical addition. Key to the success is the use of readily available, bench-stable, and unprotected glycosyl sulfinates as precursors to glycosyl radicals. The redox neutral reaction proceeds under mild and simple conditions and tolerates a broad substrate scope. A small library of carbohydrate–DNA conjugates was prepared.
Wen Torng; Ilaria Biancofiore; Sebastian Oehler; Jin Xu; Jessica Xu; Ian Watson; Brenno Masina; Luca Prati; Nicholas Favalli; Gabriele Bassi; Dario Neri; Samuele Cazzamalli; Jianwen A. Feng* ACS Omega, 2023, 8(28), 25090-25100 https://doi.org/10.1021/acsomega.3c01775AbstractDNA-Encoded Chemical Libraries (DELs) have emerged as efficient and cost-effective ligand discovery tools, which enable the generation of protein–ligand interaction data of unprecedented size. In this article, we present an approach that combines DEL screening and instance-level deep learning modeling to identify tumor-targeting ligands against carbonic anhydrase IX (CAIX), a clinically validated marker of hypoxia and clear cell renal cell carcinoma. We present a new ligand identification and hit-to-lead strategy driven by machine learning models trained on DELs, which expand the scope of DEL-derived chemical motifs. CAIX-screening datasets obtained from three different DELs were used to train machine learning models for generating novel hits, dissimilar to elements present in the original DELs. Out of the 152 novel potential hits that were identified with our approach and screened in an in vitro enzymatic inhibition assay, 70% displayed submicromolar activities (IC50 < 1 μM). To generate lead compounds that are functionalized with anticancer payloads, analogues of top hits were prioritized for synthesis based on the predicted CAIX affinity and synthetic feasibility. Three lead candidates showed accumulation on the surface of CAIX-expressing tumor cells in cellular binding assays. The best compound displayed an in vitro KD of 5.7 nM and selectively targeted tumors in mice bearing human renal cell carcinoma lesions. Our results demonstrate the synergy between DEL and machine learning for the identification of novel hits and for the successful translation of lead candidates for in vivo targeting applications.
Lijun Xue; Sufang Zhou; Jing Wu; Nicolas Duchemin; Bingxin Chen; Jie Zhang; Huanqing Zhang; Kexin Yang; Yuniin Hu Chembiochem, 2023, e202300206 https://doi.org/10.1002/cbic.202300206AbstractHere, we describe a novel method for the on-DNA synthesis of cyclic imides, an important class of molecules that includes several well-known medications. Significantly, the new method enabled on-DNA synthesis under mild conditions with high conversions and a broad functional group tolerance, utilizing ubiquitous bifunctional amines and bis-carboxylic acid, or alkyl halides, and therefore served as the linchpin for DEL synthesis. The mechanism study of off-DNA and on-DNA chemical transformations revealed unique insights in contrast to conventional chemical transformation.
Valerie Cavett; Alix I Chan; Christian N. Cunningham; Brian M. Paegel ACS Cent. Sci., 2023, 9(8), 1603-1610 https://doi.org/10.1021/acscentsci.3c00316AbstractEncoded combinatorial library technologies have dramatically expanded the chemical space for screening but are usually only analyzed by affinity selection binding. It would be highly advantageous to reformat selection outputs to ”one-bead-one-compound” solid-phase libraries, unlocking activity-based and cellular screening capabilities. Here, we describe hydrogel-encapsulated magnetic beads that enable such a transformation. Bulk emulsion polymerization of polyacrylamide hydrogel shells around magnetic microbeads yielded uniform particles (7 ± 2 μm diameter) that are compatible with diverse in-gel functionalization (amine, alkyne, oligonucleotides) and transformations associated with DNA-encoded library synthesis (acylation, enzymatic DNA ligation). In a case study of reformatting mRNA display libraries, transcription from DNA-templated magnetic beads encapsulated in gel particles colocalized both RNA synthesis via hybridization with copolymerized complementary DNA and translation via puromycin labeling. Two control epitope templates (V5, HA) were successfully enriched (50- and 99-fold, respectively) from an NNK5 library bead screen via FACS. Proximity-driven library synthesis in concert with magnetic sample manipulation provides a plausible means for reformatting encoded combinatorial libraries at scale.
Iryna Shcheholeva; Daniel Fernández-Remacha; Roger Estrada-Tejedor; Maria Duca; Veronique Michelet Chem. Eur. J., 2023, e202300825 https://doi.org/10.1002/chem.202300825AbstractTargeting RNA with small molecules is a major challenge of current medicinal chemistry, and the identification and design of original scaffolds able to selectively interact with an RNA target remains difficult. Various approaches have been developed based on classical medicinal chemistry strategies (fragment-based drug design, dynamic combinatorial chemistry, HTS or DNA-encoded libraries) as well as on advanced structural biology and biochemistry methodologies (such as X-ray, cryo-EM, NMR, or SHAPE). Here, we report the de novo design, synthesis, and biological evaluation of RNA ligands using a straightforward and sustainable chemistry combined with molecular docking and biochemical and biophysical studies that allowed us to identify a novel pharmacophore for RNA binding. Specifically, we focused on the targeting of biogenesis of microRNA-21, the re-known oncogene. This led us not only to promising inhibitors but also to a better understanding of the interactions formed between the small molecule compounds and the RNA target paving the way for the rational design of efficient inhibitors with potential anticancer activity.
Qingtong Zhou; Xiao Liu; Dario Neri; Wenxin Li; Nicholas Favalli; Gabriele Bassi; Su Yang; Dehua Yang; Peter K. Vogt; Mingwei Wang Proc. Natl. Acad. Sci. USA, 2023, 120(34), e2304071120 https://www.pnas.org/doi/abs/10.1073/pnas.2304071120AbstractClass IA phosphoinositide 3-kinase alpha (PI3Kα) is an important drug target because it is one of the most frequently mutated proteins in human cancers. However, small molecule inhibitors currently on the market or under development have safety concerns due to a lack of selectivity. Therefore, other chemical scaffolds or unique mechanisms of catalytic kinase inhibition are needed. Here, we report the cryo-electron microscopy structures of wild-type PI3Kα, the dimer of p110α and p85α, in complex with three Y-shaped ligands [cpd16 (compound 16), cpd17 (compound 17), and cpd18 (compound 18)] of different affinities and no inhibitory effect on the kinase activity. Unlike ATP-competitive inhibitors, cpd17 adopts a Y-shaped conformation with one arm inserted into a binding pocket formed by R770 and W780 and the other arm lodged in the ATP-binding pocket at an angle that is different from that of the ATP phosphate tail. Such a special interaction induces a conformation of PI3Kα resembling that of the unliganded protein. These observations were confirmed with two isomers (cpd16 and cpd18). Further analysis of these Y-shaped ligands revealed the structural basis of differential binding affinities caused by stereo- or regiochemical modifications. Our results may offer a different direction toward the design of therapeutic agents against PI3Kα.
Anant R. Kapdi; Rajesh Sahu; Saurav Yadav; Suvadeep Nath; Joydeep Banerjee Chem. Commun., 2023, 59, 6128-6147 https://doi.org/10.1039/D3CC01075AAbstractDrug discovery has undergone a tremendous change in the past decade as the less efficient nature of the high-throughput screening methods that were previously employed have been slowly replaced in the past few years by the DNA-encoded libraries. This review highlights the metal-mediated synthetic pathways that allow late-stage functionalisation of DNA strands to access such DEL libraries. Critical analysis of the literature and the methods employed has been done to allow readers to understand the usefulness as well as the limitations of these protocols.
Zhizheng Wang; Xingxing Shi; Guangyi Huang; Gefei Hao; Guangfu Yang Trends Biochem. Sci., 2023, 48(6), 539-552 https://doi.org/10.1016/j.tibs.2023.01.008AbstractProtein–protein interactions (PPIs) have important roles in various cellular processes, but are commonly described as ‘undruggable’ therapeutic targets due to their large, flat, featureless interfaces. Fragment-based drug discovery (FBDD) has achieved great success in modulating PPIs, with more than ten compounds in clinical trials. Here, we highlight the progress of FBDD in modulating PPIs for therapeutic development. Targeting hot spots that have essential roles in both fragment binding and PPIs provides a shortcut for the development of PPI modulators via FBDD. We highlight successful cases of cracking the ‘undruggable’ problems of PPIs using fragment-based approaches. We also introduce new technologies and future trends. Thus, we hope that this review will provide useful guidance for drug discovery targeting PPIs.
Siddhant Rath; Saswati Panda; James C. Sacchettini; Steven J. Berthel ACS Pharmacol. Transl. Sci., 2023, 6(7), 1043-1051 https://doi.org/10.1021/acsptsci.3c00034AbstractPrimitive data organization practices struggle to deliver at the scale and consistency required to meet multidisciplinary collaborations in drug discovery. For effective data sharing and coordination, a unified platform that can collect and analyze scientific information is essential. We present DAIKON, an open-source framework that integrates targets, screens, hits, and manages projects within a target-based drug discovery portfolio. Its knowledge capture components enable teams to record subsequent molecules as their properties improve, facilitate team collaboration through discussion threads, and include modules that visually illustrate the progress of each target as it advances through the pipeline. It serves as a repository for scientists sourcing data from Mycobrowser, UniProt, PDB. The goal is to globalize several variations of the drug-discovery program without compromising local aspects of specific workflows. DAIKON is modularized by abstracting the database and creating separate layers for entities, business logic, infrastructure, APIs, and frontend, with each tier allowing for extensions. Using Docker, the framework is packaged into two solutions: daikon-server-core and daikon-client. Organizations may deploy the project to on-premises servers or VPC. Active-Directory/SSO is supported for user administration. End users can access the application with a web browser. Currently, DAIKON is implemented in the TB Drug Accelerator program (TBDA).
Xing Ling; Sixiu Liu; Yixuan Yang; Qian Dong; Lisa A. Marcaurelle; Wei Huang; Yun Ding; Xuan Wang; Xiaojie Lu Bioconjugate Chem., 2023, 34(4), 616-620 https://doi.org/10.1021/acs.bioconjchem.3c00068AbstractCarbohydrates are an important class of naturally active products and play vital roles in regulating various physiological activities. To meet the demand for carbohydrate-based libraries used for the identification of potential drug candidates for pharmaceutical-related targets, we developed a set of on-DNA protocols to construct the DNA-encoded glycoconjugates, including Seyferth-Gilbert homologation, anomeric azidation, and CuAAC cyclization. These on-DNA chemistries enable the generation and modification of DNA-linked glycosyl compounds with good conversions and broad substrate scope. Finally, three DNA-linked glycoconjugate libraries were successfully generated to demonstrate their applicability and feasibility in library preparation.
Mike Bührmann; Shivakrishna Kallepu; Jonas D. Warmuth; Jan N. Wiese; Christiane Ehrt; Helge Vatheuer; Wolf Hiller; Carina Seitz; Laura Levy; Paul Czodrowski; Sonja Sievers; Matthias P. Müller; Daniel Rauh J. Med. Chem., 2023, 66(9), 6297-6314 https://doi.org/10.1021/acs.jmedchem.3c00187AbstractFragment-based drug discovery has played an important role in medicinal chemistry and pharmaceutical research. Despite numerous demonstrated successes, the limited diversity and overrepresentation of planar, sp2-rich structures in commercial libraries often hamper the full potential of this approach. Hence, the thorough design of screening libraries inevitably determines the probability for meaningful hits and subsequent structural elaboration. Against this background, we present the generation of an exclusive fragment library based on iterative entry nomination by a specifically designed computational workflow: “Fragtory”. Following a pharmacophore diversity-driven approach, we used Fragtory in an interdisciplinary academic setting to guide both tailored synthesis efforts and the implementation of in-house compounds to build a curated 288-member library of sp3-enriched fragments. Subsequent NMR screens against a model protein and hit validation by protein crystallography led to the identification of structurally novel ligands that were further characterized by isothermal titration calorimetry, demonstrating the applicability of our experimental approach.
Zhi Zhou; Yuang Gu; Liexin Wu; Yan Wang; Huiying Xu; Lei Ma; Zhaoyong Zhang; Jincun Zhao; Wei Zhang; Wei Peng; Guang Yang; Xiyong Yu; Hongtao Xu; Wei Yi SSRN, 2023 http://dx.doi.org/10.2139/ssrn.4342992AbstractOrgano-Se species represent a type of increasingly important core motifs widely found in endogenous proteins and bioactive small molecules. Consequently, exploring their structural diversity to entrust the powerful potential both in synthesis and in application has emerged as an important endeavor in recent basic research, but it remains extremely limited and highly challenging. Here we realize either racemic or asymmetric synthesis towards a novel class of organo-Se species, namely benzothiaselenazole-1-oxides (BTSAs), and reveal its diversified applications mainly via fascinating click chemistry transformation. For synthesis, we developed the first sulfoximine pharmacophore-directed and rhodium(III)-catalyzed C-H/N-H annulative selenylation directly with simple elemental Se. For application, beyond those chemodivergent derivatizations derived by classical Se-mediated reactions, we found that BTSAs can be recognized as a new kind of click selenylation reagents, which not only enabled on-plate modular construction of parallel (1066 examples) and DNA-encoded (261 examples) brand-new indole-C3-selenide libraries, but also provided the site-specific modification of sulfhydryl-containing chemicals in applying as anti-COVID-19 drug discovery and bioorthogonal labeling-based HER2 fluorescence imaging analysis. The systemic work greatly expands the space of organo-Se chemistry and, given the unique performance of BTSAs in click selenylation, it should find robust application in organic synthesis, medicinal chemistry and chemical biology.
Panpan Zhang; Grant Koch; Yankun Zhang; Kevin Yang; R. Scott Lokey ACS Omega, 2023, 8(26), 23477-23483 https://doi.org/10.1021/acsomega.3c00576AbstractDNA-encoded libraries (DELs) are a powerful platform in drug discovery. Peptides have unique properties that make them attractive pharmaceutical candidates. N-methylation of the peptide backbone can confer beneficial properties such as increased proteolytic stability and membrane permeability. Herein, we evaluate different DEL reaction systems and report a DNA-compatible protocol for forming N-methylated amide bonds. The DNA-compatible, bis(trichloromethyl)carbonate-mediated amide coupling is efficient for the formation of N-methyl peptide bonds, which promises to increase the opportunity to identify passively cell-permeable macrocyclic peptide hits by DNA-encoded technology.
Wen Torng; Ilaria Biancofiore; Sebastian Oehler; Jin Xu; Jessica Xu; Ian Watson; Brenno Masina; Luca Prati; Nicholas Favalli; Gabriele Bassi; Dario Neri; Samuele Cazzamalli; Jianwen A. Feng bioRxiv, 2023 https://doi.org/10.1101/2023.01.25.525453AbstractDNA-Encoded Chemical Libraries (DELs) emerged as efficient and cost-effective ligand discovery tools, which enable the generation of protein-ligand interaction data of unprecedented size. In this article, we present an approach that combines DEL screening and instance-level deep learning modeling to identify tumor-targeting ligands against Carbonic Anhydrase IX (CAIX), a clinically validated marker of hypoxia and clear cell Renal Cell Carcinoma. We present a new ligand identification and HIT-to-LEAD strategy driven by Machine Learning (ML) models trained on DELs, which expand the scope of DEL-derived chemical motifs. CAIX screening datasets obtained from three different DELs were used to train ML models for generating novel HITs, dissimilar to elements present in the original DELs. Out of the 152 novel potential HITs that were identified with our approach and screened in an in vitro enzymatic inhibition assay, 70% displayed submicromolar activities (IC50 < 1 μM). Based on the first HIT set, the model was further used to prioritize and generate LEAD compounds with nanomolar affinity for in vivo tumor-targeting applications. Three LEAD candidates showed accumulation on the surface of CAIX-expressing tumor cells in cellular binding assays. The best compound displayed in vitro KD of 5.7 nM and selectively targeted tumors in mice bearing human Renal Cell Carcinoma lesions. Our results demonstrate the synergy between DEL and machine learning for the identification of novel HITs and for the successful translation of LEAD candidates for in vivo targeting applications.
Juan Hu; Alix I. Chan; Emel Adaligil; Ivy Kekessie; Mifune Takahashi; Aimin Song; Christian N. Cunningham; Brian M. Paegel J. Med. Chem., 2023, 66(9), 6288-6296 https://doi.org/10.1021/acs.jmedchem.3c00138AbstractCombinatorial library screening increasingly explores chemical space beyond the Ro5 (bRo5), which is useful for investigating ”undruggable” targets but suffers compromised cellular permeability and therefore bioavailability. Moreover, structure–permeation relationships for bRo5 molecules are unclear partially because high-throughput permeation measurement technology for encoded combinatorial libraries is still nascent. Here, we present a permeation assay that is scalable to combinatorial library screening. A liposomal fluorogenic azide probe transduces permeation of alkyne-labeled molecules into small unilamellar vesicles via copper-catalyzed azide–alkyne cycloaddition. Control alkynes (e.g., propargylamine, various alkyne-labeled PEGs) benchmarked the assay. Cell-permeable macrocyclic peptides, exemplary bRo5 molecules, were alkyne labeled and shown to retain permeability. The assay was miniaturized to microfluidic droplets with high assay quality (Z′ ≥ 0.5), demonstrating excellent discrimination of photocleaved known membrane-permeable and -impermeable model library beads. Droplet-scale permeation screening will enable pharmacokinetic mapping of bRo5 libraries to build predictive models.
Kelly A. Teske; Wenji Su; Cesear R. Corona; Jing Wen; Jason Deng; Yan Ping; Zaihong Zhang; Qi Zhang; Jennifer Wilkinson; Michael T. Beck; Kendra R. Nealey; James D. Vasta; Mei Cong; Poncho L. Meisenheimer; Letian Kuai; Matthew B. Robers Cell Chem. Biolo., 2023, 30(8), 987-998 https://doi.org/10.1016/j.chembiol.2023.06.019AbstractDNA-encoded libraries (DELs) provide unmatched chemical diversity and starting points for novel drug modalities. Here, we describe a workflow that exploits the bifunctional attributes of DEL ligands as a platform to generate BRET probes for live cell target engagement studies. To establish proof of concept, we performed a DEL screen using aurora kinase A and successfully converted aurora DEL ligands as cell-active BRET probes. Aurora BRET probes enabled the validation and stratification of the chemical series identified from primary selection data. Furthermore, we have evaluated the effective repurposing of pre-existing DEL screen data to find suitable leads for BRET probe development. Our findings support the use of DEL workflows as an engine to create cell-active BRET probes independent of structure or compound SAR. The combination of DEL and BRET technology accelerates hit-to-lead studies in a live cell setting.
Yi Wang; Kaili Zhang; Yanjie Zhao; Yifan Li; Weijun Su; Shuai Li ACS Synth. Biol., 2023, 12(7), 1874-1888 https://doi.org/10.1021/acssynbio.3c00043AbstractDNA-encoded peptide/protein libraries are the starting point for protein evolutionary modification and functional peptide/antibody selection. Different display technologies, protein directed evolution, and deep mutational scanning (DMS) experiments employ DNA-encoded libraries to provide sequence variations for downstream affinity- or function-based selections. Mammalian cells promise the inherent post-translational modification and near-to-natural conformation of exogenously expressed mammalian proteins and thus are the best platform for studying transmembrane proteins or human disease-related proteins. However, due to the current technical bottlenecks of constructing mammalian cell-based large size DNA-encoded libraries, the advantages of mammalian cells as screening platforms have not been fully exploited. In this review, we summarize the current efforts in constructing DNA-encoded libraries in mammalian cells and the existing applications of these libraries in different fields.
Esra Balıkçı; Anne-Sophie M. C. Marques; Jesper S. Hansen; Kilian V. M. Huber Expert Opin. Drug Discov., 2023, 18(5), 505-513 https://doi.org/10.1080/17460441.2023.2199979AbstractABSTRACT Introduction The rational development of new therapeutics requires a thorough understanding of how aberrant signalling affects cellular homeostasis and causes human disease. Chemical probes are tool compounds with well-defined mechanism-of-action enabling modulation of, for example, domain-specific protein properties in a temporal manner, thereby complementing other target validation methods such as genetic gain- and loss-of-function approaches. Areas covered In this review, the authors summarize recent advances in chemical probe development for emerging target classes such as solute carriers and ubiquitin-related targets and highlight open resources to inform and facilitate chemical probe discovery as well as tool compound selection for target validation and phenotypic screening. Expert opinion Chemical probes are powerful tools for drug discovery that have led to fundamental insights into biological processes and have paved the way for the development of first-in-class drugs. Open resources can inform on various aspects of chemical probe development and provide access to data and recommendations on use of chemical probes to catalyse collaborative science and help accelerate drug target identification and validation.
Melissa J. Buskes; Aaron Coffin; Dawn M. Troast; Rachel Stein; Maria-Jesus Blanco ACS Med. Chem. Lett., 2023, 14(4), 376-385 https://doi.org/10.1021/acsmedchemlett.3c00012AbstractThe generation of multiple bonds in one reaction step has attracted massive interest in drug discovery and development. Multicomponent reactions (MCRs) offer the advantage of combining three or more reagents in a one-pot fashion to effectively yield a synthetic product. This approach significantly accelerates the synthesis of relevant compounds for biological testing. However, there is a perception that this methodology will only produce simple chemical scaffolds with limited use in medicinal chemistry. In this Microperspective, we want to highlight the value of MCRs toward the synthesis of complex molecules characterized by the presence of quaternary and chiral centers. This paper will cover specific examples showing the impact of this technology toward the discovery of clinical compounds and recent breakthroughs to expand the scope of the reactions toward topologically rich molecular chemotypes.
Zhaoping Pan; Yuxi Zhao; Xiaoyun Wang; Xin Xie; Mingxia Liu; Kaiyao Zhang; Lian Wang; Ding Bai; Leonard J. Foster; Rui Shu; Gu He Mol. Biomed., 2023, 4(1),13 https://doi.org/10.1186/s43556-023-00127-1AbstractBromodomain (BD) is an evolutionarily conserved protein module found in 46 different BD-containing proteins (BCPs). BD acts as a specific reader for acetylated lysine residues (KAc) and serves an essential role in transcriptional regulation, chromatin remodeling, DNA damage repair, and cell proliferation. On the other hand, BCPs have been shown to be involved in the pathogenesis of a variety of diseases, including cancers, inflammation, cardiovascular diseases, and viral infections. Over the past decade, researchers have brought new therapeutic strategies to relevant diseases by inhibiting the activity or downregulating the expression of BCPs to interfere with the transcription of pathogenic genes. An increasing number of potent inhibitors and degraders of BCPs have been developed, some of which are already in clinical trials. In this paper, we provide a comprehensive review of recent advances in the study of drugs that inhibit or down-regulate BCPs, focusing on the development history, molecular structure, biological activity, interaction with BCPs and therapeutic potentials of these drugs. In addition, we discuss current challenges, issues to be addressed and future research directions for the development of BCPs inhibitors. Lessons learned from the successful or unsuccessful development experiences of these inhibitors or degraders will facilitate the further development of efficient, selective and less toxic inhibitors of BCPs and eventually achieve drug application in the clinic.
Zaozao Xiao; Yuang Gu; Hewei Dong; Bo Liu; Weiwei Jin; Jie Li; Peixiang Ma; Hongtao Xu; Wei Hou EJMECH Reports, 2023, 9, 100113 https://doi.org/10.1016/j.ejmcr.2023.100113AbstractNatural products play a key role in the history of human drug discovery, and especially for the anticancer agents. Copper(I)-catalyzed alkyne-azide [3+2] cycloaddition (CuAAC) reaction is perhaps the most powerful method for the efficient modification of complex natural products, enabling the direct incorporation of various functional groups accompanied by the formation of the multifunctional 1,2,3-triazole motif, which could not only serve as an basic and hydrophilic connecting group but also as a bioisosteres of 5- or 6-membered heterocycles or an amide group, thus facilitating the improvement of anticancer activities and/or drug-like properties. This contribution extensively summarizes the state-of-the-art application of 1,2,3-triazole in the modification of natural products for anticancer activity. The aim is to gain a deep understanding of the fruitful achievements as well as limitations of CuAAC click chemistry in natural product modification for anticancer activity, and provide perspectives and directions regarding future studies in natural product medicinal chemistry.
Robin Dinter; Suzanne Willems; Mahdi Hachem; Yana Streltsova; Andreas Brunschweiger; Norbert Kockmann React. Chem. Eng., 2023, 8, 1334-1340 https://doi.org/10.1039/D3RE00020FAbstractSynthesis platforms are of particular interest to DNA-encoded library (DEL) technologies to facilitate chemistry development, building block validation, and high-throughput library synthesis. A liquid–liquid two-phase flow reactor was designed that enables parallel conduction of reactions on DNA-coupled substrates. The dispersed phase in capillary slug flow contained the DNA reaction mixture and allowed for spatially separated batch experiments in a microchannel. A coiled flow inverter (CFI) tubular reactor with a 3D-printed internal structure on which a capillary is coiled was used for improved mixing and compact setup. An inert continuous phase was introduced, which generated slug flow and prevented backmixing of the individual reactants. In order to enable parallelized reactions, slugs containing a variety of different carboxylic acids were successfully generated to act as individual reaction compartments representing single batch experiments. As a widely used exemplary DEL reaction, the amide coupling reaction was successfully transferred to the tailored flow reaction system and DNA was recovered.
Alice Shi Ming Li; Serah Kimani; Brian Wilson; Mahmoud Noureldin; Héctor González-Álvarez; Ahmed Mamai; Laurent Hoffer; John P. Guilinger; Ying Zhang; Moritz von Rechenberg; Jeremy S. Disch; Christopher J. Mulhern; Belinda L. Slakman; John W. Cuozzo; Aiping Dong; Gennady Poda; Mohammed Mohammed; Punit Saraon; Manish Mittal; Pratik Modh; Vaibhavi Rathod; Bhashant Patel; Suzanne Ackloo; Vijayaratnam Santhakumar; Magdalena M. Szewczyk; Dalia Barsyte-Lovejoy; Cheryl H. Arrowsmith; Richard Marcellus; Marie-Aude Guié; Anthony D. Keefe; Peter J. Brown; Levon Halabelian; Rima Al-awar; Masoud Vedadi J. Med. Chem., 2023, 66(7), 5041-5060 https://doi.org/10.1021/acs.jmedchem.2c02132AbstractDCAF1 is a substrate receptor of two distinct E3 ligases (CRL4DCAF1 and EDVP), plays a critical physiological role in protein degradation, and is considered a drug target for various cancers. Antagonists of DCAF1 could be used toward the development of therapeutics for cancers and viral treatments. We used the WDR domain of DCAF1 to screen a 114-billion-compound DNA encoded library (DEL) and identified candidate compounds using similarity search and machine learning. This led to the discovery of a compound (Z1391232269) with an SPR KD of 11 μM. Structure-guided hit optimization led to the discovery of OICR-8268 (26e) with an SPR KD of 38 nM and cellular target engagement with EC50 of 10 μM as measured by cellular thermal shift assay (CETSA). OICR-8268 is an excellent tool compound to enable the development of next-generation DCAF1 ligands toward cancer therapeutics, further investigation of DCAF1 functions in cells, and the development of DCAF1-based PROTACs.
Guixian Zhao; Huihong Wang; Jie Luo; Xun He; Feng Xiong; Yangfeng Li; Gong Zhang; Yizhou Li Org. Lett., 2023, 25(4), 665-670 https://doi.org/10.1021/acs.orglett.2c04293AbstractAnnelated benzodiazepines are attractive drug-like scaffolds with a broad spectrum of biological activities. Incorporation of this heterocyclic core into DNA-encoded chemical libraries (DELs) via multicomponent assembly is highly demanded. Herein, we developed a DNA-compatible method to generate the tricyclic benzodiazepine scaffold via catalyst-free three-component condensation using a broad range of aldehyde, o-phenylenediamine, and diketone sources. With either aldehyde or o-phenylenediamine conjugated with DNA tags, functionalized 1,5-benzodiazepine scaffolds were efficiently forged, expanding the chemical space of the diazepine-centered drug-like DEL.
Bo Cai; Amal El Daibani; Yuntian Bai; Tao Che; Casey J. Krusemark JACS Au, 2023, 3(4), 1076-1088 https://doi.org/10.1021/jacsau.2c00674AbstractG protein-coupled receptors (GPCRs) are the largest superfamily of human membrane target proteins for approved drugs. GPCR ligands can have a complex array of pharmacological activities. Among these activities, biased agonists have potential to serve as both chemical probes to understand specific aspects of receptor signaling and therapeutic leads with more specific, desired activity. Challenges exist, however, in the development of new biased activators due, in part, to the low throughput of traditional screening approaches. DNA-encoded chemical libraries (DELs) dramatically improve the throughput of drug discovery by allowing a collective selection, rather than discrete screening, of large compound libraries. The use of DELs has been largely limited to affinity-based selections against purified protein targets, which identify binders only. Herein, we report a split protein complementation approach that allows direct identification of DNA-linked molecules that induce the dimerization of two proteins. We used this selection with a DEL against opioid receptor GPCRs on living cells for the identification of small molecules that possess the specific function of activation of either β-arrestin or G protein signaling pathways. This approach was applied to δ-, μ-, and κ-opioid receptors and enabled the discovery of compound [66,66], a selective, G-protein-biased agonist of the κ-opioid receptor (EC50 = 100 nM, Emax = 82%, Gi bias factor = 6.6). This approach should be generally applicable for the direct selection of chemical inducers of dimerization from DELs and expand the utility of DELs to enrich molecules with a specific and desired biochemical function.
Anjali Dixit; Huda Barhoosh; Brian M. Paegel Acc. Chem. Res., 2023, 56(4), 489-499 https://doi.org/10.1021/acs.accounts.2c00791AbstractThe Human Genome Project ultimately aimed to translate DNA sequence into drugs. With the draft in hand, the Molecular Libraries Program set out to prosecute all genome-encoded proteins for drug discovery with automated high-throughput screening (HTS). This ambitious vision remains unfulfilled, even while innovations in sequencing technology have fully democratized access to genome-scale sequencing. Why? While the central dogma of biology allows us to chart the entirety of cellular metabolism through sequencing, there is no direct coding for chemistry. The rules of base pairing that relate DNA gene to RNA transcript and amino acid sequence do not exist for relating small-molecule structure with macromolecular binding partners and subsequently cellular function. Obtaining such relationships genome-wide is unapproachable via state-of-the-art HTS, akin to attempting genome-wide association studies using turn-of-the-millennium Sanger DNA sequencing. Our laboratory has been engaged in a multipronged technology development campaign to revolutionize molecular screening through miniaturization in pursuit of genome-scale drug discovery capabilities. The compound library was ripe for miniaturization: it clearly needed to become a consumable. We employed DNA-encoded library (DEL) synthesis principles in the development of solid-phase DELs prepared on microscopic beads, each harboring 100 fmol of a single library member and a DNA tag whose sequence describes the structure of the library member. Loading these DEL beads into 100 pL microfluidic droplets followed by online photocleavage, incubation, fluorescence-activated droplet sorting, and DNA sequencing of the sorted DEL beads reveals the chemical structures of bioactive compounds. This scalable library synthesis and screening platform has proven useful in several proof-of-concept projects involving current clinical targets. Moving forward, we face the problem of druggability and proteome-scale assay development. Developing biochemical or cellular assays for all genome-encoded targets is not scalable and likely impossible as most proteins have ill-defined or unknown activity and may not function outside of their native contexts. These are the dark undruggable expanses, and charting them will require advanced synthesis and analytical technologies that can generalize probe discovery, irrespective of mature protein function, to fulfill the Genome Project’s vision of proteome-wide control of cellular pharmacology.
Jeremy W. Mason; Yuen Ting Chow; Liam Hudson; Antonin Tutter; Gregory Michaud; Matthias V. Westphal; Wei Shu; Xiaolei Ma; Zher Yin Tan; Connor W. Coley; Paul A. Clemons; Simone Bonazzi; Frédéric Berst; Karin Briner; Shuang Liu; Frédéric J. Zécri; Stuart L. Schreiber Nat. Chem. Biol., 2023 https://doi.org/10.1038/s41589-023-01458-4AbstractSmall molecules that induce protein–protein associations represent powerful tools to modulate cell circuitry. We sought to develop a platform for the direct discovery of compounds able to induce association of any two preselected proteins, using the E3 ligase von Hippel–Lindau (VHL) and bromodomains as test systems. Leveraging the screening power of DNA-encoded libraries (DELs), we synthesized ~1 million DNA-encoded compounds that possess a VHL-targeting ligand, a variety of connectors and a diversity element generated by split-and-pool combinatorial chemistry. By screening our DEL against bromodomains in the presence and absence of VHL, we could identify VHL-bound molecules that simultaneously bind bromodomains. For highly barcode-enriched library members, ternary complex formation leading to bromodomain degradation was confirmed in cells. Furthermore, a ternary complex crystal structure was obtained for our most enriched library member with BRD4BD1 and a VHL complex. Our work provides a foundation for adapting DEL screening to the discovery of proximity-inducing small molecules.
Zhiwei Kong; Xin Zhang; Feng Zhou; Liu Tang; Yitong Chen; Saijie Li; Xiaokang Zhang; Letian Kuai; Wenji Su; Weiren Cui; Jiaxi Cai; Yuli Wang; Jun Yang; Youliang Peng; Dongli Wang; Junfeng Liu mBio., 2023, 14(2), e02883-22 https://doi.org/10.1128/mbio.02883-22AbstractBlast disease caused by Magnaporthe oryzae threatens rice production worldwide, and chemical control is one of the main methods of its management. The high mutation rate of the M. oryzae genome results in drug resistance, which calls for novel fungicide targets. Fungal proteins that function during the infection process might be potential candidates, and Mps1 (M. oryzae mitogen-activated protein kinase 1) is such a protein that plays a critical role in appressorium penetration of the plant cell wall. Here, we report the structure-aided identification of a small-molecule inhibitor of Mps1. High-throughput screening was performed with Mps1 against a DNA-encoded compound library, and one compound, named A378-0, with the best performance was selected for further verification. A378-0 exhibits a higher binding affinity than the kinase cosubstrate ATP and can inhibit the enzyme activity of Mps1. Cocrystallization of A378-0 with Mps1 revealed that A378-0 binds to the catalytic pocket of Mps1, while the three ring-type substructures of A378-0 constitute a triangle that squeezes into the pocket. In planta assays showed that A378-0 could inhibit both the appressorium penetration and invasive growth but not the appressorium development of M. oryzae, which is consistent with the biological function of Mps1. Furthermore, A378-0 exhibits binding and activity inhibition abilities against Mpk1, the Mps1 ortholog of the soilborne fungal pathogen Fusarium oxysporum. Collectively, these results show that Mps1 as well as its orthologs can be regarded as fungicide targets, and A378-0 might be used as a hit compound for the development of a broad-spectrum fungicide.
Yannic Altrichter; Peter Bou-Dib; Christina Kuznia; Oliver Seitz J. Pept. Sci., 2023, e3477 https://doi.org/10.1002/psc.3477AbstractNucleic acid-templated chemistry opens the intriguing prospect of triggering the synthesis of drugs only in diseased cells. Herein, we explore the feasibility of using RNA-templated chemical reactions for the activation of a known Smac peptidomimetic compound (SMC), which has proapoptotic activity. Two peptide nucleic acid (PNA) conjugates were used to enable conditional activation of a masked SMC by reduction of an azide either by Staudinger reduction or catalytic photoreduction using a ruthenium complex. The latter provided ~135 nM SMC-PNA on as little as 10 nM (0.01 eq.) template. For the evaluation of the templated azido-SMC reduction system in cellulo, a stable HEK 293 cell line was generated, which overexpressed a truncated, non-functional form of the XIAP mRNA target. We furthermore describe the development of electroporation protocols that enable a robust delivery of PNA-conjugates into HEK293 cells. The action of the reactive PNA conjugates was evaluated by viability and flow cytometric apoptosis assays. In addition, electroporated probes were re-isolated and analyzed by UPLC. Unfortunately, the ruthenium-PNA conjugate proved phototoxic, and treatment of cells with PNA-linked reducing agent and the azido-masked SMC conjugate did not result in a greater viability loss than treatment with scrambled sequence controls. Intracellular product formation was not detectable. A control experiment in total cellular RNA isolate indicated that the templated reaction can in principle proceed in a complex system. The results of this first-of-its-kind study reveal the numerous hurdles that must be overcome if RNA molecules are to trigger the synthesis of pro-apoptotic drugs inside cells.
Xin Wen; Xinyuan Wu; Rui Jin; Xiaojie Lu E. J. Med. Chem., 2023, 115079 https://doi.org/10.1016/j.ejmech.2022.115079AbstractIt is well known that heterocyclic compounds play a key role in improving drug activity, target selectivity, physicochemical properties as well as reducing toxicity. In this review, we summarized the representative heterocyclic structures involved in hit compounds which were obtained from DNA-encoded library from 2013 to 2021. In some examples, the state of the art in heterocycle-based DEL synthesis and hit-to-lead optimization are highlighted. We hope that more and more novel heterocycle-based DEL toolboxes and in-depth pharmaceutical research on these lead compounds can be developed to accelerate the discovery of new drugs.
Rui Hou; Chao Xie; Yuhan Gui; Gang Li; Xiaoyu Li ACS Omega, 2023, 8(21), 19057-19071 https://doi.org/10.1021/acsomega.3c02152AbstractDNA-encoded library (DEL) is a powerful ligand discovery technology that has been widely adopted in the pharmaceutical industry. DEL selections are typically performed with a purified protein target immobilized on a matrix or in solution phase. Recently, DELs have also been used to interrogate the targets in complex biological environment, such as membrane proteins on live cells. However, due to the complex landscape of the cell surface, the selection inevitably involves significant non-specific interactions, and the selection data is much noisier than the ones with purified proteins, making reliable hit identification highly challenging. Researchers have developed several approaches to denoise DEL datasets, but it remains unclear whether they are suitable for cell-based DEL selections. Here, we propose a new machine-learning (ML)-based approach to process cell-based DEL selection datasets by using a Maximum A Posteriori (MAP) estimation loss function, a probabilistic framework that can account for and quantify uncertainties of noisy data. We applied the approach to a DEL selection dataset, where a library of 7,721,415 compounds was selected against a purified carbonic anhydrase 2 (CA-2) and a cell line expressing the membrane protein carbonic anhydrase 12 (CA-12). The Extended-Connectivity Fingerprint (ECFP)-based regression model using the MAP loss function was able to identify the true binders and also reliable structure-activity relationship (SAR) from the noisy cell-based selection datasets. In addition, the regularized enrichment metric (known as MAP enrichment) could also be calculated directly without involving the specific machine learning model, effectively suppressing low-confidence outliers and enhancing the signal-to-noise ratio.
Yashoda Krishna Sunkari; Vijay Kumar Siripuram; Marc Flajolet Chem. Eur. J., 2023, e202203037 https://doi.org/10.1002/chem.202203037AbstractGraphical Abstract On-DNA! Aspartic acid-derived Wittig ylides were generated onto DNA and have been validated for Wittig olefination with various aldehydes to develop on-DNA α, β-unsaturated peptidomimetics. These were further converted into heterocyclic core-containing peptidomimetics through diversity-oriented synthesis (DOS). Abstract The DNA-encoded library (DEL) technology represents a revolutionary drug-discovery tool with unprecedented screening power originating from the association of combinatorial chemistry and DNA barcoding. The chemical diversity of DELs and its chemical space will be further expanded as new DNA-compatible reactions are introduced. This work introduces the use of DOS in the context of on-DNA peptidomimetics. Wittig olefination of aspartic acid-derived on-DNA Wittig ylide, combined with a broad substrate scope of aldehydes, led to formation of on-DNA α, β-unsaturated ketones. The synthesis of on-DNA multi-peptidyl-ylides was performed by incorporating sequential amino acids onto a monomeric ylide. Di-, tri- and tetrameric peptidyl-ylides were validated for Wittig olefination and led to on-DNA α, β-unsaturated-based peptidomimetics, an important class of intermediates. One on-DNA aryl Wittig ylide was also developed and applied to Wittig olefination for synthesis of on-DNA chalcone-based molecules. Furthermore, DOS was used successfully with electron-deficient peptidomimetics and led to the development of different heterocyclic cores containing on-DNA peptidomimetics.
Christopher J. Giacoletto; Martin R. Schiller BioEssays, 2023, 2200191 https://doi.org/10.1002/bies.202200191AbstractSince the 16th century, assays and screens have been essential for scientific investigation. However, most methods could be significantly improved, especially in accuracy, scalability, and often lack adequate comparisons to negative controls. There is a lack of consistency in distinguishing assays, in which accuracy is the main goal, from screens, in which scalability is prioritized over accuracy. We dissected and modernized the original definitions of assays and screens based upon recent developments and the conceptual framework of the original definitions. All methods have three components: design/measurement, performance, and interpretation. We propose a model of method development in which reproducible observations become new methods, initially assessed by sensitivity. Further development can proceed along a path to either screens or assays. The screen path focuses on scalability first, but can later prioritize analysis of negatives. Alternatively, the assay path first compares results to negative controls, assessing specificity and accuracy, later adding scalability. Both pathways converge on a high-accuracy and throughput (HAT) assay, like next generation sequencing, which we suggest should be the ultimate goal of all testing methods. Our model will help scientists better select among available methods, as well as improve existing methods, expanding their impact on science.
Aaron A. Thompsona; Michael B. Harbut; Pei-Pei Kung; Nathan K. Karpowich; Jeffrey D. Branson; Joanna C. Grant; Deborah Hagan; Heather A. Pascual; Guoyun Bai; Reza Beheshti Zavareh; Heather R. Coate; Bernard C. Collins; Marjorie Côte; Christine F. Gelin; Kelly L. Damm-Ganamet; Hadi Gholami; Adam R. Huff; Luis Limon; Kevin J. Lumb; Puiying A. Mak; Kohki M. Nakafuku; Edmund V. Price; Amy Y. Shih; Mandana Tootoonchi; Nadeem A. Vellore; Jocelyn Wang; Na Wei; Jeannie Ziff; Scott B. Berger; James P. Edwards; Agnès Gardet; Siquan Sun; Jennifer E. Towne; Jennifer D. Venable; Zhicai Shi; Hariharan Venkatesan; Marie-Laure Rives; Sujata Sharma; Brock T. Shireman; Samantha J. Allen Proc. Natl. Acad. Sci. USA, 2023, 120(18), e2216342120 https://doi.org/10.1073/pnas.2216342120AbstractSignificance The disruption of protein–protein interactions (PPI) represents a major opportunity for therapeutic intervention, as illustrated by many FDA-approved antibodies. However, the discovery of small-molecule PPI inhibitors remains challenging due to the complex nature and large size of most PPI interfaces. We report two distinct small-molecule classes targeting NKG2D (Naturalkiller group 2D), an immune receptor considered to be intractable for small-molecule inhibition. Inhibitor discovery and optimization were facilitated by large chemically diverse libraries combined with a comprehensive biochemical and cell-based assay suite. Both chemical series inhibited ligand-induced signaling by allosterically altering the NKG2D dimer conformation. This indicates that by targeting protein–protein interfaces involving oligomers, it is possible to increase the probability of discovering small-molecule PPI modulators. Abstract NKG2D (natural-killer group 2, member D) is a homodimeric transmembrane receptor that plays an important role in NK, γδ+, and CD8+ T cell-mediated immune responses to environmental stressors such as viral or bacterial infections and oxidative stress. However, aberrant NKG2D signaling has also been associated with chronic inflammatory and autoimmune diseases, and as such NKG2D is thought to be an attractive target for immune intervention. Here, we describe a comprehensive small-molecule hit identification strategy and two distinct series of protein–protein interaction inhibitors of NKG2D. Although the hits are chemically distinct, they share a unique allosteric mechanism of disrupting ligand binding by accessing a cryptic pocket and causing the two monomers of the NKG2D dimer to open apart and twist relative to one another. Leveraging a suite of biochemical and cell-based assays coupled with structure-based drug design, we established tractable structure–activity relationships with one of the chemical series and successfully improved both the potency and physicochemical properties. Together, we demonstrate that it is possible, albeit challenging, to disrupt the interaction between NKG2D and multiple protein ligands with a single molecule through allosteric modulation of the NKG2D receptor dimer/ligand interface.
Yiwei Zhang; Jiabei Guo; Jiongjia Cheng; Zhenghua Zhang; Fenghua Kang; Xiaoxing Wu; Qian Chu J. Med. Chem., 2023, 66(1), 95-106 https://doi.org/10.1021/acs.jmedchem.2c01541AbstractTherapeutic peptides have revolutionized treatment for a number of human diseases. In particular, the past two decades have witnessed rapid progress of stapled helical peptides in drug discovery. Stapled helical peptides are chemically modified and constrained in their bioactive α-helical conformation. Compared to unstabilized linear peptides, stapled helical peptides exhibit superior binding affinity and selectivity, enhanced membrane permeability, and improved metabolic stability, presenting exciting promise for targeting otherwise challenging protein–protein interfaces. In this Perspective, we summarize recent applications of high-throughput screening technologies for identification of potent stapled helical peptides with optimized binding properties. We expect to provide a broad reference to accelerate the development of stapled helical peptides as the next generation of therapeutic peptides for various human diseases.
Jiankun Lyu; John J. Irwin; Brian K. Shoichet Nat. Chem. Biol., 2023, 19, 712-718 https://doi.org/10.1038/s41589-022-01234-wAbstractRecently, ‘tangible’ virtual libraries have made billions of molecules readily available. Prioritizing these molecules for synthesis and testing demands computational approaches, such as docking. Their success may depend on library diversity, their similarity to bio-like molecules and how receptor fit and artifacts change with library size. We compared a library of 3 million ‘in-stock’ molecules with billion-plus tangible libraries. The bias toward bio-like molecules in the tangible library decreases 19,000-fold versus those ‘in-stock’. Similarly, thousands of high-ranking molecules, including experimental actives, from five ultra-large-library docking campaigns are also dissimilar to bio-like molecules. Meanwhile, better-fitting molecules are found as the library grows, with the score improving log-linearly with library size. Finally, as library size increases, so too do rare molecules that rank artifactually well. Although the nature of these artifacts changes from target to target, the expectation of their occurrence does not, and simple strategies can minimize their impact.
Peter Bellotti; Huan-Ming Huang; Teresa Faber; Frank Glorius Chem. Rev., 2023, 123(8), 4237-4352 https://doi.org/10.1021/acs.chemrev.2c00478AbstractThe emergence of modern photocatalysis, characterized by mildness and selectivity, has significantly spurred innovative late-stage C–H functionalization approaches that make use of low energy photons as a controllable energy source. Compared to traditional late-stage functionalization strategies, photocatalysis paves the way toward complementary and/or previously unattainable regio- and chemoselectivities. Merging the compelling benefits of photocatalysis with the late-stage functionalization workflow offers a potentially unmatched arsenal to tackle drug development campaigns and beyond. This Review highlights the photocatalytic late-stage C–H functionalization strategies of small-molecule drugs, agrochemicals, and natural products, classified according to the targeted C–H bond and the newly formed one. Emphasis is devoted to identifying, describing, and comparing the main mechanistic scenarios. The Review draws a critical comparison between established ionic chemistry and photocatalyzed radical-based manifolds. The Review aims to establish the current state-of-the-art and illustrate the key unsolved challenges to be addressed in the future. The authors aim to introduce the general readership to the main approaches toward photocatalytic late-stage C–H functionalization, and specialist practitioners to the critical evaluation of the current methodologies, potential for improvement, and future uncharted directions.
Yingying Wang; Xiaomin Li; Yahui Cai; Xiaoyu Li; Bingbing Shi Chem. J. Chinese Universities, 2023, 44(3), 20220438 https://doi.org/10.7503/cjcu20220438AbstractDNA-encoded library（DEL） has become a nearly indispensable technology platform for the development of new drugs in the pharmaceutical industry. The DEL technology has become matured over the past three decades and the rapid development of DEL-compatible chemistries has tremendously improved the chemical diversity of DELs and promoted its application in drug discovery. DEL technology has been widely adopted by pharmaceutical companies， and numerous clinical candidates have been identified from DEL selection. Usually， the selection of DEL is mainly affinity-based selection on solid-phase. The purified target protein is immobilized on the matrix， and then the binders are separated from non-binders by physical washes. In recent years， a series of in-solution selection methods for DEL have been developed， which has further expanded the target scope suitable for DEL and revealing the potential of DEL as an effective tool to explore fundamental biology. This review mainly introduces the selection methods of DEL in-solution phase and their applications， and finally gives a brief outlook on the application of DEL technology to complex biological targets and functional selection.
Yashoda Krishna Sunkari; Thu-Lan Nguyen; Vijay Kumar Siripuram; Marc Flajolet iScience, 2023, 107573 https://doi.org/10.1016/j.isci.2023.107573AbstractHigh-power screening (HPS) technologies, such as DNA-encoded library (DEL) technology, could exponentially increase the dimensions of the chemical space accessible for drug discovery. The intrinsic fragile nature of DNA is associated with cumbersome limitations and DNA durability (e.g., depurination, loss of phosphate groups, adduct formation) is compromised in numerous organic chemistry conditions that require empirical testing. An atlas of reaction conditions (temperature, pH, solvent/buffer, ligands, oxidizing reagents, catalysts, scavengers in function of time) that have been systematically tested in multiple combinations, indicates precisely limits useful for DEL construction. More importantly, this approach could be used broadly to effectively evaluate DNA-compatibility of any novel on-DNA chemical reaction, and it is compatible with different molecular methodologies. This atlas and the general approach presented, by allowing novel reaction conditions to be performed in presence of DNA, should greatly help expanding the DEL chemical space as well as any field involving DNA durability.
Xiaofang Li; Pingping Fang; Pei Xu Chin. J. Mod. Appl. Pharm., 2023, 40(5), 712-720 https://doi.org/10.13748/j.cnki.issn1007-7693.20220854AbstractWith the increasing demand of discovering new molecules that specifically modulate certain biological processes in pharmaceutical industry and life sciences, a novel small molecule screening technology for efficient and cost-effective discovery of affinity ligand molecules, the DNA encoded compound library(DEL) screening technology has been developed. DEL is a combinatorial chemical library comprising an unprecedented number of either simple chemicals or those with highly sophisticated structures. Each compound in the library is attached to a unique DNA tag in a covalent manner, allowing the compounds to be pooled and screened against a target protein. The binding hits can subsequently be identified based on the DNA codes through high-throughput sequencing. In recent years, the application of DEL technology in the development of clinical drug candidates has been widely reported. This paper reviews and summarizes the process and progress of DEL technology, especially recent advancements in DEL library construction and affinity screening methods, demonstrates the latest affinity ligand molecules screened and developed by DEL technology. The perspectives of the application of DEL in life sciences including biomedicine and other fields are discussed.
Anand Divakaran; Daniel A. Harki; William C. K. Pomerantz Med. Res. Rev., 2023, 43(4), 972-1018 https://doi.org/10.1002/med.21942AbstractEpigenetic mechanisms for controlling gene expression through heritable modifications to DNA, RNA, and proteins, are essential processes in maintaining cellular homeostasis. As a result of their central role in human diseases, the proteins responsible for adding, removing, or recognizing epigenetic modifications have emerged as viable drug targets. In the case of lysine-ε-N-acetylation (Kac), bromodomains serve as recognition modules (“readers”) of this activating epigenetic mark and competition of the bromodomain-Kac interaction with small-molecule inhibitors is an attractive strategy to control aberrant bromodomain-mediated gene expression. The bromodomain and extra-terminal (BET) family proteins contain eight similar bromodomains. These BET bromodomains are among the more commonly studied bromodomain classes with numerous pan-BET inhibitors showing promising anticancer and anti-inflammatory efficacy. However, these results have yet to translate into Food and Drug Administration-approved drugs, in part due to a high degree of on-target toxicities associated with pan-BET inhibition. Improved selectivity within the BET-family has been proposed to alleviate these concerns. In this review, we analyze the reported BET-domain selective inhibitors from a structural perspective. We highlight three essential characteristics of the reported molecules in generating domain selectivity, binding affinity, and mimicking Kac molecular recognition. In several cases, we provide insight into the design of molecules with improved specificity for individual BET-bromodomains. This review provides a perspective on the current state of the field as this exciting class of inhibitors continue to be evaluated in the clinic.
Ying Zhang; Minyu Qi; Dongyao Wang; Yanting Wang; Diya Lyu; Yan Cao J. Shenyang Pharm. Univ., 2023, 40(4), 529-538 https://doi.org/10.14066/j.cnki.cn21-1349/r.2022.0100AbstractObjective: To summarize the strategies of DNA-encoded compound library ( DEL) technology for drug screening． Methods: Ｒelevant domestic and international literatures were searched, and DEL screening strategies were categorized and elaborated according to the different screening strategy. Results: The DEL technology mainly included three parts: library construction，compound screening，and hit decoding and reconstitution. The technology of compounds had been a hot research topic in recent years. In order to expand the target scope of DEL screening strategies application，the initial solid-phase of the DEL screening system was developed to liquid-phase，and the target was gradually extended from purified proteins to cell lysates and even live cells． In recent years，various modern technologies were combined with DEL technology，and the affinity-based binding screening were converted to the biochemical activity screening． Conclusion: DEL technology has a broad prospect，and many research groups and commercial companies are currently conducting further research for DEL screening． It is believed that more breakthroughs in DEL technology and more implementations in drug discovery will be achieved in the future．
Micah Steffek; Elizabeth Helgason; Nataliya Popovych; Lionel Rougé; John M. Bruning; Ke Sherry Li; Daniel J. Burdick; Jianping Cai; Terry Crawford; Jing Xue; Willy Decurtins; Chunlin Fang; Felix Grubers; Michael J. Holliday; Allyson Langley; Ann Petersen; Alexander Lee Satz; Aimin Song; Daniel Stoffler; Quentin Strebel; Jeffrey Y. K. Tom; Nicholas Skelton; Steven T. Staben; Moreno Wichert; Melinda M. Mulvihill; Erin C. Dueber Biochemistry, 2023, 62(3), 633–644 https://doi.org/10.1021/acs.biochem.1c00682AbstractAutophagy-related proteins (Atgs) drive the lysosome-mediated degradation pathway, autophagy, to enable the clearance of dysfunctional cellular components and maintain homeostasis. In humans, this process is driven by the mammalian Atg8 (mAtg8) family of proteins comprising the LC3 and GABARAP subfamilies. The mAtg8 proteins play essential roles in the formation and maturation of autophagosomes and the capture of specific cargo through binding to the conserved LC3-interacting region (LIR) sequence within target proteins. Modulation of interactions of mAtg8 with its target proteins via small-molecule ligands would enable further interrogation of their function. Here we describe unbiased fragment and DNA-encoded library (DEL) screening approaches for discovering LC3 small-molecule ligands. Both strategies resulted in compounds that bind to LC3, with the fragment hits favoring a conserved hydrophobic pocket in mATG8 proteins, as detailed by LC3A-fragment complex crystal structures. Our findings demonstrate that the malleable LIR-binding surface can be readily targeted by fragments; however, rational design of additional interactions to drive increased affinity proved challenging. DEL libraries, which combine small, fragment-like building blocks into larger scaffolds, yielded higher-affinity binders and revealed an unexpected potential for reversible, covalent ligands. Moreover, DEL hits identified possible vectors for synthesizing fluorescent probes or bivalent molecules for engineering autophagic degradation of specific targets.
Peiyuan Zhang; Xiyun Ye; John C.K. Wang; Hannah T. Baddock; Zena Jensvold; Ian Foe; Andrei Loas; Dan L. Eaton; Qi Hao; Aaron H. Nile; Bradley L. Pentelute ChemRxiv, 2023 https://doi.org/10.26434/chemrxiv-2023-2ts17AbstractCovalent peptides have found widespread applications as activity-based probes and as irreversible therapeutic inhibitors. Currently, there is no rapid, label-free, and highly tunable affinity selection method to enrich covalent reactive peptides from synthetic libraries. We address this challenge by developing a reversibly reactive affinity selection platform enabled by tandem high resolution mass spectrometry (MS/MS) to identify covalent peptide binders to native protein targets. It uses mixed disulfides to build reversible peptide-protein conjugates that can enrich crosslinked peptides that after reduction can be sequenced with MS/MS. Using this platform, we achieved maturation of covalent peptide binders against two oncoproteins, human papillomavirus 16 early protein 6 (HPV16 E6) and peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1). The resulting peptides selectively covalently crosslink Cys58 of E6 with 96% yield in 4 hours at 37 °C and Cys113 of Pin1 with >99% yield in 1 hour at room temperature, respectively. This approach enables the identification of highly selective covalent peptide inhibitors for diverse molecular targets, introducing an applicable method to assist pre-clinical therapeutic development pipelines.
Erin C. Day; Supraja S. Chittari; Matthew P. Bogen; Abigail S. Knight ACS Polym. Au, 2023, 3, 6, 406–427 https://doi.org/10.1021/acspolymersau.3c00025AbstractSynthetic polymers are highly customizable with tailored structures and functionality, yet this versatility generates challenges in the design of advanced materials due to the size and complexity of the design space. Thus, exploration and optimization of polymer properties using combinatorial libraries has become increasingly common, which requires careful selection of synthetic strategies, characterization techniques, and rapid processing workflows to obtain fundamental principles from these large data sets. Herein, we provide guidelines for strategic design of macromolecule libraries and workflows to efficiently navigate these high-dimensional design spaces. We describe synthetic methods for multiple library sizes and structures as well as characterization methods to rapidly generate data sets, including tools that can be adapted from biological workflows. We further highlight relevant insights from statistics and machine learning to aid in data featurization, representation, and analysis. This Perspective acts as a “user guide” for researchers interested in leveraging high-throughput screening toward the design of multifunctional polymers and predictive modeling of structure–property relationships in soft materials.
Benson Chen; Mohammad M. Sultan; Theofanis Karaletsos arXiv, 2023, 2310.13769 https://doi.org/10.48550/arXiv.2310.13769AbstractDNA-Encoded Library (DEL) has proven to be a powerful tool that utilizes combinatorially constructed small molecules to facilitate highly-efficient screening assays. These selection experiments, involving multiple stages of washing, elution, and identification of potent binders via unique DNA barcodes, often generate complex data. This complexity can potentially mask the underlying signals, necessitating the application of computational tools such as machine learning to uncover valuable insights. We introduce a compositional deep probabilistic model of DEL data, DEL-Compose, which decomposes molecular representations into their mono-synthon, di-synthon, and tri-synthon building blocks and capitalizes on the inherent hierarchical structure of these molecules by modeling latent reactions between embedded synthons. Additionally, we investigate methods to improve the observation models for DEL count data such as integrating covariate factors to more effectively account for data noise. Across two popular public benchmark datasets (CA-IX and HRP), our model demonstrates strong performance compared to count baselines, enriches the correct pharmacophores, and offers valuable insights via its intrinsic interpretable structure, thereby providing a robust tool for the analysis of DEL data.
Renaud Prudent; Hugues Lemoine; Jarrod Walsh; Didier Roche Drug Discov. Today, 2023, 103760 https://doi.org/10.1016/j.drudis.2023.103760AbstractAffinity selection mass spectrometry (AS-MS) has gained momentum in drug discovery. This review summarizes how this technology has slowly risen as a new paradigm in hit identification and its potential synergy with DNA encoded library technology. It presents an overview of the recent results on challenging targets and perspectives on new areas of research, such as RNA targeting with small molecules. The versatility of the approach is illustrated and strategic drivers discussed in terms of the experience of a small-medium CRO and a big pharma organization. Teaser: This review highlights the interest aroused by AS-MS as a screening technology, assessing the shared experience of a small-medium contract research organization (CRO) and a big pharma organization.
Bo Cai; Amol Mhetre; Casey Krusemark ChemRxiv, 2022 https://doi.org/10.26434/chemrxiv-2022-l097fAbstractHerein, we report a selection approach to enrich ligands from DNA-encoded libraries (DELs) based on proximity to an enzymatic tag on the target protein. This method involves uncaging or installation of a biotin purification tag on the DNA construct either through photodeprotection of a protected biotin group using a light emitting protein tag (Nanoluciferase) or by acylation using an engineered biotin ligase (UltraID). This selection does not require purification of the target protein and results in improved recovery and enrichment of DNA-linked ligands. This approach should serve as a general and convenient tool for molecular discovery with DELs.
Bo Cai; Amol B. Mhetre; Casey J. Krusemark Chem. Sci., 2022, 14(2), 245-250 https://doi.org/10.1039/D2SC05495GAbstractHerein, we report a selection approach to enrich ligands from DNA-encoded libraries (DELs) based on proximity to an enzymatic tag on the target protein. This method involves uncaging or installation of a biotin purification tag on the DNA construct either through photodeprotection of a protected biotin group using a light emitting protein tag (nanoluciferase) or by acylation using an engineered biotin ligase (UltraID). This selection does not require purification of the target protein and results in improved recovery and enrichment of DNA-linked ligands. This approach should serve as a general and convenient tool for molecular discovery with DELs.
Simon L. Rössler; Nathalie M. Grob; Stephen L. Buchwald; Bradley L. Pentelute ChemRxiv, 2022 https://doi.org/10.26434/chemrxiv-2022-7jp5gAbstractInformation storage in DNA forms the fundamental basis of cellular life, but inherent limitations of information stability and density hamper innovative applications such as data storage or drug discovery. Here, we establish abiotic peptides for next-generation information storage and apply them for the encoding of diverse small molecule synthesis. The chemical stability of the peptide-based tag allows the use of palladium-mediated reactions to efficiently synthesize peptide-encoded libraries (PELs) with large chemical diversity and excellent purity. We demonstrate the successful application of PELs in drug discovery by affinity selection yielding small molecules with nanomolar affinity toward their target protein. Collectively, this work establishes abiotic peptides as carriers of information, leveraged herein for the encoding of small molecule synthesis.
Xinyuan Wu; Yujie Chen; Weiwei Lu; Rui Jin; Xiaojie Lu Bioconjugate Chem., 2022, 33(10), 1818–1824 https://doi.org/10.1021/acs.bioconjchem.2c00421AbstractThe DNA-encoded compound library (DEL) technology has accelerated the target hits discovery in new drug development. While affinity-based DEL selection can distinguish high-affinity ligands, moderate-affinity ligands are also potential drug candidates with further modifications. Herein, we designed a photo-cross-linking selection method for DELs with double-stranded DNA (dsDELs) to screen moderate-affinity ligands. We constructed two photo-cross-linking libraries with linkers of different lengths that connect a diazirine group to the DNA encoded compound. The diazirine group can be activated by UV irradiation and thus bond with the target protein in a reachable distance. In the model selection, the feasibility of the photo-cross-linking screening system was verified by qPCR and NGS technology. Both high-affinity and moderate-affinity ligands were successfully selected from the libraries.
Sebastian B. Beil; Tiffany Q. Chen; Nicholas E. Intermaggio; David W. C. MacMillan Acc. Chem. Res., 2022, 55(23), 3481-3494 https://doi.org/10.1021/acs.accounts.2c00607AbstractThe development of palladium-catalyzed cross-coupling methods for the activation of C(sp2)–Br bonds facilitated access to arene-rich molecules, enabling a concomitant increase in the prevalence of this structural motif in drug molecules in recent decades. Today, there is a growing appreciation of the value of incorporating saturated C(sp3)-rich scaffolds into pharmaceutically active molecules as a means to achieve improved solubility and physiological stability, providing the impetus to develop new coupling strategies to access these challenging motifs in the most straightforward way possible. As an alternative to classical two-electron chemistry, redox chemistry can enable access to elusive transformations, most recently, by interfacing abundant first-row transition-metal catalysis with photoredox catalysis. As such, the functionalization of ubiquitous and versatile functional handles such as (aliphatic) carboxylic acids via metallaphotoredox catalysis has emerged as a valuable field of research over the past eight years. In this Account, we will outline recent progress in the development of methodologies that employ aliphatic and (hetero)aromatic carboxylic acids as adaptive functional groups. Whereas recent decarboxylative functionalization methodologies often necessitate preactivated aliphatic carboxylic acids in the form of redox-active esters or as ligands for hypervalent iodine reagents, methods that enable the direct use of the native carboxylic acid functionality are highly desired and have been accomplished through metallaphotoredox protocols. As such, we found that bench-stable aliphatic carboxylic acids can undergo diverse transformations, such as alkylation, arylation, amination, and trifluoromethylation, by leveraging metallaphotoredox catalysis with prevalent first-row transition metals such as nickel and copper. Likewise, abundant aryl carboxylic acids are now able to undergo halogenation and borylation, enabling new entry points for traditional, primarily palladium- or copper-catalyzed cross-coupling strategies. Given the breadth of the functional group tolerance of the employed reaction conditions, the late-stage functionalization of abundant carboxylic acids toward desired targets has become a standard tool in reaction design, enabling the synthesis of various diversified drug molecules. The rapid rise of this field has positively inspired pharmaceutical discovery and will be further accelerated by novel reaction development. The achievement of generality through reaction optimization campaigns allows for future breakthroughs that can render protocols more reliable and applicable for industry. This article is intended to highlight, in particular, (i) the employment of aliphatic and (hetero)aryl carboxylic acids as powerful late-stage adaptive functional handles in drug discovery and (ii) the need for the further development of still-elusive and selective transformations. We strongly believe that access to native functionalities such as carboxylic acids as adaptive handles will further inspire researchers across the world to investigate new methodologies for complex molecular targets.
Jian Yin; Jialin Yu; Weiren Cui; Junkun Lei; Alex L. Satz; Yifan Zhou; Hua Feng; Jason Deng; Wenji Su; Letian Kuai Molecules, 2022, 27(18), 5797 https://doi.org/10.3390/molecules27185797AbstractThe recent success of AlphaFold and RoseTTAFold has demonstrated the values of AI methods in predicting highly accurate protein structures. Despite the advances, their roles in the context of small-molecule drug discovery need to be thoroughly explored. In this study, we evaluated the possibility whether the AI-based models can lead to reliable 3D structures of protein-ligand complexes. With the AI-generated protein structure, we were able to confidently predict the binding modes of small-molecule inhibitors for NLRP3, a challenging protein target in terms of obtaining the 3D model both experimentally and computationally. We therefore concluded that through careful evaluation, AI-predicted protein structures can be trusted and useful for small-molecule drug discovery projects.
Lydia Boike; Nathaniel J. Henning; Daniel K. Nomura Nat. Rev. Drug Discov., 2022, 1-18 https://doi.org/10.1038/s41573-022-00542-zAbstractCovalent drugs have been used to treat diseases for more than a century, but tools that facilitate the rational design of covalent drugs have emerged more recently. The purposeful addition of reactive functional groups to existing ligands can enable potent and selective inhibition of target proteins, as demonstrated by the covalent epidermal growth factor receptor (EGFR) and Bruton’s tyrosine kinase (BTK) inhibitors used to treat various cancers. Moreover, the identification of covalent ligands through ‘electrophile-first’ approaches has also led to the discovery of covalent drugs, such as covalent inhibitors for KRAS(G12C) and SARS-CoV-2 main protease. In particular, the discovery of KRAS(G12C) inhibitors validates the use of covalent screening technologies, which have become more powerful and widespread over the past decade. Chemoproteomics platforms have emerged to complement covalent ligand screening and assist in ligand discovery, selectivity profiling and target identification. This Review showcases covalent drug discovery milestones with emphasis on the lessons learned from these programmes and how an evolving toolbox of covalent drug discovery techniques facilitates success in this field.
Qiuji Ye; Hassane Belabed; Yong Wang; Zhifeng Yu; Murugesan Palaniappan; Jianyuan Li; Stacey A. Kalovidouris; Kevin R. MacKenzie; Mingxing Teng; Damian W. Young; Yoshitaka Fujihara; Martin M. Matzuk Andrology, 2022 https://doi.org/10.1111/andr.13309AbstractBackground: A safe, effective, and reversible nonhormonal male contraceptive drug is greatly needed for male contraception as well as for circumventing the side effects of female hormonal contraceptives. Phosducin-like 2 (PDCL2) is a testis-specific phosphoprotein in mice and humans. We recently found that male PDCL2 knockout mice are sterile due to globozoospermia caused by impaired sperm head formation, indicating that PDCL2 is a potential target for male contraception. Herein, our study for the first time developed a biophysical assay for PDCL2 allowing us to screen a series of small molecules, to study structure-activity relationships (SAR), and to discover two PDCL2 binders with novel chemical structure. Objective: To identify a PDCL2 ligand for therapeutic male contraception, we performed DNA-encoded chemical library (DECL) screening and off-DNA hit validation using a unique affinity selection mass spectrometry (ASMS) biophysical profiling strategy. Materials and methods: We employed the screening process of DECL which contain billions of chemically unique DNA-barcoded compounds generated through individual sequences of reactions and different combinations of functionalized building blocks. The structures of the PDCL2 binders are proposed based on the sequencing analysis of the DNA barcode attached to each individual DECL compound. The proposed structure is synthesized through multi-step reactions. To confirm and determine binding affinity between the DECL identified molecules and PDCL2, we developed an ASMS assay that incorporates liquid chromatography with tandem mass spectrometry (LC-MS/MS). Results: After a screening process of PDCL2 with DECLs containing >440 billion compounds, we identified a series of hits. The selected compounds were synthesized as off-DNA small molecules, characterized by spectroscopy data, and subjected to our ASMS/LC-MS/MS binding assay. By this assay, we discovered two novel compounds that showed good binding affinity for PDCL2, which was further confirmed by thermal shift assay (TSA). Discussion and conclusion and relevance: With the ASMS/LC-MS/MS assay developed in this paper, we successfully discovered a PDCL2 ligand that warrants further development as a male contraceptive. This article is protected by copyright. All rights reserved.
Qing Hu; Yuan Xiong; Guang-Hao Zhu; Ya-Ni Zhang; Yi-Wen Zhang; Ping Huang; Guang-Bo Ge MedComm, 2022, 3(3), e151 https://doi.org/10.1002/mco2.151AbstractThe main proteases (Mpro), also termed 3-chymotrypsin-like proteases (3CLpro), are a class of highly conserved cysteine hydrolases in β-coronaviruses. Increasing evidence has demonstrated that 3CLpros play an indispensable role in viral replication and have been recognized as key targets for preventing and treating coronavirus-caused infectious diseases, including COVID-19. This review is focused on the structural features and biological function of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease Mpro (also known as 3CLpro), as well as recent advances in discovering and developing SARS-CoV-2 3CLpro inhibitors. To better understand the characteristics of SARS-CoV-2 3CLpro inhibitors, the inhibition activities, inhibitory mechanisms, and key structural features of various 3CLpro inhibitors (including marketed drugs, peptidomimetic, and non-peptidomimetic synthetic compounds, as well as natural compounds and their derivatives) are summarized comprehensively. Meanwhile, the challenges in this field are highlighted, while future directions for designing and developing efficacious 3CLpro inhibitors as novel anti-coronavirus therapies are also proposed. Collectively, all information and knowledge presented here are very helpful for understanding the structural features and inhibitory mechanisms of SARS-CoV-2 3CLpro inhibitors, which offers new insights or inspiration to medicinal chemists for designing and developing more efficacious 3CLpro inhibitors as novel anti-coronavirus agents.
Samantha M. Meyer; Toru Tanaka; Patrick R. A. Zanon; Jared T. Baisden; Daniel Abegg; Xueyi Yang; Yoshihiro Akahori; Zainab Alshakarchi; Michael D. Cameron; Alexander Adibekian; Matthew D. Disney J. Am. Chem. Soc., 2022, 144(46), 21096–21102 https://doi.org/10.1021/jacs.2c07217AbstractRibonuclease targeting chimeras (RiboTACs) induce degradation of an RNA target by facilitating an interaction between an RNA and a ribonuclease (RNase). We describe the screening of a DNA-encoded library (DEL) to identify binders of monomeric RNase L to provide a compound that induced dimerization of RNase L, activating its ribonuclease activity. This compound was incorporated into the design of a next-generation RiboTAC that targeted the microRNA-21 (miR-21) precursor and alleviated a miR-21-associated cellular phenotype in triple-negative breast cancer cells. The RNA-binding module in the RiboTAC is Dovitinib, a known receptor tyrosine kinase (RTK) inhibitor, which was previously identified to bind miR-21 as an off-target. Conversion of Dovitinib into this RiboTAC reprograms the known drug to selectively affect the RNA target. This work demonstrates that DEL can be used to identify compounds that bind and recruit proteins with effector functions in heterobifunctional compounds.
Sheila Dakhel; Andrea Galbiati; Francesca Migliorini; Claudia Comacchio; Sebastian Oehler; Luca Prati; Jörg Scheuermann, Samuele Cazzamalli; Dario Neri, Gabriele Bassi; Nicholas Favalli ChemMedChem, 2022, e202200350 https://doi.org/10.1002/cmdc.202200350AbstractNatural Killer Group 2D (NKG2D) is a homo-dimeric transmembrane protein which is typically expressed on the surface of natural killer (NK) cells, natural killer T (NKT) cells, gamma delta T (γδT) cells, activated CD8 positive T-cells and activated macrophages. Bispecific molecules, capable of bridging NKG2D with a target protein expressed on the surface of tumor cells, may be used to redirect the cytotoxic activity of NK-cells towards antigen-positive malignanT-cells. In this work, we report the discovery of a novel NKG2D small molecule binder [K D = (410±60) nM], isolated from a DNA-Encoded Chemical Library (DEL). The discovery of small organic NKG2D ligands may facilitate the generation of fully synthetic bispecific adaptors, which may serve as an alternative to bispecific antibody products and which may benefit from better tumor targeting properties.
Feng Xiong; Mingao Yu; Honggui Xu; Zhenmin Zhong; Zhenwei Li; Yuhan Guo; Tianyuan Zhang; Zhixuan Zeng; Feng Jin; Xun He Front. Chem., 2022, 10, 982539 https://doi.org/10.3389/fchem.2022.982539AbstractDrug discovery has entered a new period of vigorous development with advanced technologies such as DNA-encoded library (DEL) and artificial intelligence (AI). The previous DEL-AI combination has been successfully applied in the drug discovery of classical kinase and receptor targets mainly based on the known scaffold. So far, there is no report of the DEL-AI combination on inhibitors targeting protein-protein interaction, including those undruggable targets with few or unknown active scaffolds. Here, we applied DEL technology on the T cell immunoglobulin and ITIM domain (TIGIT) target, resulting in the unique hit compound 1 (IC50 = 20.7 μM). Based on the screening data from DEL and hit derivatives a1-a34, a machine learning (ML) modeling process was established to address the challenge of poor sample distribution uniformity, which is also frequently encountered in DEL screening on new targets. In the end, the established ML model achieved a satisfactory hit rate of about 75% for derivatives in a high-scored area.
Huiyong Ma; James B. Murray; Huadong Luo; Xuemin Cheng; Qiuxia Chen; Chao Song; Cong Duan; Ping Tan; Lifang Zhang; Jian Liu; Barry A. Morgan; Jin Li; Jinqiao Wan; Lisa M. Baker; William Finnie; Lucie Guetzoyan; Richard Harris; Nicole Hendrickson; Natalia Matassova; Heather Simmonite; Julia Smith; Roderick E. Hubbard; Guansai Liu RSC Med. Chem., 2022, 13, 1341-1349 https://doi.org/10.1039/D2MD00197GAbstractWe describe a novel approach for screening fragments against a protein that combines the sensitivity of DNA-encoded library technology with the ability of fragments to explore what will bind. Each of the members of the library consists of a fragment which is linked to a photoactivatable diazirine moiety. Split and pool synthesis combines each fragment with a set of linkers with the version of the library reported here containing some 70k different compounds, each with an individual DNA code. Incubation of the library with a protein sample is followed by photoactivation, washing and subsequent PCR and sequencing which allows the individual fragment hits to be identified. We illustrate how the approach allows successful hit fragment identification using only microgram quantities of material for two targets. PAK4 is a kinase for which conventional fragment screening has generated many advance leads. The as yet undrugged target, 2-epimerase, presents a more challenging active site for identification of hit compounds. In both cases, PAC-FragmentDEL identified fragments validated as hits by ligand-observed NMR measurements and crystal structure determination of off-DNA sample binding to the proteins.
Bianca Matsuo; Albert Granados; Jadab Majhi; Mohammed Sharique; Guillaume Levitre; Gary A. Molander ACS Org. Inorg. Au, 2022, 2(6), 435-454 https://doi.org/10.1021/acsorginorgau.2c00032AbstractThe exploration of 1,2-radical shift (RS) mechanisms in photoinduced organic reactions has provided efficient routes for the generation of important radical synthons in many chemical transformations. In this Review, the basic concepts involved in the traditional 1,2-spin-center shift (SCS) mechanisms in recently reported studies are discussed. In addition, other useful 1,2-RSs are addressed, such as those proceeding through 1,2-group migrations in carbohydrate chemistry, via 1,2-boron shifts, and by the generation of α-amino radicals. The discussion begins with a general overview of the basic aspects of 1,2-RS mechanisms, followed by a demonstration of their applicability in photoinduced transformations. The sections that follow are organized according to the mechanisms operating in combination with the 1,2-radical migration event. This contribution is not a comprehensive review but rather aims to provide an understanding of the topic, focused on the more recent advances in the field, and establishes a definition for the nomenclature that has been used to describe such mechanisms.
Jessica A. Bush; Samantha M. Meyer; Rita Fuerst; Yuquan Tong; Yue Li; Raphael I. Benhamou; Haruo Aikawa; Patrick R. A. Zanon; Quentin M. R. Gibaut; Alicia J. Angelbello; Tania F. Gendron; Yong-Jie Zhang; Leonard Petrucelli; Torben Heick Jensen; Jessica L. Childs-Disney; Matthew D. Disney Proc. Natl. Acad. Sci. USA, 2022, 119(48), e2210532119 https://doi.org/10.1073/pnas.2210532119AbstractA hexanucleotide repeat expansion in intron 1 of the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia, or c9ALS/FTD. The RNA transcribed from the expansion, r(G4C2)exp, causes various pathologies, including intron retention, aberrant translation that produces toxic dipeptide repeat proteins (DPRs), and sequestration of RNA-binding proteins (RBPs) in RNA foci. Here, we describe a small molecule that potently and selectively interacts with r(G4C2)exp and mitigates disease pathologies in spinal neurons differentiated from c9ALS patient-derived induced pluripotent stem cells (iPSCs) and in two c9ALS/FTD mouse models. These studies reveal a mode of action whereby a small molecule diminishes intron retention caused by the r(G4C2)exp and allows the liberated intron to be eliminated by the nuclear RNA exosome, a multi-subunit degradation complex. Our findings highlight the complexity of mechanisms available to RNA-binding small molecules to alleviate disease pathologies and establishes a pipeline for the design of brain penetrant small molecules targeting RNA with novel modes of action in vivo.
Wei Hou; Hewei Dong; Ying Yao; Kangyin Pan; Guang Yang; Peixiang Ma; Hongtao Xu ChemMedChem, 2022, 17(17), e202200324 https://doi.org/10.1002/cmdc.202200324AbstractSelenium (Se) is an emerging versatile player in medicinal chemistry. The incorporation of Se into small molecules and natural products could have multiple benefits. However, the lack of efficient methods for the synthesis of Se-containing chemical library has greatly hindered the development of seleno-medicinal chemistry. With the aim to address this issue, we proposed the development of “clickable selenylation” reactions, which can be used in the synthesis of Se-containing in situ library and DNA-encoded library (SeDEL), thereby quickly producing ultra-large collections of Se-containing compounds and boosting the development of seleno-medicinal chemistry. This research paradigm can be concluded as “clickable selenylation chemistry development→in situ library construction/SeDEL synthesis→phenotype- or target-based screening→seleno-hit compound”.
Solji Eom; Taeyeon Kwon; Da Yeon Lee; Chi Hoon Park; Hyun Jin Kim Org. Lett., 2022, 24(27), 4881-4885 https://doi.org/10.1021/acs.orglett.2c01675AbstractThe DNA-encoded library (DEL) technology is a new method for discovering hit compounds for target proteins in the pharmaceutical industry. The N-acylsulfonamide functional group has been reported to exhibit various pharmacological activities, and based on this, the demand for a method that allows its introduction into the DEL platform has increased. In this report, a procedure for synthesizing N-acylsulfonamide functional groups applicable to DEL construction was developed in the presence of a copper reagent and water as a nucleophile from simple alkynes or sulfonyl azides, which are widely commercially available. Furthermore, we prove that a new alternative procedure can be used to construct a DNA-encoded library.
Hongtao Xu; Tingting Tan; Yiyuan Zhang; Yan Wang; Kangyin Pan; Ying Yao; Shuning Zhang; Yuang Gu; Wanting Chen; Jie Li; Hewei Dong; Yu Meng; Peixiang Ma; Wei Hou; Guang Yang Adv. Sci., 2022, 2202790 https://doi.org/10.1002/advs.202202790AbstractA successful DNA-encoded library (DEL) will consist of diverse skeletons and cover chemical space as comprehensive as possible to fully realize its potential in drug discovery and chemical biology. However, the lack of versatile on-DNA arylation methods for phenols that are less nucleophilic and reactive poses a great hurdle for DEL to include diaryl ether, a privileged chemotype in pharmaceuticals and natural products. This work describes the use of “substrate activation” approach to address the arylation of DNA-conjugated phenols. Diaryliodonium salt, a highly electrophilic and reactive arylation reagent, is employed as Ar+ sources to ensure highly selective on-DNA arylation of phenols and oximes with both high yields and DNA fidelity. Notably, the new on-DNA arylation reaction can be applied to the late-stage modification of peptides containing tyrosine side-chain and to synthesize DNA-tagged analogues of existing drug molecules such as sorafenib, a known pan-kinase inhibitor. The new on-DNA diaryliodonium salts chemistry affords a greater flexibility in DEL design and synthesis.
Xinrong Tian; Dominic Suarez; Douglas Thomson; William Li; Elizabeth A. King; Louis LaFrance; Jeffrey Boehm; Linda Barton; Christina Di Marco; Cuthbert Martyr; Reema Thalji; Jesus Medina; Steven Knight; Dirk Heerding; Enoch Gao; Eldridge Nartey; Ted Cecconie; Christopher Nixon; Guofeng Zhang; Thomas J. Berrodin; Christopher Phelps; Amish Patel; Xiaopeng Bai; Ken Lind; Ninad Prabhu; Jeffrey Messer; Zhengrong Zhu; Lisa Shewchuk; Rob Reid; Alan P. Graves; Charles McHugh; Biju Mangatt J. Med. Chem., 2022, 65(21), 14391–14408 https://doi.org/10.1021/acs.jmedchem.2c00670AbstractE1A binding protein (p300) and CREB binding protein (CBP) are two highly homologous and multidomain histone acetyltransferases. These two proteins are involved in many cellular processes by acting as coactivators of a large number of transcription factors. Dysregulation of p300/CBP has been found in a variety of cancers and other diseases, and inhibition has been shown to decrease Myc expression. Herein, we report the identification of a series of highly potent, proline-based small-molecule p300/CBP histone acetyltransferase (HAT) inhibitors using DNA-encoded library technology in combination with high-throughput screening. The strategy of reducing ChromlogD and fluorination of metabolic soft spots was explored to improve the pharmacokinetic properties of potent p300 inhibitors. Fluorination of both cyclobutyl and proline rings of 22 led to not only reduced clearance but also improved cMyc cellular potency.
Li-Hao Chang; Oliver Seitz Bioorg. Med. Chem., 2022, 66, 116786 https://doi.org/10.1016/j.bmc.2022.116786AbstractNucleic acid-programmed reactions find application in drug screening and nucleic acid diagnosis, and offer prospects for a RNA-sensitive prodrug approach. We aim for the development of a nucleic acid-templated reaction providing nucleic acid-linked molecules that can act on intracellular protein targets. Such reactions would be useful for in situ drug synthesis and activity-based DNA-encoded library screening. In this report, we show native chemical ligation-like chemical peptidyl transfer reactions between peptide-PNA conjugates. The reaction proceeds on RNA templates. As a chemical alternative to ribosomal peptide synthesis access to both L- and d-peptides is provided. In reactions affording 9 to 14 amino acid long pro-apoptotic L- and d-peptides, we found that certain PNA sequence motifs and combinations of cell penetrating peptides (CPPs) cause surprisingly high reactivity in absence of a template. Viability measurements demonstrate that the products of templated peptidyl transfer act on HeLa cells and HEK293 cells. Of note, the presence of cysteine, which is required for NCL chemistry, can enhance the bioactivity. The study provides guidelines for the application of peptide-PNA conjugates in templated synthesis and is of interest for in situ drug synthesis and activity-based DNA-encoded library screening.
Dávid Bajusz; György Miklós Keserű Expert Opin. Drug Discov., 2022, 1-12 https://doi.org/10.1080/17460441.2022.2085685AbstractExperimental and virtual screening contributes to the discovery of more than 50% of clinical candidates. Considering the similar concept and goals, early phase drug discovery would benefit from the effective integration of these approaches. After reviewing the recent trends in both experimental and virtual screening, the authors discuss different integration strategies from parallel, focused, sequential and iterative screening. Strategic considerations are demonstrated in a number of real-life case studies. Experimental and virtual screening are complementary approaches that should be integrated in lead discovery settings. Virtual screening can access extremely large synthetically feasible chemical space that can be effectively searched on GPU clusters or cloud architectures. Experimental screening provides reliable datasets by quantitative HTS applications, and DNA-encoded libraries (DEL) have enlarged the chemical space covered by these technologies. These developments, together with the use of artificial intelligence methods, represent new options for their efficient integration. The case studies discussed here demonstrate the benefits of complementary strategies such as focused and iterative screening.
Wendy A. Warr; Marc C. Nicklaus; Christos A. Nicolaou; Matthias Rarey J. Chem. Inf. Model., 2022, 62(9), 2021-2034 https://doi.org/10.1021/acs.jcim.2c00224AbstractDesigning new medicines more cheaply and quickly is tightly linked to the quest of exploring chemical space more widely and efficiently. Chemical space is monumentally large, but recent advances in computer software and hardware have enabled researchers to navigate virtual chemical spaces containing billions of chemical structures. This review specifically concerns collections of many millions or even billions of enumerated chemical structures as well as even larger chemical spaces that are not fully enumerated. We present examples of chemical libraries and spaces and the means used to construct them, and we discuss new technologies for searching huge libraries and for searching combinatorially in chemical space. We also cover space navigation techniques and consider new approaches to de novo drug design and the impact of the “autonomous laboratory” on synthesis of designed compounds. Finally, we summarize some other challenges and opportunities for the future.
Hartmut Beck; Michael Härter; Bastian Haß; Carsten Schmeck; Lars Baerfacker Drug Discov. Today, 2022, 27(6), 1560-1574 https://doi.org/10.1016/j.drudis.2022.02.015AbstractThe year 2021 marks the 125th anniversary of the Bayer Chemical Research Laboratory in Wuppertal, Germany. A significant number of prominent small-molecule drugs, from aspirin to Xarelto, have emerged from this research site. In this review, we shed light on historic cornerstones of small-molecule drug research, discussing current and future trends in drug discovery as well as providing a personal outlook on the future of drug research with a focus on small molecules.
Xiang Fu; Jie Tang; Ruyu Hua; Xiaoqian Li; Zhenghui Kang; Huang Qiu; Wenhao Hu Org. Lett., 2022, 24(11), 2208-2213 https://doi.org/10.1021/acs.orglett.2c00516AbstractTo explore potential chemical space using DNA-encoded library (DEL) technology, the development of various types of robust DNA-compatible reactions is urgently needed. Diazo compounds, which serve as valuable building blocks and important synthons in synthetic chemistry, have been rarely applied in DEL synthesis, probably because of their potential modifications of the bases and phosphate backbone of DNA. Herein we report two cases of DNA-compatible reactions with alkenes and diazo compounds, providing corresponding hydroalkylation and cyclopropanation products in moderate to excellent yields. Notably, these transformations not only provide new access to C(sp3)–C(sp3) bond formation in DELs with excellent functional group tolerance but also represent practical ligation methods to introduce functionalized molecules into DNA.
Eriko Mitsuno; Tatsuro Endo; Hideaki Hisamoto; Kenji Sueyoshi Anal. Sci., 2022, 38, 851-859 https://doi.org/10.1007/s44211-022-00101-xAbstractRecently, high-throughput analysis with minimal reagent consumption has been desired to assess interactions between drug candidates and disease-related oligonucleotides. To realize an ideal assay for drug screening, a rapid assay based on affinity capillary electrophoresis was generated to reduce the consumption of samples/reagents by a partial-filling technique under nonequilibrium conditions. In the proposed method, the first sample, oligonucleotide as a ligand, and second sample zones were injected into a capillary with spacers of background solution between the samples and oligonucleotide zones. After applying voltages, only the second sample zone passed through the partially filled oligonucleotide zone, resulting in variations in the peak parameters owing to this interaction. The electropherograms obtained were analyzed using equilibrium, reaction kinetics, and moment theories. In the interaction analyses between small molecules and DNA aptamers, only the small molecules binding to the aptamer showed significant changes in their peak heights and shapes. The estimated kinetic parameters were in good agreement with the reported values, indicating the applicability of the proposed method for drug screening. When interactions between drug candidates and disease-related RNAs were analyzed, one of the candidates showed remarkable variation in the peak parameters upon the addition of potassium ions. Consequently, the proposed method could be one of the ideal assays for drug screening.
Juan Yang; Shengdi Xia; Jiaxiang Liu; Zikun Yu; Lianghe Mei; Weiwei Lu; Yulin Chen; Shaopeng Chen; Xuan Wang; Xiaojie Lu Tetrahedron Lett., 2022, 153732 https://doi.org/10.1016/j.tetlet.2022.153732AbstractIncorporating various heteroaryl motifs into small-molecule moieties of DNA-encoded libraries is beneficial to support hit identification. More privileged heterocycles based libraries are urgently needed to expand the chemical space of the DNA encoded library. Because currently published on-DNA C-N cross-coupling methods mainly focused on exocyclic aromatic amines, lacking systematic investigation for incorporating pharmaceutically valuable N-aryl hetero-aromatic moiety into DNA-encoded libraries, in this paper, we have reported an on-DNA protocol to attach the DNA-linked electrophiles to the (H)N-heteroaryl rings. This method could translate the monofunctional heteroaryl agents into “bifunctional building blocks”, which have been applied for the DNA encoded focused indazole library synthesis.
Wang Liu; Xiaopeng Bai; Liping Song; Xuan Wang; Xiaojie Lu Bioconjugate Chem., 2022, 33(4), 560-565 https://doi.org/10.1021/acs.bioconjchem.2c00078AbstractMacrocyclic peptides are an important class of therapeutic agents for the biological targets that are difficult to modulate by small-molecule compounds. Meanwhile, DNA-encoded library technology (DELT) provides a powerful platform for hits discovery. The unity of both fields has proven highly productive in finding cyclic peptide hits against diverse pharmaceutical proteins. Many researchers have extended the chemical toolbox for constructing head-to-tail macrocyclic DNA-encoded libraries with various ring sizes. However, the linear peptides of different lengths necessitate tuning the distance between closing sites and DNA-linked sites to perform the macrocyclization process, presumably due to the constrained conformation of linear precursors. To tackle this issue and streamline the synthetic workflow, we report a two-directional synthesis strategy. This method starts from a trifunctional reagent and prepares DNA-linked macrocyclic peptides of ring size between 15 (5-mer) and 24 (8-mer) via amide bond formation reaction, a common method to create macrocyclic peptides.
Sebastian Dengler; Ryan T. Howard; Vasily Morozov; Christos Tsiamantas; Zhiwei Liu; Christopher Dobrzanski; Vojislava Pophristic; Sophie Brameyer; Céline Douat; Hiroaki Suga; Ivan Huc ChemRxiv, 2022 https://doi.org/10.26434/chemrxiv-2022-5rlmvAbstractA helical aromatic foldamer was identified that undergoes tRNA acylation by a flexizyme and ribosomal peptide initiation with yields sufficiently high to perform an mRNA display selection of macrocyclic foldamer-peptide hybrids. A hybrid macrocyle binder to the C-lobe of the E6AP HECT domain was selected that showed highly converged peptide residues. A crystal structure and molecular dynamics simulations revealed that both the peptide and foldamer are helical in an intriguing reciprocal stapling fashion. The strong residue convergence could be rationalized based on their involvement in specific interactions. The foldamer stabilizes the peptide helix through stapling and through contacts with key residues. It appears to also contribute to protein binding by direct protein interactions. The results altogether highlight possible benefits in inserting an aromatic foldamer into a peptide macrocycle for the purpose of protein recognition.
Carlos B. P. Oliveira; Sérgio R. S. Veloso; Pedro R. Figueiredo; Alexandra T. P. Carvalho; Loic Hilliou; Renato Pereira; David M. Pereira; Jose Alberto Martins; Paula Margarida Ferreira; Peter John Jervis Soft Matter, 2022, 18, 3955-3966 https://doi.org/10.1039/D2SM00121GAbstractShort peptides capped on the N-terminus with aromatic groups are often able to form supramolecular hydrogels - self-assembled networks of fibrils able to trap water molecules. Typically, these hydrogelators can form stiff gels at concentrations of 0.1 to 1.0 wt% - i.e. they consist of mainly water. The properties of these soft materials mimic those of the extracellular matrix (ECM) of biological tissue and therefore they have found many biomedical uses in tissue engineering, wound healing, drug delivery, biosensing and bioprinting applications. In drug delivery strategies related to cancer therapy, injectable hydrogels can serve as a depot formulation, where a sustained release of the chemotherapeutic from near the tumour site allows reduced doses and, therefore, decreased side effects. To further target cancer cells, folic acid-conjugated hydrogels and nanostructures are often sought, to exploit the overexpression of folate receptors on cancer cells - an approach which can allow the selective cellular uptake of an encapsulated drug. In this present study, two known dipeptide folate receptor ligands (1 and 2) recently identified from a screen of a DNA-encoded compound library, were synthesised and investigated for their hydrogelation ability and cytotoxicity. Compound 1, containing a naproxen capping group, rapidly forms hydrogels at concentrations as low as 0.03 wt% - one of the lowest critical gelation concentrations (CGCs) known for a supramolecular hydrogelator. In contrast, compound 2, which contains a 3-indolepropionic acid capping group, was unable to form hydrogels under a range of conditions and concentrations, instead forming nanospheres with diameters of 0.5 μm. Hydrogels of 1 were characterised by STEM microscopy, rheology, fluorescence spectroscopy and circular dichroism. Both compounds 1 and 2 had no impact on the proliferation of kerotinocytes (HaCaT cells) at concentrations up to 100 μM. Compound 1, containing the NSAID, was tested for anti-inflammatory activity in a human cyclooxygenase-1/2 model. The rate of the release of model drug compounds from within hydrogels of 1 was also investigated.
Mateja Klika Škopić; Florian Losch; Angus E. McMillan; Niklas Willeke; Mateo Malenica; Luis Bering; Jeffrey Bode; Andreas Brunschweiger Org. Lett., 2022, 24(6), 1383-1387 https://doi.org/10.1021/acs.orglett.2c00228AbstractReactions that require strictly dry conditions are challenging to translate to a DNA-encoded library format. Controlled pore glass solid support-connected DNA oligonucleotide–aldehyde conjugates could be condensed with SnAP reagents and cyclized to various sp3-rich heterocycles. The Boc-group of products provided a handle for product purification, and its facile removal under acidic conditions was tolerated by a chemically stabilized barcode. The reaction provides reagent-based scaffold diversity with functionalities for further library synthesis.
Li Quan Koh; Yi Wee Lim; Zachary P. Gates Int. J. Pept. Res. Ther., 2022, 28, 62 https://doi.org/10.1007/s10989-022-10370-9AbstractRecently, de novo MS/MS peptide sequencing has enabled the application of affinity selections to synthetic peptide mixtures that approach the diversity of phage libraries (> 108 random peptides). In conjunction with ‘split-mix’ solid phase synthesis to access equimolar peptide mixtures, this approach provides a straightforward means to examine synthetic peptide libraries of considerably higher diversity than has been feasible historically. Here, we offer a critical perspective on this work, report emerging data, and highlight opportunities for further methods refinement. With continued development, ‘affinity selection–mass spectrometry’ may become a complimentary approach to phage display, in vitro selection, and DNA-encoded libraries for the discovery of synthetic ligands that modulate protein function.
Samuel J. Roberts; Ziwei Liu; John D. Sutherland J. Am. Chem. Soc., 2022, 144(9), 4254-4259 https://doi.org/10.1021/jacs.2c00772AbstractTranslation according to the genetic code is made possible by selectivity both in aminoacylation of tRNA and in anticodon/codon recognition. In extant biology, tRNAs are selectively aminoacylated by enzymes using high-energy intermediates, but how this might have been achieved prior to the advent of protein synthesis has been a largely unanswered question in prebiotic chemistry. We have now elucidated a novel, prebiotically plausible stereoselective aminoacyl-RNA synthesis, which starts from RNA-amino acid phosphoramidates and proceeds via phosphoramidate-ester intermediates that subsequently undergo conversion to aminoacyl-esters by mild acid hydrolysis. The chemistry avoids the intermediacy of high-energy mixed carboxy-phosphate anhydrides and is greatly favored under eutectic conditions, which also potentially allow for the requisite pH fluctuation through the variable solubility of CO2 in solid/liquid water.
Donatien de Rochambeau; Serhii Hirka; Daniel Saliba; Shaun Anderson; Violeta Toader; Michael D. Dore; Felix J. Rizzuto; Maureen McKeague; Hanadi F. Sleiman ChemRxiv, 2022 https://doi.org/10.26434/chemrxiv-2022-zms0gAbstractPolymers with sequence definition allow access to programmable morphologies and applications, but directly correlating polymer structure to function currently requires case-by-case analysis: high throughput methods that identify promising species from entire chemical families are required. Here, we show that the discovery of effective protein target-recognition molecules can be achieved using DNA-encoded libraries of chemically diverse sequence-defined oligomers, generated on an automated DNA synthesizer. These structures are ALENOMERs – Aptamer-Like ENcoded OligoMERs – that are read and sequenced using a DNA code that branches from, and corresponds to, the target-binding oligomer. By incorporating nucleosidic and non-nucleosidic components into alenomers at specific locations, we unlock new supramolecular interactions for biomolecule binding, and directly correlate their effectiveness at each site. Our alenomer library screening removes the low throughput bottleneck of analyzing individual sequence-defined polymers, improving the binding efficacies of natural systems and enabling wide chemical spaces to be sampled for biomolecule sensing, therapy, and diagnostics.
Felix Meissner; Jennifer Geddes-McAlister; Matthias Mann; Marcus Bantscheff Nat. Rev. Drug Discov., 2022, 21, 637–654 https://doi.org/10.1038/s41573-022-00409-3AbstractProteins are the main targets of most drugs; however, system-wide methods to monitor protein activity and function are still underused in drug discovery. Novel biochemical approaches, in combination with recent developments in mass spectrometry-based proteomics instrumentation and data analysis pipelines, have now enabled the dissection of disease phenotypes and their modulation by bioactive molecules at unprecedented resolution and dimensionality. In this Review, we describe proteomics and chemoproteomics approaches for target identification and validation, as well as for identification of safety hazards. We discuss innovative strategies in early-stage drug discovery in which proteomics approaches generate unique insights, such as targeted protein degradation and the use of reactive fragments, and provide guidance for experimental strategies crucial for success.
Rui Ge; Zuyuan Shen; Jian Yin; Wenhua Chen; Qi Zhang; Yulong An; Dewei Tang; Alexander L. Satz; Wenji Su; Letian Kuai SLAS Discov., 2022, 27(2), 79-85 https://doi.org/10.1016/j.slasd.2022.01.001AbstractCovalent inhibitors targeting the main protease (Mpro, or 3CLpro) of SARS-CoV-2 have shown promise in preclinical investigations. Herein, we report the discovery of two new series of molecules that irreversibly bind to SARS-CoV-2 Mpro. These acrylamide containing molecules were discovered using our DNA-encoded library (DEL) screening platform. Following selection against SARS-CoV-2 Mpro, off-DNA compounds were synthesized and investigated to determine their inhibitory effects, the nature of their binding, and generate preliminary structure-activity relationships. LC-MS analysis indicates a 1:1 (covalent) binding stoichiometry between our hit molecules and SARS-CoV-2 Mpro. Fluorescent staining assay for covalent binding in the presence of cell lysate suggests reasonable selectivity for SARS-CoV-2 Mpro. And lastly, inhibition of enzymatic activity was also observed against a panel of 3CLpro enzymes from different coronavirus strains, with IC50 values ranging from inactive to single digit micromolar. Our results indicate that DEL selection is a useful approach for identifying covalent inhibitors of cysteine proteases.
Alexander A. Peterson; Aziz M. Rangwala; Manish K. Thakur; Patrick S. Ward; Christie Hung; Ian R. Outhwaite; Alix I. Chan; Dmitry L. Usanov; Vamsi K. Mootha; Markus A. Seeliger; David R. Liu Nat. Chem. Biol., 2022, 18(11), 1184-1195 https://doi.org/10.1038/s41589-022-01116-1AbstractAlthough cyclophilins are attractive targets for probing biology and therapeutic intervention, no subtype-selective cyclophilin inhibitors have been described. We discovered novel cyclophilin inhibitors from the in vitro selection of a DNA-templated library of 256,000 drug-like macrocycles for cyclophilin D (CypD) affinity. Iterated macrocycle engineering guided by ten X-ray co-crystal structures yielded potent and selective inhibitors (half maximal inhibitory concentration (IC50) = 10 nM) that bind the active site of CypD and also make novel interactions with non-conserved residues in the S2 pocket, an adjacent exo-site. The resulting macrocycles inhibit CypD activity with 21- to >10,000-fold selectivity over other cyclophilins and inhibit mitochondrial permeability transition pore opening in isolated mitochondria. We further exploited S2 pocket interactions to develop the first cyclophilin E (CypE)-selective inhibitor, which forms a reversible covalent bond with a CypE S2 pocket lysine, and exhibits 30- to >4,000-fold selectivity over other cyclophilins. These findings reveal a strategy to generate isoform-selective small-molecule cyclophilin modulators, advancing their suitability as targets for biological investigation and therapeutic development.
Yue Pan; Mary M. Mader J. Med. Chem., 2022, 65(7), 5288-5299 https://doi.org/10.1021/acs.jmedchem.2c00073AbstractWith several marketed drugs, allosteric inhibition of kinases has translated to pharmacological effects and clinical benefits comparable to those from orthosteric inhibition. However, despite much effort over more than 20 years, the number of kinase targets associated with FDA-approved allosteric drugs is limited, suggesting the challenges in identifying and validating allosteric inhibitors. Here we review the principles of allosteric inhibition, summarize the discovery of allosteric MEK1/2 and BCR-ABL1 inhibitors, and discuss the approaches to screening and demonstrating the functional activity of allosteric pocket ligands.
Guixian Zhao; Xiaohong Fan; Yangfeng Li; Gong Zhang; Yizhou Li ChemBioChem, 2022, e202200025 https://doi.org/10.1002/cbic.202200025AbstractDNA-encoded chemical libraries (DEL) have attracted substantial attention due to the infinite possibility for hit discovery in both pharmaceutical companies and academia. The encoding method is the initial step of DEL construction and one of the cornerstones of DEL applications. Classified by the DNA format, the existing DEL encoding strategies were categorized into single-stranded DNA-based strategies and double-stranded DNA-based strategies. The two DEL formats have their unique advantages but are usually incompatible with each other. To address this issue, we propose the concept of interconversion between double- and single-stranded DEL based on the “reversible covalent headpiece (RCHP)” design, which combines maximum robustness of synthesis with extraordinary flexibility of applications in distinct setups. Future opportunities in this field are also proposed to advance DEL technology to a comprehensive drug discovery platform.
Bingbing Shi; Yu Zhou; Xiaoyu Li RSC Chem. Biol., 2022, 3, 407-419 https://doi.org/10.1039/D2CB00007EAbstractDNA-encoded chemical library (DEL) has emerged to be a powerful technology platform in drug discovery and is also gaining momentum in academic research. The rapid development of DNA-/DEL-compatible chemistries has greatly expanded the chemical space accessible to DELs. The DEL technology has been widely adopted in the pharmaceutical industry and a number of clinical drug candidates have been identified from DEL selections. Recent innovations have combined DELs with other legacy and emerging techniques. Among them, DNA-encoded dynamic library (DEDL) introduces DNA encoding into the classic dynamic combinatorial libraries (DCLs) and also integrates the principle of fragment-based drug discovery (FBDD), making DEDL a novel approach with distinct features from static DELs. In this review, we provide a summary of the recently developed DEDL methods and their applications. Future developments in DEDLs are expected to extend the application scope of DELs to complex biological systems with unique ligand discovery capabilities.
Kunliang Cai; Yuzhao Ran; Wenbo Sun; Sen Gao; Jin Li; Jinqiao Wan; Guansai Liu Front. Chem., 2022, 10, 851674 https://doi.org/10.3389/fchem.2022.851674AbstractC-N bond formation is one of the most commonly used reactions in medicinal chemistry. Herein, we report an efficient Pd-promoted hydroamination reaction between DNA-conjugated aryl alkenes and a wide scope of aliphatic amines. The described reactions are demonstrated in good to excellent conversions to furnish C (sp3)–N bonds on DNA. This DNA-compatible transformation has strong potentials for the application into DNA-encoded library synthesis.
Zhaomei Sun; Shaoguang Yang; Lijun Xue; Jie Zhang; Kexin Yang; Yunjin Hu Chem. Asian J., 2022, e202200016 https://doi.org/10.1002/asia.202200016AbstractA series of novel N-alkyl linkers that connect small-molecule library members with their encoding DNA oligonucleotides has been developed. In comparison with the standard amide linker (usually constructed with oligo-AOP-NH2), the N-alkyl linker is not only more chemically stable, but also provides better structural diversity at the linkage point. Chemical variety in the vicinity of the polyglycol terminus, in particular, could affect binding interactions with the target protein. It could have been neglected in previous DNA-encoded chemical library (DEL) synthesis and screening studies due to the limited linkage alternatives. With these linkers, one can produce versatile key intermediates as Cycle 1 products directly amenable to Cycle 2 chemistry without the use of protecting groups. As a result, a DEL synthesis process that uses the fewest chemical conversions, such as 3-step, 3-cycle DELs, can achieve higher synthetic efficiency while creating less DNA tag degradation, resulting in higher quality DELs.
Peter Schultz Science, 2022, 375, 6576 https://doi.org/10.1126/science.abn6377AbstractRichard Lerner, physician-scientist and president of Scripps Research from 1987 to 2012, died on 2 December 2021. He was 83. Richard had an enormous impact on science, both through his powerful intellect and creativity in the laboratory and as the bold, inspirational leader who grew Scripps Research into one of the world’s preeminent research institutes.
Celina Reverdy; Gaetan Gitton; Xiangying Guan; Indranil Adhya; Rama Dumpati; Samir Roy; Gauthier Errasti; Thomas Delacroix; Raj Chakrabarti Bioorg. Med. Chem., 2022, 116999 https://doi.org/10.1016/j.bmc.2022.116999AbstractAmong the sirtuin members, Sirt3 is one of the most important deacetylases as it regulates acetylation levels in mitochondria, which are linked to the metabolism of multiple organs and therefore involved in many types of human diseases such as age-related diseases, cancer, heart disease and metabolic diseases. In the current absence of any direct activator of Sirt3, the identification of new modulators could be a key step in the development of new therapeutics. Here we report the discovery of Sirt3 modulators thanks to DNA encoded library technology (ELT). The most enriched compounds after DEL selection against SIRT3 were evaluated according to their activity and affinity. Our best activator seems at least as potent as Honokiol (HKL) while the docking studies tend to show that our modulators probably interact with Sirt3 at an atypical site.
Yoshihiro Nishimoto; Naoki Sugihara; Makoto Yasuda Synthesis, 2022, 54(12), 2765-2777 https://doi.org/10.1055/a-1755-3476AbstractPrecisely controllable C–F bond activation is a significant challenge in organic synthesis due to its large bond energy. A single C(sp3)–F bond transformation of perfluoroalkyl groups is particularly desirable to supply functionalized perfluoroalkyl compounds offering properties that could be significantly useful in pharmaceutical and materials chemistry. Recently, the single defluoronative transformation of perfluoroalkyl compounds was developed via visible-light photocatalysis. Herein, we summarize this field via two main topics. Topic 1 involves the transformation of C(sp3)–F bonds in either perfluoroalkylarenes or perfluoroalkane carbonyl compounds via a defluorinative spin-center shift in the radical anion intermediates. Topic 2 addresses the defluorinative transformation of α-trifluoromethyl alkenes to gem-difluoroalkenes via a radical/polar crossover process.
Ram K. Modukuri; Zhifeng Yu; Zhi Tan; Hai Minh Ta; Melek Nihan Ucisik; Zhuang Jin; Justin L. Anglin; Kiran L. Sharma; Pranavanand Nyshadham; Feng Li; Kevin Riehle; John C. Faver; Kevin Duong; Sureshbabu Nagarajan; Nicholas Simmons; Stephen S. Palmer; Mingxing Teng; Damian W. Young; Joanna S. Yi; Choel Kim; Martin M. Matzuk Proc. Natl. Acad. Sci. USA, 2022, 119(22), e2122506119 https://doi.org/10.1073/pnas.2122506119AbstractSignificanceBET bromodomain inhibition is therapeutic in multiple diseases; however, pan-BET inhibitors have induced significant myelosuppression and gastrointestinal toxicity, perhaps due to inhibition of both tandem bromodomains (BD) of all BET family members. However, selective inhibition of just the first BD (BD1) phenocopies pan-BET inhibitor activity in preclinical models of cancer, other diseases, and, for BRDT, in the testes for a contraceptive effect. Here, we leveraged our multibillion-molecule collection of DNA-encoded chemical libraries (DECLs) to identify BET BD1-selective inhibitors of specific chirality with high potency, stability, and good cellular activity. Our findings highlight the robustness and efficiency of the DECL platform to identify specific, potent protein binders that have promise as potential anticancer and anti-inflammatory agents and as male contraceptives.
Fabio Urbina; Sean Ekins AILSCI, 2022, 2, 100031 https://doi.org/10.1016/j.ailsci.2022.100031AbstractAnyone involved in designing or finding molecules in the life sciences over the past few years has witnessed a dramatic change in how we now work due to the COVID-19 pandemic. Computational technologies like artificial intelligence (AI) seemed to become ubiquitous in 2020 and have been increasingly applied as scientists worked from home and were separated from the laboratory and their colleagues. This shift may be more permanent as the future of molecule design across different industries will increasingly require machine learning models for design and optimization of molecules as they become “designed by AI”. AI and machine learning has essentially become a commodity within the pharmaceutical industry. This perspective will briefly describe our personal opinions of how machine learning has evolved and is being applied to model different molecule properties that crosses industries in their utility and ultimately suggests the potential for tight integration of AI into equipment and automated experimental pipelines. It will also describe how many groups have implemented generative models covering different architectures, for de novo design of molecules. We also highlight some of the companies at the forefront of using AI to demonstrate how machine learning has impacted and influenced our work. Finally, we will peer into the future and suggest some of the areas that represent the most interesting technologies that may shape the future of molecule design, highlighting how we can help increase the efficiency of the design-make-test cycle which is currently a major focus across industries.
Guilherme M. Lima; Alexey Atrazhev; Susmita Sarkar; Mirat Sojitra; Revathi Reddy; Matthew S. Macauley; Gisele Monteiro; Ratmir Derda; Karin Torres-Obreque; Carlota de Oliveira Rangel-Yagui ACS Chem. Biol., 2022, 17(11), 3024–3035 https://doi.org/10.1021/acschembio.1c00835AbstractPhage display links phenotype of displayed polypeptides with DNA sequence in phage genome and offers a universal method for discovery of proteins with novel properties. Injection of phage-displayed libraries in living organisms further provides a unique and powerful approach to optimize biochemical, pharmacological and biological properties of the displayed peptides, antibodies and other proteins in vivo. However, over 60% of the proteome is comprised of multi-domain proteins, and display of large multi-subunit proteins on phages remains a challenge. Majority of protein display systems are based on monovalent phagemid constructs but methods for robust display of multiple copies of large proteins are scarce. Here, we describe a DNA-encoded display of a ∼200 kDa tetrameric protein tetrameric L-asparaginase on M13 phage produced by ligation of SpyCatcher-Asparaginase fusion (ScA) to prospectively barcoded phage clones displaying SpyTag peptide. Starting from the SpyTag display on p3 minor coat protein or p8 major coat protein yielded constructs with five copies of ScA displayed on p3 (ScA5-phage) and 50 copies of ScA on p8 protein (ScA50-phage). ScA remained active after conjugation. It could be easily produced directly from lysates of bacteria that express ScA. Display constructs of different valency can be injected into mice and analyzed by deep-sequencing of the DNA barcodes associated phage clones. In these multiplexed studies, we observed a density-dependent clearance rate in vivo. A known clearance mechanism of L-asparaginase is endocytosis by phagocytic cells. Our observations, thus, link the increase in density of the displayed protein with the increased rate of the endocytosis by cells in vivo. In conclusion, we demonstrate that a multivalent display of L-asparaginase on phage could be used to study the circulation life of this protein in vivo and such approach opens the possibility to use DNA sequencing to investigate multiplexed libraries of other multi-subunit proteins in vivo.
Eric Koesema; Animesh Roy; Nicholas G. Paciaroni; Thomas Kodadek Angew. Chem. Int. Ed. Engl., 2022, 61(18), e202116999 https://doi.org/10.1002/anie.202116999AbstractThere is considerable interest in the development of libraries of non-peptidic macrocycles as a source of ligands for difficult targets. We report here the solid-phase synthesis of a DNA-encoded library of several hundred thousand thioether-linked macrocycles. The library was designed to be highly diverse with respect to backbone scaffold diversity and to minimize the number of amide N-H bonds, which compromise cell permeability. The utility of the library as a source of protein ligands is demonstrated through the isolation of compounds that bind streptavidin, a model target, with high affinity.
Qi Liang; Haiyue Zuo; Tian Yang; Jiatai Yin; Xiaomin Huang; Jing Wang; Qian Li; Guowei Yin; Xinfeng Zhao Eur. J. Med. Chem., 2022, 114212 https://doi.org/10.1016/j.ejmech.2022.114212AbstractThe design, synthesis, and discovery of dual-target compounds are considered as a promising strategy to develop new drugs with improved safety and efficacy compared with single-target drugs. This increasingly necessitates the methodologies that enable us to rapidly and accurately achieve the dual-target leads. Applying rosmarinic acid, 18β-glycyrrhetinic acid, rhein, and ferulic acid as template building blocks, we introduced the self-assembling DNA encoded technique to build the library containing 1,000 compounds. These compounds were screened by receptor chromatography with immobilized beta2-adrenoceptor (β2-AR) and cysteinyl-leukotriene receptor (CysLT), whereby we obtained a derivative of 18β-glycyrrhetinic acid (XC267) that specifically bind to the two receptors. In vitro assessment demonstrated the desired binding affinity of 6.57×104 M−1 to β2-AR, 2.82×104 M−1 to CysLT, and the dissociation rate constant of 7.52 s−1 to β2-AR, 17.2 s−1 to CysLT. Pharmacological examination with ovalbumin-induced mice demonstrated that XC267 significantly reduced the levels of IL-4, IL-13, and IgE after oral administration of 10 mg/kg. By western blot analysis, we observed an up-regulated expression of β2-AR and a blocked level of CysLT with a dose-dependent manner in pulmonary bronchial. Our results suggest XC627 is a potential candidate to treat asthma by simultaneously regulating the signaling pathway of the two receptors.
Isaline F.S.F. Castan; Andrew Madin; Garry Pairaudeau; Michael J. Waring Bioorg. Med. Chem., 2022, 116688 https://doi.org/10.1016/j.bmc.2022.116688AbstractDNA-Encoded Libraries (DEL) represent a promising hit finding strategy for drug discovery. Nonetheless, the available DNA-compatible chemistry remains of limited scope. Nucleophilic aromatic substitution (SNAr) has been extensively used in DEL synthesis but has generally been restricted to highly activated (hetero)arenes. Herein, we report an optimised procedure for of the SNAr reaction through the use of factorial experimental design (FED) on-DNA using 15% THF as a co-solvent. This method gave conversions of > 95% for pyridine and pyrazine scaffolds for 36 secondary cyclic amines. This analysis provides a new DNA-compatible SNAr reaction to produce high yielding libraries. The scope of this reaction on other amines is described. This work identifies challenges for the further development for DNA-compatible SNAr reactions.
Paddy R. A. Melsen; Ryoji Yoshisada; Seino A. K. Jongkees ChemBioChem, 2022, e202100685 https://doi.org/10.1002/cbic.202100685AbstractDNA-encoded small-molecule libraries and mRNA displayed peptide libraries both use numerically large pools of oligonucleotide-tagged molecules to identify potential hits for protein targets. They differ dramatically, however, in the 'drug-likeness' of the molecules that each can be used to discover. We give here an overview of the two techniques, comparing some advantages and disadvantages of each, and suggest areas where particularly mRNA display can benefit from adopting advances developed with DNA-encoded small molecule libraries. We outline cases where chemical modification of the peptide library has already been used in mRNA display, and survey opportunities to expand this using examples from DNA-encoded small molecule libraries. We also propose potential opportunities for encoding such reactions within the mRNA/cDNA tag of an mRNA-displayed peptide library to allow a more diversity-oriented approach to library modification. Finally, we outline alternate approaches for enriching target-binding hits from a pooled and tagged library, and close by detailing several examples of how an adjusted mRNA-display based approach could be used to discover new 'drug-like' modified small peptides.
Ryosuke Kita; Takashi Osawa; Satoshi Obika RSC Chem. Biol., 2022, 3, 728-738 https://doi.org/10.1039/D1CB00240FAbstractThe DNA-encoded library (DEL) is a powerful tool for drug discovery. As a result, to obtain diverse DELs, many DNA-compatible chemical reactions have been developed over the past decade. Among the most commonly used reactions in medicinal chemistry, multicomponent reactions (MCRs) can lead to the generation of various compounds in a one-step reaction. In particular, the Ugi reaction can easily provide a peptoid library. Thus, we herein report a solution-phase DEL synthesis based on the Ugi reaction. Using 6-(4-nitrophenoxycarbonylamino)hexanoic acid and N-4-nitrophenoxycarbonylgylcine as carboxylic acids, peptoids with activated carbamate moieties were obtained as the products of the Ugi reaction. These peptoids were then treated with oligonucleotides bearing a 5ʹ- or 3ʹ-terminal aminohexyl linker to give various oligonucleotide-tagged peptoids in good yields. Moreover, the obtained peptoids could be substituted by a Suzuki cross-coupling reaction and by hydrolysis of the carboxylate ester, followed by condensation with amines. These advances should therefore promote pharmaceutical and medicinal research using DELs.
Chong Lu; Shien Liu; Weihua Shi; Jun Yu; Zhou Zhou; Xiaoxiao Zhang; Xiaoli Lu; Faji Cai; Ning Xia; Yikai Wang J. Cheminformatics, 2022, 14, 19 https://doi.org/10.1186/s13321-022-00598-4AbstractChemical space exploration is a major task of the hit-finding process during the pursuit of novel chemical entities. Compared with other screening technologies, computational de novo design has become a popular approach to overcome the limitation of current chemical libraries. Here, we reported a de novo design platform named systemic evolutionary chemical space explorer (SECSE). The platform was conceptually inspired by fragment-based drug design, that miniaturized a “lego-building” process within the pocket of a certain target. The key to virtual hits generation was then turned into a computational search problem. To enhance search and optimization, human intelligence and deep learning were integrated. Application of SECSE against phosphoglycerate dehydrogenase (PHGDH), proved its potential in finding novel and diverse small molecules that are attractive starting points for further validation. This platform is open-sourced and the code is available at http://github.com/KeenThera/SECSE.
Shuting Zhong; Xianfu Fang; Yiting Wang; Gong Zhang; Yangfeng Li; Yizhou Li Org. Lett., 2022, 24(4), 1022-1026 https://doi.org/10.1021/acs.orglett.1c04169AbstractIndole-based diversification is highly desired in the DNA-encoded chemical library construction. Herein, we present a general strategy for on-DNA synthesis of diverse C3-functionalized indole derivatives via indole π-activated alcohol formation followed by direct dehydrative coupling. Highly efficient bond linkages of C–C, C–N, and C–S were achieved to fuse building blocks that are widely commercially available. DNA-encoding compatibility of the method has been further demonstrated to pave an avenue for application in constructing indole-focused three-dimensional libraries.
Peter S. Dragovich; Wolfgang Haap; Melinda M. Mulvihill; Jean-Marc Plancher; Antonia F. Stepan J. Med. Chem., 2022, 65(4), 3606-3615 https://doi.org/10.1021/acs.jmedchem.1c02106AbstractThe origin of small-molecule leads that were pursued across the independent research organizations Roche and Genentech from 2009 to 2020 is described. The identified chemical series are derived from a variety of lead-finding methods, which include public information, high-throughput screening (both full file and focused), fragment-based design, DNA-encoded library technology, use of legacy internal data, in-licensing, and de novo design (often structure-based). The translation of the lead series into in vivo tool compounds and development candidates is discussed as are the associated biological target classes and corresponding therapeutic areas. These analyses identify important trends regarding the various lead-finding approaches, which will likely impact their future application in the Roche and Genentech research groups. They also highlight commonalities and differences across the two independent research organizations. Several caveats associated with the employed data collection and analysis methodologies are included to enhance the interpretation of the presented information.
Yuting Gao; Guixian Zhao; Pengyang He; Gong Zhang; Yangfeng Li; Yizhou Li Bioconjugate Chem., 2022, 33(1), 105-110 https://doi.org/10.1021/acs.bioconjchem.1c00567AbstractAs a powerful platform in drug discovery, the DNA-encoded chemical library technique enables the generation of numerous chemical members with high structural diversity. Epoxides widely exist in a variety of approved drugs and clinical candidates, eliciting multiple pharmaceutical activities. Herein, we report a non-oxidative DNA-compatible synthesis of di-/trisubstituted α,β-epoxyketones by implementing aldehydes and α-chlorinated ketones as abundant building blocks. This methodology was demonstrated to cover a broad substrate scope with medium-to-excellent conversions. Further structural diversification and transformation were also successfully explored to fully leverage α,β-epoxyketone moiety.
Philip B. Cox; Rishi Gupta ACS Med. Chem. Lett., 2022, 13(7), 1016–1029 https://doi.org/10.1021/acsmedchemlett.1c00662AbstractIn the past decade or so there has been a dramatic increase in the number of computational applications and tools that have been developed to enable medicinal chemists to prosecute modern drug discovery programs more efficiently. The upsurge of user-friendly, well-designed computational tools that enable structure-based drug design (SBDD) and cheminformatics (CI)-based drug design has equipped the medicinal chemist with an arsenal of tools and applications that significantly augments the entire design process, thereby enhancing the speed and efficiency of the design–make–test–analyze cycle. Modern computational applications and tools transcend all areas of drug discovery, and most savvy medicinal chemists can employ them effectively in a myriad of drug discovery applications. Indeed, the sheer scope and breadth of tools available to the medicinal chemist is vast and, to our knowledge, has not been comprehensively reviewed. In this article we have catalogued many computational tools, platforms, and applications that are currently available, with four main areas highlighted: commercially available tools/platforms, open-source applications, internally developed platforms (software tools developed within a pharma or biotech organization), and artificial intelligence/machine learning-based platforms. For ease of interpretation, for these categories we provide tables organized by vendor or organization name, the name of the application, whether the tool/application is employed predominantly for SBDD or CI-based design, and a summary of the main function of the tools, with associated hyperlinks to vendor Web sites. We have tried to be as comprehensive and as inclusive as possible; however, the pace of development of new and existing tools is so rapid that there may be omissions with respect to newly developed tools and current versions of the software.
Fei Ma; Jie Li; Shuning Zhang; Yuang Gu; Tingting Tan; Wanting Chen; Shuyue Wang; Hongtao Xu; Guang Yang; Richard A. Lerner ACS Catal., 2022, 12(3), 1639-1649 https://doi.org/10.1021/acscatal.1c05338AbstractMetal catalysis, a common approach in conventional organic synthesis, poses a challenge in DEL chemistry due to the vulnerability of DNA fragments and the requirement of aqueous media. Here, we describe a facile one-pot palladium-catalyzed reaction for the formation of C(sp2)–C(sp3) and C(sp3)–S bonds in the presence of DNA encoding. Using 3, 4-dimethoxybenzenesulfonohydrazide (L8) as a bridging reactant, our studies showed that DNA-conjugated benzaldehyde (HP-ArCHO-1), serving as a common precursor, reacted with derivatives of iodine, bromine, trifluoromethanesulfonate, and disulfides in metal-catalyzed one-pot chemical transformation to afford on-DNA diarylmethanes and thioethers. Notably, all reactions displayed wide substrate scopes and moderate to excellent yields under mild reaction conditions. These chemical reactions greatly expand the chemical space of DNA-compatible reactions and the molecular scaffold diversity of DNA-encoded libraries.
Alexander L. Satz; Andreas Brunschweiger; Mark E. Flanagan; Andreas Gloger; Nils J. V. Hansen; Letian Kuai; Verena B. K. Kunig; Xiaojie Lu; Daniel Madsen; Lisa A. Marcaurelle; Carol Mulrooney; Gary O' Donovan; Sylvia Sakata; Jörg Scheuermann Nat. Rev. Methods Primers, 2022, 2, 3 https://doi.org/10.1038/s43586-021-00084-5AbstractDNA-encoded chemical library (DECL) technology is used by the pharmaceutical industry to discover small molecules capable of modulating biologically relevant targets. DECL synthesis starts with an oligonucleotide that contains a chemical linker moiety, and proceeds through iterative cycles of DNA barcode elongation and chemical synthesis. DECL selections require little protein, minimal assay development and no specialized instrumentation. Parallel DECL selections can be easily conducted, making it possible to directly compare results across different conditions. The acquisition of building blocks is a large impediment when setting up a successful DECL platform. A potential solution is the sharing of building blocks between different labs, or the high-throughput parallel synthesis of novel building blocks. DNA-compatible reactions are required to join the building blocks together, and numerous academic labs have recently taken up this challenge. DECLs exist as unpurified mixtures, complicating data analysis. Machine learning may provide an improved ability to interrogate these data. DECL selections are largely limited to soluble purified proteins. However, progress has been made towards cell surface and in-cell selections. Publication guidelines are needed to better enable reproducibility; for example, the quantification of amplifiable DNA by quantitative PCR, and more complete datasets and building block lists, should be provided.
Yashoda Krishna Sunkari; Vijay Kumar Siripuram; Thu-Lan Nguyen; Marc Flajolet Trends Pharmacol. Sci., 2022, 43(1), 4-15 https://doi.org/10.1016/j.tips.2021.10.008AbstractThe world is totally dependent on medications. As science progresses, new, better, and cheaper drugs are needed more than ever. The pharmaceutical industry has been predominantly dependent on high-throughput screening (HTS) for the past three decades. Considering that the discovery rate has been relatively constant, can one hope for a much-needed sudden trend uptick? DNA-encoded libraries (DELs) and similar technologies, that have several orders of magnitude more screening power than HTS, and that we propose to group together under the umbrella term of high-power screening (HPS), are very well positioned to do exactly that. HPS also offers novel screening options such as parallel screening, ex vivo and in vivo screening, as well as a new path to druggable alternatives such as proteolysis targeting chimeras (PROTACs). Altogether, HPS unlocks novel powerful drug discovery avenues.
Shilian Yang; Guixian Zhao; Yuting Gao; Yang Sun; Xiaohong Fan; Gong Zhang; Yangfeng Li; Yizhou Li Chem. Sci., 2022, 13, 2604-2613 https://doi.org/10.1039/D1SC06268AAbstractSulfur/selenium-containing electron-rich arenes (ERAs) exist in a wide range of both approved and investigational drugs with diverse pharmacological activities. These unique chemical structures and bioactive properties, if combined with the emerging DNA-encoded chemical library (DEL) technique, would facilitate drug and chemical probe discovery. However, it remains challenging, as there is no general DNA-compatible synthetic methodology available for the formation of C-S and C-Se bonds in aqueous solution. Herein, an in-solution direct oxidative coupling procedure that could efficiently integrate sulfur/selenium to the ERA under mild conditions is presented. This method is featured by simple DNA-conjugated electron-rich arenes with a broad substrate scope and a transition-metal free process. Furthermore, this synthetic methodology, examined by a scale-up reaction test and late-stage precise modification in a mock peptide-like DEL synthesis, will render its utility for the synthesis of sulfur/selenium-containing DNA-encoded libraries and the discovery of bioactive agents.
Daniel W. Weisgerber; Makiko Hatori; Xiangpeng Li; Adam R. Abate Anal. Chem., 2022, 94(21), 7475-7482 https://doi.org/10.1021/acs.analchem.1c04884AbstractCurrent methods for fabricating microparticles offer limited control over size and shape. Here, we demonstrate a droplet microfluidic method to form polyhedral microparticles with controlled concavity. By manipulating Laplace pressure, buoyancy, and particle rheology, we generate microparticles with diverse shapes and curvatures. Additionally, we demonstrate the particles provide increased capture efficiency when used for particle-templated emulsification. Our approach enables microparticles with enhanced chemical and biological functionality.
Malliga R. Iyer; Biswajit Kundu; Casey M. Wood Expert Opin. Ther. Patents, 2022, 32(6), 629-647 https://doi.org/10.1080/13543776.2022.2054329AbstractBiological effects mediated by the CYP450 arm of arachidonate cascade implicate the enzyme-soluble epoxide hydrolase (sEH) in hydrolyzing anti-inflammatory epoxy fatty acids to pro-inflammatory diols. Hence, inhibiting the sEH offers a therapeutic approach to treating inflammatory diseases. Over three decades of work has shown the role of sEH inhibitors (sEHis) in treating various disorders in rodents and larger veterinary subjects. Novel chemical strategies to enhance the efficacy of sEHi have now appeared.
Yizhou Li; Guixian Zhao; Shuting Zhong; Gong Zhang; Yangfeng Li Angew. Chem. Int. Ed. Engl., 2022, 61(7), e202115157 https://doi.org/10.1002/anie.202115157AbstractThe use of a proper encoding methodology is one of the most important aspects when practicing DEL technology. A “headpiece”-based double-stranded DEL encoding method is currently the most widely used for productive DEL. However, the robustness of double-stranded DEL construction conflicts with the versatility presented by single-stranded DEL applications. We here report a novel encoding method, which is based on a “reversible covalent headpiece (RCHP)”. The RCHP allows reversible interconversion between double- and single-stranded DNA formats, providing an avenue to robust synthesis and allowing for the applications in distinct setups. We have validated the versatility of this encoding method with encoded self-assembled chemical library and DNA-encoded dynamic library technology. Notably, based on the RCHP-settled library construction, a unique “ternary covalent complex” mediating ligand isolation methodologies against non-immobilized targets was developed.
Zoltán Novák; Réka Adamik; Balázs Buchholcz; Ferenc Darvas; Gellért Sipos Chem. Eur. J. 2022, e202103967 https://doi.org/10.1002/chem.202103967AbstractDNA-encoded library (DEL) technology has become widely used in drug discovery research. The construction of DELs require robust organic transformations which proceed in aqueous media under mild conditions. Unfortunately, the application of water as reaction media for organic synthesis is not evident due to generally limited solubility of organic reagents. However, the use of surfactants can offer a solution to this issue. Oil-in-water microemulsions formed by surfactant micelles are able localize hydrophobic reagents inside them, resulting in high local concentration of the organic substances in an otherwise poorly solvated environment. This review provides a conceptual and critical summary of micellar synthesis possibilities that are well-suited for DEL synthesis. Existing examples on micellar DEL approaches, together with a selection of micellar organic transformations fundamentally suitable for DEL are discussed.
Wei Hou; Hongtao Xu J. Med. Chem., 2022, 65(6), 4436-4456 https://doi.org/10.1021/acs.jmedchem.1c01859AbstractSelenium (Se)-containing compounds have emerged as potential therapeutic agents for the treatment of a range of diseases. Through tremendous effort, considerable knowledge has been acquired to understand the complex chemical properties and biological activities of selenium, especially after its incorporation into bioactive molecules. From this perspective, we compiled extensive literature evidence to summarize and critically discuss the relationship between the pharmacological activities and chemical properties of selenium compounds and the strategic incorporation of selenium into organic molecules, especially bioactive heterocycles and natural products. We also provide perspectives regarding the challenges in selenium-based medicinal chemistry and future research directions.
Juncheng Chen; Pan Tang; Yuxi Wang; Jiaxing Wang; Chengcan Yang; Yang Li; Gaoxia Yang; Fengbo Wu; Jifa Zhang; Liang Ouyang J. Med. Chem., 2022, 65(7), 5184-5211 https://doi.org/10.1021/acs.jmedchem.1c01835AbstractBlocking the interactions between bromodomain and extraterminal (BET) proteins and acetylated lysines of histones by small molecules has important implications for the treatment of cancers and other diseases. Many pan-BET inhibitors have shown satisfactory results in clinical trials, but their potential for poor tolerability and toxicity persist. However, recently reported studies illustrate that some BET bromodomain (BET-BD1 or BET-BD2)-selective inhibitors have advantage over pan-inhibitors, including reduced toxicity concerns. Furthermore, some selective BET inhibitors have similar or even better therapeutic efficacy in inflammatory diseases or cancers. Therefore, the development of selective BET inhibitors has become a hot spot for medicinal chemists. Here, we summarize the known selective BET-BD1 and BET-BD2 inhibitors and review the methods for enhancing the selectivity and potency of these inhibitors based on their different modes of interactions with BET-BD1 or BET-BD2. Finally, we discuss prospective strategies that selectively target the bromodomains of BET proteins.
Michael Grigalunas; Susanne Brakmann; Herbert Waldmann J. Am. Chem. Soc., 2022, 144(8), 3314-3329 https://doi.org/10.1021/jacs.1c11270AbstractNatural products are the result of Nature’s exploration of biologically relevant chemical space through evolution and an invaluable source of bioactive small molecules for chemical biology and medicinal chemistry. Novel concepts for the discovery of new bioactive compound classes based on natural product structure may enable exploration of wider biologically relevant chemical space. The pseudo-natural product concept merges the relevance of natural product structure with efficient exploration of chemical space by means of fragment-based compound development to inspire the discovery of new bioactive chemical matter through de novo combination of natural product fragments in unprecedented arrangements. The novel scaffolds retain the biological relevance of natural products but are not obtainable through known biosynthetic pathways which can lead to new chemotypes that may have unexpected or unprecedented bioactivities. Herein, we cover the workflow of pseudo-natural product design and development, highlight recent examples, and discuss a cheminformatic analysis in which a significant portion of biologically active synthetic compounds were found to be pseudo-natural products. We compare the concept to natural evolution and discuss pseudo-natural products as the human-made equivalent, i.e. the chemical evolution of natural product structure.
Regina Pikalyova; Yuliana Zabolotna; Dmitriy Volochnyuk; Dragos Horvath; Marcou Gilles; Alexandre Varnek Mol. Inform., 2022, 41(6), 2100289 https://doi.org/10.1002/minf.202100289AbstractDNA-Encoded Library (DEL) technology has emerged as an alternative method for bioactive molecule discovery in medicinal chemistry. It enables simple synthesis and screening of compound libraries of enormous size. Even though it gains more and more popularity each day, there are almost no reports of chemoinformatics analysis of DEL chemical space. Therefore, in this project we aimed to generate and analyze theultra-large chemical space of DEL. Around 2500 DELs were designed using commercially available BBs resulting in 2,5B DEL compounds that were compared to biologically relevant compounds from ChEMBL using Generative Topographic Mapping. This allowed to choose several optimal DELs covering the chemical space of ChEMBL to the highest extent and thus containing the maximum possible percentage of biologically relevant chemotypes. Different combinations of DELs were also analyzed to identify a set of mutually complementary libraries allowing to attain even higher coverage of ChEMBL than it is possible with one single DEL.
Xiaofang Li; Pingping Fang; Zhuoyi Wang; Pei Xu Chin. J. Agric. Biotechnol., 2022, 30(11), 2119-2127 https://doi.org/10.3969/j.issn.1674-7968.2022.11.006AbstractThe expression of purified and active target protein is prerequisite for the screening of proteinspecific inhibitors. In this study, the prokaryotic expression, purification and activity analysis of tryptophan aminotransferase of Arabidopsis 1 (TAA1) were carried out. First, the TAA1 coding sequence was retrieved from GenBank. After codon optimization according to the preferred codon usage in Escherichia coli the target sequence was synthesized and inserted into the prokaryotic expression vector pET24 with a His tag to construct the recombinant expression vector pET24-GST-EK-His-TAA1. The vector was then transformed into the E. coli BL21(DE3) cell line followed by induction and cultivation with different methods (IPTG induction/TB medium, IPTG induction/LB medium, and auto-induction). The products were analyzed with sodium lauryl sulfate polyacrylamide gel electrophoresis to identify the size, amount and existing form of the expressed protein. It was found that the auto-inducible expression system combined with incubation at 37 ℃ for 16 h yielded the greatest amount of protein product in the soluble fraction. This optimized expression method was then used for large-scale expression and purification of the proteins. The collected fusion protein was further cleaved by the enterokinase (EK) protease and purified with a nickel ion affinity column. Finally, purified recombined TAA1 protein at a purity of 78.7% and an amount of 0.5 mg was obtained. In vitro enzymatic activity assay proved that this protein had a transaminase activity. In conclusion, a high-efficient prokaryotic expression system for the Arabidopsis TAA1 protein was established successfully, from which the obtained purified and bioactive His-TAA1 protein are very useful for the screening of TAA1-specific inhibitors based on molecular interaction.
Iwan J. P. de Esch; Daniel A. Erlanson; Wolfgang Jahnke; Christopher N. Johnson; Louise Walsh J. Med. Chem., 2022, 65(1), 84-99 https://doi.org/10.1021/acs.jmedchem.1c01803AbstractFragment-based drug discovery (FBDD) continues to evolve and make an impact in the pharmaceutical sciences. We summarize successful fragment-to-lead studies that were published in 2020. Having systematically analyzed annual scientific outputs since 2015, we discuss trends and best practices in terms of fragment libraries, target proteins, screening technologies, hit-optimization strategies, and the properties of hit fragments and the leads resulting from them. As well as the tabulated Fragment-to-Lead (F2L) programs, our 2020 literature review identifies several trends and innovations that promise to further increase the success of FBDD. These include developing structurally novel screening fragments, improving fragment-screening technologies, using new computer-aided design and virtual screening approaches, and combining FBDD with other innovative drug-discovery technologies.
Louise Plais; Alice Lessing; Michelle Keller; Adriano Martinelli; Sebastian Oehler; Gabriele Bassi; Dario Neri; Jörg Scheuermann Chem. Sci., 2022, 13, 967-974 https://doi.org/10.1039/D1SC05721AAbstractDNA-encoded chemical libraries (DELs) are useful tools for the discovery of small molecule ligands to protein targets of pharmaceutical interest. Compared with single-pharmacophore DELs, dual-pharmacophore DELs simultaneously display two chemical moieties on both DNA strands, and allow for the construction of highly diverse and pure libraries, with a potential for targeting larger protein surfaces. Although methods for the encoding of simple, fragment-like dual-display libraries have been established, more complex libraries require a different encoding strategy. Here, we present a robust and convenient “large encoding design” (LED), which facilitates the PCR-amplification of multiple codes distributed among two partially complementary DNA strands. We experimentally implemented multiple coding regions and we compared the new DNA encoding scheme with previously reported dual-display DEL modalities in terms of amplifiability and performance in test selections against two target proteins. With the LED methodology in place, we foresee the construction and screening of DELs of unprecedented sizes and designs.
Matthias Krumb; Lisa Marie Kammer; Shorouk O. Badir; María Jesús Cabrera-Afonso; Victoria E. Wu; Minxue Huang; Adam Csakai; Lisa A. Marcaurelle; Gary A Molander Chem. Sci., 2022, 13, 1023-1029 https://doi.org/10.1039/D1SC05683BAbstractDNA-encoded library (DEL) technology has emerged as a time- and cost-efficient technique for the identification of therapeutic candidates in the pharmaceutical industry. Although several reaction classes have been successfully validated in DEL environments, there remains a paucity of DNA-compatible reactions that harness building blocks (BBs) from readily available substructures bearing multifunctional handles for further library diversification under mild, dilute, and aqueous conditions. In this study, the direct C–H carbofunctionalization of medicinally-relevant heteroarenes can be accomplished via the photoreduction of DNA-conjugated (hetero)aryl halides to deliver reactive aryl radical intermediates in a regulated fashion within minutes of blue light illumination. A broad array of electron-rich and electron-poor heteroarene scaffolds undergo transformation in the presence of sensitive functional groups.
Qigui Nie; Xianfu Fang; Changyang Liu; Gong Zhang; Xiaohong Fan; Yangfeng Li; Yizhou Li J. Org. Chem., 2022, 87(5), 2551-2558 https://doi.org/10.1021/acs.joc.1c02496AbstractThe incorporation of the isoindole core into the DNA-encoded chemical library is highly desirable for the great potential pharmacological characters exampled by molecules like lenalidomide. Herein, we reported a DNA-compatible protocol for the OPA-mediated transformation of amines into drug-like moieties represented by isoindolinone and thio-2-isoindole, respectively. The high conversion and wide substrate-scope property of our protocol render its feasibility in the manipulation of terminal amines on oligonucleotide conjugates, including “cap-and-catch” purification, sequential synthesis during DEL construction, and on-DNA macrocyclization.
Zhanghong Wang; Yu Sun; Liu-Yu Shen; Wen-Chao Yang; Fei Meng; Pinhua Li Org. Chem. Front., 2022, 9, 853-873 https://doi.org/10.1039/D1QO01512EAbstractThe activation and functionalization of C-F bonds under mild conditions can serve as an important tool for organic syntheses including the modification of pharmaceuticals or agrochemicals and the synthesis of value-added chemicals. The C–F bond with high energy and low activity, however, is difficult to be activated. Thus, the direct activation of the C-F bond under mild or transition-metal free conditions is a challenging research topic. Recently, the photochemical or electrochemical C−F bond activation and functionalization has been evaluated as a scalable and sustainable approach that applies photo or electricity to replace dangerous chemical oxidants or metal catalysts. To date, the achivement made in this field has attracted great attention, however, reviews on the photochemical or electrochemical C−F bond activation and functionalization are scarce. In this review, we summarize recent significant developments in this rapidly growing area, mainly focusing on reaction design and reaction mechanisms.
Bo Cai; Casey J. Krusemark Angew. Chem. Int. Ed. Engl., 2022, 61(3), e202113515 https://doi.org/10.1002/anie.202113515AbstractSmall-molecule binding assays to target proteins are a core component of drug discovery and development. While a number of assay formats are available, significant drawbacks still remain in cost, sensitivity, and throughput. To improve assays by capitalizing on the power of DNA sequence analysis, we have developed an assay method that combines DNA encoding with split-and-pool sample handling. The approach involves affinity labeling of DNA-linked ligands to a protein target. Critically, the labeling event assesses ligand binding and enables subsequent pooling of several samples. Application of a purifying selection on the pool for protein-labeled DNAs allows detection of ligand binding by quantification of DNA barcodes. We demonstrate the approach in both ligand displacement and direct binding formats and demonstrate its utility in determination of relative ligand affinity, profiling ligand specificity, and high-throughput small-molecule screening.
Jolita Bruzgulienė; Greta Račkauskienė; Aurimas Bieliauskas; Vaida Milišiūnaitė; Miglė Dagilienė; Gita Matulevičiūtė; Vytas Martynaitis; Sonata Krikštolaitytė; Frank A. Sløk; Algirdas Šačkus Beilstein J. Org. Chem., 2022, 18, 102-109 https://doi.org/10.3762/bjoc.18.11AbstractA convenient and efficient synthesis of novel achiral and chiral heterocyclic amino acid-like building blocks was developed. Regioisomeric methyl 5-(N-Boc-cycloaminyl)-1,2-oxazole-4-carboxylates were prepared by the reaction of β-enamino ketoesters (including azetidine, pyrrolidine or piperidine enamines) with hydroxylamine hydrochloride. Unambiguous structural assignments were based on chiral HPLC analysis, 1H, 13C, and 15N NMR spectroscopy, HRMS, and single-crystal X-ray diffraction data.
Justina Medžiūnė; Žana Kapustina; Simona Žeimytė; Jevgenija Jakubovska; Rūta Sindikevičienė; Inga Čikotienė; Arvydas Lubys Commun. Chem., 2022, 5, 34 https://doi.org/10.1038/s42004-022-00649-9AbstractThe ever-growing demand for inexpensive, rapid, and accurate exploration of genomes calls for refinement of existing sequencing techniques. The development of next-generation sequencing (NGS) was a revolutionary milestone in genome analysis. While modified nucleotides already were inherent tools in sequencing and imaging, further modification of nucleotides enabled the expansion into even more diverse applications. Herein we describe the design and synthesis of oligonucleotide-tethered 2′,3′-dideoxynucleotide (ddONNTP) terminators bearing universal priming sites attached to the nucleobase, as well as their enzymatic incorporation and performance in read-through assays. In the context of NGS library preparation, the incorporation of ddONNTP fulfills two requirements at once: the fragmentation step is integrated into the workflow and the obtained fragments are readily labeled by platform-specific adapters. DNA polymerases can incorporate ddONNTP nucleotides, as shown by primer extension assays. More importantly, reading through the unnatural linkage during DNA synthesis was demonstrated, with 25-30% efficiency in single-cycle extension.
Qiuxia Chen; You Li; Chunrong Lin; Liu Chen; Hao Luo; Shuai Xia; Chuan Liu; Xuemin Cheng; Chengzhong Liu; Jin Li; Dengfeng Dou Nucleic Acids Research, 2022, 50(12), e67 https://doi.org/10.1093/nar/gkac173AbstractDNA-encoded library (DEL) technology is a powerful tool for small molecule identification in drug discovery, yet the reported DEL selection strategies were applied primarily on protein targets in either purified form or in cellular context. To expand the application of this technology, we employed DEL selection on an RNA target HIV-1 TAR (trans-acting responsive region), but found that the majority of signals were resulted from false positive DNA–RNA binding. We thus developed an optimized selection strategy utilizing RNA patches and competitive elution to minimize unwanted DNA binding, followed by k-mer analysis and motif search to differentiate false positive signal. This optimized strategy resulted in a very clean background in a DEL selection against Escherichia coli FMN Riboswitch, and the enriched compounds were determined with double digit nanomolar binding affinity, as well as similar potency in functional FMN competition assay. These results demonstrated the feasibility of small molecule identification against RNA targets using DEL selection. The developed experimental and computational strategy provided a promising opportunity for RNA ligand screening and expanded the application of DEL selection to a much wider context in drug discovery.
Hongbao Sun; Qinghe Xue; Chang Zhang; Haoxing Wu; Ping Feng Org. Chem. Front., 2022, 9, 481-498 https://doi.org/10.1039/D1QO01324FAbstractBioorthogonal chemistry is widely used in biological systems and has been trialed in patients, attracting a lot of attention in this century. Tetrazine-based bioorthogonal reactions are essential in chemical biology applications, including cellular labeling, live-cell imaging, diagnosis, drug release, and oncotherapy, due to their tunable rapid reaction kinetics and unique fluorogenic characteristics. However, the scope of de novo tetrazine synthesis is restricted due to the limited supply of commercial starting materials. Therefore, derivatization based on tetrazine scaffold has been used to synthesize various tetrazine derivatives to enhance applications of tetrazine bioorthogonal reactions. Herein, the recent advances in tetrazine scaffold-based derivatizations, including tetrazine skeleton, aromatic substituents and alkyl substituents of tetrazines have been summarized. The advantages and limitations of derivatization methods and applications of developed tetrazine derivatives in biorthogonal chemistry have also been highlighted.
Kai Tang; Bo Wang; Bin Yu; Hong-Min Liu Eur. J. Med. Chem., 2022, 227, 113967 https://doi.org/10.1016/j.ejmech.2021.113967AbstractIndoleamine 2,3-dioxygenase 1 (IDO1), a known immunosuppressive enzyme that catalyzes the rate-limiting step in the oxidation of tryptophan (Trp) to kynurenine (Kyn), has received increasing attention as an attractive immunotherapeutic target for cancer therapy. Up to now, eleven small-molecule IDO1 inhibitors have entered clinical trials for the treatment of cancers. In addition, proteolysis targeting chimera (PROTAC) based degraders also provide prospects for cancer therapy. Herein we present a comprehensive overview of the medicinal chemistry strategies and potential therapeutic applications of IDO1 inhibitors in nonclinical trials and IDO1-PROTAC degraders.
Ricardo A. M. Serafim; Jonathan M. Elkins; William J. Zuercher; Stefan A. Laufer; Matthias Gehringer J. Med. Chem., 2022, 65(2), 1132-1170 https://doi.org/10.1021/acs.jmedchem.1c00980AbstractOver 20 years after the approval of the first-in-class protein kinase inhibitor imatinib, the biological function of a significant fraction of the human kinome remains poorly understood while most research continues to be focused on few well-validated targets. Given the strong genetic evidence for involvement of many kinases in health and disease, the understudied fraction of the kinome holds a large and unexplored potential for future therapies. Specific chemical probes are indispensable tools to interrogate biology enabling proper preclinical validation of novel kinase targets. In this Perspective, we highlight recent case studies illustrating the development of high-quality chemical probes for less-studied kinases and their application in target validation. We spotlight emerging techniques and approaches employed in the generation of chemical probes for protein kinases and beyond and discuss the associated challenges and opportunities.
Harriet A. Stanway-Gordon; Jessica S. Graham; Michael James Waring Angew. Chem. Int. Ed. Engl., 2022, 61(3), e202111927 https://doi.org/10.1002/anie.202111927AbstractDNA-encoded libraries (DELs) are an increasingly popular approach to finding small molecule ligands for proteins. Many DEL synthesis protocols hinge on sequential additions of monomers using split-pool combinatorial methods. Therefore, compatible protecting group strategies that allow the unmasking of reactive functionality (e.g. amines and alcohols) prior to monomer coupling, or the removal of less desirable functionality (e.g. alkenes and alkynes) are highly desirable. Hydrogenation / hydrogenolysis procedures would achieve these ends but has not been amenable to DEL chemistry. We report a catalytic hydrogen transfer reaction using Pd / C, HCONH 4 and the micelle-forming surfactant, TPGS-750-M, which gives highly efficient conversions for hydrogenolysis of Cbz-protected amines and benzyl protected alcohols and hydrogenation of nitros, halides, nitriles, aldehydes, alkenes and alkynes. Application to multicycle synthesis of an encoded compound was fully compatible with DNA-amplification and sequencing, demonstrating its applicability to DEL synthesis. This method will enable synthetic DEL sequences using orthogonal protecting groups.
Samir Bouayad-Gervais; Daniel J. St-Cyr; Mathieu Courcelles; Éric Bonneil; Florence H. Gohard; Pierre Thibault; William C. Earnshaw; Mike Tyers Pept. Sci., 2022, e24254 https://doi.org/10.1002/pep2.24254AbstractGenetically-encoded cyclic peptide libraries allow rapid in vivo screens for inhibitors of any target protein of interest. In particular, the Split Intein Circular Ligation of Protein and Peptides (SICLOPPS) system exploits spontaneous protein splicing of inteins to produce intracellular cyclic peptides. A previous SICLOPPS screen against Aurora B kinase, which plays a critical role during chromosome segregation, identified several candidate inhibitors that we sought to recapitulate by chemical synthesis. We describe the syntheses of cyclic peptide hits and analogs via solution-phase macrocyclization of side chain-protected linear peptides obtained from standard solid-phase peptide synthesis. Cyclic peptide targets, including cyclo-[CTWAR], were designed to match both the variable portions and conserved cysteine residue of their genetically-encoded counterparts. Synthetic products were characterized by tandem high-resolution mass spectrometry to analyze a combination of exact mass, isotopic pattern, and collisional dissociation-induced fragmentation pattern. The latter analyses facilitated the distinction between targets and oligomeric side products, and served to confirm peptidic sequences in a manner that can be readily extended to analyses of complex biological samples. This alternative chemical synthesis approach for cyclic peptides allows cost-effective validation and facile chemical elaboration of hit candidates from SICLOPPS screens.
Hongyao Zhu; Timothy L. Foley; Justin I. Montgomery; Robert V. Stanton J. Chem. Inf. Model., 2022, 62(9), 2239-2247 https://doi.org/10.1021/acs.jcim.1c00986AbstractBy analyzing data sets of replicate DNA-Encoded Library (DEL) selections, an approach for estimating the noise level of the experiment has been developed. Using a logarithm transformation of the number of counts associated with each compound and a subset of compounds with the highest number of counts, it is possible to assess the quality of the data through normalizing the replicates and use this same data to estimate the noise in the experiment. The noise level is seen to be dependent on sequencing depth as well as specific selection conditions. The noise estimation is independent of any cutoff used to remove low frequency compounds from the data analysis. The removal of compounds with only 1–5 read counts greatly reduces some of the challenges encountered in DEL data analysis as it can reduce the data set by greater than 100-fold without impacting the interpretation of the results.
Raphael I. Benhamou; Blessy M. Suresh; Yuquan Tong; Wesley G. Cochrane; Valerie Cavett; Simon Vezina-Dawod; Daniel Abegg; Jessica L. Childs-Disney; Alexander Adibekian; Brian M. Paegel; Matthew D. Disney Proc. Natl. Acad. Sci. USA, 2022, 119(6), e2114971119 https://doi.org/10.1073/pnas.2114971119AbstractNature evolves molecular interaction networks through persistent perturbation and selection, in stark contrast to drug discovery, which evaluates candidates one at a time by screening. Here, nature’s highly parallel ligand-target search paradigm is recapitulated in a screen of a DNA-encoded library (DEL; 73,728 ligands) against a library of RNA structures (4,096 targets). In total, the screen evaluated ∼300 million interactions and identified numerous bona fide ligand–RNA three-dimensional fold target pairs. One of the discovered ligands bound a 5′GAG/3′CCC internal loop that is present in primary microRNA-27a (pri-miR-27a), the oncogenic precursor of microRNA-27a. The DEL-derived pri-miR-27a ligand was cell active, potently and selectively inhibiting pri-miR-27a processing to reprogram gene expression and halt an otherwise invasive phenotype in triple-negative breast cancer cells. By exploiting evolutionary principles at the earliest stages of drug discovery, it is possible to identify high-affinity and selective target–ligand interactions and predict engagements in cells that short circuit disease pathways in preclinical disease models.
Qigui Nie; Shuting Zhong; Yangfeng Li; Gong Zhang; Yizhou Li Chin. Chem. Lett., 2022, 33(5), 2559-2563 https://doi.org/10.1016/j.cclet.2021.09.041AbstractDNA-encoded chemical library (DEL) represents an emerging drug discovery technology to construct compound libraries with abundant chemical combinations. While drug-like small molecule DELs facilitate the discovery of binders against targets with defined pockets, macrocyclic DELs harboring extended scaffolds enable targeting of the protein–protein interaction (PPI) interface. We previously demonstrated the design of the first-generation DNA-encoded multiple display based on a constant macrocyclic scaffold, which harvested binders against difficult targets such as tumor necrosis factor-α (TNF-α). Here, we developed a novel strategy which utilized four orthogonal amine-protecting groups on DNA, to explore larger chemical combinations on the same constant macrocyclic scaffold, following the parallel paradigm to mimic the versatile antibody-like multivalent epitope recognition patterns. We successfully integrated these orthogonal protecting groups with acylation and made a mock second-generation DNA-encoded display combination. This work illustrates a strategy to produce larger encoded multiple display on a constant macrocyclic scaffold, which could potentially discover binders with enhanced affinity to clinically significant PPI targets.
Huiya Zhang; Shiyu Chen RSC Chem. Biol., 2022, 3, 18-31 https://doi.org/10.1039/D1CB00154JAbstractIn contrast to the major families of small molecules and antibodies, cyclic peptides, as a family of synthesizable macromolecules, have distinct biochemical and therapeutic properties for pharmaceutical applications. Cyclic peptide-based drugs have increasingly been developed in the past two decades, confirming the common perception that cyclic peptides have high binding affinities and low metabolic toxicity as antibodies, good stability and ease of manufacture as small molecules. Natural peptides were the major source of cyclic peptide drugs in the last century, and cyclic peptides derived from novel screening and cyclization strategies are the new source. In this review, we will discuss and summarize 18 cyclic peptides approved for clinical use in the past two decades to provide a better understanding of cyclic peptide development and to inspire new perspectives. The purpose of the present review is to promote efforts to resolve the challenges in the development of cyclic peptide drugs that are more effective.
Yiran Huang; Yizhou Li; Xiaoyu Li Nat. Chem., 2022, 14, 129-140 https://doi.org/10.1038/s41557-021-00877-xAbstractDNA-encoded chemical libraries (DELs) have emerged as a powerful technology in drug discovery. The wide adoption of DELs in the pharmaceutical industry and the rapid advancements of DEL-compatible chemistry have further fuelled its development and applications. In general, a DEL has been considered as a massive binding assay to identify physical binders for individual protein targets. However, recent innovations demonstrate the capability of DELs to operate in the complex milieu of biological systems. In this Perspective, we discuss the recent progress in using DNA-encoded chemical libraries to interrogate complex biological targets and their potential to identify structures that elicit function or possess other useful properties. Future breakthroughs in these aspects are expected to catapult DEL to become a momentous technology platform not only for drug discovery but also to explore fundamental biology.
Jonathan C. Chen; Jonathan P. Chen; Max W. Shen; Michael Wornow; Minwoo Bae; Wei-Hsi Yeh; Alvin Hsu; David R. Liu Nat. Commun., 2022, 13, 4541 https://doi.org/10.1038/s41467-022-31955-4AbstractIn vitro selection queries large combinatorial libraries for sequence-defined polymers with target binding and reaction catalysis activity. While the total sequence space of these libraries can extend beyond 1022 sequences, practical considerations limit starting sequences to ≤~1015 distinct molecules. Selection-induced sequence convergence and limited sequencing depth further constrain experimentally observable sequence space. To address these limitations, we integrate experimental and machine learning approaches to explore regions of sequence space unrelated to experimentally derived variants. We perform in vitro selections to discover highly side-chain-functionalized nucleic acid polymers (HFNAPs) with potent affinities for a target small molecule (daunomycin KD = 5–65 nM). We then use the selection data to train a conditional variational autoencoder (CVAE) machine learning model to generate diverse and unique HFNAP sequences with high daunomycin affinities (KD = 9–26 nM), even though they are unrelated in sequence to experimental polymers. Coupling in vitro selection with a machine learning model thus enables direct generation of active variants, demonstrating a new approach to the discovery of functional biopolymers.
Meiying Cui; Xueping Zhao; Francesco V. Reddavide; Michelle Patino Gaillez; Stephan Heiden; Luca Mannocci; Michael Thompson; Yixin Zhang Nucleic Acids Research, 2022, gkac672 https://doi.org/10.1093/nar/gkac672AbstractInformation processing functions are essential for organisms to perceive and react to their complex environment, and for humans to analyze and rationalize them. While our brain is extraordinary at processing complex information, winner-take-all, as a type of biased competition is one of the simplest models of lateral inhibition and competition among biological neurons. It has been implemented as DNA-based neural networks, for example, to mimic pattern recognition. However, the utility of DNA-based computation in information processing for real biotechnological applications remains to be demonstrated. In this paper, a biased competition method for nonlinear manipulation and analysis of mixtures of DNA sequences was developed. Unlike conventional biological experiments, selected species were not directly subjected to analysis. Instead, parallel computation among a myriad of different DNA sequences was carried out to reduce the information entropy. The method could be used for various oligonucleotide-encoded libraries, as we have demonstrated its application in decoding and data analysis for selection experiments with DNA-encoded chemical libraries against protein targets.
Samantha Stone; David J. Newman; Steven L. Collettic; Derek S. Tan Nat. Prod. Rep., 2022, 39, 20-32 https://doi.org/10.1039/D1NP00039JAbstractNatural products continue to play a major role in drug discovery, with half of new chemical entities based structurally on a natural product. Herein, we report a cheminformatic analysis of the structural and physicochemical properties of natural product-based drugs in comparison to top-selling brand-name synthetic drugs, and a selection of chemical probes recently discovered from diversity-oriented synthesis libraries. In this analysis, natural product-based drugs covered a broad range of chemical space based on size, polarity, and three-dimensional structure. Natural product-based structures were also more prevalent in top-selling drugs of 2018 compared to 2006. Further, the drugs clustered well according to biosynthetic origins, but less so based on therapeutic classes. Macrocycles occupied distinctive and relatively underpopulated regions of chemical space, while chemical probes largely overlapped with synthetic drugs. This analysis highlights the continued opportunities to leverage natural products and their pharmacophores in modern drug discovery.
Lisa E. J. Douglas; James A. Reihill; Melisa W. Y. Ho; Jeffrey M. Axten; Nino Campobasso; Jessica L. Schneck; Alan R. Rendina; Keith M. Wilcoxen; S. Lorraine Martin Cell Chem. Biol., 2022, 29, 947–957 https://doi.org/10.1016/j.chembiol.2022.02.001AbstractIn cystic fibrosis (CF), excessive furin activity plays a critical role in the activation of the epithelial sodium channel (ENaC), dysregulation of which contributes to airway dehydration, ineffective mucociliary clearance (MCC), and mucus obstruction. Here, we report a highly selective, cell-permeable furin inhibitor, BOS-318, that derives selectivity by eliciting the formation of a new, unexpected binding pocket independent of the active site catalytic triad. Using human ex vivo models, BOS-318 showed significant suppression of ENaC, which led to enhanced airway hydration and an ∼30-fold increase in MCC rate. Furin inhibition also protected ENaC from subsequent activation by neutrophil elastase, a soluble protease dominant in CF airways. Additional therapeutic benefits include protection against epithelial cell death induced by Pseudomonas aeruginosa exotoxin A. Our findings demonstrate the utility of selective furin inhibition as a mutation-agnostic approach that can correct features of CF airway pathophysiology in a manner expected to deliver therapeutic value.
Gong Zhang; Juan Zhang; Yuting Gao; Yangfeng Li; Yizhou Li Expert Opin. Drug Discov., 2022, 17(1), 55-69 https://doi.org/10.1080/17460441.2021.1969359AbstractUndruggable targets refer to clinically meaningful therapeutic targets that are ‘difficult to drug’ or ‘yet to be drugged’ via traditional approaches. Featuring characteristics of lacking defined ligand-binding pockets, non-catalytic protein–protein interaction functional modes and less-investigated 3D structures, these undruggable targets have been targeted with novel therapeutic entities developed with the progress of unconventional drug discovery approaches, such as targeted degradation molecules and display technologies.
Lisa Candish; Karl D. Collins; Gemma C. Cook; James J. Douglas; Adrián Gómez-Suárez; Anais Jolit; Sebastian Keess Chem. Rev., 2022, 122(2), 2907-2980 https://doi.org/10.1021/acs.chemrev.1c00416AbstractIn the pursuit of new pharmaceuticals and agrochemicals, chemists in the life science industry require access to mild and robust synthetic methodologies to systematically modify chemical structures, explore novel chemical space, and enable efficient synthesis. In this context, photocatalysis has emerged as a powerful technology for the synthesis of complex and often highly functionalized molecules. This Review aims to summarize the published contributions to the field from the life science industry, including research from industrial-academic partnerships. An overview of the synthetic methodologies developed and strategic applications in chemical synthesis, including peptide functionalization, isotope labeling, and both DNA-encoded and traditional library synthesis, is provided, along with a summary of the state-of-the-art in photoreactor technology and the effective upscaling of photocatalytic reactions.
Vivian M. Lechner; Manuel Nappi; Patrick J. Deneny; Sarah Folliet; John C. K. Chu; Matthew J. Gaunt Chem. Rev., 2022, 122(2), 1752-1829 https://doi.org/10.1021/acs.chemrev.1c00357AbstractChemically modified biomacromolecules—i.e., proteins, nucleic acids, glycans, and lipids—have become crucial tools in chemical biology. They are extensively used not only to elucidate cellular processes but also in industrial applications, particularly in the context of biopharmaceuticals. In order to enable maximum scope for optimization, it is pivotal to have a diverse array of biomacromolecule modification methods at one’s disposal. Chemistry has driven many significant advances in this area, and especially recently, numerous novel visible-light-induced photochemical approaches have emerged. In these reactions, light serves as an external source of energy, enabling access to highly reactive intermediates under exceedingly mild conditions and with exquisite spatiotemporal control. While UV-induced transformations on biomacromolecules date back decades, visible light has the unmistakable advantage of being considerably more biocompatible, and a spectrum of visible-light-driven methods is now available, chiefly for proteins and nucleic acids. This review will discuss modifications of native functional groups (FGs), including functionalization, labeling, and cross-linking techniques as well as the utility of oxidative degradation mediated by photochemically generated reactive oxygen species. Furthermore, transformations at non-native, bioorthogonal FGs on biomacromolecules will be addressed, including photoclick chemistry and DNA-encoded library synthesis as well as methods that allow manipulation of the activity of a biomacromolecule.
Xing Ling; Weiwei Lu; Lin Miao; Lisa A. Marcaurelle; Xuan Wang; Yun Ding; Xiaojie Lu J. Org. Chem., 2022, 87(4), 1971-1976 https://doi.org/10.1021/acs.joc.1c00670AbstractA group of highly efficient and divergent transformations for constructing multiple DNA-linked chemotypes based on a piperidone core were successfully developed. We reported the first procedure for the synthesis of a DNA-conjugated piperidine intermediate under basic conditions. Subsequently, this substructure was subjected to additional reactions to generate several privileged scaffolds, including 4-aminopiperidine, fused [1,2,4]triazolo[1,5-a]pyrimidine, and a quinoline derivative. These transformations paved the way for constructing focused scaffold-based DNA-encoded libraries with druglike properties.
Hansol Choi; Yeongjae Choi; Jaewon Choi; Amos Chungwon Lee; Huiran Yeom; Jinwoo Hyun; Taehoon Ryu; Sunghoon Kwon Nat. Biotech., 2022, 40, 47-53 https://doi.org/10.1038/s41587-021-00988-3AbstractComplex oligonucleotide (oligo) libraries are essential materials for diverse applications in synthetic biology, pharmaceutical production, nanotechnology and DNA-based data storage. However, the error rates in synthesizing complex oligo libraries can be substantial, leading to increment in cost and labor for the applications. As most synthesis errors arise from faulty insertions and deletions, we developed a length-based method with single-base resolution for purification of complex libraries containing oligos of identical or different lengths. Our method—purification of multiplex oligonucleotide libraries by synthesis and selection—can be performed either step-by-step manually or using a next-generation sequencer. When applied to a digital data-encoded library containing oligos of identical length, the method increased the purity of full-length oligos from 83% to 97%. We also show that libraries encoding the complementarity-determining region H3 with three different lengths (with an empirically achieved diversity >106) can be simultaneously purified in one pot, increasing the in-frame oligo fraction from 49.6% to 83.5%.
Balayeshwanth R. Vummidi; Lluc Farrera-Soler; Jean-Pierre Daguer; Millicent Dockerill; Sofia Barluenga; Nicolas Winssinger Nat. Chem., 2022, 14, 141-152 https://doi.org/10.1038/s41557-021-00829-5AbstractDNA-encoded library technologies enable the screening of synthetic molecules but have thus far not tapped into the power of Darwinian selection with iterative cycles of selection, amplification and diversification. Here we report a simple strategy to rapidly assemble libraries of conformationally constrained peptides that are paired in a combinatorial fashion (suprabodies). We demonstrate that the pairing can be shuffled after each amplification cycle in a process similar to DNA shuffling or mating to regenerate diversity. Using simulations, we show the benefits of this recombination in yielding a more accurate correlation of selection fitness with affinity after multiple rounds of selection, particularly if the starting library is heterogeneous in the concentration of its members. The method was validated with selections against streptavidin and applied to the discovery of PD-L1 binders. We further demonstrate that the binding of self-assembled suprabodies can be recapitulated by smaller (∼7 kDa) synthetic products that maintain the conformational constraint of the peptides.
Ayun Luo; Hongxia Zhou; Qi Zhou; Qini Hua; Xue Zhao; Xiaobing Yu; Kexin Yang; Yunjin Hu Bioconjugate Chem., 2022, 33(12), 2299-2306 https://doi.org/10.1021/acs.bioconjchem.2c00471Abstract1-Iodoalkynes and 1,3-diynes are versatile chemical intermediates and pharmaceutically valuable ingredients. In this study, copper mediated on-DNA alkyne iodination and Cadiot-Chodkiewicz coupling are developed for the first time. This generates diverse, systematic, and unprecedented topographic structural features, which could be invaluable as molecular recognition agents for drug discovery in DEL screening.
Yurong Shen; Guanyu Yang; Wei Huang; Alex Shaginian; Qian Lin; Jinqiao Wan; Jin Li; Yun Deng; Guansai Liu Org. Lett., 2022, 24(14), 2650-2654 https://doi.org/10.1021/acs.orglett.2c00697AbstractHerein, we report an on-DNA photoredox-mediated deaminative alkylation method for diversifying DNA-tagged acrylamide substrate with amine-derived radicals. The radicals can be conveniently generated from sterically hindered primary amines, and the deaminative alkylation can tolerate a broad array of radical precursors. Furthermore, the methodology is applicable to Boc-protected diamines, free amino acids, and aryl halides, which bear functional groups enabling additional rounds of diversification. The method is believed to offer a high potential for constructing DNA-encoded libraries, as was demonstrated by the production of a mock library in a 2 × 3 matrix format and confirmation of DNA stability by UPLC–MS and qPCR experiments.
Yuhan Gui; Clara Shania Wong; Guixian Zhao; Chao Xie; Rui Hou; Yizhou Li; Gang Li; Xiaoyu Li ACS Omega, 2022, 7(13), 11491-11500 https://doi.org/10.1021/acsomega.2c01152AbstractDNA-encoded library (DEL) is an efficient high-throughput screening technology platform in drug discovery and is also gaining momentum in academic research. Today, the majority of DELs are assembled and encoded with double-stranded DNA tags (dsDELs) and has been selected against numerous biological targets; however, dsDELs are not amendable to some of the recently developed selection methods, such as the cross-linking-based selection against immobilized targets and live-cell-based selections, which require DELs encoded with single-stranded DNAs (ssDELs). Herein, we present a simple method to convert dsDELs to ssDELs using exonuclease digestion without library redesign and resynthesis. We show that dsDELs could be efficiently converted to ssDELs and used for affinity-based selections either with purified proteins or on live cells.
Katherine S. Lim; Andrew G. Reidenbach; Bruce K. Hua; Jeremy W. Mason; Christopher J. Gerry; Paul A. Clemons; Connor W. Coley arXiv, 2021, 2108.12471 https://arxiv.org/abs/2108.12471AbstractDNA-encoded library (DEL) screening and quantitative structure-activity relationship (QSAR) modeling are two techniques used in drug discovery to find small molecules that bind a protein target. Applying QSAR modeling to DEL data can facilitate the selection of compounds for off-DNA synthesis and evaluation. Such a combined approach has been shown recently by training binary classifiers to learn DEL enrichments of aggregated "disynthons" to accommodate the sparse and noisy nature of DEL data. However, a binary classifier cannot distinguish between different levels of enrichment, and information is potentially lost during disynthon aggregation. Here, we demonstrate a regression approach to learning DEL enrichments of individual molecules using a custom negative log-likelihood loss function that effectively denoises DEL data and introduces opportunities for visualization of learned structure-activity relationships (SAR). Our approach explicitly models the Poisson statistics of the sequencing process used in the DEL experimental workflow under a frequentist view. We illustrate this approach on a dataset of 108k compounds screened against CAIX, and a dataset of 5.7M compounds screened against sEH and SIRT2. Due to the treatment of uncertainty in the data through the negative log-likelihood loss function, the models can ignore low-confidence outliers. While our approach does not demonstrate a benefit for extrapolation to novel structures, we expect our denoising and visualization pipeline to be useful in identifying SAR trends and enriched pharmacophores in DEL data. Further, this approach to uncertainty-aware regression is applicable to other sparse or noisy datasets where the nature of stochasticity is known or can be modeled; in particular, the Poisson enrichment ratio metric we use can apply to other settings that compare sequencing count data between two experimental conditions.
Andreas Brunschweiger Bioorg. Med. Chem., 2021, 116582 https://doi.org/10.1016/j.bmc.2021.116582AbstractAt first glance, the idea of providing 500 Da drug-like compounds with a molecular identifier that is about 100 times larger and a biomacromolecule takes some getting used to. However, over the past decade, and especially in recent years, DNA-encoded libraries, or DELs, have gained increasing attention from both academic and industrial researchers as a generic small molecule screening technology for a wide range of biological targets. There are several reasons for massively increased research activities in this field. These are ranging from (a) purely technical underpinnings, namely the ready availability of synthetic DNA oligonucleotides, affordable sequencing technology, and computational power to analyze massively large data sets; (b) breakthrough research by a few pioneers that have demonstrated the proof of concept; (c) the promise of the technology, namely harnessing the data storage capability of DNA for efficiently handling and screening vast chemical space on biological targets by a conceptually simple selection assay; to (d) relevance, given the lack of drug development entry points for a large number of disease-relevant biological targets.
Rasel A. Al-Amin; Lars Johansson; Eldar Abdurakhmanov; Nils Landegren; Liza Löf; Linda Arngården; Andries Blokzijl; Richard Svensson; Maria Hammond; Peter Lönn; Johannes Haybaeck; Masood Kamali-Moghaddam; Annika Jenmalm Jensen; Helena Danielson; Per Artursson; Thomas Lundbäck; Ulf Landegren Research Square, 2021 https://doi.org/10.21203/rs.3.rs-1109577/v1AbstractDrugs are designed to bind their target proteins in physiologically relevant tissues and organs to modulate biological functions and elicit desirable clinical outcomes. Information about target engagement at cellular and subcellular resolution is therefore critical for guiding compound optimization in drug discovery, and for probing resistance mechanisms to targeted therapies in clinical samples. We describe a target engagement-mediated amplification (TEMA) technology, where oligonucleotide-conjugated drugs are used to visualize and measure target engagement in situ, amplified via rolling-circle replication of circularized oligonucleotide probes. We illustrate the TEMA technique using dasatinib and gefitinib, two kinase inhibitors with distinct selectivity profiles. In vitro binding by dasatinib probe to arrays of displayed proteins accurately reproduced known selectivity profiles, while their differential binding to a panel of fixed adherent cells agreed with expectations from expression profiles of the cells. These findings were corroborated by competition experiments using kinase inhibitors with overlapping and non-overlapping target specificities, and translated to pathology tissue sections. We also introduce a proximity ligation variant of TEMA in which these drug-DNA conjugates are combined with antibody-DNA conjugates to selectively investigate binding to specific target proteins of interest. This form of the assay serves to improve resolution of binding to on- and off-target proteins. In conclusion, TEMA has the potential to aid in drug development and clinical routine by conferring valuable insights in drug-target interactions at spatial resolution in protein arrays, cells and tissues.
Kate A. Nicastri; Soren A. Zappia; Jared C. Pratt; Julia M. Duncan; Ilia A. Guzei; Israel Fernández; Jennifer M. Schomaker ChemRxiv, 2021 https://doi.org/10.26434/chemrxiv-2021-0g6f0AbstractMethods for rapid preparation of densely functionalized and stereochemically complex N-heterocyclic scaffolds are in demand for exploring potential new bioactive chemical space. This work describes experimental and computational studies to better understand the features of aziridinium ylides as intermediates for the synthesis of highly substituted dehydromorpholines. The development of this chemistry has enabled the extension of aziridinium ylide chemistry to the concomitant formation of both a C–N and a C–O bond in a manner that preserves the stereochemical information embedded in the substrate. The chemistry is tolerant of a wide range of functionalities that can be employed for DNA-encoded library (DEL) synthesis to prepare diverse libraries of heterocycles with potential bioactivity. In addition, we have uncovered several key insights that describe the importance of steric effects, rotational barriers around the C–N bond of the aziridinium ylide, and non-covalent interactions (NCIs) on the ultimate reaction outcome. These critical insights will assist in the further development of this chemistry to generate novel and complex N-heterocycles that will further expand complex amine chemical space.
Xiang Zhang; Shuning Zhang; Songfeng Zhao; Xuan Wang; Bo Liu; Hongtao Xu Front. Chem., 2021, 9, 774977 https://doi.org/10.3389/fchem.2021.774977AbstractClick chemistry is perhaps the most powerful synthetic toolbox that can efficiently access the molecular diversity and unique functions of complex natural products up to now. It enables the ready synthesis of diverse sets of natural product derivatives either for the optimization of their drawbacks or for the construction of natural product-like drug screening libraries. This paper showcases the state-of-the-art development of click chemistry in natural product modification and summarizes the pharmacological activities of the active derivatives as well as the mechanism of action. The aim of this paper is to gain a deep understanding of the fruitful achievements and to provide perspectives, trends, and directions regarding further research in natural product medicinal chemistry.
Ann E. Cleves; Stephen R. Johnson; Ajay N. Jain J. Chem. Inf. Model., 2021, 61(12), 5948-5966 https://doi.org/10.1021/acs.jcim.1c01382AbstractWe present results on the extent to which physics-based simulation (exemplified by FEP+) and focused machine learning (exemplified by QuanSA) are complementary for ligand affinity prediction. For both methods, predictions of activity for LFA-1 inhibitors from a medicinal chemistry lead optimization project were accurate within the applicable domain of each approach. A hybrid model that combined predictions by both approaches by simple averaging performed better than either method, with respect to both ranking and absolute pKi values. Two publicly available FEP+ benchmarks, covering 16 diverse biological targets, were used to test the generality of the synergy. By identifying training data specifically focused on relevant ligands, accurate QuanSA models were derived using ligand activity data known at the time of the original series publications. Results across the 16 benchmark targets demonstrated significant improvements both for ranking and for absolute pKi values using hybrid predictions that combined the FEP+ and QuanSA predicted affinity values. The results argue for a combined approach for affinity prediction that makes use of physics-driven methods as well as those driven by machine learning, each applied carefully on appropriate compounds, with hybrid prediction strategies being employed where possible.
Simone Brogi; Vincenzo Calderone Preprints, 2021, 2021100242 https://doi.org/10.20944/preprints202110.0242.v1AbstractThe huge advancement of Internet web facilities as well as the progress in computing and algorithm development, along with current innovations regarding high-throughput techniques enables the scientific community to gain access to biological datasets, clinical data, and several databases containing billions of information concerning scientific knowledge. Consequently, during the last decade the system for managing, analyzing, processing and extrapolating information from scientific data has been considerably modified in several fields including the medical one. As a consequence of the mentioned scenario, scientific vocabulary was enriched by novel lexicons such as Machine Learning (ML)/Deep Learning (DL) and overall Artificial Intelligence (AI). Beyond the terminology, these computational techniques are revolutionizing the scientific research in drug discovery pitch, from the preclinical studies to clinical investigation. Interestingly, between preclinical and clinical research, the translational research is benefitting from computer-based approaches, transforming the design and execution of the translational research, resulting in breakthroughs for advancing human health. Accordingly, in this review article, we analyze the most advanced applications of AI in translational medicine, providing an up-to-date outlook regarding this emerging field.
Edward B. Pimentel; Trenton M. Peters-Clarke; Joshua J. Coon; Jeffrey D. Martell J. Am. Chem. Soc., 2021, 143(50), 21402-21409 https://doi.org/10.1021/jacs.1c10757AbstractWe report DNA-scaffolded synergistic catalysis, a concept that combines the diverse reaction scope of synergistic catalysis with the ability of DNA to precisely preorganize abiotic groups and undergo stimuli-triggered conformational changes. As an initial demonstration of this concept, we focus on Cu-TEMPO-catalyzed aerobic alcohol oxidation, using DNA as a scaffold to hold a copper cocatalyst and an organic radical cocatalyst (TEMPO) in proximity. The DNA-scaffolded catalyst maintained a high turnover number upon dilution and exhibited 190-fold improvement in catalyst turnover number relative to the unscaffolded cocatalysts. By incorporating the cocatalysts into a DNA hairpin-containing scaffold, we demonstrate that the rate of the synergistic catalytic reaction can be controlled through a reversible DNA conformational change that alters the distance between the cocatalysts. This work demonstrates the compatibility of synergistic catalytic reactions with DNA scaffolding, opening future avenues in reaction discovery, sensing, responsive materials, and chemical biology.
Ying Yang; Kun Yao; Matthew P. Repasky; Karl Leswing; Robert Abel; Brian K. Shoichet; Steven V. Jerome J. Chem. Theory Comput., 2021, 17, 11, 7106-7119 https://doi.org/10.1021/acs.jctc.1c00810AbstractWith the advent of make-on-demand commercial libraries, the number of purchasable compounds available for virtual screening and assay has grown explosively in recent years, with several libraries eclipsing one billion compounds. Today’s screening libraries are larger and more diverse, enabling the discovery of more-potent hit compounds and unlocking new areas of chemical space, represented by new core scaffolds. Applying physics-based in silico screening methods in an exhaustive manner, where every molecule in the library must be enumerated and evaluated independently, is increasingly cost-prohibitive. Here, we introduce a protocol for machine learning-enhanced molecular docking based on active learning to dramatically increase throughput over traditional docking. We leverage a novel selection protocol that strikes a balance between two objectives: (1) identifying the best scoring compounds and (2) exploring a large region of chemical space, demonstrating superior performance compared to a purely greedy approach. Together with automated redocking of the top compounds, this method captures almost all the high scoring scaffolds in the library found by exhaustive docking. This protocol is applied to our recent virtual screening campaigns against the D4 and AMPC targets that produced dozens of highly potent, novel inhibitors, and a blind test against the MT1 target. Our protocol recovers more than 80% of the experimentally confirmed hits with a 14-fold reduction in compute cost, and more than 90% of the hit scaffolds in the top 5% of model predictions, preserving the diversity of the experimentally confirmed hit compounds.
Huiying Xu; Weijie Chen; Mengyao Bian; Hongtao Xu; Hui Gao; Ting Wang; Zhi Zhou; Wei Yi ACS Catal. 2021, 11, 14694-14701 https://doi.org/10.1021/acscatal.1c04508AbstractCp*Rh(III)-catalyzed [4 + 3] annulation of N-methoxy amides for the direct assembly of seven-numbered 2H-azepin-2-one frameworks has been realized with gem-difluorocyclopropenes acting as innovative β-monofluorinated three sp2 carbon sources. Either annular arylamides or linear acrylamides with the embedment of various functional groups, including DNA-tagged substrates, were found to be compatible with the established [4 + 3] reaction mode. A redox-neutral Rh(III)–Rh(V)–Rh(III) catalytic cycle, specifically via HOAc-assisted tandem site-/regioselective oxidative addition/reductive elimination/C–F bond cleavage-enabled ring-scission involving the unprecedented olefinic C(sp2)–C(sp2) bond cleavage, has been deduced based on experimental and computational mechanistic studies. Taken together, our findings not only identified gem-difluorocyclopropenes as potent and efficient coupling partners for C–H activation development but also provided a sound basis for the organic integration of transition-metal-catalyzed C–H functionalization with cyclopropene and fluorine chemistries.
Adrián Gironda-Martínez; Émile M. D. Gorre; Luca Prati; Jean-François Gosalbes; Sheila Dakhel; Samuele Cazzamalli; Florent Samain; Etienne J. Donckele; Dario Neri J. Med. Chem., 2021, 64(23), 17496-17510 https://doi.org/10.1021/acs.jmedchem.1c01693AbstractInterleukin-2 (IL2) is a pro-inflammatory cytokine that plays a crucial role in immunity, which is increasingly being used for therapeutic applications. There is growing interest in developing IL2-based therapeutics which do not interact with the alpha subunit of the IL2 receptor (CD25) as this protein is primarily found on immunosuppressive regulatory T cells (Tregs). Screenings of a new DNA-encoded library, comprising 669,240 members, provided a novel series of IL2 ligands, subsequently optimized by medicinal chemistry. One of these molecules (compound 18) bound to IL2 with a dissociation constant of 0.34 μM was able to form a kinetically stable complex with IL2 in size-exclusion chromatography and recognized the CD25-binding site as evidenced by competition experiments with the NARA1 antibody. Compound 18 and other members of the series may represent the starting point for the discovery of potent small-molecule modulators of IL2 activity, abrogating the binding to CD25.
Jialu Wang; Biswaranjan Pani; Ilhan Gokhan; Xinyu Xiong; Alem W. Kahsai; Haoran Jiang; Seungkirl Ahn; Robert J. Lefkowitz; Howard A Rockman Mol. Pharmacol., 2021, 100(6), 568-579 https://doi.org/10.1124/molpharm.121.000359Abstractβ1 adrenergic receptors (β1ARs) are central regulators of cardiac function and a drug target for cardiac disease. As a member of G protein-coupled receptor family, β1ARs activate cellular signaling by primarily coupling to Gs proteins to activate adenylyl cyclase and cAMP-dependent pathways, and the multifunctional adaptor-transducer protein β-arrestin. Carvedilol, a traditional β-blocker widely used in treating high blood pressure and heart failure by blocking βAR-mediated G-protein activation, can selectively stimulate Gs-independent β-arrestin signaling of βARs, a process known as β-arrestin-biased agonism. Recently a DNA-encoded small molecule library screen against agonist-occupied β2 adrenergic receptors (β2AR) identified Compound-6 (Cmpd-6) to be a positive allosteric modulator for agonists on β2ARs. Intriguingly, it was further discovered that Cmpd-6 is positively cooperative with the β-arrestin biased ligand carvedilol at β2ARs. Here we describe the surprising finding that at β1ARs, unlike the case of β2ARs, Cmpd-6 is cooperative only with carvedilol and not agonists. Cmpd-6 increases the binding affinity of carvedilol for β1ARs and potentiates carvedilol-stimulated, β-arrestin-dependent β1AR signaling such as epidermal growth factor receptor transactivation and extracellular signal-regulated kinase activation, while having no effect on Gs-mediated cAMP generation. In vivo, Cmpd-6 enhances the anti-apoptotic cardioprotective effect of carvedilol in response to myocardial ischemia/reperfusion injury. This anti-apoptotic role of carvedilol is dependent on β-arrestins, since it is lost in mice with myocyte-specific deletion of β-arrestins. Our findings demonstrate that Cmpd-6 is a selective β-arrestin-biased allosteric modulator of β1ARs and highlight its potential clinical utility in enhancing carvedilol-mediated cardioprotection against ischemic injury.
Pinwen Cai; Lukas A. Schneider; Cedric Stress; Dennis Gillingham Org. Lett., 2021, 23(22), 8772-8776 https://doi.org/10.1021/acs.orglett.1c03262AbstractDNA-encoded library (DEL) technology uses DNA tags to track the synthetic history of individual members in a split-and-pool combinatorial synthesis scheme. DEL synthesis hinges on robust methodologies that tolerate combinatorial synthesis schemes while not destroying the information in DNA. We introduce here a DEL-compatible reaction that assembles a boron-containing pyridazine heterocycle. The heterocycle is unique because it can engage in reversible covalent interactions with alcohols─a feature that, until now, has not been deliberately engineered into DELs
Jessica S. Graham; James H. Hunter; Michael J. Waring J. Org. Chem., 2021, 86(23), 17257-17264 https://doi.org/10.1021/acs.joc.1c02325AbstractDNA-encoded libraries are a very efficient means of identifying ligands for protein targets in high throughput. To fully maximize their use, it is essential to be able to carry out efficient reactions on DNA-conjugated substrates. Arylamines are privileged motifs in druglike molecules, and methods for their incorporation into DNA-encoded libraries are highly desirable. One of the preferred methods for their preparation, the Buchwald–Hartwig coupling, does not perform well on DNA conjugates using current approaches. We report the application of our recently developed micellar technology for on-DNA chemistry to the Buchwald–Hartwig reaction. Optimization of conditions led to a robust, high-yielding method for the synthesis of DNA-conjugated aryl and heteroarylamines, which is broad in substrate scope for both the arylamine and the DNA-conjugated aryl halide and is fully compatible with DNA-encoding and decoding procedures. This method will enable the preparation of diverse, high-fidelity libraries of biarylamines.
Juan Zhang; Xianfeng Li; Haimei Wei; Yangfeng Li; Gong Zhang; Yizhou Li Org. Lett., 2021, 23(21), 8429-8433 https://doi.org/10.1021/acs.orglett.1c03145AbstractThe construction of chemical libraries containing polysubstituted pyrazoline scaffolds is highly desirable for the discovery of novel chemical ligands for biological targets. Herein, we report a sequential DNA-encoded synthesis strategy for polysubstituted pyrazoline heterocycles, which fuses a broad panel of aldehydes, aryl amines, and alkenes as building blocks. Furthermore, mock library synthesis and selection demonstrated the ability of the method to produce DNA-encoded focused libraries with highly functionalized pyrazoline cores.
James H. Hunter; Marco Potowski; Harriet A. Stanway-Gordon; Andrew Madin; Garry Pairaudeau; Andreas Brunschweiger; Michael J. Waring J. Org. Chem., 2021, 86(24), 17930-17935 https://doi.org/10.1021/acs.joc.1c02259AbstractDNA-encoded libraries (DELs) offer great promise for the discovery of new ligands for proteins. Many current reactions used for DEL synthesis do not proceed efficiently over a wide range of substrates. Combining a diverse array of multicomponent reactions with micellar-promoted Suzuki–Miyaura cross-coupling provides a strategy for synthesizing highly diverse DELs with exceptionally high fidelity. These results demonstrate that the micellar Suzuki–Miyaura reaction has exceptional functional group tolerance and broad applicability.
Weilong Liu; Emma E. Watson; Nicolas Winssinger Helv. Chim. Acta, 2021, 104, e202100179 https://doi.org/10.1002/hlca.202100179AbstractBiomedical research has been empowered by tools that enable spatial and temporal control of biological systems. These have predominantly come from photocaged bioactive molecules (optochemical control; N. Ankenbruck, T. Courtney, Y. Naro, A. Deiters, Angew. Chem. Int. Ed. 2018, 57, 2768–2798) and light-dependent proteins (optogenetic control; L. Fenno, O. Yizhar, K. Deisseroth, Annu. Rev. Neurosci. 2011, 34, 389–412). Recent advances in photocatalysis offer the opportunity to amplify these strategies by providing new dimension of biorthogonality. Photocatalysis also empowers bioconjugation with unprecedented reactivities enabling new crosslinking chemistry or biomolecule functionalization, while merging photocatalysis with biocatalysis extends the scope of both of these powerful classes of transformation.
Wesley G. Cochrane; Patrick R. Fitzgerald; Brian M. Paegel ACS Chem. Biol., 2021, 16(12), 2752-2756 https://doi.org/10.1021/acschembio.1c00714AbstractThe global rise of multidrug resistant infections poses an imminent, existential threat. Numerous pipelines have failed to convert biochemically active molecules into bona fide antibacterials, owing to a lack of chemical material with antibacterial-like physical properties in high-throughput screening compound libraries. Here, we demonstrate scalable design and synthesis of an antibacterial-like solid-phase DNA-encoded library (DEL, 7488 members) and facile hit deconvolution from whole-cell Escherichia coli and Bacillus subtilis cytotoxicity screens. The screen output identified two low-micromolar inhibitors of B. subtilis growth and recapitulated known structure–activity relationships of the fluoroquinolone antibacterial class. This phenotypic DEL screening strategy is also potentially applicable to adherent cells and will broadly enable the discovery and optimization of cell-active molecules.
Daniella Bar-Lev; Itai Orr; Omer Sabary; Tuvi Etzion; Eitan Yaakobi arXiv, 2021, 2109.00031 https://arxiv.org/abs/2109.00031AbstractThe concept of DNA storage was first suggested in 1959 by Richard Feynman who shared his vision regarding nanotechnology in the talk "There is plenty of room at the bottom". Later, towards the end of the 20-th century, the interest in storage solutions based on DNA molecules was increased as a result of the human genome project which in turn led to a significant progress in sequencing and assembly methods. DNA storage enjoys major advantages over the well-established magnetic and optical storage solutions. As opposed to magnetic solutions, DNA storage does not require electrical supply to maintain data integrity and is superior to other storage solutions in both density and durability. Given the trends in cost decreases of DNA synthesis and sequencing, it is now acknowledged that within the next 10-15 years DNA storage may become a highly competitive archiving technology and probably later the main such technology. With that said, the current implementations of DNA based storage systems are very limited and are not fully optimized to address the unique pattern of errors which characterize the synthesis and sequencing processes. In this work, we propose a robust, efficient and scalable solution to implement DNA-based storage systems. Our method deploys Deep Neural Networks (DNN) which reconstruct a sequence of letters based on imperfect cluster of copies generated by the synthesis and sequencing processes. A tailor-made Error-Correcting Code (ECC) is utilized to combat patterns of errors which occur during this process. Since our reconstruction method is adapted to imperfect clusters, our method overcomes the time bottleneck of the noisy DNA copies clustering process by allowing the use of a rapid and scalable pseudo-clustering instead. Our architecture combines between convolutions and transformers blocks and is trained using synthetic data modelled after real data statistics.
Jingjing Qi; Sixiu Liu; Mengnisa Seydimemet; Xuan Wang; Xiaojie Lu Bioconjugate Chem., 2021, 32(11), 2290-2294 https://doi.org/10.1021/acs.bioconjchem.1c00427AbstractDNA-encoded library (DEL) technology provided a powerful screening platform for identifying potential bioactive small molecules with high affinity to biologically interesting targets. Essential to a successful DEL campaign are the drug-like small molecular moieties of DNA-encoded libraries with expanded chemical space. Our laboratory has been working on developing and producing novel DNA-encoded libraries that complement current reported DELs. Herein, we demonstrated a general set of DNA-compatible reactions that enable the preparation of pyrrole-based DNA-encoded libraries in which the DNA tags are linked to the N position of the pyrrole central core. Further diversification could be rapidly incorporated into the pyrrole scaffold by robust iodination and Suzuki coupling reactions.
Wendy Warr ChemRxiv, 2021 https://doi.org/10.33774/chemrxiv-2021-x5sj7AbstractThe virtual workshop took place on May 18-20, 2021. It was a follow-up from the December 2020 NIH Workshop on Ultra Large Chemistry Databases. A major theme emerging from the December 2020 workshop was the fact that all the databases of a billion or more structures are virtual. For each virtual molecule the question then arises of whether, or how, it can be synthesized. The organizers therefore assembled speakers to give presentations about how reaction-related data are represented, captured, managed in databases, analyzed, used for drug design, applied in robotics, and exchanged locally as well as globally. This report summarizes talks from 27 practitioners in the reaction informatics field. The aim is to represent as accurately as possible the information that was delivered by the speakers; the report does not seek to be evaluative. The themes, in the order used for this report, were reaction representations, file formats, and standards; sources of reaction data; AI and machine learning applications of reaction-related data in de novo drug design, synthetic accessibility, synthesis planning, reaction prediction etc.; and automation and progression toward autonomous synthesis.
Louise Plais; Jörg Scheuermann RSC Chem. Biol., 2021, 3(1), 7-17 https://doi.org/10.1039/D1CB00161BAbstractWhile macrocyclic peptides are extensively researched for therapeutically relevant protein targets, DNA-encoded chemical libraries (DELs) are developed at a quick pace to discover novel small molecule binders. The combination of both fields has been explored since 2004 and the number of macrocyclic peptide DELs is steadily increasing. Macrocycles with high affinity and potency were identified for diverse classes of proteins, revealing DEL's huge potential. By giving a historical perspective, we would like to review the methods which permitted the rise of macrocyclic peptide DELs, describe the different DELs which were created and discuss the achievements and challenges of this emerging field.
Sebastian Oehler; Louise Plais; Gabriele Bassi; Deri Nario; Jörg Scheuermann Chem. Commun., 2021 ,57, 12289-12292 https://doi.org/10.1039/D1CC04306DAbstractDNA-encoded chemical libraries (DELs) are increasingly being used for the discovery of protein ligands and can be constructed displaying either one or two molecules at the extremities of the two complementary DNA strands. Here, we describe that DELs, featuring the simultaneous display of two molecules, can be encoded using various types of DNA structures, which go beyond the use of conventional double-stranded DNA fragments. Specifically, we compared dual-display methodologies in hairpin, circular or linear formats in terms of polymerase chain reaction (PCR) amplifiability and performance in affinity capture selections. The methods described in this article highlight the feasibility and modularity of the described encoding strategies and may thus further expand the scope of DNA-encoded chemistry, particularly for the identification of compounds which recognize adjacent epitopes on the surface of target proteins of interest.
Bizhen Lin; Weiwei Lu; Zhen-yu Chen; Yue Zhang; Yin-zhe Duan; Xiaojie Lu; Ming Yan; Xue-jing Zhang Org. Lett., 2021, 23(19), 7381-7385 https://doi.org/10.1021/acs.orglett.1c02562AbstractDNA-encoded library (DEL) technology is a powerful tool in the discovery of bioactive probe molecules and drug leads. Mostly, the success in DEL technology stems from the molecular diversity of the chemical libraries. However, the construction of DELs has been restricted by the idiosyncratic needs and the required low concentration (∼1 mM or less) of the library intermediate. Here, we report visible-light-promoted on-DNA radical coupling reactions via an electron donor–acceptor (EDA) complex and a reversible adsorption to solid support (RASS) strategy. This protocol provides a unique solution to the challenges of increasing the reactivity of highly diluted DNA substrates and reducing the residues of heavy metals from photocatalysts. A series of on-DNA indole sulfone and selenide derivatives were obtained with good to quantitative conversions. It is anticipated that these mild-condition on-DNA radical reactions will significantly improve the chemical diversity of DELs and find widespread utility to DEL construction.
Hiroki Gazuma Pharmacia, 2021, 57(8), 771 https://doi.org/10.14894/faruawpsj.57.8_771AbstractScreening using a DNA-encoded library (DEL) is a useful drug discovery method. DEL is a trillion-scale compound library that binds compounds and DNA and associates compound information with DNA sequences. The DEL compound is assayed against the immobilized target protein and then washed away to select the compound that interacts with the target protein. DNA is excised from these compounds and their sequences are read to identify the chemical structure. Since DNA can be amplified by PCR, it is a great merit that even a very small amount of compound can be evaluated. On the other hand, in order to construct a large-scale DEL, it is necessary to select a reaction that proceeds in water in the presence of alcohols and amines contained in DNA, even under the condition of using a low concentration (1 mM) compound. Therefore, the substrates and reaction conditions that can be used are limited, and there is a problem that the diversity of DEL is difficult to increase, and the development of reaction conditions to solve this problem is required. Coupling reactions using metal catalysts are useful for DEL construction, but are limited to sp2 carbon coupling. Yu et al. Reported the C-H bond activation reaction of sp3 carbon, which is effective for the construction of various DELs, and will be introduced in this volume. This paper introduces the research results based on the following documents.
Kejia Yan; Vivian Triana; Sunil Vasu Kalmady; Kwami Aku-Dominguez; Sharyar Memon; Alex Brown; Russell Greinerbd; Ratmir Derda Chem. Sci., 2021, 12, 14301-14308 https://doi.org/10.1039/D1SC04146KAbstractThe Wittig reaction can be used for late stage functionalization of proteins and peptides to ligate glycans, pharmacophores, and many other functionalities. In this manuscript, we modified 160 000 N-terminal glyoxaldehyde peptides displayed on phage with the Wittig reaction by using a biotin labeled ylide under conditions that functionalize only 1% of the library population. Deep-sequencing of the biotinylated and input populations estimated the rate of conversion for each sequence. This “deep conversion” (DC) from deep sequencing correlates with rate constants measured by HPLC. Peptide sequences with fast and slow reactivity highlighted the critical role of primary backbone amides (N–H) in accelerating the rate of the aqueous Wittig reaction. Experimental measurement of reaction rates and density functional theory (DFT) computation of the transition state geometries corroborated this relationship. We also collected deep-sequencing data to build structure–activity relationship (SAR) models that can predict the DC value of the Wittig reaction. By using these data, we trained two classifier models based on gradient boosted trees. These classifiers achieved area under the ROC (receiver operating characteristic) curve (ROC AUC) of 81.2 ± 0.4 and 73.7 ± 0.8 (90–92% accuracy) in determining whether a sequence belonged to the top 5% or the bottom 5% in terms of its reactivity. This model can suggest new peptides never observed experimentally with ‘HIGH’ or ‘LOW’ reactivity. Experimental measurement of reaction rates for 11 new sequences corroborated the predictions for 8 of them. We anticipate that phage-displayed peptides and related mRNA or DNA-displayed substrates can be employed in a similar fashion to study the substrate scope and mechanisms of many other chemical reactions.
Ranjan Jana; Hasina Mamataj Begam; Enakshi Dinda Chem. Commun., 2021, 57, 10842-10866 https://doi.org/10.1039/D1CC04083AAbstractOwing to the market competitiveness and urgent societal need, an optimum speed of drug discovery is an important criteria for successful implementation. Despite the rapid ascent of artificial intelligence, computational and bioanalytical techniques to accelerate drug discovery in big pharma; organic synthesis of privileged scaffolds predicted in silico for in vitro and in vivo studies is still considered as the rate-limiting step. C−H activation is the latest technology added into an organic chemist’s toolbox for the rapid construction and late-stage modification of functional molecules to achieve required chemical and physical properties. Particularly, elimination of prefunctionalization steps, exceptional functional group tolerability, complexity-to-diversity oriented synthesis, late-stage functionalization of privileged medicinal scaffolds expand the dimension of chemical space. It has immense potential for the rapid synthesis of a library of molecules, structural modification to achieve required pharmacological properties such as absorption, distribution, metabolism, excretion, toxicology (ADMET) and attachment of chemical reporters for proteome profiling, metabolite synthesis, etc. for preclinical studies. Although, heterocycle synthesis, late-stage drug modification, 18F labelling, methylation, etc. via C−H functionalization have been reviewed from synthetic standpoint; a general overview of these protocols from medicinal and drug discovery aspects has not been reviewd. In this feature article, we will discuss recent trends of C−H activation methodologies such as synthesis of medicinal scaffolds through C–H activation/annulation cascade; C–H arylation for sp2–sp2 and sp2–sp3 cross-coupling; C–H borylation/silylation to introduce functional linchpin for further manipulations; C–H amination for N-heterocycle and hydrogen bond acceptor; C–H fluorination/fluoroalkylation to tune polarity and lipophilicity; C–H methylation: methyl magic in drug discovery; peptide modification and macrocyclization for therapeutics and biologics; fluorescent and radiolabelling for bioimaging; bioconjugation for chemical biology studies; drug-metabolite synthesis for biodistribution and exctretion studies; late-stage diversification of drug-molecules to incraese efficacy and safety; cutting-edge DNA encoded library synthesisand improved synthesis of drug molecules via C–H activation in medicinal chemistry and drug discovery.
Jing Chai; Xiaojie Lu; Christopher C. Arico-Muendel; Yun Ding; Michael P. Pollastri Bioconjugate Chem., 2021, 32(9), 1973-1978 https://doi.org/10.1021/acs.bioconjchem.1c00363AbstractEnzymatic catalysis is a highly attractive approach to the DNA encoded library technology (DEL) that has not been widely explored. In this paper, we report an l-threonine aldolase (l-TA)-catalyzed on-DNA aldol reaction to form β-hydroxy-α-amino acids, and its diastereoselectivity determination. l-TAs from three species show good on-DNA aldehyde scope and complementary stereoselectivity. The formed aldol product can be further diversified via various reactions, which demonstrates the utility of this reaction in DEL.
Basilius Sauter; Lukas Schneider; Cedric Stress; Dennis Gillingham Bioorg. Med. Chem., 2021, 116508 https://doi.org/10.1016/j.bmc.2021.116508AbstractDNA encoded libraries have became an essential hit-finding tool in early drug discovery. Revent advances in synthetic methods for DNA encoded libraries have expanded the available chemical space, but precisely how each type of chemistry affects the DNA is unstudied. Available assays to quantify the damage are limited to write efficiency, where the ability to ligate DNA onto a working encoded library strand is measured, or qPCR is performed to measure the amplifiability of the DNA. These measures read signal quantity and overall integrity, but do not report on specific damages in the encoded information. Herein, we use next generation sequencing (NGS) to measure the quality of the read signal in order to quantify the truthfulness of the retrieved information. We identify CuAAC to be the worst offender in terms of DNA damage amonst commonly used reactions in DELs, causing an increase of G→T transversions. Furthermore, we show that the analysis provides useful information even in fully elaborated DELs; indeed we see that vestiges of the synthetic history, both chemical and biochemical, are written into the mutational spectra of NGS datasets.
Giulia Pacchioni Nat. Rev. Mater., 2021, 6, 648-649 https://doi.org/10.1038/s41578-021-00349-1AbstractGoogle Applied Science is a division of Google Research that applies computational methods, and in particular machine learning, to a broad range of scientific problems. Patrick Riley, until recently one of their software engineers and now head of artificial intelligence at Relay Therapeutics, talks to Nature Reviews Materials about his experience working on machine-learning projects in an industrial setting.
Yue Zhang; Huadong Luo; Huiyong Ma; Jinqiao Wan; Yue Ji; Alex Shaginian; Jin Li; Yun Deng; Guansai Liu Bioconjugate Chem., 2021, 32(8), 1576-1580 https://doi.org/10.1021/acs.bioconjchem.1c00346AbstractAn efficient visible-light-induced alkylation of DNA-tagged quinoxaline-2-ones was described. The methodology demonstrated moderate-to-excellent conversions under mild conditions. The reaction was found to be tolerant with both N-protected α-amino acids and aliphatic carboxylic acids and could be applied to the synthesis of focused DNA-encoded quinoxalin-2-one libraries.
Yuliana Zabolotna; Dmitriy M. Volochnyuk; Sergey V. Ryabukhin; Dragos Horvath; Kostiantyn Gavrylenko; Gilles Marcou; Yurii S. Moroz; Oleksandr Oksiuta; Alexandre Varnek ChemRxiv, 2021 https://doi.org/10.33774/chemrxiv-2021-clq4hAbstractThe ability to efficiently synthesize desired compounds can be a limiting factor for chemical space exploration in drug discovery. This ability is conditioned not only by the existence of well-studied synthetic protocols but also by the availability of corresponding reagents, so-called building blocks (BB). In this work, we present a detailed analysis of the chemical space of 400K purchasable BB. The chemical space was defined by corresponding synthons – fragments contributed to the final molecules upon reaction. They allow an analysis of BB physicochemical properties and diversity, unbiased by the leaving and protective groups in actual reagents. The main classes of BB were analyzed in terms of their availability, rule-of-two-defined quality, and diversity. Available BBs were eventually compared to a reference set of biologically relevant synthons derived from ChEMBL fragmentation, in order to illustrate how well they cover the actual medicinal chemistry needs. This was performed on a newly constructed universal generative topographic map of synthon chemical space, allowing to visualize both libraries and analyze their overlapping and library-specific regions.
Srinivas Chamakuri; Mee-Kyung Chung; Errol L. G. Samuel; Kevin A. Tran; Ying-Chu Chen; Pranavanand Nyshadham; Conrad Santini; Martin M. Matzuk; Damian W. Young Bioorg. Med. Chem., 2021, 116387 https://doi.org/10.1016/j.bmc.2021.116387AbstractHere we report the successful construction of a novel, stereochemically diverse DNA-Encoded Chemical Library (DECL) by utilizing 24 enantiomerically pure trifunctional 2, 6- di-substituted piperazines as central cores. We introduce the concept of positional diversity by placing the DNA attachment at either of two possible sites on the piperazine scaffold. Using a wide range of building blocks, a diverse library of 77 million compounds was produced. Cheminformatic analysis demonstrates that this library occupies a wide swath of chemical space, and that the piperazines scaffold confers different shape diversity compared to the commonly used triazine core.
Pramod M. Sabale; Mateusz Imiołek; Pierre Raia; Sofia Barluenga; Nicolas Winssinger J. Am. Chem. Soc., 2021, 143(45), 18932-18940 https://doi.org/10.1021/jacs.1c07013AbstractStapled peptides with an enforced α-helical conformation have been shown to overcome major limitations in the development of short peptides targeting protein–protein interactions (PPIs). While the growing arsenal of methodologies to staple peptides facilitates their preparation, stapling methodologies are not broadly embraced in synthetic library screening. Herein, we report a strategy leveraged on hybridization of short PNA–peptide conjugates wherein nucleobase driven assembly facilitates ligation of peptide fragments and constrains the peptide’s conformation into an α-helix. Using native chemical ligation, we show that a mixture of peptide fragments can be combinatorially ligated and used directly in affinity selection against a target of interest. This approach was exemplified with a focused library targeting the p-53/MDM2 interaction. One hundred peptides were obtained in a one-pot ligation reaction, selected by affinity against MDM2 immobilized on beads, and the best binders were identified by mass spectrometry.
Angela F. De Fazio; Doxi Misatziou; Ysobel R. Baker; Otto L. Muskens; Tom Brown; Antonios G. Kanaras Chem. Soc. Rev., 2021, 50, 13410-13440 https://doi.org/10.1039/D1CS00632KAbstractThe self-assembly of inorganic nanoparticles to larger structures is of great research interest as it allows the fabrication of novel materials with collective properties correlated to the nanoparticles’ individual characteristics. Recently developed methods for controlling nanoparticle organisation have enabled the fabrication of a range of new materials. Amongst these, the assembly of nanoparticles using DNA has attracted significant attention due to the highly selective recognition between complementary DNA strands, DNA nanostructure versatility, and ease of DNA chemical modification. In this review we discuss the application of various chemical DNA modifications and molecular intercalators as tools for the manipulation of DNA-nanoparticle structures. In detail, we discuss how DNA modifications and small molecule intercalators have been employed in the chemical and photochemical DNA ligation in nanostructures; DNA rotaxanes and catenanes associated with reconfigurable nanoparticle assemblies; and DNA backbone modifications including locked nucleic acids, peptide nucleic acids and borane nucleic acids, which affect the stability of nanostructures in complex environments. We conclude by highlighting the importance of maximising the synergy between the communities of DNA chemistry and nanoparticle self-assembly with the aim to enrich the library of tools available for the manipulation of nanostructures.
Yinan Song; Xiaoyu Li Acc. Chem. Res., 2021, 54(17), 3491-3503 https://doi.org/10.1021/acs.accounts.1c00375AbstractIn the past two decades, a DNA-encoded chemical library (DEL or DECL) has emerged and has become a major technology platform for ligand discovery in drug discovery as well as in chemical biology research. Although based on a simple concept, i.e., encoding each compound with a unique DNA tag in a combinatorial chemical library, DEL has been proven to be a powerful tool for interrogating biological targets by accessing vast chemical space at a fraction of the cost of traditional high-throughput screening (HTS). Moreover, the recent technological advances and rapid developments of DEL-compatible reactions have greatly enhanced the chemical diversity of DELs. Today, DELs have been adopted by nearly all major pharmaceutical companies and are also gaining momentum in academia. However, this field is heavily biased toward library encoding and synthesis, and an underexplored aspect of DEL research is the selection methods. Generally, DEL selection is considered to be a massive binding assay conducted over an immobilized protein to identify the physical binders using the typical bind–wash–elute procedure. In recent years, we and other research groups have developed new approaches that can perform DEL selections in the solution phase, which has enabled the selection against complex biological targets beyond purified proteins. On the one hand, these methods have significantly widened the target scope of DELs; on the other hand, they have enabled the functional and potentially phenotypic assays of DELs beyond simple binding. An overview of these methods is provided in this Account. Our laboratory has been using DNA-programmed affinity labeling (DPAL) as the main strategy to develop new DEL selection methods. DPAL is based on DNA-templated synthesis; by using a known ligand to guide the target binding, DPAL is able to specifically establish a stable linkage between the target protein and the ligand. The DNA tag of the target-ligand conjugates serves as a programmable handle for protein characterization or hit compound decoding in the case of DEL selections. DPAL also takes advantage of the fast reaction kinetics of photo-cross-linking to achieve high labeling specificity and fidelity, especially in the selection of DNA-encoded dynamic libraries (DEDLs). DPAL has enabled DEL selections not only in buffer and cell lysates but also with complex biological systems, such as large protein complexes and live cells. Moreover, this strategy has also been employed in other biological applications, such as site-specific protein labeling, protein detection, protein profiling, and target identification. In the Account, we describe these methods, highlight their underlying principles, and conclude with perspectives of the development of the DEL technology.
Gabriele Bassi; Nicholas Favalli; Christian Pellegrino; Yuichi Onda; Jörg Scheuermann; Samuele Cazzamalli; Markus G. Manz; Dario Neri J. Med. Chem., 2021, 64(21), 15799-15809 https://doi.org/10.1021/acs.jmedchem.1c01103AbstractPlacental alkaline phosphatase (PLAP) is an abundant surface antigen in the malignancies of the female reproductive tract. Nevertheless, the discovery of PLAP-specific small organic ligands for targeting applications has been hindered by ligand cross-reactivity with the ubiquitous tissue non-specific alkaline phosphatase (TNAP). In this study, we used DNA-encoded chemical libraries to discover a potent (IC50 = 32 nM) and selective PLAP inhibitor, with no detectable inhibition of TNAP activity. Subsequently, the PLAP ligand was conjugated to fluorescein; it specifically bound to PLAP-positive tumors in vitro and targeted cervical cancer in vivo in a mouse model of the disease. Ultimately, the fluorescent derivative of the PLAP inhibitor functioned as a bispecific engager redirecting the killing of chimeric antigen receptor-T cells specific to fluorescein on PLAP-positive tumor cells.
Richard J. Fair; Ryan T. Walsh; Christopher D. Hupp Bioorg. Med. Chem. Lett., 2021, 128339 https://doi.org/10.1016/j.bmcl.2021.128339AbstractOver the past decade, DNA-encoded libraries (DELs) have emerged as a leading platform for small molecule drug discovery among pharmaceutical companies, biotech companies and academic drug hunters alike. This revolutionary technology has tremendous potential that is yet to be fully realized, as the exploration of therapeutically relevant chemical space is fueled by the ever-expanding repertoire of DNA-compatible reactions used to construct the libraries. Advances in direct coupling reactions, like photo-catalytic cross couplings, unique cyclizations such as the formation of 1,2,4–oxadiazoles, and new functional group transformations are valuable contributions to the DEL reaction toolkit, and indicate where future reaction development efforts should focus in order to maximize the productivity of DELs.
Srinivas Chamakuri; Shuo Lu; Melek Nihan Ucisik; Kurt M. Bohren; Ying-Chu Chen; Huang-Chi Du; John C. Faver; Ravikumar Jimmidi; Feng Li; Jian-Yuan Li; Pranavanand Nyshadham; Stephen S. Palmer; Jeroen Pollet; Xuan Qin; Shannon E. Ronca; Banumathi Sankaran; Kiran L. Sharma; Zhi Tan; Leroy Versteeg; Zhifeng Yu; Martin M. Matzuk; Timothy Palzkill; Damian W. Young Proc. Natl. Acad. Sci. USA, 2021, 118(36), e2111172118 https://doi.org/10.1073/pnas.2111172118AbstractSARS-CoV-2 has had a crippling impact on human life globally. Vaccine development has been used as a first-line strategy for COVID-19 prevention and mitigation; however, small-molecule drugs are still vitally needed to extend treatment options. Traditional screening methods for identifying biologically active small molecules are sluggish and often sample an insufficient number of compounds to identify suitable hits. Here, we applied a screening method known as DNA-encoded chemistry technology (DEC-Tec) to screen billions of compounds against a critical viral protein, Mpro. In rapid fashion, we identified the compound CDD-1713 as a potent and selective Mpro inhibitor. This study illuminates DEC-Tec as a highly expeditious strategy toward generating small molecules against critical targets of infectious agents.
Jingwei Shao; Yuqian Yan; Donglin Ding; Dejie Wang; Yundong He; Yunqian Pan; Wei Yan; Anupreet Kharbanda; Hong-yu Li; Haojie Huang Adv. Sci., 2021, 8(20), 2102555 https://doi.org/10.1002/advs.202102555AbstractDNA-binding proteins, including transcription factors (TFs), play essential roles in various cellular processes and pathogenesis of diseases, deeming to be potential therapeutic targets. However, these proteins are generally considered undruggable as they lack an enzymatic catalytic site or a ligand-binding pocket. Proteolysis-targeting chimera (PROTAC) technology has been developed by engineering a bifunctional molecule chimera to bring a protein of interest (POI) to the proximity of an E3 ubiquitin ligase, thus inducing the ubiquitination of POI and further degradation through the proteasome pathway. Here, the development of oligonucleotide-based PROTAC (O'PROTACs), a class of noncanonical PROTACs in which a TF-recognizing double-stranded oligonucleotide is incorporated as a binding moiety of POI is reported. It is demonstrated that O'PROTACs of lymphoid enhancer-binding factor 1 (LEF1) and ETS-related gene (ERG), two highly cancer-related transcription factors, successfully promote degradation of these proteins, impede their transcriptional activity, and inhibit cancer cell growth in vitro and in vivo. The programmable nature of O'PROTACs indicates that this approach is also applicable to destruct other TFs. O'PROTACs not only can serve as a research tool but also can be harnessed as a therapeutic arsenal to target DNA binding proteins for effective treatment of diseases such as cancer.
Shorouk O. Badir; Alexander Lipp; Matthias Krumb; María Jesús Cabrera-Afonso; Lisa Marie Kammer; Victoria E. Wu; Minxue Huang; Adam Csakai; Lisa A. Marcaurelle; Gary A Molander Chem. Sci., 2021, 12, 12036-12045 https://doi.org/10.1039/D1SC03191KAbstractDNA-encoded library (DEL) technology features a time- and cost-effective interrogation format for the discovery of therapeutic candidates in the pharmaceutical industry. To develop DEL platforms, the implementation of water-compatible transformations that facilitate the incorporation of multifunctional building blocks (BBs) with high C(sp3) carbon counts is integral for success. In this report, a decarboxylative-based hydroalkylation of DNA-conjugated trifluoromethyl-substituted alkenes enabled by single-electron transfer (SET) and subsequent hydrogen atom termination through electron donor-acceptor (EDA) complex activation is detailed. In a further photoredox-catalyzed hydroarylation protocol, the coupling of functionalized, electronically unbiased olefins is achieved under air and within minutes of blue light irradiation through the intermediacy of reactive (hetero)aryl radical species with full retention of the DNA tag integrity. Notably, these processes operate under mild reaction conditions, furnishing complex structural scaffolds with a high density of pendant functional groups.
James H. Hunter; Matthew J. Anderson; Isaline F. S. F. Castan; Jessica S. Graham; Catherine L. A. Salvini; Harriet A. Stanway-Gordon; James J. Crawford; Andrew Madin; Garry Pairaudeau; Michael J. Waring Chem. Sci., 2021, 12(27), 9475-9484 https://doi.org/10.1039/D1SC03007HAbstractDNA encoded libraries (DELs) represent powerful new technology for finding small molecule ligands for proteins and are increasingly being applied to hit finding in medicinal chemistry. Crucial to the synthesis of high quality DELs is the identification of chemical reactions for their assembly that proceed with very high conversion across a range of different substrates, under conditions compatible with DNA-tagged substrates. Many current chemistries used in DEL synthesis do not meet this requirement, resulting in libraries of low fidelity. Amide couplings are the most commonly used reaction in synthesis of screening libraries and also in DELs. The ability to carry out highly efficient, widely applicable amide couplings in DEL synthesis would therefore be highly desirable. We report a method for amide coupling using micelle forming surfactants, promoted by a modified linker, that is broadly applicable across a wide range of substrates. Most significantly, this works exceptionally well for coupling of DNA-conjugated carboxylic acids (N-to-C) with amines in solution, a procedure that is currently very inefficient. The optimisation of separate procedures for coupling of DNA-conjugated acids and amines by reagent screening and statistically driven optimisation is described. The generality of the method is illustrated by the application to a wide range of examples with unprecedented levels of conversion. The utility of the (N-to-C) coupling of DNA-conjugated acids in DEL synthesis is illustrated by the three cycle synthesis of a fully DNA-encoded compound by two cycles of coupling of an aminoester, with intermediate ester hydrolysis, followed by capping with an amine. This methodology will be of great utility in the synthesis of high fidelity DELs.
Árpád Furka MedComm, 2021, 2(3), 481-489 https://doi.org/10.1002/mco2.84AbstractCombinatorial chemistry invented nearly 40 years ago was welcomed with enthusiasm in the drug research community. The method offered access to a practically unlimited number of new compounds. The new compounds however are mixtures, and methods had to be developed for the identification of the bioactive components. This was one of the reasons why the method could not providethe expected cornucopia of new drugs. Among the different screening methods, two approaches seem to offer the best results. One of them is based on the intrinsic property of the combinatorial split and pool solid-phase synthesis: One compound forms on each bead of the solid support. Different methods have been developed to encode the beads and identify the structure of compounds formed on them. The most important method applies DNA oligomers for encoding. As a second approach in screening, DNA-encoded combinatorial libraries are synthesized omitting the solid support and the mixtures are screened in solution using affinity binding methods. Libraries containing billions and even trillions of components are synthesized and successfully tested, which led to the identification of a significant number of new leads.
Marco Potowski; Ricarda Lüttig; Alexandros Vakalopoulos; Andreas Brunschweiger Org. Lett., 2021, 23(14), 5480-5484 https://doi.org/10.1021/acs.orglett.1c01834AbstractDNA-encoded libraries designed around heterocyclic scaffolds have proven highly productive in target-based screening. Here, we show the synthesis of imidazopyridines on a controlled pore glass-coupled DNA oligonucleotide for solid phase-initiated encoded library synthesis. The target compounds were synthesized by a variant of the A3 coupling reaction from aminopyridines, alkynes, and aldehydes promoted by copper(I/II) and furnished diverse substituted scaffolds with functionalities for library design. Although proceeding under forcing conditions, it produced minimal DNA damage.
Minsoo Song; Gil Tae Hwang Nat. Chem., 2021, 13, 515–517 https://doi.org/10.1038/s41557-021-00690-6AbstractDNA-encoded libraries are a powerful tool to identify hit compounds for drug discovery. Now, two papers have reported new advances in this technology. One paper reports a method to screen for binders inside a living cell, and the other investigates the effects of stereo- and regiochemistry on ligand discovery.
William McCoull; Scott Boyd; Martin R. Brown; Muireann Coen; Olga Collingwood; Nichola L. Davies; Ann Doherty; Gary Fairley; Kristin Goldberg; Elizabeth Hardaker; Guang He; Edward J. Hennessy; Philip Hopcroft; George Hodgson; Anne Jackson; Xiefeng Jiang; Ankur Karmokar; Anne-Laure Lainé; Nicola Lindsay; Yumeng Mao; Roshini Markandu; Lindsay McMurray; Neville McLean; Lorraine Mooney; Helen Musgrove; J. Willem M. Nissink; Alexander Pflug; Venkatesh Pilla Reddy; Philip B. Rawlins; Emma Rivers; Marianne Schimpl; Graham F. Smith; Sharon Tentarelli; Jon Travers; Robert I. Troup; Josephine Walton; Cheng Wang; Stephen Wilkinson; Beth Williamson; Jon Winter-Holt; Dejian Yang; Yuting Zheng; Qianxiu Zhu; Paul D. Smith J. Med. Chem., 2021, 64(18), 13524-13539 https://doi.org/10.1021/acs.jmedchem.1c00920AbstractInhibition of Mer and Axl kinases has been implicated as a potential way to improve the efficacy of current immuno-oncology therapeutics by restoring the innate immune response in the tumor microenvironment. Highly selective dual Mer/Axl kinase inhibitors are required to validate this hypothesis. Starting from hits from a DNA-encoded library screen, we optimized an imidazo[1,2-a]pyridine series using structure-based compound design to improve potency and reduce lipophilicity, resulting in a highly selective in vivo probe compound 32. We demonstrated dose-dependent in vivo efficacy and target engagement in Mer- and Axl-dependent efficacy models using two structurally differentiated and selective dual Mer/Axl inhibitors. Additionally, in vivo efficacy was observed in a preclinical MC38 immuno-oncology model in combination with anti-PD1 antibodies and ionizing radiation.
Kejia Yan; Vivian Triana; Sunil Vasu Kalmady; Kwami Aku-Dominguez; Sharyar Memon; Alex Brown; Russell Greiner; Ratmir Derda ChemRxiv, 2021 https://doi.org/10.26434/chemrxiv.14609670.v1AbstractThe Wittig reaction can be used for late stage functionalization of proteins and peptides to ligate glycans, pharmacophores, and many other functionalities. In this manuscript, we modified 160,000 N-terminal glyoxaldehyde peptides displayed on phage with the Wittig reaction by biotin labeled ylide under conditions that functionalize only 1% of the library population. Deep-sequencing of the biotinylated and input populations estimated the rate of conversion for each sequence. This “deep conversion” (DC) from deep sequencing correlates with rate constants measured by HPLC. Peptide sequences with fast and slow reactivity highlighted a critical role of primary backbone amides (N-H) in accelerating the rate of the aqueous Wittig reaction. Experimental measurement of reaction rates and density functional theory (DFT) computation of the transition state geometries corroborated this relationship. We also collected deep-sequencing data to build structure activity relationship (SAR) models that can predict DC value of the Wittig reaction. By using this data, we trained two classifier models based on Gradient Boosted trees. These classifiers achieved area under the ROC (Receiver Operating Characteristic) Curve (ROC AUC) of 81.2 ± 0.4 and 73.7 ± 0.8 (90–92% accuracy) in determining whether a sequence belonged to the top 5% or the bottom 5% in terms of its reactivity. We have deployed our learned models as a publicly available web app: http://44.226.164.95/ We anticipate that phage-displayed peptides and related mRNA or DNA-displayed substrates can be employed in a similar fashion to study the substrate scope and mechanisms of many other chemical reactions.
M. Dominic Ryan; Alastair L. Parkes; David Corbett; Anthony P. Dickie; Michelle Southey; Ole A. Andersen; Daniel B. Stein; Olivier R. Barbeau; Angelo Sanzone; Pia Thommes; John Barker; Ricky Cain; Christel Compper; Magali Dejob; Alain Dorali; Donnya Etheridge; Sian Evans; Adele Faulkner; Elise Gadouleau; Timothy Gorman; Denes Haase; Maisie Holbrow-Wilshaw; Thomas Krulle; Xianfu Li; Christopher Lumley; Barbara Mertins; Spencer Napier; Rajesh Odedra; Kostas Papadopoulos; Vasileios Roumpelakis; Kate Spear; Emily Trimby; Jennifer Williams; Michael Zahn; Anthony D. Keefe; Ying Zhang; Holly T. Soutter; Paolo A. Centrella; Matthew A. Clark; John W. Cuozzo; Christoph E. Dumelin; Boer Deng; Avery Hunt; Eric A. Sigel; Dawn M. Troast; Boudewijn L. M. DeJonge J. Med. Chem., 2021, 64(19), 14377-14425 https://doi.org/10.1021/acs.jmedchem.1c00888AbstractThis study describes a novel series of UDP-N-acetylglucosamine acyltransferase (LpxA) inhibitors that was identified through affinity-mediated selection from a DNA-encoded compound library. The original hit was a selective inhibitor of Pseudomonas aeruginosa LpxA with no activity against Escherichia coli LpxA. The biochemical potency of the series was optimized through an X-ray crystallography-supported medicinal chemistry program, resulting in compounds with nanomolar activity against P. aeruginosa LpxA (best half-maximal inhibitory concentration (IC50) <5 nM) and cellular activity against P. aeruginosa (best minimal inhibitory concentration (MIC) of 4 μg/mL). Lack of activity against E. coli was maintained (IC50 > 20 μM and MIC > 128 μg/mL). The mode of action of analogues was confirmed through genetic analyses. As expected, compounds were active against multidrug-resistant isolates. Further optimization of pharmacokinetics is needed before efficacy studies in mouse infection models can be attempted. To our knowledge, this is the first reported LpxA inhibitor series with selective activity against P. aeruginosa.
Sandra Röhm; Benedict-Tilman Berger; Martin Schröder; Deep Chatterjee; Sebastian Mathea; Andreas C. Joerger; Daniel M. Pinkas; Joshua C. Bufton; Amelie Tjaden; Lohitesh Kovooru; Mark Kudolo; Christian Pohl; Alex N. Bullock; Susanne Müller; Stefan Laufer; Stefan Knapp J. Med. Chem., 2021, 64(18), 13451-13474 https://doi.org/10.1021/acs.jmedchem.1c00868AbstractDiscoidin domain receptors 1 and 2 (DDR1/2) play a central role in fibrotic disorders, such as renal and pulmonary fibrosis, atherosclerosis, and various forms of cancer. Potent and selective inhibitors, so-called chemical probe compounds, have been developed to study DDR1/2 kinase signaling. However, these inhibitors showed undesired activity on other kinases such as the tyrosine protein kinase receptor TIE or tropomyosin receptor kinases, which are related to angiogenesis and neuronal toxicity. In this study, we optimized our recently published p38 mitogen-activated protein kinase inhibitor 7 toward a potent and cell-active dual DDR/p38 chemical probe and developed a structurally related negative control. The structure-guided design approach used provided insights into the P-loop folding process of p38 and how targeting of non-conserved amino acids modulates inhibitor selectivity. The developed and comprehensively characterized DDR/p38 probe, 30 (SR-302), is a valuable tool for studying the role of DDR kinase in normal physiology and in disease development.
Kang Ju Lee; Geul Bang; Yong Wook Kim; Min Hyeon Shin; Hyun-Suk Lim Bioorg. Med. Chem., 2021, 116423 https://doi.org/10.1016/j.bmc.2021.116423AbstractHere we describe the design and synthesis of a DNA-encoded library of bicyclic peptoids. We show that our solid-phase strategy is facile and DNA-compatible, yielding a structurally diverse combinatorial library of bicyclic peptoids of various ring sizes. We also demonstrate that affinity-based screening of a DNA-encoded library of bicyclic peptoids enables to efficiently identify high-affinity ligands for a target protein. Given their highly constraint structures, as well as increased cell permeability and proteolytic stability relative to native peptides, bicyclic peptoids could be an excellent source of protein capture agents. As such, our DNA-encoded library of bicyclic peptoids will serve as versatile tools that facilitate the generation of potent ligands against many challenging targets, such as such as intracellular protein-protein interactions.
Kaifeng Chen; Weijie Chen; Fangyuan Chen; Haiman Zhang; Huiying Xu; Zhi Zhou; Wei Yi Org. Chem. Front., 2021, 8, 4452-4458 https://doi.org/10.1039/D1QO00709BAbstractEfficient, mild and metal-free [3 + 2] annulation of N-phenoxy amides with gem-difluoroalkenes has been realized via a base-mediated tandem [3,3]-sigmatropic rearrangement, which gives direct access to 2-aminobenzofuran derivatives involved in the one-pot cleavage of multiple bonds including C–H, O–N and twofold C–F bonds. The subsequent success of the on-DNA compatible synthesis and the application of the obtained products as potential anticancer agents further demonstrates the versatility of this transformation.
Yulong An; Hao Yan; Zhenzhen Dong; Alexander L. Satz Curr. Protoc., 2021, 1(5), e125 https://doi.org/10.1002/cpz1.125AbstractAn efficient method for the synthesis of DNA‐conjugated 1,2,3‐triazoles is copper (II) [Cu(II)‐β‐cyclodextrin]‐mediated Huisgen cycloaddition (“click reaction”) of DNA‐conjugated alkynes with azides. However, a diverse array of building blocks is required to produce useful DNA encoded libraries, and the commercial availability of azides is limited. The method described herein generates azides in situ from aryl borates and TMSN3, which then further react with DNA‐conjugated terminal alkynes.
Beilei Wang; Hong Wu; Chen Hu; Haizhen Wang; Jing Liu; Wenchao Wang; Qingsong Liu Sig. Transduct. Target. Ther., 2021, 6, 423 https://doi.org/10.1038/s41392-021-00826-7AbstractSince the clinical approval of imatinib, the discovery of protein kinase downregulators entered a prosperous age. However, challenges still exist in the discovery of kinase downregulator drugs, such as the high failure rate during development, side effects, and drug-resistance problems. With the progress made through multidisciplinary efforts, an increasing number of new approaches have been applied to solve the above problems during the discovery process of kinase downregulators. In terms of in vitro and in vivo drug evaluation, progress was also made in cellular and animal model platforms for better and more clinically relevant drug assessment. Here, we review the advances in drug design strategies, drug property evaluation technologies, and efficacy evaluation models and technologies. Finally, we discuss the challenges and perspectives in the development of kinase downregulator drugs.
Yu Zhou; Wenyin Shen; Jianzhao Peng; Yuqing Deng; Xiaoyu Li Bioorg. Med. Chem., 2021, 116328 https://doi.org/10.1016/j.bmc.2021.116328AbstractDNA-encoded chemical library (DEL) has emerged to be a powerful ligand screening technology in drug discovery. Recently, we reported a DNA-encoded dynamic library (DEDL) approach that combines the principle of traditional dynamic combinatorial library (DCL) with DEL. DEDL has shown excellent potential in fragment-based ligand discovery with a variety of protein targets. Here, we further tested the utility of DEDL in identifying low molecular weight fragments that are selective for different isoforms or domains of the same protein family. A 10,000-member DEDL was selected against sirtuin-1, 2, and 5 (SIRT1, 2, 5) and the BD1 and BD2 domains of bromodomain 4 (BRD4), respectively. Albeit with modest potency, a series of isoform/domain-selective fragments were identified and the corresponding inhibitors were derived by fragment linking.
Miguel M. Vaidergorn; Flavio da Silva Emery; A. Ganesan J. Med. Chem., 2021, 64(19), 13980-14010 https://doi.org/10.1021/acs.jmedchem.1c00787AbstractWe review progress in the application of fragment-based drug discovery (FBDD) to epigenetic drug discovery (EPIDD) targeted at epigenetic writer and eraser enzymes as well as reader domains over the last 15 years. The greatest successes to date are in prospecting for bromodomain binding ligands. From a diverse array of fragment hits, multiple potent and selective compounds ensued, including the oncology clinical candidates mivebresib, ABBV-744, pelabresib, and PLX51107.
Ilaria Proietti Silvestri; Paul J. J. Colbon ACS Med. Chem. Lett., 2021, 12(8), 1220-1229 https://doi.org/10.1021/acsmedchemlett.1c00251AbstractModern-day drug discovery is now blessed with a wide range of high-throughput hit identification (hit-ID) strategies that have been successfully validated in recent years, with particular success coming from high-throughput screening, fragment-based lead discovery, and DNA-encoded library screening. As screening efficiency and throughput increases, this enables the viable exploration of increasingly complex three-dimensional (3D) chemical structure space, with a realistic chance of identifying highly specific hit ligands with increased target specificity and reduced attrition rates in preclinical and clinical development. This minireview will explore the impact of an improved design of multifunctionalized, sp3-rich, stereodefined scaffolds on the (virtual) exploration of 3D chemical space and the specific requirements for different hit-ID technologies.
Adrián Gironda-Martínez; Etienne J. Donckele; Florent Samain; Dario Neri ACS Pharmacol. Transl. Sci., 2021, 4(4), 1265-1279 https://doi.org/10.1021/acsptsci.1c00118AbstractDNA-encoded chemical libraries (DELs) represent a versatile and powerful technology platform for the discovery of small-molecule ligands to protein targets of biological and pharmaceutical interest. DELs are collections of molecules, individually coupled to distinctive DNA tags serving as amplifiable identification barcodes. Thanks to advances in DNA-compatible reactions, selection methodologies, next-generation sequencing, and data analysis, DEL technology allows the construction and screening of libraries of unprecedented size, which has led to the discovery of highly potent ligands, some of which have progressed to clinical trials. In this Review, we present an overview of diverse approaches for the generation and screening of DEL molecular repertoires. Recent success stories are described, detailing how novel ligands were isolated from DEL screening campaigns and were further optimized by medicinal chemistry. The goal of the Review is to capture some of the most recent developments in the field, while also elaborating on future challenges to further improve DEL technology as a therapeutic discovery platform.
Kyle W. Knouse; Dillon T. Flood; Julien C. Vantourout; Michael A. Schmidt; Ivar M. Mcdonald; Martin D. Eastgate; Phil S. Baran ACS Cent. Sci., 2021, 7(9), 1473-1485 https://doi.org/10.1021/acscentsci.1c00487AbstractPhosphate linkages govern life as we know it. Their unique properties provide the foundation for many natural systems from cell biology and biosynthesis to the backbone of nucleic acids. Phosphates are ideal natural moieties; existing as ionized species in a stable P(V)-oxidation state, they are endowed with high stability but exhibit enzymatically unlockable potential. Despite intense interest in phosphorus catalysis and condensation chemistry, organic chemistry has not fully embraced the potential of P(V) reagents. To be sure, within the world of chemical oligonucleotide synthesis, modern approaches utilize P(III) reagent systems to create phosphate linkages and their analogs. In this Outlook, we present recent studies from our laboratories suggesting that numerous exciting opportunities for P(V) chemistry exist at the nexus of organic synthesis and biochemistry. Applications to the synthesis of stereopure antisense oligonucleotides, cyclic dinucleotides, methylphosphonates, and phosphines are reviewed as well as chemoselective modification to peptides, proteins, and nucleic acids. Finally, an outlook into what may be possible in the future with P(V) chemistry is previewed, suggesting these examples represent just the tip of the iceberg.
Jonas Persson; Emmi Puuvuori; Bo Zhang; Irina Velikyan; Ola Åberg; Malin Müller; Per-Åke Nygren; Stefan Ståhl; Olle Korsgren; Olof Eriksson; John Löfblom Sci. Rep., 2021,11, 19151 https://doi.org/10.1038/s41598-021-97694-6AbstractDue to the wide scale of inflammatory processes in different types of disease, more sensitive and specific biomarkers are required to improve prevention and treatment. Cluster of differentiation 69 (CD69) is one of the earliest cell surface proteins expressed by activated leukocytes. Here we characterize and optimize potential new imaging probes, Affibody molecules targeting CD69 for imaging of activated immune cells. Analysis of candidates isolated in a previously performed selection from a Z variant E. coli library to the recombinant extracellular domain of human CD69, identified one cross-reactive Z variant with affinity to murine and human CD69. Affinity maturation was performed by randomization of the primary Z variant, followed by selections from the library. The resulting Z variants were evaluated for affinity towards human and murine CD69 and thermal stability. The in vivo biodistribution was assessed by SPECT/CT in rats following conjugation of the Z variants by a DOTA chelator and radiolabeling with Indium-111. A primary Z variant with a Kd of approximately 50 nM affinity to human and murine CD69 was identified. Affinity maturation generated 5 additional Z variants with improved or similar affinity. All clones exhibited suitable stability. Radiolabeling and in vivo biodistribution in rat demonstrated rapid renal clearance for all variants, while the background uptake and washout varied. The variant ZCD69:4 had the highest affinity for human and murine CD69 (34 nM) as well as the lowest in vivo background binding. In summary, we describe the discovery, optimization and evaluation of novel Affibody molecules with affinity for CD69. Affibody molecule ZCD69:4 is suitable for further development for imaging of activated immune cells.
Wei Wang; S. Cyrus Khojasteh; Dian Su Molecules, 2021, 26(11), 3338 https://doi.org/10.3390/molecules26113338AbstractMacrocyclic peptides are predominantly peptide structures bearing one or more rings and spanning multiple amino acid residues. Macrocyclization has become a common approach for improving the pharmacological properties and bioactivity of peptides. A variety of ribosomal-derived and non-ribosomal synthesized cyclization approaches have been established. The biosynthesis of backbone macrocyclic peptides using seven new emerging methodologies will be discussed with regard to the features and strengths of each platform rather than medicinal chemistry tools. The mRNA display variant, known as the random nonstandard peptide integrated discovery (RaPID) platform, utilizes flexible in vitro translation (FIT) to access macrocyclic peptides containing nonproteinogenic amino acids (NAAs). As a new discovery approach, the ribosomally synthesized and post-translationally modified peptides (RiPPs) method involves the combination of ribosomal synthesis and the phage screening platform together with macrocyclization chemistries to generate libraries of macrocyclic peptides. Meanwhile, the split-intein circular ligation of peptides and proteins (SICLOPPS) approach relies on the in vivo production of macrocyclic peptides. In vitro and in vivo peptide library screening is discussed as an advanced strategy for cyclic peptide selection. Specifically, biosynthetic bicyclic peptides are highlighted as versatile and attractive modalities. Bicyclic peptides represent another type of promising therapeutics that allow for building blocks with a heterotrimeric conjugate to address intractable challenges and enable multimer complexes via linkers. Additionally, we discuss the cell-free chemoenzymatic synthesis of macrocyclic peptides with a non-ribosomal catalase known as the non-ribosomal synthetase (NRPS) and chemo-enzymatic approach, with recombinant thioesterase (TE) domains. Novel insights into the use of peptide library tools, activity-based two-hybrid screening, structure diversification, inclusion of NAAs, combinatorial libraries, expanding the toolbox for macrocyclic peptides, bicyclic peptides, chemoenzymatic strategies, and future perspectives are presented. This review highlights the broad spectrum of strategy classes, novel platforms, structure diversity, chemical space, and functionalities of macrocyclic peptides enabled by emerging biosynthetic platforms to achieve bioactivity and for therapeutic purposes.
Shilian Yang; Yizhou Li SciencePaper, 2021.4.14 http://www.paper.edu.cn/releasepaper/content/202104-110.AbstractThis article reports a method of constructing 1,2,4,5-tetrazine compounds by reacting oligonucleotides coupled with nitrile building blocks and hydrazine monohydrate with the participation of thiols. This method has the characteristics of mild conditions and compatibility with water. This research has the following significance: (1) It lays the foundation for the study of oligonucleotide-compatible bioorthogonal reactions based on 1,2,4,5-tetrazine compounds; (2) It is the DNA coding molecular library technology ( DNA encoded library, DEL) to construct a 1,2,4,5-tetrazine molecular library to provide a synthetic method, which increases the structural diversity of the molecular library.
Isaline F. S. F. Castan; Jessica S. Graham; Catherine L. A. Salvini; Harriet A. Stanway-Gordon; Michael J. Waring Bioorg. Med. Chem., 2021, 116273 https://doi.org/10.1016/j.bmc.2021.116273AbstractDNA-encoded libraries (DELs) are becoming an established technology for finding ligands for protein targets. We have abstracted and analysed libraries from the literature to assess the synthesis strategy, selections of reactions and monomers and their propensity to reveal hits. DELs have led to hit compounds across a range of diverse protein classes. The range of reactions and monomers utilised has been relatively limited and the hits are often higher in molecular weight than might be considered ideal. Considerations for future library designs with reference to chemical diversity and lead-like properties are discussed.
Zhennan Fang; Huiqiang Wei; Wenfeng Gou; Leyuan Chen; Changfen Bi; Wenbin Hou; Yiliang Li Future Med. Chem., 2021, 13(9), 817-837 https://doi.org/10.4155/fmc-2020-0386AbstractNonapoptotic types of regulated cell death have attracted widespread interest since the discovery that certain forms of cell necrosis can be regulated. In particular, research into cell necroptosis has made significant progress in connection with kidney, inflammatory, degenerative and neoplastic diseases. Inhibitors targeting the critical necroptosis-associated proteins RIPK1/3 and MLKL have been in development for more than a decade. Herein the authors compile a list of the known small-molecule inhibitors of these enzymes and representative structures of compounds co-crystallized with these proteins and put forward some thoughts regarding their future development.
Natalia Milosevich; Chelsea Wilson; Tyler Brown; Aktan Alpsoy; Sijie Wang; Katelyn E. Connelly; Kirsten Sinclair; Felino Ponio; Rebecca Hof; Emily Dykhuizen; Fraser Hof ChemRxiv, 2021 https://doi.org/10.26434/chemrxiv.14390672.v1AbstractMethyllysine reader proteins bind to methylated lysine residues and alter gene transcription by changing the compaction state of chromatin or by the recruitment of other multiprotein complexes. The polycomb paralog family of methyllysine readers bind to trimethylated lysine on the tail of histone 3 via a highly conserved aromatic cage located in their chromodomains. Each of the polycomb paralogs are implicated in several disease states. CBX6 and CBX8 are members of the polycomb paralog family with two structurally similar chromodomains. By exploring the structure-activity relationships of a previously reported CBX6 inhibitor we have discovered more potent and cell permeable analogs. Our current report includes potent, dual-selective inhibitors of CBX6 and CBX8. We have shown that the –2 position in our scaffold is an important residue for selectivity amongst the polycomb paralogs. Preliminary cell-based studies show that the new inhibitors impact cell proliferation in a rhabdoid tumor cell line. This report includes data on inhibitor design, inhibitor synthesis, compound characterization by LCMS, compound activity by fluorescence polarization, analysis of structure-activity relationships, rhabdoid tumor cell line activity.
Jeffrey Y. K. Wong; Raja Mukherjee; Jiayuan Miao; Olena Bilyk; Vivian Triana; Mark Miskolzie; Antoine Henninot; John J. Dwyer; Serhii Kharchenko; Anna Iampolska; Dmitriy M. Volochnyuk; Yu-Shan Lin; Lynne-Marie Postovit; Ratmir Derda Chem. Sci., 2021, 12(28), 9694-9703 https://doi.org/10.1039/D1SC01916CAbstractIn this manuscript, we developed a two-fold symmetric linchpin (TSL) that converts readily available phage-displayed peptides libraries made of 20 common amino acids to genetically-encoded libraries of bicyclic peptides displayed on phage. TSL combines an aldehyde-reactive group and two thiol-reactive groups; it bridges two side chains of cysteine [C] with an N-terminal aldehyde group derived from the N-terminal serine [S], yielding a novel bicyclic topology that lacks a free N-terminus. Phage display libraries of SX1CX2X3X4X5X6X7C sequences, where X is any amino acid but Cys, were converted to a library of bicyclic TSL-[S]X1[C]X2X3X4X5X6X7[C] peptides in 45 ± 15% yield. Using this library and protein morphogen NODAL as a target, we discovered bicyclic macrocycles that specifically antagonize NODAL-induced signaling in cancer cells. At a 10 μM concentration, two discovered bicyclic peptides completely suppressed NODAL-induced phosphorylation of SMAD2 in P19 embryonic carcinoma cells. The TSL-[S]Y[C]KRAHKN[C] bicycle inhibited NODAL-induced proliferation of NODAL-TYK-nu ovarian carcinoma cells with apparent IC50 of 1 μM. The same bicycle at 10 μM concentration did not affect the growth of the control TYK-nu cells. TSL-bicycles remained stable over the course of the 72 hour-long assays in a serum-rich cell-culture medium. We further observed general stability in mouse serum and in a mixture of proteases (Pronase™) for 21 diverse bicyclic macrocycles of different ring sizes, amino acid sequences, and cross-linker geometries. TSL-constrained peptides to expand the previously reported repertoire of phage-displayed bicyclic architectures formed by cross-linking Cys side chains. We anticipate that it will aid the discovery of proteolytically stable bicyclic inhibitors for a variety of protein targets.
Bart Vanhaesebroeck; Matthew W. D. Perry; Jennifer R. Brown; Fabrice André; Klaus Okkenhaug Nat. Rev. Drug Discov., 2021, 20(10), 741-769 https://doi.org/10.1038/s41573-021-00209-1AbstractOveractive phosphoinositide 3-kinase (PI3K) in cancer and immune dysregulation has spurred extensive efforts to develop therapeutic PI3K inhibitors. Although progress has been hampered by issues such as poor drug tolerance and drug resistance, several PI3K inhibitors have now received regulatory approval — the PI3Kα isoform-selective inhibitor alpelisib for the treatment of breast cancer and inhibitors mainly aimed at the leukocyte-enriched PI3Kδ in B cell malignancies. In addition to targeting cancer cell-intrinsic PI3K activity, emerging evidence highlights the potential of PI3K inhibitors in cancer immunotherapy. This Review summarizes key discoveries that aid the clinical translation of PI3Kα and PI3Kδ inhibitors, highlighting lessons learnt and future opportunities
Francesco Fiorentino; Antonello Mai; Dante Rotili J. Med. Chem., 2021, 64(14), 9732-9758 https://doi.org/10.1021/acs.jmedchem.1c00601AbstractSirtuin 6 (SIRT6) is an NAD+-dependent protein deacylase and mono-ADP-ribosyltransferase of the sirtuin family with a wide substrate specificity. In vitro and in vivo studies have indicated that SIRT6 overexpression or activation has beneficial effects for cellular processes such as DNA repair, metabolic regulation, and aging. On the other hand, SIRT6 has contrasting roles in cancer, acting either as a tumor suppressor or promoter in a context-specific manner. Given its central role in cellular homeostasis, SIRT6 has emerged as a promising target for the development of small-molecule activators and inhibitors possessing a therapeutic potential in diseases ranging from cancer to age-related disorders. Moreover, specific modulators allow the molecular details of SIRT6 activity to be scrutinized and further validate the enzyme as a pharmacological target. In this Perspective, we summarize the current knowledge about SIRT6 pharmacology and medicinal chemistry and describe the features of the activators and inhibitors identified so far.
Marco Potowski; Verena Kunig; Lukas Eberlein; Alexandros Vakalopoulos; Stefan Kast; Andreas Brunschweiger Angew. Chem. Int. Ed. Engl., 2021, 60(36), 19744-19749 https://doi.org/10.1002/anie.202104348AbstractDNA-encoded compound libraries are a widely used small molecule screening technology. One important aim in library design is the coverage of chemical space through structurally diverse molecules. Yet, the chemical reactivity of native DNA barcodes limits the toolbox of reactions for library design. Substituting the chemically vulnerable purines by 7-deazaadenine, which exhibits tautomerization stability similar to natural adenine with respect to the formation of stable Watson-Crick pairs, yielded ligation-competent, amplifiable, and readable DNA barcodes for encoded chemistry with enhanced stability against protic acid- and metal ion-promoted depurination. The barcode stability allowed for straightforward translation of 16 exemplary reactions that included isocyanide multicomponent reactions, acid-promoted Pictet-Spengler and Biginelli reactions, and metal-promoted pyrazole syntheses on controlled pore glass-coupled barcodes for diverse DEL design. The Boc protective group of reaction products offered a convenient handle for encoded compound purification.
Wenji Su; Rui Ge; Duanchen Ding; Wenhua Chen; Wenqing Wang; Hao Yan; Weikun Wang; Youlang Yuan; Huan Liu; Meng Zhang; Jiyuan Zhang; Qisheng Shu; Alexander L. Satz; Letian Kuai Bioconjugate Chem., 2021, 32(5), 1001–1007 https://doi.org/10.1021/acs.bioconjchem.1c00170AbstractDNA encoded library (DEL) technology allows for rapid identification of novel small-molecule ligands and thus enables early-stage drug discovery. DEL technology is well-established, numerous cases of discovered hit molecules have been published, and the technology is widely employed throughout the pharmaceutical industry. Nonetheless, DEL selection results can be difficult to interpret, as library member enrichment may derive from not only desired products, but also DNA-conjugated byproducts and starting materials. Note that DELs are generally produced using split-and-pool combinatorial chemistry, and DNA-conjugated byproducts and starting materials cannot be removed from the library mixture. Herein, we describe a method for high-throughput parallel resynthesis of DNA-conjugated molecules such that byproducts, starting materials, and desired products are produced in a single pot, using the same chemical reactions and reagents as during library production. The low-complexity mixtures of DNA–conjugate are then assessed for protein binding by affinity selection mass spectrometry and the molecular weights of the binding ligands ascertained. This workflow is demonstrated to be a practical tool to triage and validate potential hits from DEL selection data.
Matthew J. Henley; Angela N. Koehler Nat. Rev. Drug Discov., 2021, 20, 669-688 https://doi.org/10.1038/s41573-021-00199-0AbstractTranscription factors (TFs) represent key biological players in diseases including cancer, autoimmunity, diabetes and cardiovascular disease. However, outside nuclear receptors, TFs have traditionally been considered ‘undruggable’ by small-molecule ligands due to significant structural disorder and lack of defined small-molecule binding pockets. Renewed interest in the field has been ignited by significant progress in chemical biology approaches to ligand discovery and optimization, especially the advent of targeted protein degradation approaches, along with increasing appreciation of the critical role a limited number of collaborators play in the regulation of key TF effector genes. Here, we review current understanding of TF-mediated gene regulation, discuss successful targeting strategies and highlight ongoing challenges and emerging approaches to address them.
Yiran Huang; Xiaoyu Li ChemBioChem, 2021, 22(14), 2384-2397 https://doi.org/10.1002/cbic.202100144AbstractDNA-encoded library (DEL) has come of age and become a major technology platform for ligand discovery in both academia and the pharmaceutical industry. Technological maturation in the past two decades and the recent explosive developments of DEL-compatible chemistries have greatly improved the chemical diversity of DELs and fuelled its applications in drug discovery. A relatively less-covered aspect of DELs is the selection method. Typically, DEL selection is considered as a binding assay and the selection is conducted with purified protein targets immobilized on a matrix, and the binders are separated from the non-binding background via physical washes. However, the recent innovations in DEL selection methods have not only expanded the target scope of DELs, but also revealed the potential of the DEL technology as a powerful tool in exploring fundamental biology. In this Review, we first cover the "classic" DEL selection methods with purified proteins on solid phase, and then we discuss the strategies to realize DEL selections in solution phase. Finally, we focus on the emerging approaches for DELs to interrogate complex biological targets.
Jean-Pierre Daguer; Arthur Gonse; Yevhenii Shchukin; Lluc Farrera-Soler; Sofia Barluenga; Nicolas Winssinger Bioorg. Med. Chem., 2021, 116282 https://doi.org/10.1016/j.bmc.2021.116282AbstractA dual Bcl-XL / Bcl-2 inhibitor was discovered from DNA-encoded libraries using a two steps process. In the first step, DNA was used to pair PNA-encoded fragments exploring > 250 000 combinations. In the second step, a focused library combining the selected fragments with linkers of different lengths and geometries led to the identification of tight binding adducts that were further investigated for their selective target engagement in pull-down assays, for their affinity by SPR, and their selectivity in a cytotoxicity assay. The best compound showed comparable cellular activity to venetoclax, the first-in-class therapeutic targeting Bcl-2.
An T. H. Le; Svetlana M. Krylova; Stanislav S. Beloborodov; Tong Y. Wang; Ryan Hili; Philip E. Johnson; Feng Li; Rakesh N. Veedu; Svetlana Belyanskaya; Sergey N. Krylov Anal. Chem., 2021, 93(13), 5343–5354 http://www.yorku.ca/skrylov/publications.htmlAbstractScreening molecular libraries for ligands capable of binding proteins is widely used for hit identification in the early drug discovery process. Oligonucleotide libraries provide very high diversity of compounds, while the combination of the polymerase chain reaction and DNA sequencing allow the identification of ligands in low copy numbers selected from such libraries. Ligand selection from oligonucleotide libraries requires mixing the library with the target followed by the physical separation of the ligand–target complexes from the unbound library. Cumulatively, the low abundance of ligands in the library and the low efficiency of available separation methods necessitate multiple consecutive rounds of partitioning. Multiple rounds of inefficient partitioning make the selection process ineffective and prone to failures. There are continuing efforts to develop a separation method capable of reliably generating a pure pool of ligands in a single round of partitioning; however, none of the proposed methods for single-round selection have been universally adopted. Our analysis revealed that the developers’ efforts are disconnected from each other and hindered by the lack of quantitative criteria of selection quality assessment. Here we present a formalism that describes single-round selection mathematically and provides parameters for quantitative characterization of selection quality. We use this formalism to define a universal strategy for development and validation of single-round selection methods. Finally, we analyze the existing partitioning methods, the published single-round selection reports, and some pertinent practical considerations through the prism of this formalism. This formalism is not an experimental protocol, but a framework for correct development of experimental protocols. While single-round selection is not a goal by itself and may not always suffice selection of good-quality ligands, our work will help developers of highly-efficient selection approaches to consolidate their efforts under an umbrella of universal quantitative criteria of method development and assessment.
Nicholas Favalli; Gabriele Bassi; Davide Bianchi; Jörg Scheuermann; Dario Neri Bioorg. Med. Chem., 2021, 116206 https://doi.org/10.1016/j.bmc.2021.116206AbstractProgress in DNA-encoded chemical library synthesis and screening crucially relies on the availability of DNA-compatible reactions, which proceed with high yields and excellent purity for a large number of possible building blocks. In the past, experimental conditions have been presented for the execution of Suzuki and Sonogashira cross-coupling reactions on-DNA. In this article, our aim was to optimize Suzuki and Sonogashira reactions, comparing our results to previously published procedures. We have tested the performance of improved conditions using 606 building blocks (including boronic acids, pinacol boranes and terminal alkynes), achieving > 70% conversion for 84% of the tested molecules. Moreover, we describe efficient experimental conditions for the on-DNA synthesis of amide bonds, starting from DNA derivatives carrying a carboxylic acid moiety and 300 primary, secondary and aromatic amines, as amide bonds are frequently found in DNA-encoded chemical libraries thanks to their excellent DNA compatibility.
Bing Xia; G. Joseph Franklin; Xiaojie Lu; Katie L. Bedard; LaShadric C. Grady; Jennifer D. Summerfield; Eric X. Shi; Bryan W. King; Kenneth E. Lind; Cynthia Chiu; Eleanor Watts; Vera Bodmer; Xiaopeng Bai; Lisa A. Marcaurelle ACS Med. Chem. Lett., 2021, 12(7), 1166-1172 https://doi.org/10.1021/acsmedchemlett.1c00156AbstractDNA-encoded library (DEL) technology is a powerful platform for hit identification in academia and the pharmaceutical industry. When conducting off-DNA resynthesis hit confirmation after affinity selection, PCR/sequencing, and data analysis, one typically assumes a “one-to-one” relationship between the DNA tag and the chemical structure of the attached small-molecule it encodes. Because library synthesis often yields a mixture, this approximation increases the risk of overlooking positive discoveries and valuable information. To address this issue, we apply a library synthesis “recipe” strategy for on-DNA resynthesis using a cleavable linker, followed by direct affinity selection mass spectrometry (AS-MS) evaluation and identification of binder(s) from the released small-molecule mixture. We validate and showcase this approach employing the receptor-interacting-protein kinase 2 (RIP2) DEL campaign. We also designed and developed two cleavable linkers to enable this method, a photocleavable linker (nitrophenyl-based) and acid-labile linker (tetrahydropyranyl ether). The strategy provides an effective means of hit identification and rapid determination of key active component(s) of the mixture.
Rongfeng Wu; Tian Du; Wenbo Sun; Alex Shaginian; Sen Gao; Jin Li; Jinqiao Wan; Guansai Liu Org. Lett., 2021, 23(9), 3486–3490 https://doi.org/10.1021/acs.orglett.1c00924AbstractA highly efficient approach to C(sp3)–C(sp3) bond construction via on-DNA photoredox catalysis between on-DNA alkenes and N-aryl tertiary amines was developed. The methodology demonstrated 55%–95% conversions without obvious DNA damage, as seen by qPCR tests. Furthermore, various functional groups, such as carboxylic acids, aldehydes, and aryl halides, that can be used to create library diversities were shown to be tolerant of the C–H activation conditions.
Mateja Klika Skopic; Christian Gramse; Rosario Oliva; Sabrina Pospich; Laura Neukirch; Magiliny Manisegaran; Stefan Raunser; Roland Winter; Ralf Weberskirch; Andreas Brunschweiger Chem. Eur. J., 2021, 27(39), 10048-10057 https://doi.org/10.1002/chem.202100980AbstractThe development of DNA‐compatible reaction methodologies is a central theme to advance DNA‐encoded screening library technology. Recently, we could show that sulfonic acid functionalized block copolymer micelles facilitated Brønsted acid promoted reactions such as the Povarov reaction on DNA‐coupled starting materials with minimal DNA degradation. Here, we investigated the impact of polymer composition on micelle shape, and reaction conversion. A dozen sulfonic acid‐functionalized block copolymers of different molar mass and composition were prepared by RAFT‐polymerization and were tested in the Povarov reaction, removal of the Boc protective group, and the Biginelli reaction. The results showed trends of polymer structure ‐ micellar catalytic activity relationship. For instance, micelles composed of block copolymers with shorter acrylate ester chains formed smaller particles and tended to provide faster reaction kinetics. Moreover, fluorescence quenching experiments as well as circular dichroism spectroscopy showed that DNA‐oligomer‐conjugates, although highly water‐soluble, accumulated very effectively in the micellar compartments as an essential prerequisite for carrying out a DNA‐encoded reaction in the presence of polymer micelles.
Ying Zhang; Matthew A. Clark Bioorg. Med. Chem., 2021, 116189 https://doi.org/10.1016/j.bmc.2021.116189AbstractAn approach of building block (BB) inclusivity and atom efficient library schemes deliver the quality and diversity of DNA-encoded libraries best suited for small molecule drug discovery. In this Perspective, we offer key learnings in DEL design from a decade’s worth of DEL-driven screening. It has been over 10 years since synthesis and screening of non-templated, non-bead based DNA-encoded libraries (DELs) were first reported.1 Since that time, numerous groups have sought to increase the scope of synthetic chemistry that can be conducted on DNA. Many successes have been reported such as metal-catalyzed C-C bond formation,2 photoredox chemistry,3–5 enzymatic catalysis,6 solid-phase immobilization,7 and multi-component condensations.8 Despite this growing body of work, there has been less attention paid to the design considerations of DELs.9 In this Perspective, we share our opinions around DEL library design. These opinions are based on a decade of work in applying DELs to drug hit generation. We articulate why the quality and diversity of DELs is driven primarily by the building blocks, and how this consideration influences the kinds of chemistry that are most useful for DEL synthesis. X-Chem has a long history in discovery of peptides, macrocycles and covalent inhibitors. Our libraries contain over a hundred billion compounds in these classes. This Perspective, however, focuses on conventional small molecule hit discovery. Our opinions are rooted in a conservative approach to physicochemical properties of drugs and leads. While recent work has shown that there can be promise “beyond the Rule of 5”,10 we believe that compact and ligand efficient starting points have the best chance of surviving the attrition of medicinal chemistry.
Sijie Wang; Aktan Alpsoy; Surbhi Sood; Sandra Carolina Ordonez-Rubiano; Alisha Dhiman; Yixing Sun; Guanming Jiao; Casey J. Krusemark; Emily Dykhuizen ChemBioChem, 2021, 22, 2335-2344 https://doi.org/10.1002/cbic.202100118AbstractPolycomb group (PcG) proteins are epigenetic regulators that facilitate both embryonic development and cancer progression. PcG proteins form Polycomb repressive complexes 1 and 2 (PRC1 and PRC2). PRC2 trimethylates histone H3 lysine 27 (H3K27me3), a histone mark recognized by the N‐terminal chromodomain (ChD) of the CBX subunit of canonical PRC1. There are five PcG CBX paralogs in humans. CBX2, in particular, is upregulated in a variety of cancers, particularly in advanced prostate cancers. Using CBX2 inhibitors to understand and target CBX2 in prostate cancer is highly desirable; however, high structural similarity among the CBX ChDs has been challenging for developing selective CBX ChD inhibitors. Here, we utilize selections of focused DNA encoded libraries (DELs) for the discovery of a selective CBX2 chromodomain probe, SW2_152F. SW2_152F binds to CBX2 ChD with a K d of 80 nM and displays 24‐1000‐fold selectivity for CBX2 ChD over other CBX paralogs in vitro . SW2_152F is cell permeable, selectively inhibits CBX2 chromatin binding in cells, and blocks neuroendocrine differentiation of prostate cancer cell lines in response to androgen deprivation.
Clément Bechtler; Christina Lamers RSC Med. Chem., 2021,12, 1325-1351 https://doi.org/10.1039/D1MD00083GAbstractPeptides are a growing therapeutic class due to their unique spatial characteristics that can target traditionally “undruggable” protein–protein interactions and surfaces. Despite their advantages, peptides must overcome several key shortcomings to be considered as drug leads, including their high conformational flexibility and susceptibility to proteolytic cleavage. As a general approach for overcoming these challenges, macrocyclization of a linear peptide can usually improve these characteristics. Their synthetic accessibility makes peptide macrocycles very attractive, though traditional synthetic methods for macrocyclization can be challenging for peptides, especially for head-to-tail cyclization. This review provides an updated summary of the available macrocyclization chemistries, such as traditional lactam formation, azide–alkyne cycloadditions, ring-closing metathesis as well as unconventional cyclization reactions, and it is structured according to the obtained functional groups. Keeping peptide chemistry and screening in mind, the focus is given to reactions applicable in solution, on solid supports, and compatible with contemporary screening methods.
Elena Lenci; Lorenzo Baldini; Andrea Trabocchi Bioorg. Med. Chem., 2021, 116218 https://doi.org/10.1016/j.bmc.2021.116218AbstractDNA-encoded libraries (DEL) represent a powerful technology for generating compound collections for drug discovery campaigns, that have allowed for the selection of many hit compounds over last three decades. However, the application of split-and-pool combinatorial methodologies, as well as the limitation imposed by DNA-compatible chemistry, has often brought to a limited exploration of the chemical space, with an over-representation of flat aromatic or peptide-like structures, whereas a higher scaffold complexity is generally associated with a more successful biological of the library. In this context, the application of Diversity-Oriented Synthesis, capable of creating sp3-rich molecular entities even starting from simple flat building blocks, can represent an efficient strategy to significantly broaden the chemical space explored by DELs. In this review, we present selected examples of DNA-compatible complexity-generating reactions that can be applied for the generation of DNA-encoded DOS libraries, including: (i) multicomponent reactions; (ii) C-H/C-X functionalization; (iii) tandem approaches; (iv) cycloadditions; (v) reactions introducing privileged elements. Also, selected case studies on the generation of DELs with high scaffold diversity are discussed, reporting their application in drug discovery programs.
Francesco Rianjongdee; Stephen J. Atkinson; Chun-wa Chung; Paola Grandi; James R. J. Gray; Laura J. Kaushansky; Patricia Medeiros; Cassie Messenger; Alex Phillipou; Alex Preston; Rab K. Prinjha; Inmaculada Rioja; Alexander L. Satz; Simon Taylor; Ian D. Wall; Robert J. Watson; Gang Yao; Emmanuel H. Demont J. Med. Chem., 2021, 64(15), 10806-10833 https://doi.org/10.1021/acs.jmedchem.1c00412AbstractSecond-generation bromodomain and extra terminal (BET) inhibitors, which selectively target one of the two bromodomains in the BET proteins, have begun to emerge in the literature. These inhibitors aim to help determine the roles and functions of each domain and assess whether they can demonstrate an improved safety profile in clinical settings compared to pan-BET inhibitors. Herein, we describe the discovery of a novel BET BD2-selective chemotype using a structure-based drug design from a hit identified by DNA-encoded library technologies, showing a structural differentiation from key previously reported greater than 100-fold BD2-selective chemotypes GSK620, GSK046, and ABBV-744. Following a structure-based hypothesis for the selectivity and optimization of the physicochemical properties of the series, we identified 60 (GSK040), an in vitro ready and in vivo capable BET BD2-inhibitor of unprecedented selectivity (5000-fold) against BET BD1, excellent selectivity against other bromodomains, and good physicochemical properties. This novel chemical probe can be added to the toolbox used in the advancement of epigenetics research.
Christopher P. Austin Clin. Transl. Sci., 2021, 14, 1629–1647 https://doi.org/10.1111/cts.13055AbstractThe mission of translational science is to bring predictivity and efficiency to the development and dissemination of interventions that improve human health. Ten years ago this year, the National Center for Advancing Translational Sciences was founded to embody, conduct, and support this new discipline. The Center’s first decade has brought substantial progress across a broad range of translational areas, from diagnostic and drug development to clinical trials to implementation science to education. The origins of the translational science and advances to this point are reviewed here and allow the establishment of an ambitious future research agenda for the field.
Fei Ma; Jie Li; Shuning Zhang; Yuang Gu; Tingting Tan; Wanting Chen; Shuyue Wang; Peixiang Ma; Hongtao Xu; Guang Yang; Richard Alan Lerner Chem. Eur. J., 2021, 27(31), 8214-8220 https://doi.org/10.1002/chem.202100850AbstractDNA‐encoded combinatorial chemical library (DEL) technology, an approach that combines the power of genetics and chemistry, has emerged as an invaluable tool in drug discovery. Skeletal diversity plays a fundamental importance in DEL applications, and relies heavily on novel DNA‐compatible chemical reactions. We report herein a phylogenic chemical transformation strategy using DNA‐conjugated benzoyl hydrazine as a common versatile precursor in azole chemical expansion of DELs. DNA‐compatible reactions deriving from the common benzoyl hydrazine precursor showed excellent functional group tolerance with exceptional efficiency in the synthesis of various azoles, including oxadiazoles, thiadiazoles, and triazoles, under mild reaction conditions. The phylogenic chemical transformation strategy provides DELs a facile way to expand into various unique chemical spaces with privileged scaffolds and pharmacophores.
Sebastian Oehler; Marco Catalano; Ilario Scapozza; Martina Bigatti; Gabriele Bassi; Nicholas Favalli; Michael R. Mortensen; Florent Samain; Jörg Scheuermann; Dario Neri Chem. Eur. J., 2021, 27(35), 8985-8993 https://doi.org/10.1002/chem.202100816AbstractDNA‐encoded chemical libraries are typically screened against purified protein targets. Recently, cell‐based selections with encoded chemical libraries have been described, commonly revealing suboptimal performance due to insufficient recovery of binding molecules. We used carbonic anhydrase IX (CAIX)‐expressing tumor cells as a model system to optimize selection procedures with code‐specific quantitative polymerase chain reaction (qPCR) as selection readout. Salt concentration and performing PCR on cell suspension had the biggest impact on selection performance, leading to 15‐fold enrichment factors for high‐affinity monovalent CAIX binders (acetazolamide; K D = 8.7 nM). Surprisingly, the homobivalent display of acetazolamide at the extremities of both complementary DNA strands led to a substantial improvement of both ligand recovery and enrichment factors (above 100‐fold). The optimized procedures were used for selections with a DNA‐encoded chemical library comprising 1 million members against tumor cell lines expressing CAIX, leading to a preferential recovery of known and new ligands against this validated tumor‐associated target. This work may facilitate future affinity selections on cells against target proteins which might be difficult to express otherwise.
Ying Yang; Kun Yao; Matthew P. Repasky; Karl Leswing; Robert Abel; Brian Shoichet; Steven Jerome ChemRxiv, 2021 https://doi.org/10.26434/chemrxiv.14153819.v1AbstractWith the advent of make-on-demand commercial libraries, the number of purchasable compounds available for virtual screening and assay has grown explosively in recent years, with several libraries eclipsing one billion compounds. Today’s screening libraries are larger and more diverse, enabling discovery of more potent hit compounds and unlocking new areas of chemical space, represented by new core scaffolds. Applying physics-based in-silico screening methods in an exhaustive manner, where every molecule in the library must be enumerated and evaluated independently, is increasingly cost-prohibitive. Here, we introduce a protocol for machine learning-enhanced molecular docking based on active learning to dramatically increase throughput over traditional docking. We leverage a novel selection protocol that strikes a balance between two objectives: (1) Identifying the best scoring compounds and (2) exploring a large region of chemical space, demonstrating superior performance compared to a purely greedy approach. Together with automated redocking of the top compounds, this method captures nearly all the high scoring scaffolds in the library found by exhaustive docking. This protocol is applied to our recent virtual screening campaigns against the D4 and AMPC targets that produced dozens of highly potent, novel inhibitors, and a blinded test against the MT1 target. Our protocol recovers more than 80% of the experimentally confirmed hits with a 14-fold reduction in compute cost, and more than 90% of the hit scaffolds in the top 5% of model predictions, preserving the diversity of the experimentally confirmed hit compounds.
Muhammad Jbara; Jacob Rodriguez; Heemal H. Dhanjee; Andrei Loas; Stephen L. Buchwald; Bradley L. Pentelute Angew. Chem. Int. Ed., 2021, 60(21), 12109-12115 https://doi.org/10.1002/anie.202103180AbstractOrganometallic reagents enable practical strategies for bioconjugation. Innovations in the design of water‐soluble ligands and the enhancement of reaction rates have allowed for chemoselective cross‐coupling reactions of peptides and proteins to be carried out in water. There are currently no organometallic‐based methods for oligonucleotide bioconjugation to other biomolecules. Here we report bifunctional palladium(II)‐oxidative addition complexes (OACs) as reagents for high‐yielding oligonucleotide bioconjugation reactions. These bifunctional OACs react chemoselectively with amine‐modified oligonucleotides to generate the first isolable, bench stable oligonucleotide‐palladium(II) OACs. These complexes undergo site‐selective C‐S arylation with a broad range of native thiol‐containing biomolecules at low micromolar concentrations in under one hour. This approach provided oligonucleotide‐peptide, oligonucleotide‐protein, oligonucleotide‐small molecule, and oligonucleotide‐oligonucleotide conjugates in >80% yield and afforded conjugation of multiple copies of oligonucleotides onto a monoclonal antibody.
Franco Scalambra; Pablo Lorenzo-Luis; Isaac de los Rios; Antonio Romerosa Coord. Chem. Rev., 2021, 443, 213997 https://doi.org/10.1016/j.ccr.2021.213997AbstractThe C–C coupling is the best strategy to obtain new organic molecules and building blocks from economical and abundant small molecules. Traditionally these reactions are accomplished in organic solvents, which are often dried before use as water molecules can react quickly with both the catalyst or the intermediate of the reactions, blocking the catalytic cycle. Nevertheless, the efforts of the researchers are providing new synthetic procedures in water that can substitute some of those, previously developed, “dry reactions”. In the last years, really useful processes to obtain C–C bonds in water were proposed. These reactions usually are catalysed with 4d-metals but also examples with metals of the first transition row were proposed. This review aims to summarize and order by types the C–C coupling catalytic reactions in water published during the last five years. The abundance of reactions and the significance of the published results shows that a long way is needed to achieve a good knowledge of catalytic C–C coupling reactions in water. Nevertheless, accomplish this objective can provide significant and valuable synthetic processes, more economical, cleaner, more selective, therefore friendlier and more respectful with nature.
Anokha S. Ratnayake; Mark E. Flanagan; Timothy L. Foley; Scott L. Hultgren; Justin Bellenger; Justin I. Montgomery; Manjinder S. Lall; Bo Liu; Tim Ryder; Dominik K. Kölmel; Andre Shavnya; Xidong Feng; Bruce Lefker; Laura J. Byrnes; Parag V. Sahasrabudhe; Kathleen A. Farley; Shi Chen; Jinqiao Wan Bioorg. Med. Chem., 2021, 116205 https://doi.org/10.1016/j.bmc.2021.116205AbstractThe ability to predict chemical structure from DNA sequence has to date been a necessary cornerstone of DNA-encoded library technology. DNA-encoded libraries (DELs) are typically screened by immobilized affinity selection and enriched library members are identified by counting the number of times an individual compound’s sequence is observed in the resultant dataset. Those with high signal reads (DEL hits) are subsequently followed up through off-DNA synthesis of the predicted small molecule structures. However, hits followed-up in this manner often fail to translate to confirmed ligands. To address this low conversion rate of DEL hits to off-DNA ligands, we have developed an approach that eliminates the reliance on chemical structure prediction from DNA sequence. Here we describe our method of combining non-combinatorial resynthesis on-DNA following library procedures as a rapid means to assess the probable molecules attached to the DNA barcode. Furthermore, we apply our Bead-Assisted Ligand Isolation Mass Spectrometry (BALI-MS) technique to identify the true binders found within the mixtures of on-DNA synthesis products. Finally, we describe a Normalized Enrichment (NE) metric that allows for the quantitative assessment of affinity selection in these studies. We exemplify how this combined approach enables the identification of putative hit matter against a clinically relevant therapeutic target bisphosphoglycerate mutase, BPGM.
John P. Guilinger; Archna Archna; Martin Augustin; Andreas Bergmann; Paolo A. Centrella; Matthew A. Clark; John W. Cuozzo; Maike Däther; MarieAude Guié; Sevan Habeshian; Reiner Kiefersauer; Stephan Krapp; Alfred Lammens; Lukas Lercher; Julie Liu; Yanbin Liu; Klaus Maskos; Michael Mrosek; Klaus Pflügler; Markus Siegert; Heather A. Thomson; Xia Tian; Ying Zhang; Debora L. Konz Makino; Anthony D. Keefe Bioorg. Med. Chem., 2021, 116223 https://doi.org/10.1016/j.bmc.2021.116223AbstractLibraries of DNA-Encoded small molecules created using combinatorial chemistry and synthetic oligonucleotides are being applied to drug discovery projects across the pharmaceutical industry. The majority of reported projects describe the discovery of reversible, i.e. non-covalent, target modulators. We synthesized multiple DNA-encoded chemical libraries terminated in electrophiles and then used them to discover covalent irreversible inhibitors and report the successful discovery of acrylamide- and epoxide-terminated Bruton’s Tyrosine Kinase (BTK) inhibitors. We also demonstrate their selectivity, potency and covalent cysteine engagement using a range of techniques including X-ray crystallography, thermal transition shift assay, reporter displacement assay and intact protein complex mass spectrometry. The epoxide BTK inhibitors described here are the first ever reported to utilize this electrophile for this target.
Jinming Shan; Xing Ling; JiaXiang Liu; Xuan Wang; Xiaojie Lu Bioorg. Med. Chem., 2021, 116234 https://doi.org/10.1016/j.bmc.2021.116234AbstractWe described a mode of catalytic activation that accomplished the α-alkylation of N-Boc saturated heterocycles with DNA-linked acrylamide via photoredox-mediated hydrogen atom transfer (HAT) catalysis. This C(sp3)-C(sp3) bond formation reaction tolerated five-, six- and seven-membered cyclic substrates, substantially streamline synthetic efforts to functionalize the α-position of heterocycles with native C-H functional handle. This photoredox catalyzed C-H functionalization proceeded in mild DNA-compatible condition, and suited for the construction of DNA-encoded libraries.
Tuo Yang; Sabine Ottilie; Eva S. Istvan; Karla P. Godinez-Macias; Amanda K. Lukens; Beatriz Baragaña; Brice Campo; Chris Walpole; Jacquin C. Niles; Kelly Chibale; Koen J. Dechering; Manuel Llinás; Marcus C.S. Lee; Nobutaka Kato; Susan Wyllie; Case W. McNamara; Francisco Javier Gamo; Jeremy Burrows; David A. Fidock; Daniel E. Goldberg; Ian H. Gilbert; Dyann F. Wirth; Elizabeth A. Winzeler; The Malaria Drug Accelerator Consortium Trends Parasitol., 2021, 37(6), 493-507 https://doi.org/10.1016/j.pt.2021.01.009AbstractThe Malaria Drug Accelerator (MalDA) is a consortium of 15 leading scientific laboratories. The aim of MalDA is to improve and accelerate the early antimalarial drug discovery process by identifying new, essential, druggable targets. In addition, it seeks to produce early lead inhibitors that may be advanced into drug candidates suitable for preclinical development and subsequent clinical testing in humans. By sharing resources, including expertise, knowledge, materials, and reagents, the consortium strives to eliminate the structural barriers often encountered in the drug discovery process. Here we discuss the mission of the consortium and its scientific achievements, including the identification of new chemically and biologically validated targets, as well as future scientific directions.
Selahattin Ede; Mandy Schenk; Donald Bierer; Hilmar Weinmann; Keith Graham Molecules, 2021, 26(6), 1790 https://doi.org/10.3390/molecules26061790AbstractDNA-encoded libraries (DEL) are increasingly being used to identify new starting points for medicinal chemistry in drug discovery. Herein, we discuss the development of methods that allow the conversion of both primary amines and anilines, attached to DNA, to their corresponding azides in excellent yields. The scope of these diazo-transfer reactions was investigated, and a proof-of-concept has been devised to allow for the synthesis of macrocycles on DNA.
Jing Zhu; Meng Xin; Congcong Xu; Yuan He; Wannian Zhang; Zhibin Wang; Chunlin Zhuang Acta Pharm. Sin. B, 2021, 11(10), 3193-3205 https://doi.org/10.1016/j.apsb.2021.05.017AbstractReceptor-interacting protein (RIP) kinase 1 is involved in immune-mediated inflammatory diseases including ulcerative colitis (UC) by regulating necroptosis and inflammation. Our group previously identified TAK-632 (5) as an effective necroptosis inhibitor by dual-targeting RIP1 and RIP3. In this study, using ligand-based substituent-anchoring design strategy, we focused on the benzothiazole ring to obtain a series of TAK-632 analogues showing significantly improving on the anti-necroptosis activity and RIP1 selectivity over RIP3. Among them, a conformational constrained fluorine-substituted derivative (25) exhibited 333-fold selectivity for RIP1 (Kd = 15 nmol/L) than RIP3 (Kd > 5000 nmol/L). This compound showed highly potent activity against cell necroptosis (EC50 = 8 nmol/L) and systemic inflammatory response syndrome (SIRS) induced by TNF-α in vivo. Especially, it was able to exhibit remarkable anti-inflammatory treatment efficacy in a DSS-induced mouse model of UC. Taken together, the highly potent, selective, orally active anti-necroptosis inhibitor represents promising candidate for clinical treatment of UC.
Keun Ah Ryu; Christina M. Kaszuba; Noah B. Bissonnette; Rob C. Oslund; Olugbeminiyi O. Fadeyi Nat. Rev. Chem., 2021, 5, 322-337 https://doi.org/10.1038/s41570-021-00265-6AbstractLight-powered catalysis has found broad utility as a chemical transformation strategy, with widespread impact on energy, environment, drug discovery and human health. A noteworthy application impacting human health is light-induced sensitization of cofactors for photodynamic therapy in cancer treatment. The clinical adoption of this photosensitization approach has inspired the search for other photochemical methods, such as photoredox catalysis, to influence biological discovery. Over the past decade, light-mediated catalysis has enabled the discovery of valuable synthetic transformations, propelling it to become a highly utilized chemical synthesis strategy. The reaction components required to achieve a photoredox reaction are identical to photosensitization (catalyst, light source and substrate), making it ideally suited for probing biological environments. In this Review, we discuss the therapeutic application of photosensitization and advancements made in developing next-generation catalysts. We then highlight emerging uses of photoredox catalytic methods for protein bioconjugation and probing complex cellular environments in living cells.
Qi Liang; Xue Zhao; Xiaoying Fu; Jing Wang; Qian Li; Xinfeng Zhao Bioorg. Chem., 2021, 104986 https://doi.org/10.1016/j.bioorg.2021.104986AbstractThe rapid growth of demands for drug discovery has necessitated the ongoing pursuit of new methods for specific ligands screening and identification. This work combined receptor-affinity chromatography (RAC) with high-throughput sequencing techniques to rapidly screen and identify the specific ligands. By this method, immobilized angiotensin II type I receptor(AT1R) and endothelin receptor A(ETAR) based on RAC were utilized for lead screening from a DNA-encoded library. The specific ligands of AT1R (ligand A1, A2) and ETAR (ligand B1, B2) were synthesized after decoding by high-throughput sequencing techniques. The dissociation rate constants(kd) of ligand A1, A2 to AT1R and B1, B2 to ETAR were 9.65×10-4, 31.1×10-4 and 0.66, 1.22 s-1 by peak profiling assay. The association constant (KA) to the receptors of four ligands was 5.4×106, 3.3×106 and 1.6×106, 2.2×105 by injection amount dependent method. The kinetic and thermodynamic parameters of the four specific ligands are similar to those of the positive drugs. This indicates that they are promising to a drug candidate. The druggability of the four ligands through pharmacokinetic investigation by HPLC-MS/MS presented desired pharmacokinetic behavior including the fast absorption, the relatively slow elimination. These results, taking together, indicated that the RAC combined with high-throughput sequencing techniques can screen and identify the specific ligands according to various proteins, thus creating a general strategy for rapid discovery of promising drug candidates.
Yiran Huang; Yuqing Deng; Jianfu Zhang; Ling Meng; Xiaoyu Li Chem. Commun., 2021, 57, 3769-3772 https://doi.org/10.1039/D1CC00961CAbstractMembrane proteins are important drug targets; however, ligand discovery for membrane proteins is highly challenging due to their hydrophobic nature. We show that membrane proteins may be specifically labelled with a DNA tag by DNA-programmed affinity labelling (DPAL), thereby enabling the screening of chemical compounds against membrane proteins directly on live cells.
Yiyuan Zhang; Wanting Chen; Tingting Tan; Yuang Gu; Shuning Zhang; Jie Li; Yan Wang; Wei Hou; Guang Yang; Peixiang Ma; Hongtao Xu Chem. Commun., 2021, 57, 4588-4591 https://doi.org/10.1039/D1CC00769FAbstractWe report a general palladium-catalyzed one-pot procedure for the synthesis of phosphonates, phosphinates and phosphine oxides from phenols mediated by sulfuryl fluoride. It features mild conditions, broad substrate scope, high functionality tolerance and water insensitive. Its utility has been well demonstrated by gram-scale synthesis, sequential synthesis of click chemistry building blocks, late-stage decoration of drugs and natural products and on-DNA synthesis of phosphine oxide for DNA-encoded library (DEL).
Yue Ji; Dongliang Dai; Huadong Luo; Simin Shen; Jing Fan; Zhao Wang; Min Chen; Jinqiao Wan; Jin Li; Huiyong Ma; Guansai Liu Bioconjugate Chem., 2021, 32(4), 685–689 https://doi.org/10.1021/acs.bioconjchem.1c00076AbstractThioethers have been widely found in biologically active compounds, including pharmaceuticals. In this report, a highly efficient approach to on-DNA construction of thioethers via Cu-promoted Ullmann cross-coupling between DNA-conjugated aryl iodides and thiols is developed. This methodology was demonstrated with medium to high yields, without obvious DNA damage. This reported reaction has strong potential for application in DNA-encoded chemical library synthesis.
Peng Yang; Xuan Wang; Bo Li; Yixuan Yang; Jinfeng Yue; Yanrui Suo; Huarong Tong; Gang He; Xiaojie Lu; Gong Chen Chem. Sci., 2021, 12, 5804-5810 https://doi.org/10.1039/D1SC00789KAbstractA highly efficient and versatile method for construction of peptide macrocycles via palladium-catalyzed intramolecular S-arylation of alkyl and aryl thiols with aryl iodides under mild conditions is developed. The method exhibits a broad substrate scope for thiols, aryl iodides and amino acid units. Peptide macrocycles of a wide range of size and composition can be readily assembled in high yield from various easily accessible building blocks. This method has been successfully employed to prepare an 8-million-membered tetrameric cyclic peptide DNA-encoded library (DEL). Preliminary screening of the DEL library against protein p300 identified compounds with single digit micromolar inhibition activity.
Peter Stacey; Hannah Lithgow; Xiao Lewell; Agnieszka Konopacka; Stephen Besley; Georgina Green; Ryan Whatling; Robert Law; Sascha Röth; Gopal P. Sapkota; Ian E. D. Smith; Glenn A. Burley; John Harling; Andrew B. Benowitz; Markus A. Queisser; Marcel Muelbaier SLAS Discov., 2021, 26(7), 885-895 https://doi.org/10.1177/24725552211017517AbstractTargeted protein degradation is an emerging new strategy for the modulation of intracellular protein levels with applications in chemical biology and drug discovery. One approach to enable this strategy is to redirect the ubiquitin–proteasome system to mark and degrade target proteins of interest (POIs) through the use of proteolysis targeting chimeras (PROTACs). Although great progress has been made in enabling PROTACs as a platform, there are still a limited number of E3 ligases that have been employed for PROTAC design. Herein we report a novel phenotypic screening approach for the identification of E3 ligase binders. The key concept underlying this approach is the high-throughput modification of screening compounds with a chloroalkane moiety to generate HaloPROTACs in situ, which were then evaluated for their ability to degrade a GFP-HaloTag fusion protein in a cellular context. As proof of concept, we demonstrated that we could generate and detect functional HaloPROTACs in situ, using a validated Von Hippel–Lindau (VHL) binder that successfully degraded the GFP-HaloTag fusion protein in living cells. We then used this method to prepare and screen a library of approximately 2000 prospective E3 ligase-recruiting molecules.
Silke Miller; Maria-Jesus Blanco RSC Med. Chem., 2021, 12, 871-886 https://doi.org/10.1039/D1MD00036EAbstractChronically activated microglia and the resulting cascade of neuroinflammatory mechanisms have been postulated to play a critical role in neurodegenerative disorders. Microglia are the main component of the brain's innate immune system and become activated by infection, injury, misfolded proteins or a multitude of other stimuli. Activated microglia release pro-inflammatory and cytotoxic factors that can damage neurons and transform astrocytes to become toxic to neurons as well. Therapeutic approaches aiming to modulate microglia activation may be beneficial to mitigate the progression of inflammatory-mediated neurodegenerative diseases. In this literature review, we provide an overview of recent progress on key microglia targets and discovery of small molecule compounds advancing in clinical trials to minimize neuroinflammation.
Wentao Liu; Wei Huang; Qian Lin; Mei-Hsuan Tsai; Rui Zhang; Lijun Fan; Jack D. Scott; Guansai Liu; Jinqiao Wan Bioorg. Med. Chem., 2021, 116118 https://doi.org/10.1016/j.bmc.2021.116118AbstractA robust palladium-catalyzed hydroxycarbonylation of aryl halides on DNA has been developed. Instead of Mo(CO)6 as a source of carbon monoxide as previously described in the literature, chloroform was used as a surrogate in this report for the purpose of avoiding to use a large excess of molybdenum reagent which is not totally soluble in aqueous reaction mixtures.
Arunika I. Ekanayake; Lena Sobze; Payam Kelich; Jihea Youk; Nicholas J. Bennett; Raja Mukherjee; Atul Bhardwaj; Frank Wuest; Lela Vukovic; Ratmir Derda J. Am. Chem. Soc., 2021, 143(14), 5497-5507 https://doi.org/10.1021/jacs.1c01186AbstractGenetically encoded macrocyclic peptide libraries with unnatural pharmacophores are valuable sources for the discovery of ligands for many targets of interest. Traditionally, generation of such libraries employs “early stage” incorporation of unnatural building blocks into the chemically or translationally produced macrocycles. Here, we describe a divergent late-stage approach to such libraries starting from readily available starting material: genetically encoded libraries of peptides. A diketone linchpin 1,5-dichloropentane-2,4-dione converts peptide libraries displayed on phage to 1,3-diketone bearing macrocyclic peptides (DKMP): shelf-stable precursors for Knorr pyrazole synthesis. Ligation of diverse hydrazine derivatives onto DKMP libraries displayed on phage that carries silent DNA-barcodes yields macrocyclic libraries in which the amino acid sequence and the pharmacophore are encoded by DNA. Selection of this library against carbonic anhydrase enriched macrocycles with benzenesulfonamide pharmacophore and nanomolar Kd. The methodology described in this manuscript can graft diverse pharmacophores into many existing genetically encoded phage libraries and significantly increase the value of such libraries in molecular discoveries.
Grégoire J.B. Philippe; David J. Craik; Sónia T. Henriques Drug Discov. Today, 2021, 26(6), 1521-1531 https://doi.org/10.1016/j.drudis.2021.01.022AbstractPeptides are gaining increasing attention as therapeutics to target intracellular protein–protein interactions that are involved in disease progression. In this review, we discuss how peptides that are able to bind and inhibit a therapeutic target can be translated into drug leads. We discuss the advantages of using peptides as therapeutics to target intracellular protein–protein interactions, chemical strategies to generate macrocyclic peptides that are resistant to proteolytic enzymes, high-throughput screening approaches to identify peptides that have high affinity for therapeutic targets, strategies that permit these peptides to cross cell membranes and so reach intracellular targets, and the importance of investigating their mode-of-action in guiding the development of novel therapeutics.
Yi Li; Jianping Chen; Andrew A Bolinger; Haiying Chen; Zhiqing Liu; Yingzi Cong; Allan R Brasier; Irina V Pinchuk; Bing Tian; Jia Zhou Inflamm. Bowel Dis., 2021, 27, S38–S62 https://doi.org/10.1093/ibd/izab190AbstractInflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn’s disease (CD), is a class of severe and chronic diseases of the gastrointestinal (GI) tract with recurrent symptoms and significant morbidity. Long-term persistence of chronic inflammation in IBD is a major contributing factor to neoplastic transformation and the development of colitis-associated colorectal cancer. Conversely, persistence of transmural inflammation in CD is associated with formation of fibrosing strictures, resulting in substantial morbidity. The recent introduction of biological response modifiers as IBD therapies, such as antibodies neutralizing tumor necrosis factor (TNF)-α, have replaced nonselective anti-inflammatory corticosteroids in disease management. However, a large proportion (~40%) of patients with the treatment of anti-TNF-α antibodies are discontinued or withdrawn from therapy because of (1) primary nonresponse, (2) secondary loss of response, (3) opportunistic infection, or (4) onset of cancer. Therefore, the development of novel and effective therapeutics targeting specific signaling pathways in the pathogenesis of IBD is urgently needed. In this comprehensive review, we summarize the recent advances in drug discovery of new small molecules in preclinical or clinical development for treating IBD that target biologically relevant pathways in mucosal inflammation. These include intracellular enzymes (Janus kinases, receptor interacting protein, phosphodiesterase 4, IκB kinase), integrins, G protein-coupled receptors (S1P, CCR9, CXCR4, CB2) and inflammasome mediators (NLRP3), etc. We will also discuss emerging evidence of a distinct mechanism of action, bromodomain-containing protein 4, an epigenetic regulator of pathways involved in the activation, communication, and trafficking of immune cells. We highlight their chemotypes, mode of actions, structure-activity relationships, characterizations, and their in vitro/in vivo activities and therapeutic potential. The perspectives on the relevant challenges, new opportunities, and future directions in this field are also discussed.
Guanyu Yang; Dou He; Yijun Zhu; Weiwei Zhu; Yang Tan; Xingwen Long; Jinqiao Wan; Zhicai Shi; David Schuman; Pratik Chheda; Nicholas Simmons; Guansai Liu Bioconjugate Chem., 2021, 32(4), 667–671 https://doi.org/10.1021/acs.bioconjchem.1c00045AbstractWe report two cholesterol-modified oligonucleotides for use as internal controls for on-DNA reactions during the pooled stages of a DNA-encoded chemical library (DECL) synthesis. As these cholesterol-tagged oligonucleotides are chromatographically separable from normal DECL intermediates, they can be directly monitored by mass spectrometry to track reaction progression within a complex pool of DNA. We observed similar product conversions for reactions on substrates linked to a standard DECL DNA headpiece, to the cholesterol-modified oligonucleotides, and to the cholesterol-modified oligonucleotides while in the presence of pooled DECL synthetic intermediates—validating their use as a representative control. We also highlight an example from a DECL production in which the use of the cholesterol-modified oligonucleotides provided quality control information that guided synthetic decisions. We conclude that the use of cholesterol-modified oligonucleotides as a regular control will significantly improve the quality of DECL productions.
Robert B. A. Quinlan; Paul E. Brennan RSC Chem. Biol., 2021, 2, 759-795 https://doi.org/10.1039/D1CB00016KAbstractIn recent years chemical probes have proved valuable tools for the validation of disease-modifying targets, facilitating investigation of target function, safety, and translation. Whilst probes and drugs often differ in their properties, there is a belief that chemical probes are useful for translational studies and can accelerate the drug discovery process by providing a starting point for small molecule drugs. This review seeks to describe clinical candidates that have been inspired by, or derived from, chemical probes, and the process behind their development. By focusing primarily on examples of probes developed by the Structural Genomics Consortium, we examine a variety of epigenetic modulators along with other classes of probe.
Richard Houska; Marvin Björn Stutz; Oliver Seitz Chem. Sci., 2021, 12, 13450-13457 https://doi.org/10.1039/D1SC00513HAbstractWe describe a reaction system that enables the synthesis of Bcr–Abl tyrosine kinase inhibitors (TKI) via benzanilide formation in water. The reaction is based on native chemical ligation (NCL). In contrast to previous applications, we used the NCL chemistry to establish aromatic rather than aliphatic amide bonds in coupling reactions between benzoyl and o-mercaptoaniline fragments. The method was applied for the synthesis of thiolated ponatinib and GZD824 derivatives. Acid treatment provided benzothiazole structures, which opens opportunities for diversification. Thiolation affected the affinity for Abl1 kinase only moderately. Of note, a ponatinib-derived benzothiazole also showed nanomolar affinity. NCL-enabled benzanilide formation may prove useful for fragment-based drug discovery. To show that benzanilide synthesis can be put under the control of a template, we connected the benzoyl and o-mercaptoaniline fragments to DNA and peptide nucleic acid (PNA) oligomers. Complementary RNA templates enabled adjacent binding of reactive conjugates triggering a rapid benzoyl transfer from a thioester-linked DNA conjugate to an o-mercaptoaniline-DNA or -PNA conjugate. We evaluated the influence of linker length and unpaired spacer nucleotides within the RNA template on the product yield. The data suggest that nucleic acid-templated benzanilide formation could find application in the establishment of DNA-encoded combinatorial libraries (DEL).
Chunnian Xia; Zhengguang Yao; Lijuan Xu; Wannian Zhang; Haihu Chen; Chunlin Zhuang Eur. J. Med. Chem., 2021, 220, 113484 https://doi.org/10.1016/j.ejmech.2021.113484AbstractNecroptosis is reported to play a critical role in contributing to a variety of human pathologies. The benzoxazepinone GSK′772 is a potent necroptosis inhibitor optimized using a hit from a DNA-encoded library, which is currently in phase II clinical trials for psoriasis, rheumatoid arthritis, and ulcerative colitis. In the present study, the bioisosterism strategy was applied to replace the amide and benzene ring of GSK′772 based on the co-crystal structure of GSK′772 with its binding target RIPK1. As a result, the novel thio-benzoxazepinones exhibited higher anti-necroptosis activity in a human HT-29 cell necroptosis model. The effect on anti-necroptosis activity by the chirality was significantly reduced in the thio-benzoxazepinones, which was explained by the ligand conformation calculation. Among these analogues, compound 11 (S) and 12 (R) specifically inhibited necroptosis rather than apoptosis with EC50 values of 2.8 and 22.6 nM. They blocked necrosome formation by inhibiting the phosphorylation of RIPK1, RIPK3 and MLKL in necroptotic cells. Collectively, the highly potent thio-benzoxazepinones represent promising lead structures for further development of necroptosis-related diseases.
Jeremy S. Disch; Jennifer M. Duffy; Esther C. Y. Lee; Diana Gikunju; Betty Chan; Benjamin Levin; Michael I. Monteiro; Sarah A. Talcott; Anthony C. Lau; Fei Zhou; Anton Kozhushnyan; Neil E. Westlund; Patrick B. Mullins; Yan Yu; Moritz von Rechenberg; Junyi Zhang; Yelena A. Arnautova; Yanbin Liu; Ying Zhang; Andrew J. McRiner; Anthony D. Keefe; Anna Kohlmann; Matthew A. Clark; John W. Cuozzo; Christelle Huguet; Shilpi Arora J. Med. Chem., 2021, 64(8), 5049–5066 https://doi.org/10.1021/acs.jmedchem.1c00127AbstractBispecific degraders (PROTACs) of ERα are expected to be advantageous over current inhibitors of ERα signaling (aromatase inhibitors/SERMs/SERDs) used to treat ER+ breast cancer. Information from DNA-encoded chemical library (DECL) screening provides a method to identify novel PROTAC binding features as the linker positioning, and binding elements are determined directly from the screen. After screening ∼120 billion DNA-encoded molecules with ERα WT and 3 gain-of-function (GOF) mutants, with and without estradiol to identify features that enrich ERα competitively, the off-DNA synthesized small molecule exemplar 7 exhibited nanomolar ERα binding, antagonism, and degradation. Click chemistry synthesis on an alkyne E3 ligase engagers panel and an azide variant of 7 rapidly generated bispecific nanomolar degraders of ERα, with PROTACs 18 and 21 inhibiting ER+ MCF7 tumor growth in a mouse xenograft model of breast cancer. This study validates this approach toward identifying novel bispecific degrader leads from DECL screening with minimal optimization.
Mark Wigglesworth; Peter Hodder SLAS Discov., 2021, 26(2), 165-167 https://doi.org/10.1177/2472555220982257AbstractThis special issue was conceived with the intent of highlighting the use of existing and progressive technologies in hit discovery and how these technologies may be combined to generate vital hits for drug discovery. These aspects now cross both small-molecule and large-molecule discovery in many organizations. In addition, the issue is intended to remind readers of the heritage of SLAS journals, and how they have supported the use and application of innovative hit identification approaches.
Qi Liang; Jianyu He; Xue Zhao; Yan Xue; Haiyue Zuo; Ru Xu; Yan Jin; Jing Wang; Qian Li; Xinfeng Zhao J. Med. Chem., 2021, 64(7), 4196–4205 https://doi.org/10.1021/acs.jmedchem.1c00123AbstractNatural products have failed to meet the urgent need for drug discovery in recent decades due to limited resources, necessitating new strategies for re-establishing the key role of natural products in hit screening. This work introduced DNA-encoding techniques into the synthesis of phenolic acid-focused libraries containing 32 000 diverse compounds. Online selection of the library using immobilized angiotensin II type I receptor (AT1R) resulted in seven phenolic acid derivatives. The half-maximal concentration (IC50) of hit 1 for the right shift of the [125I]-Sar1-AngII competition curve was 19.6 nM. Pharmacological examination of renovascular hypertensive rats demonstrated that hit 1 significantly lowered the blood pressure of the animals without changing their heart rates. These results were used to create a general strategy for rapid and unbiased discovery of hits derived from natural products with high throughput and efficiency.
Ramón Rama-Garda; Jorge Amigo; Julián Priego; Manuel Molina-Martin; Leticia Cano; Eduardo Domínguez; María Isabel Loza; Alfonso Rivera-Sagredo; Jesúsde Blas Bioorg. Med. Chem., 2021, 116178 https://doi.org/10.1016/j.bmc.2021.116178AbstractThe output of an affinity selection screening results in a huge amount of valuable data that, after conducting the appropriate analysis, lead to the correct identification of the compounds enriched in the target of interest. The approach chosen to perform these analyses has become a key step in the development of a successful DNA Encoded Library platform. In this paper, we describe the combination of High Performance Liquid Chromatography purification during the library production with the Next Generation Sequencing analysis of the libraries to assess the yield of the chemical reactions prior to the affinity selection. This process allows us, apart from achieving higher quality libraries, to enable a normalization analysis of the affinity selection output, thus minimizing the bias induced by the chemical yield of each reaction as a misleading factor within the analysis and subsequent compound short-listing for off-DNA synthesis.
Jimmy Smadja; Agnès Quéméner; Mike Maillasson; Benoit Sicard; Aurélien Leray; Laurence Arzel; Jacques Lebreton; Erwan Mortier; Didier Dubreuil; Monique Mathé-Allainmat Bioorg. Med. Chem., 2021, 39, 116161 https://doi.org/10.1016/j.bmc.2021.116161AbstractInterleukin (IL)-15 is a pleiotropic cytokine structurally close to IL-2 and sharing with the IL-2Rβ and γc receptor (R) subunits. IL-15 plays important roles in innate and adaptative immunity, supporting the activation and proliferation of NK, NK-T, and CD8+ T cells. Over-expression of IL-15 has been shown to participate to the development of inflammatory and autoimmune diseases and diverse T cell malignancies. This study is in continuity of our previous work through which a family of small-molecule inhibitors impeding IL-15/IL-2Rβ interaction with sub-micromolar activity has been identified using pharmacophore-based virtual screening and hit optimization methods. With the aim to improve the efficacy and selectivity of our lead inhibitor, specific modifications have been introduced on the basis of optimized SAR and modelisation. The new series of compounds generated have been evaluated for their capacity to inhibit the proliferation as well as the down-stream signaling of IL-15-dependent cells and to bind to IL-15.
Brian J. Bender; Stefan Gahbauer; Andreas Luttens; Jiankun Lyu; Chase M. Webb; Reed M. Stein; Elissa A. Fink; Trent E. Balius; Jens Carlsson; John J. Irwin; Brian K. Shoichet Nat. Protoc., 2021, 16, 4799-4832 https://doi.org/10.1038/s41596-021-00597-zAbstractStructure-based docking screens of large compound libraries have become common in early drug and probe discovery. As computer efficiency has improved and compound libraries have grown, the ability to screen hundreds of millions, and even billions, of compounds has become feasible for modest-sized computer clusters. This allows the rapid and cost-effective exploration and categorization of vast chemical space into a subset enriched with potential hits for a given target. To accomplish this goal at speed, approximations are used that result in undersampling of possible configurations and inaccurate predictions of absolute binding energies. Accordingly, it is important to establish controls, as are common in other fields, to enhance the likelihood of success in spite of these challenges. Here we outline best practices and control docking calculations that help evaluate docking parameters for a given target prior to undertaking a large-scale prospective screen, with exemplification in one particular target, the melatonin receptor, where following this procedure led to direct docking hits with activities in the subnanomolar range. Additional controls are suggested to ensure specific activity for experimentally validated hit compounds. These guidelines should be useful regardless of the docking software used. Docking software described in the outlined protocol (DOCK3.7) is made freely available for academic research to explore new hits for a range of targets.
Martins S. Oderinde; Soomin Jin; T. G. Murali Dhar; Nicholas A. Meanwell; Arvind Mathur; James Kempson Tetrahedron, 2021, 132087 https://doi.org/10.1016/j.tet.2021.132087AbstractThe dearomatizing photocycloaddition reaction is a powerful and effective strategy for synthesizing complex, three-dimensional, polycyclic scaffolds from simple aromatic precursors. Generally, the dearomatizing photocycloaddition reaction is promoted by visible light and occurs via an energy transfer (EnT) process. This mini-review provides an overview of recent advances in this area (2018-2020), encompassing both intramolecular and intermolecular transformations. While the majority of the studies are centered on intramolecular processes due to their predictable regio- and stereo-selectivity, intermolecular transformations that show an exceptionally broad substrate scope are beginning to emerge.
Gisele Nishiguchi; Sourav Das; Jason Ochoada; Heather Long; Richard E. Lee; Zoran Rankovic; Anang A. Shelat Drug Discov. Today, 2021, 26(4), 1060-1069 https://doi.org/10.1016/j.drudis.2021.01.005AbstractThe quality of lead compounds is a key factor for determining the success of chemical probe and drug discovery programs. Given that high-throughput screening (HTS) continues to be a dominant lead generation paradigm, access to high-quality screening libraries is crucial for such efforts in both industry and academia. Here, we discuss the strategy implemented a decade ago to build from scratch one of the largest compound collections in academia, containing ∼575 000 carefully annotated small molecules, and a recent multidisciplinary effort designed to further enhance the collection to meet our research demands for the next decade.
Michael J. Bond; Craig M. Crews RSC Chem. Biol., 2021, 2, 725-742 https://doi.org/10.1039/D1CB00011JAbstractWith the discovery of PROteolysis TArgeting Chimeras (PROTACs) twenty years ago, targeted protein degradation (TPD) has changed the landscape of drug development. PROTACs have evolved from cell-impermeable peptide-small molecule chimeras to orally bioavailable clinical candidate drugs that degrade oncogenic proteins in humans. As we move into the third decade of TPD, the pace of discovery will only accelerate. Improved technologies are enabling the development of ligands for “undruggable” proteins and the recruitment of new E3 ligases. Moreover, enhanced computing power will expedite identification of active degraders. Here we discuss the strides made in these areas and what advances we can look forward to as the next decade in this exciting field begins.
Ramu Manjula; Kumari Anuja; Francisco J. Alcain Front. Pharmacol., 2021, 11, 585821 https://doi.org/10.3389/fphar.2020.585821AbstractSirtuins are NAD+ dependent histone deacetylases (HDAC) that play a pivotal role in neuroprotection and cellular senescence. SIRT1-7 are different homologs from sirtuins. They play a prominent role in many aspects of physiology and regulate crucial proteins. Modulation of sirtuins can thus be utilized as a therapeutic target for metabolic disorders. Neurological diseases have distinct clinical manifestations but are mainly age-associated and due to loss of protein homeostasis. Sirtuins mediate several life extension pathways and brain functions that may allow therapeutic intervention for age-related diseases. There is compelling evidence to support the fact that SIRT1 and SIRT2 are shuttled between the nucleus and cytoplasm and perform context-dependent functions in neurodegenerative diseases including Alzheimer’s disease (AD), Parkinson’s disease (PD), and Huntington’s disease (HD). In this review, we highlight the regulation of SIRT1 and SIRT2 in various neurological diseases. This study explores the various modulators that regulate the activity of SIRT1 and SIRT2, which may further assist in the treatment of neurodegenerative disease. Moreover, we analyze the structure and function of various small molecules that have potential significance in modulating sirtuins, as well as the technologies that advance the targeted therapy of neurodegenerative disease.
Timothy L. Foley; Woodrow Burchett; Qiuxia Chen; Mark E. Flanagan; Brendon Kapinos; Xianyang Li; Justin I. Montgomery; Anokha S. Ratnayake; Hongyao Zhu; Marie-Claire Peakman SLAS Discov., 2021, 26(2), 263-280 https://doi.org/10.1177/2472555220979589AbstractOver the past 20 years, the toolbox for discovering small-molecule therapeutic starting points has expanded considerably. Pharmaceutical researchers can now choose from technologies that, in addition to traditional high-throughput knowledge-based and diversity screening, now include the screening of fragment and fragment-like libraries, affinity selection mass spectrometry, and selection against DNA-encoded libraries (DELs). Each of these techniques has its own unique combination of advantages and limitations that makes them more, or less, suitable for different target classes or discovery objectives, such as desired mechanism of action. Layered on top of this are the constraints of the drug-hunters themselves, including budgets, timelines, and available platform capacity; each of these can play a part in dictating the hit identification strategy for a discovery program. In this article, we discuss some of the factors that we use to govern our building of a hit identification roadmap for a program and describe the increasing role that DELs are playing in our discovery strategy. Furthermore, we share our learning during our initial exploration of DEL and highlight the approaches we have evolved to maximize the value returned from DEL selections. Topics addressed include the optimization of library design and production, reagent validation, data analysis, and hit confirmation. We describe how our thinking in these areas has led us to build a DEL platform that has begun to deliver tractable matter to our global discovery portfolio.
Jonathan B Baell Future Med. Chem., 2021, 13(2), 91-93 https://doi.org/10.4155/fmc-2021-0005AbstractWelcome to this Early Career Special Focus Issue of Future Medicinal Chemistry. Assembled within is a veritable panoply of chemical biology discourses, showcasing the brilliance of our next generation of medicinal chemists and chemical biologists. In ’Abiotic scaffolds in medicinal chemistry: not a waste of chemical space', Ronald W Brown (Bayer, Frankfurt, Germany) discusses ’abiotic’ and ’natural product-like’ scaffolds, presenting the overarching questions: Do complex abiotic scaffolds still hold a place in medicinal chemistry, and, if so, do they present unique advantages? Nicely, answers to these questions are to an extent left up to the reader to formulate based on the information provided. What is clear is that the European Lead Factory has encouraged application of newer synthetic methods that can provide 3D topologies previously inaccessible by Nature to populate chemical screening space with compounds that might have drug development characteristics. One imagines that compound libraries derived therefrom would tend to be focused and bespoke. At the other extreme, new technologies are enabling construction of high-throughput screening libraries numbering in the billions. In ’The maturation of DNA encoded libraries: opportunities for new users', Conole et al. (Imperial College London, London, UK) pose the question of whether DNA-encoded libraries are feasible in academia and provide us a useful how-to guide for those outside pharmacology and interested in developing capability in this interesting technology. Or perhaps I should say "interesting technologies," as it becomes clear that this increasingly validated approach is blossoming with new developments. We are told that, conventionally, DNA-encoded libraries generally do not exceed more than three synthetic modifications, and so one could argue that a library that consists of a constant central scaffold lacks structural diversity. But then this is countered with the intelligent proposition that perhaps the ability to screen far larger numbers of compounds means that such constraints can be relaxed in DNA-encoded libraries, as screening may lead to higher affinity and a more elaborated hit. Nevertheless, the authors, recognizing the enormity of the theoretical chemical diversity space, sensibly view that library design would be best implemented by adopting the established guidelines for lead-like libraries.
Laura Guasch; Michael Reutlinger; Daniel Stoffler; Moreno Wichert Chimia, 2021, 75(1), 105-107 https://doi.org/10.2533/chimia.2021.105AbstractDNA-encoded library (DEL) technology has emerged as one of the fastest and most cost-effective screening platforms available in industry both for hit discovery as well as more recently for druggability and tractability assessments and successive prioritization of therapeutic targets in the early phase of drug discovery programs. The key principle of DELs is based on the combinatorial assembly (synthesis) of library members from chemical building blocks (BBs) and the corresponding tagging of each BB with unique DNA sequences (barcodes) in an alternating fashion of chemical reactions and DNA ligations. In analogy to phage display technology, this physical linkage of small organic molecules with distinctive DNA barcodes enables to deconvolute the chemical identity (structure) of each and every molecule by next-generation sequencing (NGS) at any time.
Bingjie Zhang; Wenqing Shui Chin. J. Nat., 2021, 43(1), 32-38 https://doi.org/10.3969/j.issn.0253-9608.2021.01.005AbstractAffinity mass spectrometry (affinity MS) based on the detection of protein target-ligand interaction is a powerful technology complementary to conventional high-throughput screening (HTS) assays and has been successfully incorporated into the early-phase lead discovery pipeline. Owing to the special biochemical properties of G protein-coupled receptors (GPCRs), tremendous technical challenges are posed to affinity MS-based ligand screening for GPCR targets. Here we briefly summarize the principle and features of the affinity MS technique adapted to GPCR ligand screening, and review the recent progress in screening both large-scale compound libraries and natural herb extracts with affinity MS for discovery of unreported GPCR ligands and modulators.
Filip Bošković; Alexander Ohmann; Ulrich F. Keyser; Kaikai Chen Small Struct., 2021, 2000144 https://doi.org/10.1002/sstr.202000144AbstractDigitally encoded deoxyribonucleic acid (DNA) nanostructures built via DNA self‐assembly have established applications in multiplexed biosensing and storing digital information. However, a key challenge is that DNA structures are not easily copied which is of vital importance for their large‐scale production and access to desired molecules by target‐specific amplification. Herein, DNA structural barcodes are built and the copying and random access of the barcodes from a library of molecules is demonstrated using a modified polymerase chain reaction (PCR). The structural barcodes are assembled by annealing a single‐stranded DNA scaffold with complementary short oligonucleotides containing protrusions as digital bits at defined locations. DNA nicks in these structures are ligated to facilitate barcode copying using PCR. To randomly access a target from a library of barcodes, a non‐complementary end in the DNA construct that serves as a barcode‐specific primer‐template is used. Readout of the DNA structural barcodes is performed with nanopore measurements. The study provides a roadmap for the convenient production of large quantities of self‐assembled DNA nanostructures. In addition, this strategy offers access to specific targets, a crucial capability for multiplexed single‐molecule sensing, and DNA data storage.
Yun Ding; Svetlana Belyanskaya; Jennifer L. DeLorey; Jeffrey A. Messer; G. Joseph Franklin; Paolo A. Centrella; Barry A. Morgan; Matthew A. Clark; Steven R. Skinner; Jason W. Dodson; Peng Li; Joseph P. Marino Jr.; David I. Israel Bioorg. Med. Chem., 2021, 116216 https://doi.org/10.1016/j.bmc.2021.116216AbstractInhibition of soluble epoxide hydrolase (sEH) has recently emerged as a new approach to treat cardiovascular disease and respiratory disease. Inhibitors based on 1,3,5-triazine chemotype were discovered through affinity selection against two triazine-based DNA-encoded libraries. The structure and activity relationship study led to the expansion of the original 1,4-cycloalkyl series to related aniline, piperidine, quinoline, aryl-ether and benzylic series. The 1,3-cycloalkyl chemotype led to the discovery of a clinical candidate (GSK2256294) for COPD.
Esther C. Y. Lee; Andrew J. McRiner; Katy E. Georgiadis; Julie Liu; Zooey Wang; Andrew D. Ferguson; Benjamin Levin; Moritz von Rechenberg; Christopher D. Hupp; Michael I. Monteiro; Anthony D. Keefe; Allison Olszewski; Charles J. Eyermann; Paolo Centrella; Yanbin Liu; Shilpi Arora; John W. Cuozzo; Ying Zhang; Matthew A. Clark; Christelle Huguet; Anna Kohlmann J. Med. Chem., 2021, 64(10), 6730–6744 https://doi.org/10.1021/acs.jmedchem.0c02271AbstractInhibition of hydroxy acid oxidase 1 (HAO1) is a strategy to mitigate the accumulation of toxic oxalate that results from reduced activity of alanine–glyoxylate aminotransferase (AGXT) in primary hyperoxaluria 1 (PH1) patients. DNA-Encoded Chemical Library (DECL) screening provided two novel chemical series of potent HAO1 inhibitors, represented by compounds 3–6. Compound 5 was further optimized via various structure–activity relationship (SAR) exploration methods to 29, a compound with improved potency and absorption, distribution, metabolism, and excretion (ADME)/pharmacokinetic (PK) properties. Since carboxylic acid-containing compounds are often poorly permeable and have potential active glucuronide metabolites, we undertook a brief, initial exploration of acid replacements with the aim of identifying non-acid-containing HAO1 inhibitors. Structure-based drug design initiated with Compound 5 led to the identification of a nonacid inhibitor of HAO1, 31, which has weaker potency and increased permeability.
Jun Hyung Park; Hee Myeong Wang; Min Hyeon Shin; Hyun‐Suk Lim Bull. Korean Chem. Soc., 2021, 42(4), 691-698 https://doi.org/10.1002/bkcs.12243AbstractDeveloping DNA‐encoded libraries of privileged scaffolds, such as pyrrolopyrimidines, is of great interest in drug discovery and chemical biology as a powerful tool to rapidly and inexpensively discover potent drug candidates. However, it is often challenging to construct such DNA‐encoded libraries because many reaction conditions are not compatible with DNA. Here, we describe the development of a convenient solid‐phase synthetic strategy that overcomes the current limitations and allows the efficient synthesis of a DNA‐encoded combinatorial library of structurally diverse tetra‐substituted pyrrolo[2,3‐d]pyrimidines.
Rosario González-Muñiz; María Ángeles Bonache; María Jesús Pérez de Vega Molecules, 2021, 26(2), 445 https://doi.org/10.3390/molecules26020445AbstractCyclic and macrocyclic peptides constitute advanced molecules for modulating protein–protein interactions (PPIs). Although still peptide derivatives, they are metabolically more stable than linear counterparts, and should have a lower degree of flexibility, with more defined secondary structure conformations that can be adapted to imitate protein interfaces. In this review, we analyze recent progress on the main methods to access cyclic/macrocyclic peptide derivatives, with emphasis in a few selected examples designed to interfere within PPIs. These types of peptides can be from natural origin, or prepared by biochemical or synthetic methodologies, and their design could be aided by computational approaches. Some advances to facilitate the permeability of these quite big molecules by conjugation with cell penetrating peptides, and the incorporation of β-amino acid and peptoid structures to improve metabolic stability, are also commented. It is predicted that this field of research could have an important future mission, running in parallel to the discovery of new, relevant PPIs involved in pathological processes.
Shivani Patel; Shorouk O. Badir; Gary A. Molander Trends Chem., 2021, 3(3), 161-175 https://doi.org/10.1016/j.trechm.2020.11.010AbstractRecently, DNA-encoded library (DEL) technology has emerged as an innovative screening modality for the rapid discovery of therapeutic candidates in pharmaceutical settings. This platform enables a cost-effective, time-efficient, and large-scale assembly and interrogation of billions of small organic ligands against a biological target in a single experiment. An outstanding challenge in DEL synthesis is the necessity for water-compatible transformations under ambient conditions. To access uncharted chemical space, the adoption of photoredox catalysis in DELs, including Ni-catalyzed manifolds and radical/polar crossover reactions, has enabled the construction of novel structural scaffolds through regulated odd-electron intermediates. Herein, a critical discussion of the validation of photoredox-mediated alkylation in DEL environments is presented. Current synthetic gaps are highlighted and opportunities for further development are speculated upon.
Bonwoo Koo; Haneul Yoo; Ho Jeong Choi; Min Kim; Cheoljae Kim; Ki Tae Kim Molecules, 2021, 26(3), 556 https://doi.org/10.3390/molecules26030556AbstractThe expanding scope of chemical reactions applied to nucleic acids has diversified the design of nucleic acid-based technologies that are essential to medicinal chemistry and chemical biology. Among chemical reactions, visible light photochemical reaction is considered a promising tool that can be used for the manipulations of nucleic acids owing to its advantages, such as mild reaction conditions and ease of the reaction process. Of late, inspired by the development of visible light-absorbing molecules and photocatalysts, visible light-driven photochemical reactions have been used to conduct various molecular manipulations, such as the cleavage or ligation of nucleic acids and other molecules as well as the synthesis of functional molecules. In this review, we describe the recent developments (from 2010) in visible light photochemical reactions involving nucleic acids and their applications in the design of nucleic acid-based technologies including DNA photocleaving, DNA photoligation, nucleic acid sensors, the release of functional molecules, and DNA-encoded libraries.
Animesh Roy; Thomas Kodadek Angew. Chem. Int. Ed. Engl., 2021, 60(21), 11983-11990 https://doi.org/10.1002/anie.202100230AbstractThere is considerable interest in the development of libraries of scaffold-diverse macrocycles as a source of ligands for difficult targets, such as protein-protein interaction surfaces. A classic problem in the synthesis of high-quality macrocyclic libraries is that some linear precursors will cyclize efficiently while some will not, depending on their conformational preferences. We report here a powerful quality control method that can be employed to readily distinguish between scaffolds that do and do not cyclize efficiently during solid-phase synthesis of thioether macrocycles without the need for tedious liquid chromatography/mass spectrometry analysis. We demonstrate that this assay can be employed to identify largely linear “impurities” in a DNA-encoded library of macrocycles. We also use the method to establish a useful quality control protocol for re-synthesis of putative macrocyclic screening hits.
Jörg Scheuermann; Yuichi Onda; Gabriele Bassi; Abdullah Elsayed; Franziska Ulrich; Sebastian Oehler; Louise Plais; Dario Neri Chem. Eur. J., 2021, 27(24), 7160-7167 https://doi.org/10.1002/chem.202005423AbstractWe describe the synthesis and characterization of a novel DNA-encoded library of macrocyclic peptide derivatives, based on three sets of proteinogenic and non-proteinogenic amino acid building blocks and featuring the use of copper alkyne-azide cycloaddition (CuAAC) reaction for ring closure. The library (termed YO-DEL) which contains 1,254,838 compounds, was encoded with DNA in single-stranded format and was screened against target proteins of interest using affinity capture procedures and photocrosslinking. YO-DEL selections yielded specific binders against serum albumins, carbonic anhydrases and NKp46, a marker of activated Natural Killer cells.
Sixiu Liu; Jingjing Qi; Weiwei Lu; Xuan Wang; Xiaojie Lu Org. Lett., 2021, 23, 3, 908-913 https://doi.org/10.1021/acs.orglett.0c04118AbstractTaking advantage of the diversity-oriented synthesis strategy with α,β-unsaturated carbonyl compounds, we have successfully established the DNA-compatible transformations for various heterocyclic scaffolds. The ring-closure reactions for pyrrole, pyrrolidine, pyrazole, pyrazoline, isoxazoline, pyridine, piperidine, cyclohexenone, and 5,8-dihydroimidazo[1,2-a]pyrimidine were elegantly demonstrated in a DNA-compatible format. These efforts paved the way for preparing DNA-encoded libraries with more extensive chemical space.
Xuan Wang; Jiaxiang Liu; Ziqin Yan; Xiaohong Liu; Sixiu Liu; Yanrui Suo; Weiwei Lu; Jinfeng Yue; Kaixian Chen; Hualiang Jiang; Yujun Zhao; Mingyue Zheng; Dongcheng Dai; Xiaojie Lu Chem. Sci., 2021,12, 2841-2847 https://doi.org/10.1039/D0SC06696FAbstractDNA-encoded library technology (DELT) employs DNA as a barcode to track the sequence of chemical reactions and enables the design and synthesis of libraries with billions of small molecules through combinatorial expansion. This powerful technology platform has been successfully demonstrated for hit identification and target validation for many types of diseases. As a highly integrated technology platform, DEL is capable of accelerating the translation of synthetic chemistry by using on-DNA compatible reactions or off-DNA scaffold synthesis. Herein, we report the development of a series of novel on-DNA transformations based on oxindole scaffolds for the design and synthesis of diversity-oriented DNA-encoded libraries for screening. Specifically, we have developed 1,3-dipolar cyclizations, cyclopropanations, ring-opening of reactions of aziridines and Claisen-Schmidt condensations to construct diverse oxindole derivatives. The majority of these transformations enable a diversity-oriented synthesis of DNA-encoded oxindole libraries which have been used in the successful hit identification for three protein targets. We have demonstrated that a diversified strategy for DEL synthesis could accelerate the application of synthetic chemistry for drug discovery.
Julien C. Vantourout; Andrew M. Mason; Josephine Yuen; Graham L. Simpson; Ghotas Evindar; Letian Kuai; Michael Hobbs; Emma Edgar; Saul Needle; Xiaopeng Bai; Steve Wilson; Paul Scott-Stevens; William Traylen; Kim Lambert; Neil Young; Shenaz Bunally; Scott G. Summerfield; Richard Snell; Rakesh Lad; Eric Shi; Steven Skinner; Lisa Shewchuk; Allan J.B. Watson; Chun-wa Chung; Sandeep Pal; Dennis A. Holt; Lara S. Kallander; Joanne Prendergast; Katrina Rivera; David G. Washburn; Mark R. Harpel; Christopher Arico-Muendel; Albert Isidro-Llobet Bioconjugate Chem., 2021, 32, 2, 279-289 https://doi.org/10.1021/acs.bioconjchem.0c00662AbstractReducing the required frequence of drug dosing can improve the adherence of patients to chronic treatments. Hence, drugs with longer in vivo half-lives are highly desirable. One of the most promising approaches to extend the in vivo half-life of drugs is conjugation to human serum albumin (HSA). In this work, we describe the use of AlbuBinder 1, a small-molecule noncovalent HSA binder, to extend the in vivo half-life and pharmacology of small-molecule BMP1/TLL inhibitors in humanized mice (HSA KI/KI). A series of conjugates of AlbuBinder 1 with BMP1/TLL inhibitors were prepared. In particular, conjugate c showed good solubility and a half-life extension of >20-fold versus the parent molecule in the HSA KI/KI mice, reaching half-lives of >48 h with maintained maximal inhibition of plasma BMP1/TLL. The same conjugate showed a half-life of only 3 h in the wild-type mice, suggesting that the half-life extension was principally due to specific interactions with HSA. It is envisioned that conjugation to AlbuBinder 1 should be applicable to a wide range of small molecule or peptide drugs with short half-lives. In this context, AlbuBinders represent a viable alternative to existing half-life extension technologies.
John Liddle; Andrew C. Pearce; Christopher Arico-Muendel; Svetlana Belyanskaya; Andrew Brewster; Murray Brown; Chun-wa Chung; Alexis Denis; Nerina Dodic; Anthony Dossang; Peter Eddershaw; Diana Klimaszewska; Imran Haq; Duncan S. Holmes; Alistair Jagger; Toral Jakhria; Emilie Jigorel; Ken Lind; Jeff Messer; Margaret Neu; Allison Olszewski; Riccardo Ronzoni; James Rowedder; Martin Rüdiger; Steve Skinner; Kathrine J. Smith; Lionel Trottet; Iain Uings; Zhengrong Zhu; James A. Irving; David A. Lomas Bioorg. Med. Chem. Lett., 2021, 127973 https://doi.org/10.1016/j.bmcl.2021.127973Abstractα1-antitrypsin deficiency is characterised by the misfolding and intracellular polymerisation of mutant α1-antitrypsin protein within the endoplasmic reticulum (ER) of hepatocytes. Small molecules that bind and stabilise Z α1-antitrypsin were identified via a DNA-encoded library screen. A subsequent structure based optimisation led to a series of highly potent, selective and cellular active α1-antitrypsin correctors.
Alexander L. Satz; Letian Kuai; Xuanjia Peng Bioorg. Med. Chem. Lett., 2021, 127851 https://doi.org/10.1016/j.bmcl.2021.127851AbstractThe use of DNA-encoded libraries (DELs) has increased greatly over the last decade, and today a majority of pharmaceutical companies employ the technology. The technology may be applied to most soluble and purified targets. However, standard DEL technology has limitations; some targets are challenging to purify, and it is not possible to directly screen for cellular or biochemical activity. Numerous creative methods have been reported to overcome these limitations and expand DEL target scope. Reported proof-of-concept experiments include DEL selections of cell surfaces, and inside of living cells. Additional alternatives include the construction and biochemical screening of one-bead-one-compound (OBOC) DELs using picoliter aqueous droplets or microfabricated wells as containers. In these cases, the small-molecule moiety of the library member is liberated from its DNA barcode, and able to interact freely with the desired target. Lastly, patent literature suggests the ability to conduct cellular functional screens using OBOC DELs.
Yunhong Nong; Yanyan Hou; Yuting Pu; Si Li; Yan Lan SLAS Discov., 2021, 26(5), 628-641 https://doi.org/10.1177/24725552211002463AbstractThroughout recent decades, histone deacetylase (HDAC) inhibitors have shown encouraging potential in cancer treatment, and several pan-HDAC inhibitors have been approved for treating malignant cancers. Numerous adverse effects of pan-HDAC inhibitors have been reported, however, during preclinical and clinical evaluations. To avoid undesirable responses, an increasing number of investigations are focusing on the development of isotype-selective HDAC inhibitors. In this study, we present an effective and quantitative cellular assay using high-content analysis (HCA) to determine compounds’ inhibition of the activity of HDAC6 and Class I HDAC isoforms, by detecting the acetylation of their corresponding substrates (i.e., α-tubulin and histone H3). Several conditions that are critical for HCA assays, such as cell seeding number, fixation and permeabilization reagent, and antibody dilution, have been fully validated in this study. We used selective HDAC6 inhibitors and inhibitors targeting different HDAC isoforms to optimize and validate the capability of the HCA assay. The results indicated that the HCA assay is a robust assay for quantifying compounds’ selectivity of HDAC6 and Class I HDAC isoforms in cells. Moreover, we screened a panel of compounds for HDAC6 selectivity using this HCA assay, which provided valuable information for the structure–activity relationship (SAR). In summary, our results suggest that the HCA assay is a powerful tool for screening selective HDAC6 inhibitors.
Herman van Vlijmen; Jean-Yves Ortholand; Volkhart M-J Li; Jon SB de Vlieger Drug Discov. Today, 2021, 26(10), 2406-2413 https://doi.org/10.1016/j.drudis.2021.04.019AbstractThrough the European Lead Factory model, industry-standard high-throughput screening and hit validation are made available to academia, small and medium-sized enterprises, charity organizations, patient foundations, and participating pharmaceutical companies. The compound collection used for screening is built from a unique diversity of sources. It brings together compounds from companies with different therapeutic area heritages and completely new compounds from library synthesis. This generates structural diversity and combines molecules with complementary physicochemical properties. In 2019, the screening library was updated to enable another 5 years of running innovative drug discovery projects. Here, we investigate the physicochemical and diversity properties of the updated compound collection. We show that it is highly diverse, drug like, and complementary to commercial screening libraries.
J. Willem M. Nissink; Sana Bazzaz; Carolyn Blackett; Matthew A. Clark; Olga Collingwood; Jeremy S. Disch; Diana Gikunju; Kristin Goldberg; John P. Guilinger; Elizabeth Hardaker; Edward J. Hennessy; Rachael Jetson; Anthony D. Keefe; William McCoull; Lindsay McMurray; Allison Olszewski; Ross Overman; Alexander Pflug; Marian Preston; Philip B. Rawlins; Emma Rivers; Marianne Schimpl; Paul Smith; Caroline Truman; Elizabeth Underwood; Juli Warwicker; Jon Winter-Holt; Simon Woodcock; Ying Zhang J. Med. Chem., 2021, 64(6), 3165-3184 https://doi.org/10.1021/acs.jmedchem.0c01904AbstractMer is a member of the TAM (Tyro3, Axl, Mer) kinase family that has been associated with cancer progression, metastasis, and drug resistance. Their essential function in immune homeostasis has prompted an interest in their role as modulators of antitumor immune response in the tumor microenvironment. Here we illustrate the outcomes of an extensive lead-generation campaign for identification of Mer inhibitors, focusing on the results from concurrent, orthogonal high-throughput screening approaches. Data mining, HT (high-throughput), and DECL (DNA-encoded chemical library) screens offered means to evaluate large numbers of compounds. We discuss campaign strategy and screening outcomes, and exemplify series resulting from prioritization of hits that were identified. Concurrent execution of HT and DECL screening successfully yielded a large number of potent, selective, and novel starting points, covering a range of selectivity profiles across the TAM family members and modes of kinase binding, and offered excellent start points for lead development.
Christopher A. Reiher; David P. Schuman; Nicholas Simmons; Scott E. Wolkenberg ACS Med. Chem. Lett., 2021, 12, 3, 343-350 https://doi.org/10.1021/acsmedchemlett.0c00615AbstractDNA-encoded library (DEL) screens have emerged as a powerful hit-finding tool for a number of biological targets. In this Innovations article, we review published hit-to-lead optimization studies following DEL screens. Trends in molecular property changes from hit to lead are identified, and specific optimization tactics are exemplified in case studies. Across the studies, physicochemical property and structural changes post-DEL screening are similar to those which occur during hit-to-lead optimization following high throughputscreens (HTS). However, unique aspects of DEL—the combinatorial synthetic methods which enable DEL synthesis and the linker effects at the DNA attachment point—impact the strategies and outcomes of hit-to-lead optimizations.
Ying Shi; Yanran Wu; Jianqiang Yu; Wannian Zhang; Chunlin Zhuang RSC Adv., 2021, 11, 2359-2376 https://doi.org/10.1039/D0RA09889BAbstractA DNA-encoded library is a collection of small molecules covalently linked to DNA that has unique information about the identity and the structure of each library member. A DNA-encoded chemical library (DEL) is broadly adopted by major pharmaceutical companies and used in numerous drug discovery programs. The application of the DEL technology is advantageous at the initial period of drug discovery because of reduced cost, time, and storage space for the identification of target compounds. The key points for the construction of DELs comprise the development and the selection of the encoding methods, transfer of routine chemical reaction from off-DNA to on-DNA, and exploration of new chemical reactions on DNA. The limitations in the chemical space and the diversity of DEL were reduced gradually by using novel DNA-compatible reactions based on the formation and the cleavage of various bonds. Here, we summarized a series of novel DNA-compatible chemistry reactions for DEL building blocks and analysed the druggability of screened hit molecules via DELs in the past five years.
Dominik K. Kölmel; Hongyao Zhu; Mark E. Flanagan; Sylvie K. Sakata; Anthony R. Harris; Jinqiao Wan; Barry A. Morgan Chem. Rec., 2021, 21(4), 616-630 https://doi.org/10.1002/tcr.202000148AbstractThis Personal Account describes the authors' foray into DNA-encoded libraries. The article addresses several key aspects of this hit generation technology, from the development of new synthetic methodology to the subsequent conception, design, and delivery of a DNA-encoded library. In particular, we have been engaged in adapting photocatalytic reactions to the idiosyncratic requirements of DNA-encoded chemistry. We have chosen one such methodology, namely a photocatalytic [2+2] cycloaddition reaction, to showcase how we employed property-based computational analyses to guide the selection and validation of building blocks for the production of a library. Ultimately, these novel bond disconnections and design principles led to the assembly of a DNA-encoded library that is composed of structurally diverse compounds within largely desirable property space and, therefore, well positioned to deliver novel chemical matter for drug discovery programs.
Verena B. K. Kunig; Marco Potowski; Mateja Klika Skopic; Andreas Brunschweiger ChemMedChem, 2021, 16(7), 1048-1062 https://doi.org/10.1002/cmdc.202000869AbstractUnderstanding the ligandability of a target protein, defined as the capability of a protein to bind drug‐like compounds on any sites, can give important stimuli for drug development projects. For instance, inhibition on protein‐protein interaction usually depends on identification of protein surface binders. DNA‐encoded chemical libraries (DELs) allow for scanning protein surfaces with large chemical space. Encoded library selection screens uncovered several protein‐protein interaction inhibitors and compounds binding to the surface of G protein‐coupled receptors (GPCRs) and kinases. The protein surface‐binding chemotypes from DELs are predominantly chemically modified and cyclized peptides, and functional small molecule peptidomimetics. Peptoid libraries and structural peptidomimetics have been less studied in the DEL field, hinting at hitherto less populated chemical space and suggesting alternative library designs. Roughly a third of the bioactive molecules evolved from smaller, target‐focused libraries. They showcase the potential of encoded libraries to identify more potent molecules from weak, e.g. fragment‐like, starting points.
Weidong Shang; Sheng-Nan Su; Rong Shi; Ze-Dong Mou; Guo-Qiang Yu,; Xia Zhang; Dawen Niu Angew. Chem. Int. Ed. Engl., 2021, 60(1), 385-390 https://doi.org/10.1002/anie.202009828AbstractWe here report glycosyl sulfoxides appended with an aryl iodide moiety as readily available, air and moisture stable precursors to glycosyl radicals. These glycosyl sulfoxides could be converted to glycosyl radicals by way of a rapid and efficient intramolecular radical substitution event. The use of this type of precursors enabled the synthesis of various complex C-linked glycoconjugates under mild conditions. This reaction could be performed in aqueous media and is amenable to the synthesis of glycopeptidomimetics and carbohydrate-DNA conjugates.
Jiong Zhang; Jiajia Dong Chin. J. Chem., 2021, 39(4), 1025-1027 https://doi.org/10.1002/cjoc.202000596AbstractHigh throughput screening towards chemical libraries is the primary way to discover lead compounds in developing innovative drugs, especially new molecular entities. Accordingly, the preparation of chemical libraries is the key step for drug development. Now, conventional small molecule chemical libraries and DNA‐encoded chemical libraries are the main models of chemical libraries for high‐throughput screening. Recently, a new model of chemical library, called modular click chemistry library, was proposed, which gives us an alternative choice to construct chemical libraries for high throughput screening and exhibits broad prospects to accelerate drug development. Herein, this article mainly focuses on the strategy to prepare the modular click chemistry library.
Gurubasavaraja Swamy Purawarga Matada; Arka Das; Prasad Sanjay Dhiwar; Abhishek Ghara Med. Chem. Res., 2021, 30, 535-551 https://doi.org/10.1007/s00044-020-02694-2AbstractCancer is the 2nd most fatal disease around the globe. Various receptors have been showed to be overexpressed and/or mutated in numerous cancers. Discoidin domain receptors 1 (DDR1) and 2 (DDR2) are one of the novel receptor tyrosine kinases (RTKs), which have been proved to regulate various cellular signaling pathways, cell proliferation, adhesion, migration, matrix remodeling, and dysregulation of these receptors may lead to metastatic cancer progressions. These receptors belong to unique category of RTKs, which require collagen binding for its activation. Yet the mechanism of this extracellular collagen binding and activation of cytosolic kinase domain of the receptors is not clear. Like other RTKs, these receptors also showed its extensive implications in numerous cancers like lung, breast, ovarian, pancreatic cancer and many others. Therefore DDR1 and DDR2 emerge as potential therapeutic targets in preventing cancer. Various small molecule tyrosine kinase inhibitors have been developed against these two receptors and proved to be highly efficacious in reducing tumor progressions. This review would highlight the detailed structure, functions, mechanism of action, signaling pathways of DDR1 and DDR2, their roles in cancer developments and the inhibition of these receptors with numerous inhibitors can be a promising strategy to combat this hefty menace.
Adam J. Wolpaw; Richard Bayliss; Gabriele Büchel; Chi V. Dang; Martin Eilers; W. Clay Gustafson; Gwenn H. Hansen; Natalia Jura; Stefan Knapp; Mark A. Lemmon; David Levens; John M. Maris; Ronen Marmorstein; Steven J. Metallo; Julie R. Park; Linda Z. Penn; Michael Rape; Martine F. Roussel; Kevan M. Shokat; William P. Tansey; Kliment A. Verba; Seychelle M. Vos; William A. Weiss; Elmar Wolf; Yaël P. Mossé Cancer Research, 2021, 81(7), 1627-1632 https://doi.org/10.1158/0008-5472.CAN-20-3108AbstractEffective treatment of pediatric solid tumors has been hampered by the predominance of currently “undruggable” driver transcription factors. Improving outcomes while decreasing the toxicity of treatment necessitates the development of novel agents that can directly inhibit or degrade these elusive targets. MYCN in pediatric neural-derived tumors, including neuroblastoma and medulloblastoma, is a paradigmatic example of this problem. Attempts to directly and specifically target MYCN have failed due to its similarity to MYC, the unstructured nature of MYC family proteins in their monomeric form, the lack of an understanding of MYCN-interacting proteins and ability to test their relevance in vivo, the inability to obtain structural information on MYCN protein complexes, and the challenges of using traditional small molecules to inhibit protein–protein or protein–DNA interactions. However, there is now promise for directly targeting MYCN based on scientific and technological advances on all of these fronts. Here, we discuss prior challenges and the reasons for renewed optimism in directly targeting this “undruggable” transcription factor, which we hope will lead to improved outcomes for patients with pediatric cancer and create a framework for targeting driver oncoproteins regulating gene transcription.
Johan J. N. Veerman; Yorik B. Bruseker; Eddy Damen; Erik H. Heijne; Wendy van Bruggen; Koen F. W. Hekking; Rob Winkel; Christopher D. Hupp; Anthony D. Keefe; Julie Liu; Heather A. Thomson; Ying Zhang; John W. Cuozzo; Andrew J. McRiner; Mark J. Mulvihill; Peter van Rijnsbergen; Birgit Zech; Louis M. Renzetti; Lee Babiss; Gerhard Müller ACS Med. Chem. Lett., 2021, 12(4), 555-562 https://doi.org/10.1021/acsmedchemlett.0c00547AbstractHerein we report the discovery of 2,4-1H-imidazole carboxamides as novel, biochemically potent, and kinome selective inhibitors of transforming growth factor β-activated kinase 1 (TAK1). The target was subjected to a DNA-encoded chemical library (DECL) screen. After hit analysis a cluster of compounds was identified, which was based on a central pyrrole-2,4-1H-dicarboxamide scaffold, showing remarkable kinome selectivity. A scaffold-hop to the corresponding imidazole resulted in increased biochemical potency. Next, X-ray crystallography revealed a distinct binding mode compared to other TAK1 inhibitors. A benzylamide was found in a perpendicular orientation with respect to the core hinge-binding imidazole. Additionally, an unusual amide flip was observed in the kinase hinge region. Using structure-based drug design (SBDD), key substitutions at the pyrrolidine amide and the glycine resulted in a significant increase in biochemical potency.
Julián Priego; Eduardo de Pedro Beato; Jesús Benavides; Adrián Gironda-Martínez; Fernando González; Jesús Blas; María Dolores Martín-Ortega; Ramón Rama-Garda; Jesús Ezquerra; Miguel A. Toledo; Alicia Torrado Bioconjugate Chem., 2021, 32, 1, 88-93 https://doi.org/10.1021/acs.bioconjchem.0c00566AbstractHerein we describe a method to orthogonally remove on-DNA N-Cbz, N-Alloc, N-Allyl, O-Bn, and O-Allyl protecting groups in the presence of other common protecting groups to afford free amines and carboxylic acids, respectively. The developed method uses NaBH4 as the source of hydrogen in the presence of Pd(OAc)2 under DNA aqueous conditions. In addition, under the developed conditions we were able to successfully hydrogenate triple and double bonds to totally saturated compounds. Furthermore, we introduce a new alternative procedure to reduce azides and aromatic nitro compounds to primary amines.
Wajid Ali; Gaurav Prakash; Debabrata Maiti Chem. Sci., 2021, 12, 2735-2759 https://doi.org/10.1039/D0SC05555GAbstractTransition metal-catalysed functionalizations of inert C–H bonds to construct C–C bonds represent an ideal route in the synthesis of valuable organic molecules. Fine tuning of directing groups, catalysts and ligands has played a crucial role in selective C–H bond (sp2 or sp3) activation. Recent developments in these areas have assured a high level of regioselectivity in C–H olefination reactions. In this review, we have summarized the recent progress in the oxidative olefination of sp2 and sp3 C–H bonds with special emphasis on distal, atroposelective, non-directed sp2 and directed sp3 C–H olefination. The scope, limitation, and mechanism of various transition metal-catalysed olefination reactions have been described briefly.
Zhifeng Yu; Angela F. Ku; Justin L. Anglin; Rajesh Sharma; Melek Nihan Ucisik; John C. Faver; Feng Li; Pranavanand Nyshadham; Nicholas Simmons; Kiran L. Sharma; Sureshbabu Nagarajan; Kevin Riehle; Gundeep Kaur; Banumathi Sankaran; Marta Storl-Desmond; Stephen S. Palmer; Damian W. Young; Choel Kim; Martin M. Matzuk Proc. Natl. Acad. Sci. USA, 2021, 118(9), e2021102118 https://doi.org/10.1073/pnas.2021102118AbstractBromodomain testis (BRDT), a member of the bromodomain and extraterminal (BET) subfamily that includes the cancer targets BRD2, BRD3, and BRD4, is a validated contraceptive target. All BET subfamily members have two tandem bromodomains (BD1 and BD2). Knockout mice lacking BRDT-BD1 or both bromodomains are infertile. Treatment of mice with JQ1, a BET BD1/BD2 nonselective inhibitor with the highest affinity for BRD4, disrupts spermatogenesis and reduces sperm number and motility. To assess the contribution of each BRDT bromodomain, we screened our collection of DNA-encoded chemical libraries for BRDT-BD1 and BRDT-BD2 binders. High-enrichment hits were identified and resynthesized off-DNA and examined for their ability to compete with JQ1 in BRDT and BRD4 bromodomain AlphaScreen assays. These studies identified CDD-1102 as a selective BRDT-BD2 inhibitor with low nanomolar potency and >1,000-fold selectivity over BRDT-BD1. Structure–activity relationship studies of CDD-1102 produced a series of additional BRDT-BD2/BRD4-BD2 selective inhibitors, including CDD-1302, a truncated analog of CDD-1102 with similar activity, and CDD-1349, an analog with sixfold selectivity for BRDT-BD2 versus BRD4-BD2. BROMOscan bromodomain profiling confirmed the great affinity and selectivity of CDD-1102 and CDD-1302 on all BET BD2 versus BD1 with the highest affinity for BRDT-BD2. Cocrystals of BRDT-BD2 with CDD-1102 and CDD-1302 were determined at 2.27 and 1.90 Å resolution, respectively, and revealed BRDT-BD2 specific contacts that explain the high affinity and selectivity of these compounds. These BD2-specific compounds and their binding to BRDT-BD2 are unique compared with recent reports and enable further evaluation of their nonhormonal contraceptive potential in vitro and in vivo.
Favour Danladi Makurvet Med. Drug Discov., 2021, 9, 100075 https://doi.org/10.1016/j.medidd.2020.100075AbstractSignificant advances in drug research and development are herein reviewed first to set the background for a critical consideration of the economic sustainability of biologics and small molecules, why biologic drugs are more expensive, and how drug cost often influences patient access to one drug class over the other. Also strongly emphasised is the need for the drug-making, especially the biopharmaceutical, industry to consider a reassignment of priorities so that more patients can enjoy the great benefits that come with blockbuster drugs, many of which are of biological origin but extremely expensive. A balance between the efficacy of wonder-performing drugs and the patient's financial ability to access them must be established to obliterate the crippling effect of the high costs of drugs on the poor majority of patients – those who cannot afford them. The overarching point broadly emphasised is that the actual success in drug discovery and development and in healthcare delivery should be measured not only by the magnitude of scientific breakthroughs but also by the level to which they are affordable to patients as determined by their costs. To enhance patients' access to drugs and new and/or improved healthcare technologies, more research attention must be paid to such cheaper alternatives as small molecules generics, biosimilars, and antibody-drug conjugates; government policies must be established to encourage the commercialisation of biosimilars; and pharmaceutical companies must be charitable enough to run assistance programmes for eligible financially handicap patients while seeking to make profits from the drug-making business.
Favour Danladi Makurvet Med. Drug Discov., 2021, 9, 100075 https://doi.org/10.1016/j.medidd.2020.100075AbstractSignificant advances in drug research and development are herein reviewed first to set the background for a critical consideration of the economic sustainability of biologics and small molecules, why biologic drugs are more expensive, and how drug cost often influences patient access to one drug class over the other. Also strongly emphasized is the need for the drug-making, especially the biopharmaceutical, industry to consider a reassignment of priorities so that more patients can enjoy the great benefits that come with blockbuster drugs, many of which are of biological origin but extremely expensive. A balance between the efficacy of wonder-performing drugs and the patient's financial ability to access them must be established to obliterate the crippling effect of the high costs of drugs on the poor majority of patients – those who cannot afford them. The overarching point broadly emphasized is that the actual success in drug discovery and development and in healthcare delivery should be measured not only by the magnitude of scientific breakthroughs but also by the level to which they are affordable to patients as determined by their costs. To enhance patients' access to drugs and new and/or improved healthcare technologies, more research attention must be paid to such cheaper alternatives as small molecules generics, biosimilars, and antibody-drug conjugates; government policies must be established to encourage the commercialization of biosimilars; and pharmaceutical companies must be charitable enough to run assistance programs for eligible financially handicap patients while seeking to make profits from the drug-making business.
Hui Gao; Shuang Lin; Shunning Zhang; Weijie Chen; Xiawen Liu; Guang Yang; Richard A. Lerner; Hongtao Xu; Zhi Zhou; Wei Yi Angew. Chem. Int. Ed., 2021, 60(4), 1959-1966 https://doi.org/10.1002/anie.202013052AbstractUsing gem ‐difluoromethylene alkynes as effectors, unprecedented diverse C−H activation/[4+2] annulations of simple benzoic acids are reported here. Remarkably, the chemodivergent reaction outcomes are well‐tuned by Rh/Ir‐catalyzed system; in the Rh(III) catalysis, 3‐alkenyl‐1 H ‐isochromen‐1‐one and 3,4‐dialkylideneisochroman‐1‐one skeletons are afforded in a solvent‐dependent manner whereas difluoromethylene‐substituted 1 H ‐isochromen‐1‐ones are generated under the Ir(III)‐catalyzed system. Mechanistic studies revealed that unusually double β‐F eliminations and fluorine effect‐induced regioselective reductive elimination are independently involved to enable distinct reaction modes for divergent product formations. Besides, synthetic application in both the derivatization of obtained diene products and the on‐DNA synthesis of DNA‐tagged difluorinated isocoumarin have been demonstrated, which manifested great potential for synthetic utility of the developed protocols.
Fanwang Meng; Zhongjie Liang; Kehao Zhao; Cheng Luo Med. Res. Rev., 2021, 41(3), 1701-1750 https://doi.org/10.1002/med.21774AbstractModern drug design aims to discover novel lead compounds with attractable chemical profiles to enable further exploration of the intersection of chemical space and biological space. Identification of small molecules with good ligand efficiency, high activity, and selectivity is crucial toward developing effective and safe drugs. However, the intersection is one of the most challenging tasks in the pharmaceutical industry, as chemical space is almost infinity and continuous, whereas the biological space is very limited and discrete. This bottleneck potentially limits the discovery of molecules with desirable properties for lead optimization. Herein, we present a new direction leveraging posttranslational modification (PTM) protein isoforms target space to inspire drug design termed as “Post‐translational Modification Inspired Drug Design (PTMI‐DD).” PTMI‐DD aims to extend the intersections of chemical space and biological space. We further rationalized and highlighted the importance of PTM protein isoforms and their roles in various diseases and biological functions. We then laid out a few directions to elaborate the PTMI‐DD in drug design including discovering covalent binding inhibitors mimicking PTMs, targeting PTM protein isoforms with distinctive binding sites from that of wild‐type counterpart, targeting protein‐protein interactions involving PTMs, and hijacking protein degeneration by ubiquitination for PTM protein isoforms. These directions will lead to a significant expansion of the biological space and/or increase the tractability of compounds, primarily due to precisely targeting PTM protein isoforms or complexes which are highly relevant to biological functions. Importantly, this new avenue will further enrich the personalized treatment opportunity through precision medicine targeting PTM isoforms.
Renaud Prudent; D. Allen Annis; Peter J. Dandliker; Jean-Yves Ortholand; Didier Roche Nat. Rev. Chem., 2021, 5, 62-71 https://doi.org/10.1038/s41570-020-00229-2AbstractAffinity selection-mass spectrometry (AS-MS) is a high-throughput screening (HTS) technique for drug discovery that enables rapid screening of large collections of compounds to identify ligands for a specific biomolecular target. AS-MS is a binding assay that is insensitive to the functional effects a ligand might have, which is important because it lets us identify novel ligands irrespective of their binding site. This approach is gaining popularity, notably due to its role in the emergence of useful agents for targeted protein degradation. This Perspective highlights the use of AS-MS techniques to explore broad chemical space and identify small-molecule ligands for biological targets that have proven challenging to address with other screening paradigms. We present chemical structures of reported AS-MS hits to illustrate the potential of this screening approach to deliver high-quality hits for further optimization. AS-MS has, thus, evolved from being an infrequent alternative to traditional HTS or DNA-encoded library strategies to now firmly establishing itself as a HTS approach for drug discovery.
Kaiyu Nakamura; Kosuke Yasui; Phil S. Baran J. Synth. Org. Chem. Jpn, 2021, 79(4), 333-343 https://doi.org/10.5059/yukigoseikyokaishi.79.333AbstractTeleocidine B family was isolated in1960 by Sakai group and was shown to have highly potent protein kinase-C (PKC) activation, similar to that of phorbol and related natural products. In this report, we have developed a unified total synthesis of teleocidins B-1-B-4 in 11 chemical steps. The highlights of the work are 1) the Ni catalyzed electrochemical amination, 2) Cu-mediated aziridine opening followed by direct macrolactamization, and 3) the tactical combination of C-H borylation and a redox-relay chain walking (RRCW) methodology to make the carbon quaternary center.
Yuteng Wu; Jack Williams; Ewen D. D. Calder; Louise J. Walport RSC Chem. Biol., 2021, 2, 151-165 https://doi.org/10.1039/D0CB00167HAbstractCombining different compound classes gives molecular hybrids that can offer access to novel chemical space and unique properties. Peptides provide ideal starting points for such molecular hybrids, which can be easily modified with a variety of molecular entities. The addition of small molecules can improve the potency, stability and cell permeability of therapeutically relevant peptides. Furthermore, they are often applied to create peptide-based tools in chemical biology. In this review, we discuss general methods that allow the discovery of this compound class and highlight key examples of peptide–small molecule hybrids categorised by the application and function of the small molecule entity.
Marco Catalano; Gabriele Bassi; Giulia Rotondi; Lyna Khettabi; Maria Dichiara; Patrizia Murer; Jörg Scheuermann; Montserrat Soler-Lopezc; Dario Neri RSC Med. Chem., 2021,12, 363-369 https://doi.org/10.1039/D0MD00310GAbstractHuman tyrosinase (hTYR) and tyrosinase-related protein 1 (hTYRP1) are closely-related enzymes involved in the synthesis of melanin, which are selectively expressed in melanocytes and, in a pathological context, in melanoma lesions. We used a previously described tyrosinase inhibitor (Thiamidol™) and DNA-encoded library technology for the discovery of novel hTYR and hTYRP1 ligands, that could be used as vehicles for melanoma targeting. Performing de novo selections with DNA-encoded libraries, we discovered novel ligands capable of binding to both hTYR and hTYRP1. More potent ligands were obtained by multimerizing Thiamidol™ moieties, leading to homotetrameric structures that avidly bound to melanoma cells, as revealed by flow cytometry. These findings suggest that melanoma lesions may, in the future, be targeted not only by monoclonal antibody reagents but also by small organic ligands.
Hsin-Mei Huang; Philipp Stephan; Hajo Kries Cell Chem. Biol., 2021, 28(2), 221-227 https://doi.org/10.1016/j.chembiol.2020.11.004AbstractDiffusive escape of intermediates limits the rate enhancement that nanocontainers or macromolecular scaffolds can provide for artificial biocatalytic cascades. Nonribosomal peptide synthetases (NRPSs) naturally form gigantic assembly lines and prevent escape by covalently tethering intermediates. Here, we have built DNA-templated NRPS (DT-NRPS) by adding zinc-finger tags to split NRPS modules. The zinc fingers direct the NRPS modules to 9-bp binding sites on a DNA strand, where they form a catalytically active enzyme cascade. Geometric constraints of the DT-NRPSs were investigated using the template DNA as a molecular ruler. Up to four DT-NRPS modules were assembled on DNA to synthesize peptides. DT-NRPSs outperform previously reported DNA-templated enzyme cascades in terms of DNA acceleration, which demonstrates that covalent intermediate channeling is possible along the DNA template. Attachment of assembly line enzymes to a DNA scaffold is a promising catalytic strategy for the sequence-controlled biosynthesis of nonribosomal peptides and other polymers.
Takashi Motoyaji YAKUGAKU ZASSHI, 2021, 141, 511-515 https://doi.org/10.1248/yakushi.20-00204-3AbstractThe first step in small-molecule drug discovery is the identification of hit compounds via high-throughput screening (HTS). In transporter drug discovery, most HTS assays are based on the uptake of labeled substrates, but such functional assays cannot be developed for many transporters, such as intracellular organelle transporters. These transporters remain unexplored in drug discovery despite their promise as drug targets. Affinity selection-mass spectrometry (AS-MS) is a label-free binding assay technology that has been developed as an HTS technology for analyzing interactions between targets and compounds. The use of AS-MS technology enables HTS against every type of drug target, in contrast to functional assays. AS-MS technology is usually used for soluble proteins, but we have developed this technology for application to membrane proteins as well. So far, we have used AS-MS for HTS of approximately 400000 compounds. In this review, the principles and application of AS-MS technology are introduced and an HTS campaign for solute carrier type 17A8 (SLC17A8) (vesicular glutamate transporter 3) is presented as an example.
Chengpeng Sun; Xinyue Zhang; Christophe Morisseau, Sung Hee Hwang; Zhanjun Zhang; Bruce D. Hammock; Xiaochi Ma J. Med. Chem., 2021, 64(1), 184-215 https://doi.org/10.1021/acs.jmedchem.0c01507AbstractSoluble epoxide hydrolase (sEH) is an α/β hydrolase fold protein and widely distributed in numerous organs including the liver, kidney, and brain. The inhibition of sEH can effectively maintain endogenous epoxyeicosatrienoic acids (EETs) levels and reduce dihydroxyeicosatrienoic acids (DHETs) levels, resulting in therapeutic potentials for cardiovascular, central nervous system, and metabolic diseases. Therefore, since the beginning of this century, the development of sEH inhibitors is a hot research topic. A variety of potent sEH inhibitors have been developed by chemical synthesis or isolated from natural sources. In this review, we mainly summarized the interconnected aspects of sEH with cardiovascular, central nervous system, and metabolic diseases and then focus on representative inhibitors, which would provide some useful guidance for the future development of potential sEH inhibitors.
Lars K. Petersen; Allan B. Christensen; Jacob Andersen; Charlotta G. Folkesson; Ole Kristensen; Charlotte Andersen; Amaya Alzu; Frank A. Sløk; Peter Blakskjær; Daniel Madsen; Carlos Azevedo; Iolanda Micco; Nils J. V. Hansen J. Am. Chem. Soc., 2021, 143, 7, 2751-2756 https://doi.org/10.1021/jacs.0c09213AbstractDNA-encoded small molecule libraries (DELs) have facilitated the discovery of novel modulators of many different therapeutic protein targets. We report the first successful screening of a multimillion membered DEL inside a living cell. We demonstrate a novel method using oocytes from the South African clawed frog Xenopus laevis. The large size of the oocytes of 1 μL, or 100 000 times bigger than a normal somatic cell, permits simple injection of DELs, thus resolving the fundamental problem of delivering DELs across cell membranes for in vivo screening. The target protein was expressed in the oocytes fused to a prey protein, to allow specific DNA labeling and hereby discriminate between DEL members binding to the target protein and the endogenous cell proteins. The 194 million member DEL was screened against three pharmaceutically relevant protein targets, p38α, ACSS2, and DOCK5. For all three targets multiple chemical clusters were identified. For p38α, validated hits with single digit nanomolar potencies were obtained. This work demonstrates a powerful new approach to DEL screening, which eliminates the need for highly purified active target protein and which performs the screening under physiological relevant conditions and thus is poised to increase the DEL amenable target space and reduce the attrition rates.
Connor W.Coley Trends Chem., 2021, 3(2), 133-145 https://doi.org/10.1016/j.trechm.2020.11.004AbstractDesigning functional molecules with desirable properties is often a challenging, multi-objective optimization. For decades, there have been computational approaches to facilitate this process through the simulation of physical processes, the prediction of molecular properties using structure–property relationships, and the selection or generation of molecular structures. This review provides an overview of some algorithmic approaches to defining and exploring chemical spaces that have the potential to operationalize the process of molecular discovery. We emphasize the potential roles of machine learning and the consideration of synthetic feasibility, which is a prerequisite to ‘closing the loop’. We conclude by summarizing important directions for the future development and evaluation of these methods.
Jesse Q. Zhang; Christian A. Siltanen; Ata Dolatmoradi; Chen Sun; Kai-Chun Chang; Russell H. Cole; Zev J. Gartner; Adam R. Abate Sci. Rep., 2021, 11, 4351 https://doi.org/10.1038/s41598-021-83865-yAbstractDroplet libraries consisting of many reagents encapsulated in separate droplets are necessary for applications of microfluidics, including combinatorial chemical synthesis, DNA-encoded libraries, and massively multiplexed PCR. However, existing approaches for generating them are laborious and impractical. Here, we describe an automated approach using a commercial array spotter. The approach can controllably emulsify hundreds of different reagents in a fraction of the time of manual operation of a microfluidic device, and without any user intervention. We demonstrate that the droplets produced by the spotter are similarly uniform to those produced by microfluidics and automate the generation of a ~ 2 mL emulsion containing 192 different reagents in ~ 4 h. The ease with which it can generate high diversity droplet libraries should make combinatorial applications more feasible in droplet microfluidics. Moreover, the instrument serves as an automated droplet generator, allowing execution of droplet reactions without microfluidic expertise.
Patrick R. Fitzgerald, Brian M. Paegel Chem. Rev., 2021, 121, 12, 7155–7177 https://doi.org/10.1021/acs.chemrev.0c00789AbstractClick chemistry, proposed nearly 20 years ago, promised access to novel chemical space by empowering combinatorial library synthesis with a “few good reactions”. These click reactions fulfilled key criteria (broad scope, quantitative yield, abundant starting material, mild reaction conditions, and high chemoselectivity), keeping the focus on molecules that would be easy to make, yet structurally diverse. This philosophy bears a striking resemblance to DNA-encoded library (DEL) technology, the now-dominant combinatorial chemistry paradigm. This review highlights the similarities between click and DEL reaction design and deployment in combinatorial library settings, providing a framework for the design of new DEL synthesis technologies to enable next-generation drug discovery.
David A. Lomas; James A. Irving; Christopher Arico-Muendel; Svetlana Belyanskaya; Andrew Brewster; Murray Brown; Chun-wa Chung; Hitesh Dave; Alexis Denis; Nerina Dodic; Anthony Dossang; Peter Eddershaw; Diana Klimaszewska; Imran Haq; Duncan S. Holmes; Jonathan P. Hutchinson; Alistair M. Jagger; Toral Jakhria; Emilie Jigorel; John Liddle; Ken Lind; Stefan J. Marciniak; Jeff Messer; Margaret Neu; Allison Olszewski; Adriana Ordonez; Riccardo Ronzoni; James Rowedder; Martin R€udiger; Steve Skinner; Kathrine J. Smith; Rebecca Terry; Lionel Trottet; Iain Uings; Steve Wilson; Zhengrong Zhu; Andrew C. Pearce EMBO Mol. Med., 2021, e13167 https://doi.org/10.15252/emmm.202013167AbstractSevere α1‐antitrypsin deficiency results from the Z allele (Glu342Lys) that causes the accumulation of homopolymers of mutant α1‐antitrypsin within the endoplasmic reticulum of hepatocytes in association with liver disease. We have used a DNA‐encoded chemical library to undertake a high‐throughput screen to identify small molecules that bind to, and stabilise Z α1‐antitrypsin. The lead compound blocks Z α1‐antitrypsin polymerisation in vitro, reduces intracellular polymerisation and increases the secretion of Z α1‐antitrypsin threefold in an iPSC model of disease. Crystallographic and biophysical analyses demonstrate that GSK716 and related molecules bind to a cryptic binding pocket, negate the local effects of the Z mutation and stabilise the bound state against progression along the polymerisation pathway. Oral dosing of transgenic mice at 100 mg/kg three times a day for 20 days increased the secretion of Z α1‐antitrypsin into the plasma by sevenfold. There was no observable clearance of hepatic inclusions with respect to controls over the same time period. This study provides proof of principle that “mutation ameliorating” small molecules can block the aberrant polymerisation that underlies Z α1‐antitrypsin deficiency.
Petr A. Sokolov; Ruslan R. Ramazanov; Valeriy I. Rolich; Maria A. Popova; Vyacheslav E. Shalygin; Nina A. Kasyanenko Nanotechnology, 2021, 32(4), 045604 https://doi.org/10.1088/1361-6528/abc037AbstractNanostructures synthesized using DNA-conjugated gold nanoparticles have a wide range of applications in the field of biosensorics. The stability of the DNA duplex plays a critical role as it determines the final geometry of these nanostructures. The main way to control DNA stability is to maintain a high ionic strength of the buffer solution; at the same time, high salt concentrations lead to an aggregation of nanoparticles. In this study, by means of the instrumentality of DNA-bridged seeds using tris(hydroxymethyl)aminomethane as a soft reducing agent the dumbbell-like gold nanoparticles up to 35 nm were synthesized with a high concentration of sodium ions of up to 100 mM and magnesium ions up to 1 mM. We also examined at the atomic level the details of the effect of the gold nanoparticle surface, as well as Na+ and Mg2+ ions, on the stability of nucleotide pairs located in close proximity to the grafting site.
Daniel G. Rivera; Manuel G. Ricardo; Aldrin V. Vasco; Ludger A. Wessjohann; Erik V. Van der Eycken Nat. Protoc., 2021, 16, 561-578 https://doi.org/10.1038/s41596-020-00445-6AbstractSolid-phase synthesis represents the methodological showcase for technological advances such as split-and-pool combinatorial chemistry and the automated synthesis of peptides, nucleic acids and polysaccharides. These strategies involve iterative coupling cycles that do not generate functional diversity besides that incorporated by the amino acids, nucleosides and monosaccharide building blocks. In sharp contrast, multicomponent reactions (MCRs) are traditionally used to generate both skeletal and appendage diversity in short, batchwise procedures. On-resin MCRs have traditionally been employed for the construction of heterocycle and peptidomimetic libraries, but that scenario has changed recently, and today the focus is more on the solid-phase derivatization of peptides and oligonucleotides. This review presents relevant experimental details and addresses the synthetic scope of such on-resin multicomponent protocols employed to accomplish specific biopolymer covalent modifications that are practically inviable by traditional solution-phase methodologies. Recommendations are provided to facilitate the implementation of solid-supported protocols and avoid possible pitfalls associated with the selection of the polymeric resin, the solvent and the order and amount of the reagents employed. We describe procedures comprising the multicomponent lipidation, biotinylation and labeling of both termini and the side chains, as well as the use of MCRs in the traceless on-resin synthesis of ligated and cyclic peptides. Solid-phase protocols for the assembly of α-helical and parallel β-sheet peptides as well as hybrid peptide–peptoid and peptide–peptide nucleic acid architectures are described. Finally, the solid-supported multicomponent derivatization of DNA oligonucleotides is illustrated as part of the DNA-encoded library technology relying on MCR-derived heterocyclic compounds.
Nicholas Favalli; Gabriele Bassi; Christian Pellegrino; Jacopo Millul; Roberto De Luca; Samuele Cazzamalli; Su Yang; Anika Trenner; Nour L. Mozaffari; Renier Myburgh; Mustafa Moroglu; Stuart J. Conway; Alessandro A. Sartori; Markus G. Manz; Richard A. Lerner; Peter K. Vogt; Jörg Scheuermann; Dario Neri Nat. Chem., 2021, 13, 540-548 https://doi.org/10.1038/s41557-021-00660-yAbstractThe encoding of chemical compounds with amplifiable DNA tags facilitates the discovery of small-molecule ligands for proteins. To investigate the impact of stereo- and regiochemistry on ligand discovery, we synthesized a DNA-encoded library of 670,752 derivatives based on 2-azido-3-iodophenylpropionic acids. The library was selected against multiple proteins and yielded specific ligands. The selection fingerprints obtained for a set of protein targets of pharmaceutical relevance clearly showed the preferential enrichment of ortho-, meta- or para-regioisomers, which was experimentally verified by affinity measurements in the absence of DNA. The discovered ligands included novel selective enzyme inhibitors and binders to tumour-associated antigens, which enabled conditional chimeric antigen receptor T-cell activation and tumour targeting.
Shinya Mimasu; Hiroaki Yamagishi; Satoshi Kubo; Mie Kiyohara; Toshihiro Matsuda; Toshiko Yahata; Heather A. Thomson; Christopher D. Hupp; Julie Liu; Takao Okuda; Kenichi Kakefuda Commun. Biol., 2021, 4(1), 159 https://doi.org/10.1038/s42003-021-01666-5AbstractThe structural mechanisms of single-pass transmembrane enzymes remain elusive. Kynurenine 3-monooxygenase (KMO) is a mitochondrial protein involved in the eukaryotic tryptophan catabolic pathway and is linked to various diseases. Here, we report the mammalian full-length structure of KMO in its membrane-embedded form, complexed with compound 3 (identified internally) and compound 4 (identified via DNA-encoded chemical library screening) at 3.0 Å resolution. Despite predictions suggesting that KMO has two transmembrane domains, we show that KMO is actually a single-pass transmembrane protein, with the other transmembrane domain lying laterally along the membrane, where it forms part of the ligand-binding pocket. Further exploration of compound 3 led to identification of the brain-penetrant compound, 5. We show that KMO is dimeric, and that mutations at the dimeric interface abolish its activity. These results will provide insight for the drug discovery of additional blood-brain-barrier molecules, and help illuminate the complex biology behind single-pass transmembrane enzymes.
Yiran Huang; Ling Meng; Qigui Nie; Yu Zhou; Langdong Chen; Shilian Yang; Yi Man Eva Fung; Xiaomeng Li; Cen Huang; Yan Cao; Yizhou Li; Xiaoyu Li Nat. Chem., 2021, 13, 77-88 https://doi.org/10.1038/s41557-020-00605-xAbstractMembrane proteins on the cell surface perform a myriad of biological functions; however, ligand discovery for membrane proteins is highly challenging, because a natural cellular environment is often necessary to maintain protein structure and function. DNA-encoded chemical libraries (DELs) have emerged as a powerful technology for ligand discovery, but they are mainly limited to purified proteins. Here we report a method that can specifically label membrane proteins with a DNA tag, and thereby enable target-specific DEL selections against endogenous membrane proteins on live cells without overexpression or any other genetic manipulation. We demonstrate the generality and performance of this method by screening a 30.42-million-compound DEL against the folate receptor, carbonic anhydrase 12 and the epidermal growth factor receptor on live cells, and identify and validate a series of novel ligands for these targets. Given the high therapeutic significance of membrane proteins and their intractability to traditional high-throughput screening approaches, this method has the potential to facilitate membrane-protein-based drug discovery by harnessing the power of DEL.
Antonie J. van der Zouwen; Martin D. Witte Front. Chem.,2021, 9, 644811 https://doi.org/10.3389/fchem.2021.644811AbstractCombinatorial and modular methods to synthesize small molecule modulators of protein activity have proven to be powerful tools in the development of new drug-like molecules. Over the past decade, these methodologies have been adapted toward utilization in the development of activity- and affinity-based chemical probes, as well as in chemoproteomic profiling. In this review, we will discuss how methods like multicomponent reactions, DNA-encoded libraries, phage displays, and others provide new ways to rapidly screen novel chemical probes against proteins of interest.
Tuan Thanh Nguyen; Kui Cai; Kees A. Schouhamer Immink; Han Mao Kiah arXiv, 2001.02839v1 https://arxiv.org/abs/2001.02839AbstractWe propose coding techniques that limit the length of homopolymers runs, ensure the GC-content constraint, and are capable of correcting a single edit error in strands of nucleotides in DNA-based data storage systems. In particular, for given ℓ,ϵ>0, we propose simple and efficient encoders/decoders that transform binary sequences into DNA base sequences (codewords), namely sequences of the symbols A, T, C and G, that satisfy the following properties: (i) Runlength constraint: the maximum homopolymer run in each codeword is at most ℓ, (ii) GC-content constraint: the GC-content of each codeword is within [0.5−ϵ,0.5+ϵ], (iii) Error-correction: each codeword is capable of correcting a single deletion, or single insertion, or single substitution error. For practical values of ℓ and ϵ, we show that our encoders achieve much higher rates than existing results in the literature and approach the capacity. Our methods have low encoding/decoding complexity and limited error propagation
Jie Li; Juan Kong; Guang Yang; Hongtao Xu; Peixiang Ma Bio-101, 2020, e1010829 https://doi.org/10.21769/BioProtoc.1010829AbstractThe DNA-encoded compound library (DEL) is a giant compound library containing millions or even hundreds of millions of specific DNA tags. Sequencing technology enables high-throughput decoding of screening information (Brenner and Lerner, 1992; Ma et al., 2019). We successfully constructed the first natural product-based DNA-encoded compound library (nDEL) using photocrosslinking technology and click chemistry. We screened against the target protein PARP1 (polyadenosine diphosphate ribose polymerase-1) and obtained a small molecule inhibitor - luteolin (Luteolin). Once the DNA in the cell is damaged by single-strand break, the DNA-binding domain of PARP1 protein can quickly bind to the damage site, and the catalytic domain at the C-terminal catalyzes the decomposition of NAD+ into ADP ribose and nicotinamide, and then uses ADP ribose as a substrate. Long-chain modification of polyadenosine diphosphate ribose (Poly [ADP-ribose], PAR) on residues such as glutamic acid and aspartic acid of itself and other proteins, recruiting other proteins to participate in important functions such as DNA damage repair . In order to further explore the ability of luteolin to inhibit the activity of PARP1 in cells, we used high-content imaging technology to detect the generation of poly-ADP-ribose PAR in cells after luteolin treatment (Yuan et al., 2017; Li et al., 2020).
Yasushi AMANO Nihon Kessho Gakkaishi, 2020, 62(4), 241-242 https://doi.org/10.5940/jcrsj.62.241AbstractTo satisfy the unmet medical needs, innovation of the technology for structural biology is important. Three-dimensional structures of drug-target molecules and their complexes reveal the mechanism of disease and accelerate discovery of novel drugs for patients.
Vittorio Katis; William Bradshaw; Ranjita Betarbet; Carmen Jimenez‐Antunez; Nicholas T. Seyfried; Belinda Slakman; Christopher Mulhern; Marie‐Aude Guie; Anthony Keefe; John Guilinger; Ying Zhang; Christelle Huguet; Allan I. Levey; Paul E. Brennan; Lara M. Mangravite; Opher Gileadi; The Open‐AD Consortium Alzheimer's Dement, 2020, 16, e045915 https://doi.org/10.1002/alz.045915AbstractAbstract Background Investigation of the Alzheimer’s Disease (AD) brain proteome, along with weighted co‐expression network analysis, revealed a module enriched with proteins involved in inflammation (PMID: 27989508). This module progressively increases in brains from Controls to Asymptomatic AD to those with Symptomatic AD. Moesin (MSN) and CD44 emerged as key inflammation module hub proteins. MSN is a cytoskeletal protein with roles in focal adhesion‐mediated cell motility; CD44 is a transmembrane receptor that interacts with MSN. This project aims to develop chemical tools to disrupt this pathway. Method To investigate the AD‐relevant distribution of MSN, tissue sections and cells were stained with MSN‐specific antibodies as well as with Iba1, 4G8 and pHF‐Tau antibodies. Purified FERM domains of MSN and EPB41L3 were produced in E. coli and co‐crystallized or soaked with ligands after crystallization. The CD44 ectodomain was expressed in bacteria and refolded or secreted from HEK293 cells. DNA‐encoded library screens were performed as described (http://www.x‐chemrx.com/our‐science/). Result MSN‐staining in cortical human brain sections localised to endothelial cells and microglia‐like cells. MSN‐positive cells also stained for microglia‐specific marker, Iba1. In AD brains, MSN‐positive microglia‐like cells were present around amyloid plaques and tangle‐like structures. These immunohistochemical studies confirmed the association of MSN with AD pathology. Co‐crystal structures of Moesin and the related FERM protein EPB41L3 revealed the binding mode of the intracellular tail of CD44 to Moesin. This binding interaction was then used to develop HTRF‐based assays for high‐throughput screening for molecules that disrupt the CD44‐MSN interaction. In parallel, a crystal‐based fragment screen using EPB41L3 identified binding pockets and initial hits. Hits and chemical staring points will be further developed with the Open‐AD Medicinal Chemistry core at UNC. To identify inhibitors of CD44 ectodomain binding to hyaluronic acid we screened DNA‐encoded libraries. The screens identified micromolar hits which are being further developed, aiming to obtain potent brain‐penetrant inhibitors of CD44. Conclusion (1) Moesin expression is associated with AD pathology. (2) The CD44‐Moesin signalling axis was targeted for potential points of intervention. (3) We have developed the assays, structural biology and chemical starting points to develop small‐molecule inhibitors of the CD44 ectodomain and the CD44‐MSN interaction.
Daniel Conole; James H. Hunter; Michael J. Waring Future Med. Chem., 2020,13(2), 173-191 https://www.future-science.com/doi/10.4155/fmc-2020-0285AbstractDNA-encoded combinatorial libraries (DECLs) represent an exciting new technology for high-throughput screening, significantly increasing its capacity and cost–effectiveness. Historically, DECLs have been the domain of specialized academic groups and industry; however, there has recently been a shift toward more drug discovery academic centers and institutes adopting this technology. Key to this development has been the simplification, characterization and standardization of various DECL subprotocols, such as library design, affinity screening and data analysis of hits. This review examines the feasibility of implementing DECL screening technology as a first-time user, particularly in academia, exploring the some important considerations for this, and outlines some applications of the technology that academia could contribute to the field.
Ajaybabu V. Pobbati; Brian P. Rubin Molecules, 2020, 25(24), 6001 https://doi.org/10.3390/molecules25246001AbstractThe identification of protein-protein interaction disruptors (PPIDs) that disrupt the YAP/TAZ-TEAD interaction has gained considerable momentum. Several studies have shown that YAP/TAZ are no longer oncogenic when their interaction with the TEAD family of transcription factors is disrupted. The transcriptional co-regulator YAP (its homolog TAZ) interact with the surface pockets of TEADs. Peptidomimetic modalities like cystine-dense peptides and YAP cyclic and linear peptides exploit surface pockets (interface 2 and interface 3) on TEADs and function as PPIDs. The TEAD surface might pose a challenge for generating an effective small molecule PPID. Interestingly, TEADs also have a central pocket that is distinct from the surface pockets, and which small molecules leverage exclusively to disrupt the YAP/TAZ-TEAD interaction (allosteric PPIDs). Although small molecules that occupy the central pocket belong to diverse classes, they display certain common features. They are flexible, which allows them to adopt a palmitate-like conformation, and they have a predominant hydrophobic portion that contacts several hydrophobic residues and a small hydrophilic portion that faces the central pocket opening. Despite such progress, more selective PPIDs that also display favorable pharmacokinetic properties and show tolerable toxicity profiles are required to evaluate the feasibility of using these PPIDs for cancer therapy.
Naděžda Cankařová; Viktor Krchňák Int. J. Mol. Sci., 2020, 21(23), 9160 https://doi.org/10.3390/ijms21239160AbstractDrug discovery efforts largely depend on access to structural diversity. Multicomponent reactions allow for time-efficient chemical transformations and provide advanced intermediates with three or four points of diversification for further expansion to a structural variety of organic molecules. This review is aimed at solid-phase syntheses of small molecules involving isocyanide-based multicomponent reactions. The majority of all reported syntheses employ the Ugi four-component reaction. The review also covers the Passerini and Groebke-Blackburn-Bienaymé reactions. To date, the main advantages of the solid-phase approach are the ability to prepare chemical libraries intended for biological screening and elimination of the isocyanide odor. However, the potential of multicomponent reactions has not been fully exploited. The unexplored avenues of these reactions, including chiral frameworks, DNA-encoded libraries, eco-friendly synthesis, and chiral auxiliary reactions, are briefly outlined.
Jorg Scheuermann; Dario Neri Biochem. Biophys. Res. Commun., 2020, 533, 2, iii-iv https://doi.org/10.1016/j.bbrc.2020.10.055AbstractDNA-Encoded Chemical Libraries (DELs) are increasingly being used for the identification of small-molecule binders to target proteins of interest. In anticipation of the growing relevance of DEL technology we had organized in 2006 at ETH Zürich the first International Symposium on DNA-Encoded Chemical Libraries. At that time the community consisted in just a handful of groups working on DELs. However, interest in the technology increased quite dramatically, especially during the last five years. Most pharmaceutical companies have now set-up their own DEL platforms and routinely perform screening campaigns. In addition, many academic groups have realized the potential of DEL technology for hit discovery, as large libraries can be constructed and interrogated at reasonable costs. The first DEL-derived drugs are expected to begin pivotal clinical trials in the next few years, suggesting that DEL technology has now matured. Already now DELs represent the largest collections of small molecules and facilitate the isolation of specific protein ligands.
Huanan Wen; Rui Ge; Yi Qu; Jialin Sun; Xiaodong Shi; Weiren Cui; Hao Yan; Qi Zhang; Yulong An; Wenji Su; Hongfang Yang; Letian Kuai; Alexander L. Satz; Xuanjia Peng Org. Lett., 2020, 22(24), 9484-9489 https://doi.org/10.1021/acs.orglett.0c03461AbstractWe report a DNA-compatible photoredox decarboxylative coupling of α-amino acids with carbonyl compounds to access DNA-encoded sp3-rich 1,2-amino alcohols. The reaction proceeds efficiently for a wide range of DNA-conjugated aldehydes and ketones and provides the desired 1,2-amino alcohols with conversions generally >50%. Additional utility of the developed protocol is demonstrated by one-pot cyclization of DNA-conjugated 1,2-amino alcohols into oxazolidiones and morpholinones. Lastly, qPCR and sequencing data analysis indicates no significant DNA damage upon photoredox decarboxylative coupling.
Huang-Chi Du; Martin M. Matzuk; Ying-Chu Chen Org. Biomol. Chem., 2020, 18(45), 9221-9226 https://doi.org/10.1039/D0OB02021DAbstractA zinc bromide-catalyzed synthesis of 5-substituted tetrazoles via DNA-conjugated nitriles using sodium azide has been developed. The protocol offered moderate to excellent yields of tetrazoles with a broad range of substrates, including a variety of functionalized aromatic, heterocyclic, and aliphatic nitriles. In addition, the electronic effect within the substrate scope was evaluated. DNA fidelity was assessed by ligation efficiency and amplifiability analysis. The ability to generate tetrazoles expands the diversity of heterocycles in the preparation of DNA-encoded chemical libraries.
Anna Tomberg; Jonas Boström Drug Discov. Today, 2020, 25(12), 2174-2181 https://doi.org/10.1016/j.drudis.2020.09.027AbstractStatements, such as ‘you need to break free from amide-formations to improve molecular properties’ and ‘novel chemistry leads to novel biology’ are frequently encountered in the medicinal chemistry community. To verify whether truth lies in such preconceptions, we investigated whether complex, diverse, and novel molecules can be made by ‘easy’ chemistry. By analyzing the AstraZeneca screening collection, we conclude that novelty, diversity, and molecular complexity is currently not compromised by the use of the most popular reaction, amide bond formation, mainly because of a recent steady increase in unique amines available. Easy chemistry allows speedy access to a broad chemical space, facilitating progress in projects, and opens the possibility of synthesis automation and new technologies, such as DNA-encoded libraries.
Lorena Baranda Pellejero; Malihe Mahdifar; Gianfranco Ercolani; Jonathan Watson; Tom Brown; Francesco Ricci Nat. Commun., 2020, 11, 6242 https://doi.org/10.1038/s41467-020-20024-3AbstractDNA-templated synthesis takes advantage of the programmability of DNA-DNA interactions to accelerate chemical reactions under diluted conditions upon sequence-specific hybridization. While this strategy has proven advantageous for a variety of applications, including sensing and drug discovery, it has been so far limited to the use of nucleic acids as templating elements. Here, we report the rational design of DNA templated synthesis controlled by specific IgG antibodies. Our approach is based on the co-localization of reactants induced by the bivalent binding of a specific IgG antibody to two antigen-conjugated DNA templating strands that triggers a chemical reaction that would be otherwise too slow under diluted conditions. This strategy is versatile, orthogonal and adaptable to different IgG antibodies and can be employed to achieve the targeted synthesis of clinically-relevant molecules in the presence of specific IgG biomarker antibodies.
Andrea Angeli; Fabrizio Carta; Alessio Nocentini; Jean-Yves Winum; Raivis Zalubovskis; Atilla Akdemir; Valentina Onnis; Wagdy M. Eldehna; Clemente Capasso; Giuseppina De Simone; Simona Maria Monti; Simone Carradori; William A. Donald; Shoukat Dedhar; Claudiu T. Supuran Metabolites, 2020, 10(10), 412 https://doi.org/10.3390/metabo10100412AbstractThe tumor microenvironment is crucial for the growth of cancer cells, triggering particular biochemical and physiological changes, which frequently influence the outcome of anticancer therapies. The biochemical rationale behind many of these phenomena resides in the activation of transcription factors such as hypoxia-inducible factor 1 and 2 (HIF-1/2). In turn, the HIF pathway activates a number of genes including those involved in glucose metabolism, angiogenesis, and pH regulation. Several carbonic anhydrase (CA, EC 4.2.1.1) isoforms, such as CA IX and XII, actively participate in these processes and were validated as antitumor/antimetastatic drug targets. Here, we review the field of CA inhibitors (CAIs), which selectively inhibit the cancer-associated CA isoforms. Particular focus was on the identification of lead compounds and various inhibitor classes, and the measurement of CA inhibitory on-/off-target effects. In addition, the preclinical data that resulted in the identification of SLC-0111, a sulfonamide in Phase Ib/II clinical trials for the treatment of hypoxic, advanced solid tumors, are detailed.
Yihang Jing; Jose Montano; Michaella Levy; Jeff Lopez; Pei-Pei Kung; Paul Richardson; Krzysztof Krajewski; Laurence Florens; Michael Washburn; Jordan L. Meier bioRxiv, 2020, 09, 24 https://doi.org/10.1101/2020.09.24.311530AbstractChemical proteomics provides a powerful strategy for the high-throughput assignment of enzyme function or inhibitor selectivity. However, identifying optimized probes for an enzyme family member of interest and differentiating signal from background remain persistent challenges in the field. To address this obstacle, here we report a physiochemical discernment strategy for optimizing chemical proteomics based on the Coenzyme A (CoA) cofactor. First, we synthesize a pair of CoA-based Sepharose pulldown resins differentiated by a single negatively charged residue, and find this change alters their capture properties in gel-based profiling experiments. Next, we integrate these probes with quantitative proteomics and benchmark analysis of ‘probe selectivity’ versus traditional ‘competitive chemical proteomics’. This reveals the former is well-suited for the identification of optimized pulldown probes for specific enzyme family members, while the latter may have advantages in discovery applications. Finally, we apply our anionic CoA pulldown probe to evaluate the selectivity of a recently reported small molecule N-terminal acetyltransferase inhibitor. These studies further validate the use of physical discriminant strategies in chemoproteomic hit identification and demonstrate how CoA-based chemoproteomic probes can be used to evaluate the selectivity of small molecule protein acetyltransferase inhibitors, an emerging class of pre-clinical therapeutic agents.
Stanislav S. Terekhov; Igor E. Eliseev; Leyla A. Ovchinnikova; Marsel R. Kabilov; Andrey D. Prjibelski; Alexey E. Tupikin; Ivan V. Smirnov; Alexey A. Belogurov; Konstantin V. Severinov; Yakov A. Lomakin; Sidney Altman; Alexander G. Gabibov Proc. Natl. Acad. Sci. USA, 2020, 117(44), 27300-27306 https://doi.org/10.1073/pnas.2017138117AbstractDNA libraries are predisposed to template mispairing during conventional “bulk” PCR, leading to the loss of unique sequences. The latter is facilitated by the nonuniform distribution of templates frequently observed in DNA libraries. These effects result in a prominent reduction of the original diversity. The encapsulation of DNA repertoires in liquid droplets abolishes the effects of mispairing in DNA libraries. The fundamental advantages of emulsion PCR (ePCR) over bulk PCR are illustrated by deep sequencing and mathematical modeling, which provide the general strategy for ePCR rationalization. The quasi single-molecule ePCR reveals total genetic information by counteracting the degeneration of DNA libraries’ diversity.
Rongfeng Wu; Sen Gao; Tian Du; Kunliang Cai; Xuemin Cheng; Jing Fan; Jing Feng; Alex Shaginian; Jin Li; Jinqiao Wan; Guansai Liu Chem. Asian J., 2020, 15(23), 4033-4037 https://doi.org/10.1002/asia.202001105AbstractA DNA‐encoded chemical library (DECL) is built with combinatorial chemistry, which works by bringing chemical fragments together to generate diverse structures. However, chemical diversity of DNA‐encoded chemical libraries is often limited by DNA compatible synthetic reactions. This report shows a conceptual strategy to expand chemical space of DNA‐encoded chemical libraries by incorporation of diversity‐oriented synthesis in DECL synthesis. We developed Aldol reactions on DNA in a combinatorial way. After obtaining DNA‐tagged α , β ‐unsaturated ketones which represent important chemical intermediates, many distinct structures with skeletal diversities are achieved by diversity‐oriented synthesis.
Rajdip Chowdhury; Zhunzhun Yu; My Linh Tong; Stefanie V. Kohlhepp; Xiang Yin; Abraham Mendoza J. Am. Chem. Soc., 2020, 142(47), 20143-20151 https://doi.org/10.1021/jacs.0c09678AbstractPhotoexcited dihydronicotinamides like NADH and analogues have been found to generate alkyl radicals upon reductive decarboxylation of redox-active esters without auxiliary photocatalysts. This principle allowed aliphatic photocoupling between redox-active carboxylate derivatives and electron-poor olefins, displaying surprising water and air-tolerance and unusually high coupling rates in dilute conditions. The orthogonality of the reaction in the presence of other carboxylic acids and its utility in the functionalization of DNA is presented, notably using visible light in combination with NADH, the ubiquitous reductant of life.
Josephine Yuen; Jing Chai; Yun Ding Bioconjugate Chem., 2020, 31(12), 2712-2718 https://doi.org/10.1021/acs.bioconjchem.0c00508AbstractCondensation of imines with anhydrides have been proven to be a valuable method for the synthesis of tetrahydroisoquinolones. Herein, we report the application of this chemistry with DNA-conjugated imines. Condensation of DNA-conjugated imine (which can be formed in situ from DNA-conjugated amines and aldehydes or DNA-conjugated aldehyde and primary amines) with homophthalic anhydride produces isoquinolones in moderate to excellent yields. The formed isoquinolone can be further derivatized with a variety of amines through amide bond formation. Development of this chemistry on-DNA enables the synthesis of an isoquinolone core-focused DNA-encoded library.
Philip A. Harris Expert Opin. Ther. Patents, 2020, 1-15 https://doi.org/10.1080/13543776.2021.1854729AbstractIntroduction: RIP1 kinase is a serine/threonine-protein kinase that has recently emerged as a central regulator of TNF-α dependent programmed necrosis (necroptosis), an inflammatory form of cell death, with important roles in inflammation and neurodegeneration. Small molecule RIP1 kinase inhibitors may provide new opportunities for treating a variety of autoimmune, inflammatory, and neurodegenerative diseases, among others, and thus have attracted widespread drug development efforts and a corresponding large amount of patent activity in recent years. Areas covered: This review focuses on the patent literature covering small molecule inhibitors of RIP1 kinase from 2016–present. Expert opinion: Inhibition of programmed necrosis (necroptosis) by RIP1 kinase inhibitors is a new field that has attracted widespread recent interest as a possible therapeutic means to treat a number of diseases in the inflammatory, neurodegenerative, and oncology areas. The interest in the therapeutic potential of RIP1kinase is evidenced by more than 40 small molecule patent applications published since 2016. To date, only a few RIP1 kinase inhibitors have entered the clinic. An understanding of the optimal clinical setting, in terms of dosing and disease indications for RIP1 inhibition, will require further clinical readouts as the current inhibitors progress and additional molecules enter into full development.
Rahul Singh; Deepak B. Salunke E. J. Med. Chem., 2020, 113071 https://doi.org/10.1016/j.ejmech.2020.113071AbstractIndoleamine-2,3-dioxygenase 1 (IDO1) catalyses the first and rate limiting step of kynurenine pathway accounting for the major contributor of L-Tryptophan degradation. The Kynurenine metabolites are identified as essential cofactors, antagonists, neurotoxins, immunomodulators, antioxidants as well as carcinogens. The catalytic active site of IDO1 enzyme consists of hydrophobic Pocket-A positioned in the distal heme site and remains connected to a second hydrophobic Pocket-B towards the entrance of the active site. IDO1 enzyme also relates directly to the modulation of the innate and adaptive immune system. Various studies proved that the over expression of IDO1 enzyme play a predominant role in the escape of immunity during cancer progression. Recently, there has been considerable interest in evaluating the potential of IDO1 inhibitors to mobilize the body’s immune system against solid tumours. In the last two decades, enormous attempts to advance new IDO1 inhibitors are on-going both in pharmaceutical industries and in academia which resulted in the discovery of a diverse range of selective and potent IDO1 inhibitors. The IDO1 inhibitors have therapeutic utility in various diseases and in the near future, it may have utility in the treatment of COVID-19. Despite various reviews on IDO1 inhibitors in last five years, none of the reviews provide a complete overview of diverse chemical space including naturally occurring and synthetic IDO1 inhibitors with detailed structure activity relationship studies. The present work provides a complete overview on the IDO1 inhibitors known in the literature so far along with the Structure-Activity Relationship (SAR) in each class of compounds.
Arvind Negi; Paul V. Murphy Eur. J. Med. Chem., 2020, 113038 https://doi.org/10.1016/j.ejmech.2020.113038AbstractThe myeloid leukemia cell differentiation protein (Mcl-1) is an anti-apoptotic protein of the B-cell lymphoma 2 (Bcl-2) family, which regulates cellular apoptosis. Mcl-1 expression plays a key role in survival of cancer cells and therefore serves as a promising target in cancer therapy. Besides, its importance as a cancer target, various peptides and small-molecule inhibitors have been successfully designed and synthesized, yet no Mcl-1 inhibitor is approved for clinical use. However, recent development on the understanding of Mcl-1’s role in key cellular processes in cancer and an upsurge of reports highlighting its association in various anticancer drug resistance supports the view that Mcl-1 is a key target in various cancers, especially hematological cancers. This review compiles structures of a variety of inhibitors of Mcl-1 reported to date. These include inhibitors based on a diverse range of heterocycles (e.g. indole, imidazole, thiophene, nicotinic acid, piperazine, triazine, thiazole, isoindoline), oligomers (terphenyl, quaterpyridine), polyphenol, phenalene, anthranilic acid, anthraquinone, macrocycles, natural products, and metal-based complexes. In addition, an effort has been made to summarize the structure activity relationships of some important classes of Mcl-1 inhibitors, giving affinities and selectivities for Mcl-1 compared to other Bcl-2 family members. A focus has been placed on categorizing the inhibitors based on their core frameworks (scaffolds) to appeal to the chemical biologist or medicinal chemist.
Thomas C. Fessard; Kristina Goncharenko; Quentin Lefebvre; Christophe Salomé Chmia, 2020, 74(10), 803-807 https://doi.org/10.2533/chimia.2020.803AbstractIn highly competitive research environments, the ability to access more complex structural spaces efficiently is a predictor of a company's ability to generate novel IP-protected small molecule candidates with adequate properties, hence filling their development pipelines. SpiroChem is consistently developing new synthetic methodologies and strategies to access complex molecular structure, thereby facilitating and accelerating small molecule drug discovery. Pushing the limits of what are perceived as complex molecular structures allows SpiroChem and its clients to unleash creativity and explore meaningful chemical spaces, which are under-exploited sources of novel active molecules. In this article, we explain how we differentiated ourselves in a globalized R&D environment and we provide several snapshots of how efficient methodologies can generate complex structures, rapidly.
Wen Liu; Yi Zou; Kaiming Li; Shushan Ge; Haiqing Zhong; Yuanyuna Wang; Yue Hu; Yisheng Lai; Xianchi Dong; Wenjie Guo; Qiang Xu Authorea, 2020 https://doi.org/10.22541/au.159863181.10139431AbstractBACKGROUND AND PURPOSE Pharmacological inhibition of indoleamine-2,3-dioxygenase 1 activity is now considered to be a potential therapeutic tool for cancer therapy. However, the anti-cancer efficacy may be the biggest obstacle for the clinical application of current IDO1 inhibitors. EXPERIMENTAL APPROACHES HeLa cell-based IDO1/Kyn assay as well as recombinant IDO1 activity assay were used to determine the IDO1 enzyme activity. Interaction was examined by UV-visible spectra, isothermal titration calorimetry assay, cellular thermal shift assay and co-crystallization. Mouse colon cancer CT26 syngeneic model and azoxymethane/dextran sulfate sodium induced colon carcinogenesis model were employed to confirm the anti-tumor effect in vivo. KEY RESULTS B37 effectively and specifically inhibited IDO1 by targeting its heme-free conformation (apo-IDO1). By competing with heme for binding to apo-IDO1, B37 potently inhibited IDO1 activity with IC50 for 22 pM in the HeLa cell based assay. X-ray co-crystal structures of the inhibitor-enzyme complexes showed that unlike the hIDO1-BMS-986205 complex, the B37-hIDO1 complex displayed stronger hydrophobic interactions, which enhanced its binding affinity measured with ITC. Accordingly, stronger non-covalent interactions including π stacking and hydrogen bonds formed between B37 and apo-hIDO1 underlay the enthalpy-driven force for B37 to bind the enzyme. This binding model endowed B37 potent anti-tumor efficacy in mouse colon cancer CT26 syngeneic model and azoxymethane/dextran sulfate sodium induced colon carcinogenesis model by activating the host’s immune system. Moreover, the combination of B37 with a VEGFR2 inhibitor apatinib synergistically inhibited tumor growth. CONCLUSIONS AND IMPLICATIONS These results revealed that B37 may serve as a candidate for apo-IDO1 inhibition mediated immunotherapy.
Omer Sabary; Yoav Orlev; Roy Shafir; Leon Anavy; Eitan Yaakobi; Zohar Yakhini Bioinformatics, 2020, btaa740 https://doi.org/10.1093/bioinformatics/btaa740AbstractMotivation Recent years have seen a growing number and an expanding scope of studies using synthetic oligo libraries for a range of applications in synthetic biology. As experiments are growing by numbers and complexity, analysis tools can facilitate quality control andsupport better assessment and inference. Results We present a novel analysis tool, called SOLQC, which enables fast and comprehensive analysis of synthetic oligo libraries, based on NGS analysis performed by the user. SOLQC provides statistical information such as the distribution of variant representation, different error rates and their dependence on sequence or library properties. SOLQC produces graphical reports from the analysis, in a flexible format. We demonstrate SOLQC by analyzing literature libraries. We also discuss the potential benefits and relevance of the different components of the analysis. Software Availability SOLQC is a free software for non-commercial use, available at https://app.gitbook.com/@yoav-orlev/s/solqc/. For commercial use please contact the authors.
Vincent Blay; Bhairavi Tolani; Sunita P. Ho; Michelle R. Arkin Drug Discov. Today, 2020, 25(10), 1807-1821 https://doi.org/10.1016/j.drudis.2020.07.024AbstractHigh-throughput screening (HTS) provides starting chemical matter in the adventure of developing a new drug. In this review, we survey several HTS methods used today for hit identification, organized in two main flavors: biochemical and cell-based assays. Biochemical assays discussed include fluorescence polarization and anisotropy, FRET, TR-FRET, and fluorescence lifetime analysis. Binding-based methods are also surveyed, including NMR, SPR, mass spectrometry, and DSF. On the other hand, cell-based assays discussed include viability, reporter gene, second messenger, and high-throughput microscopy assays. We devote some emphasis to high-content screening, which is becoming very popular. An advisable stage after hit discovery using phenotypic screens is target deconvolution, and we provide an overview of current chemical proteomics, in silico, and chemical genetics tools. Emphasis is made on recent CRISPR/dCas-based screens. Lastly, we illustrate some of the considerations that inform the choice of HTS methods and point to some areas with potential interest for future research.
Peijun Li; Jack A. Terrett; Jason R. Zbieg ACS Med. Chem. Lett. 2020, 11, 11, 2120–2130 https://doi.org/10.1021/acsmedchemlett.0c00436AbstractVisible light-mediated photocatalysis, which relies on the ability of photocatalysts to absorb low-energy visible light and engage in single-electron transfer (SET) or energy transfer (ET) processes with organic substrates, has emerged as one of the fastest growing fields in organic synthesis. This catalytic platform enables a highly selective approach to promote radical-based organic transformations which unlocks unique reaction pathways. Due to the extremely mild conditions of these transformations and compatibility in aqueous environments, photocatalysis has emerged as an enabling technology in drug discovery. Photocatalysis is uniquely positioned for application in pharmaceutical development because of its demonstrated potential for broad functional group tolerance, biocompatibility, site-specific selectivity, and operational simplicity. This review will highlight the recent advances of visible-light photocatalysis through its application in peptide functionalization, protein bioconjugation, Csp3–Csp2 cross-coupling, late-stage functionalization, isotopic labeling, DNA-encoded library technology (DELT), and microenvironment mapping (μMap).
Andrea Canal-Martín; Ruth Pérez-Fernández ACS Omega, 2020, 5(41), 26307-26315 https://doi.org/10.1021/acsomega.0c03800AbstractProtein-directed dynamic combinatorial chemistry (P-D DCC) is considered a powerful strategy to identify ligands to pharmacologically relevant protein targets. The protein selects its affinity ligands in situ through a thermodynamic templated effect in which the library composition shifts to the formation of specific library members at the expense of other (nonbinding) species. The increase in concentration of the selected species is known as amplification and leads to the discovery of new hit compounds for protein targets. This Mini-Review contains an updated overview of the protein-directed DCC applications and the fundamental aspects to take into account when designing a P-D DCC experiment such as the most biocompatible reversible reactions and the methodology used to analyze the experiments.
Riley J. Giesler; Patrick W. Erickson; Michael S. Kay Curr. Opin. Chem. Biol., 2020, 58, 37-44 https://doi.org/10.1016/j.cbpa.2020.04.003AbstractNative chemical ligation has enabled the chemical synthesis of proteins for a wide variety of applications (e.g., mirror-image proteins). However, inefficiencies of this chemoselective ligation in the context of large or otherwise challenging protein targets can limit the practical scope of chemical protein synthesis. In this review, we focus on recent developments aimed at enhancing and expanding native chemical ligation for challenging protein syntheses. Chemical auxiliaries, use of selenium chemistry, and templating all enable ligations at otherwise suboptimal junctions. The continuing development of these tools is making the chemical synthesis of large proteins increasingly accessible.
Heidi R. Culver; Jasmine Sinha; Tania R. Prieto; Christopher J. Calo; Benjamin D. Fairbanks; Christopher N. Bowman Biomacromolecules, 2020, 21(10), 4205-4211 https://doi.org/10.1021/acs.biomac.0c00996AbstractClick nucleic acids (CNAs) are a new, low-cost class of xeno nucleic acid (XNA) oligonucleotides synthesized by an efficient and scalable thiol-ene polymerization. In this work, a thorough characterization of oligo(thymine) CNA-oligo(adenine) DNA ((dA)20) hybridization was performed to guide future implementation of CNAs in applications that rely on sequence specific interactions. Microscale thermophoresis (MST) provided a convenient platform to rapidly and systematically test the effects of several factors (i.e., sequence, length, and salt concentration) on the CNA-DNA dissociation constant (Kapp). Because CNAs have limited water solubility, all studies were performed in aqueous-DMSO mixtures. CNA-DNA hybrids between oligo(thymine) CNA (average length of 16 bases) and (dA)20 DNA have good stability despite the high organic content, a favorable attribute for many emerging applications of XNAs. Specifically, the Kapp of CNA-DNA hybrids in 65 vol% DMSO with 10 mM sodium chloride (NaCl) was 0.74 0.1 μM, while the Kapp for (dT)20-(dA)20 DNA-DNA was measured to be 45 2 μM in a buffer without DMSO but at the same NaCl concentration. CNA hybridized with DNA following Watson-Crick base pairing with excellent sequence specificity, discriminating even a single base pair mismatch, with Kapp of 0.74 0.1 and 3.7 0.6 μM for complementary and single-base-pair mismatch sequences, respectively. As with dsDNA, increasing CNA length led to more stable hybrids as a result of increased base pairing, where Kapp decreased from 5.6 0.8 to 0.27 0.1 μM as CNA average length increased from 7 to 21 bases. However, unlike DNA-DNA duplexes, which are largely unstable at low salt concentrations, CNA-DNA stability does not depend on salt concentration, with Kapp remaining consistent between 1.0 – 1.9 μM over a NaCl concentration range of 1.25 to 30 mM.
David Lomas; James A. Irving; Christopher Arico-Muendel; Svetlana Belyanskaya; Andy Brewster; Murray J. B. Brown; Chun-wa Chung; Hitesh Dave; Alexis Denis; Nerina Dodic; Anthony Dossang; Peter Eddershaw; Diana Klimaszewska; Imran Haq; Duncan Holmes; Jon Hutchinson; Alistair M. Jagger; Toral Jakhria; Emilie Jigorel; John Liddle; Ken Lind; Stefan John Marciniak; Jeff Messer; Margaret Neu; Allison Olszewski; Adriana Ordoñez; Riccardo Ronzoni; James Rowedder; Martin Rüdiger; Steven Skinner; Kate Smith; Rebecca Terry; Lionel Trottet; Iain Uings; Steve Wilson; Zhengrong Zhu; Andrew C. Pearce bioRxiv, 2020 https://doi.org/10.1101/2020.07.26.217661AbstractSevere α1-antitrypsin deficiency results from the Z allele (Glu342Lys) that causes the accumulation of homopolymers of mutant α1-antitrypsin within the endoplasmic reticulum of hepatocytes in association with liver disease. We have used a DNA-encoded chemical library to undertake a high throughput screen to identify small molecules that bind to, and stabilise Z α1-antitrypsin. The lead compound blocks Z α1-antitrypsin polymerisation in vitro, reduces intracellular polymerisation and increases the secretion of Z α1-antitrypsin three-fold in mammalian cells including an iPSC model of disease. Crystallographic and biophysical analyses demonstrate that GSK716 and related molecules bind to a cryptic binding pocket, negate the local effects of the Z mutation and stabilise the bound state against progression along the polymerization pathway. Oral dosing of transgenic mice at 100 mg/kg three times a day for 20 days increased the secretion of Z α1-antitrypsin into the plasma by 7-fold. There was no observable clearance of hepatic inclusions with respect to controls. This study provides proof-of-principle that 'mutation ameliorating' small molecules are a viable approach to treat protein conformational diseases.
Hacer Karatas; Mohammad Akbarzadeh; Hélène Adihou; Gernot Hahne; Ajaybabu V. Pobbati; Elizabeth Yihui Ng; Stéphanie M. Guéret; Sonja Sievers; Axel Pahl; Malte Metz; Sarah Zinken; Lara Dötsch; Christine Nowak; Sasikala Thavam; Alexandra Friese; Congbao Kang; Wanjin Hong; Herbert Waldmann J. Med. Chem., 2020, 63(20), 11972-11989 https://doi.org/10.1021/acs.jmedchem.0c01275AbstractTranscriptional enhanced associate domain (TEAD) transcription factors together with coactivators and corepressors modulate the expression of genes that regulate fundamental processes, as organogenesis and cell growth, and elevated TEAD activity is associated with tumorigenesis. Hence novel modulators of TEAD and methods for their identification are in high demand. We describe the development of a new “thiol conjugation assay” for identification of novel small molecules that bind to the TEAD central pocket. The assay monitors prevention of covalent binding of a fluorescence turn-on probe to a cysteine in the central pocket by small molecules. Screening of a collection of compounds revealed kojic acid analogs as TEAD inhibitors, which covalently targets the cysteine in the central pocket, blocks the interaction with coactivator Yes-Associated Protein (YAP) with nanomolar apparent IC50 values and reduces TEAD target gene expression. This methodology promises to enable new medicinal chemistry programs aimed at the modulation of TEAD activity.
Wenyu Yang; Wei Chen; Huilin Su; Rong Li; Chen Song; Zhouyu Wang; Lingling Yang RSC Adv., 2020, 10, 37382-37390 https://doi.org/10.1039/D0RA06316AAbstractSirtuin 2 (SIRT2) is an important and special member of the atypical histone deacetylase Sirtuin (SIRT) family. Due to its extensive catalytic effects, SIRT2 can regulate autophagy, myelination, immunity, inflammation and other physiological processes. Recent evidence revealed that dysregulation of human SIRT2 activity is associated with the pathogenesis and prognosis of cancers, Parkinson's disease and other disorders; thus SIRT2 is a promising target for potential therapeutic intervention. This review presents a systematic summary of nine chemotypes of small-molecule SIRT2 inhibitors, particularly including the discovery and structural optimization strategies, which will be useful for future efforts to develop new inhibitors targeting SIRT2 and associated target proteins.
Zhoulong Fan; Shuai Zhao; Tao Liu; Pengxiang Shen; Zi-Ning Cui; Zhe Zhuang; Qian Shao; Jason S. Chen; Anokha Ratnayake; Mark Flanagan; Dominik K. Kölmel; David W. Piotrowski; Paul Richardson; Jinquan Yu Chem. Sci., 2020, 11, 12282-12288 https://doi.org/10.1039/D0SC03935GAbstractDNA-encoded library (DEL) technology has the potential to dramatically expedite hit identification in drug discovery owing to its ability to perform protein affinity selection with millions or billions of molecules in a few experiments. To expand the molecular diversity of DEL, it is critical to develop different types of DNA-encoded transformations that produces billions of molecules with distinct molecular scaffolds. Sequential functionalization of multiple C–H bonds provides a unique avenue for creating diversity and complexity from simple starting materials. However, the use of water as solvent, the presence of DNA, and the extremely low concentration of DNA-encoded coupling partners (0.001 M) have hampered the development DNA-encoded C(sp3)–H activation reactions. Herein, we report the realization of palladium-catalyzed C(sp3)–H arylation of aliphatic carboxylic acids, amides and ketones with DNA-encoded aryl iodides in water. Notably, the present method enables the use of alternative sets of monofunctional building blocks, providing a linchpin to facilitate further setup for DELs. Furthermore, the C–H arylation chemistry enabled the on-DNA synthesis of structurally-diverse scaffolds containing enriched C(sp3) character, chiral centers, cyclopropane, cyclobutane, and heterocycles.
Marco Catalano; Mustafa Moroglu; Petra Balbi; Federica Mazzieri; James Clayton; Katrina H. Andrews; Martina Bigatti; Jörg Scheuermann; Stuart J. Conway; Dario Neri ChemMedChem, 2020, 15(18), 1752-1756 https://doi.org/10.1002/cmdc.202000528AbstractDNA‐encoded chemical libraries (DECLs) are collections of chemical moieties individually coupled to distinctive DNA barcodes. Compounds can be displayed either at the end of a single DNA strand (i.e., single‐pharmacophore libraries) or at the extremities of two complementary DNA strands (i.e., dual‐pharmacophore libraries). In this work, we describe the use of a dual‐pharmacophore Encoded Self‐Assembly Chemical (ESAC) library for the affinity maturation of a known 4,5‑dihydrobenzodiazepinone ring (THBD) acetyl‑lysine (KAc) mimic for the cyclic‐AMP response element binding protein (CREB) binding protein (CREBBP or CBP) bromodomain. The new pair of fragments discovered from library selections showed a sub‐micromolar affinity for the CREBBP bromodomain in fluorescence polarization and ELISA assays, and selectivity against BRD4(1).
Olivier B. C. Monty; Nicholas Simmons; Srinivas Chamakuri; Martin M. Matzuk; Damian W. Young ACS Comb. Sci., 2020, 22(12), 833-843 https://doi.org/10.1021/acscombsci.0c00144AbstractPeptide drug discovery has shown a resurgence since 2000, bringing 28 non-insulin therapeutics to the market compared to 56 since its first peptide drug, insulin, in 1923. While the main method of discovery has been biological display—phage, mRNA, and ribosome—the synthetic limitations of biological systems has restricted the depth of exploration of peptide chemical space. In contrast, DNA-encoded chemistry offers the synergy of large numbers and ribosome-independent synthetic flexibility for the fast and deeper exploration of the same space. Hence, as a bridge to building DNA-encoded chemical libraries (DECLs) of peptides, we have developed substrate-tolerant amide coupling reaction conditions for amino acid monomers, performed a coupling screen to illustrate such tolerance, developed protecting group strategies for relevant amino acids and reported the limitations thereof, developed a strategy for the coupling of α,α-disubstituted alkenyl amino acids relevant to all-hydrocarbon stapled peptide drug discovery, developed reaction conditions for the coupling of tripeptides likely to be used in DECL builds, and synthesized a fully deprotected DNA-decamer conjugate to illustrate the potency of the developed methodology for on-DNA peptide synthesis.
Mike Lemke; Hannah Ravenscroft; Nicole J. Rueb; Dmitri Kireev; Dana Ferraris; Raphael M. Franzini Bioorg. Med. Chem. Lett., 2020, 127464 https://doi.org/10.1016/j.bmcl.2020.127464AbstractTwo critical steps in drug development are 1) the discovery of molecules that have the desired effects on a target, and 2) the optimization of such molecules into lead compounds with the required potency and pharmacokinetic properties for translation. DNA-encoded chemical libraries (DECLs) can nowadays yield hits with unprecedented ease, and lead-optimization is becoming the limiting step. Here we integrate DECL screening with structure-based computational methods to streamline the development of lead compounds. The presented workflow consists of enumerating a virtual combinatorial library (VCL) derived from a DECL screening hit and using computational binding prediction to identify molecules with enhanced properties relative to the original DECL hit. As proof-of-concept demonstration, we applied this approach to identify an inhibitor of PARP10 that is more potent and druglike than the original DECL screening hit.
Alfredo Martín; Christos A. Nicolaou; Miguel A. Toledo Commun. Chem., 2020, 3, 127 https://doi.org/10.1038/s42004-020-00374-1AbstractDNA-encoded library (DEL) technology is a novel ligand identification strategy that allows the synthesis and screening of unprecedented chemical diversity more efficiently than conventional methods. However, no reports have been published to systematically study how to increase the diversity and improve the molecular property space that can be covered with DEL. This report describes the development and application of eDESIGNER, an algorithm that comprehensively generates all possible library designs, enumerates and profiles samples from each library and evaluates them to select the libraries to be synthesized. This tool utilizes suitable on-DNA chemistries and available building blocks to design and identify libraries with a pre-defined molecular weight distribution and maximal diversity compared with compound collections from other sources.
Ke Li; Yi Qu; Yulong An; Eric Breinlinger; Huanan Wen; Matthew P. Webster; Duanchen Ding; Meng Zhao; Xiaodong Shi; Jiangong Wang; Wenji su; Weiren Cui; Alexander L. Satz; Hongfang Yang; Letian Kuai; Andrew Little; Xuanjia Peng Bioconjugate Chem., 2020, 31, 2092−2097 https://doi.org/10.1021/acs.bioconjchem.0c00392AbstractWe report a DNA-compatible protocol for synthesizing amides from DNA-bound aldehydes and non-nucleophilic aryla-mines including aza-substituted anilines, 2-aminobenzimidazoles, and 3-aminopyrazoles. The reactions were carried out at room temperature and provided reasonable conversions and wide functional group compatibility. The reactions were also successful when employing aryl and aliphatic aldehydes. In addition, qPCR and NGS data suggested no negative impact on DNA integrity after the copper-mediated oxidative amidation reaction.
Melissa J. Buskes; Maria-Jesus Blanco Molecules, 2020, 25(15), 3493 https://doi.org/10.3390/molecules25153493AbstractCross-coupling reactions have played a critical role enabling the rapid expansion of structure–activity relationships (SAR) during the drug discovery phase to identify a clinical candidate and facilitate subsequent drug development processes. The reliability and flexibility of this methodology have attracted great interest in the pharmaceutical industry, becoming one of the most used approaches from Lead Generation to Lead Optimization. In this mini-review, we present an overview of cross-coupling reaction applications to medicinal chemistry efforts, in particular the Suzuki–Miyaura and Buchwald–Hartwig cross-coupling reactions as a remarkable resource for the generation of carbon–carbon and carbon–heteroatom bonds. To further appreciate the impact of this methodology, the authors discuss some recent examples of clinical candidates that utilize key cross-coupling reactions in their large-scale synthetic process. Looking into future opportunities, the authors highlight the versatility of the cross-coupling reactions towards new chemical modalities like DNA-encoded libraries (DELs), new generation of peptides and cyclopeptides, allosteric modulators, and proteolysis targeting chimera (PROTAC) approaches.
Ranjana Aggarwal; Garima Sumran Eur. J. Med. Chem., 2020, 205, 112652 https://doi.org/10.1016/j.ejmech.2020.112652AbstractThe present review aims to summarize the pharmacological profile of 1,2,4-triazole, one of the emerging privileged scaffold, as antifungal, antibacterial, anticancer, anticonvulsant, antituberculosis, antiviral, antiparasitic, analgesic and anti-inflammatory agents, etc. along with structure-activity relationship. The comprehensive compilation of work carried out in the last decade on 1,2,4-triazole nucleus will provide inevitable scope for researchers for the advancement of novel potential drug candidates having better efficacy and selectivity.
Andrew G Reidenbach; Michael F Mesleh; Dominick Casalena; View ORCID ProfileSonia M Vallabh; Jayme L Dahlin; Alison J Leed; Alix I Chan; Dmitry L Usanov; Jenna B Yehl; Christopher T Lemke; Arthur J Campbell; Rishi N Shah; Om K Shrestha; Joshua R Sacher; Victor L Rangel; Jamie A Moroco; Murugappan Sathappa; Maria Cristina Nonato; Kong T Nguyen; S Kirk Wright; David R Liu; Florence F Wagner; Virendar K Kaushik; Douglas S Auld; Stuart L Schreiber; Eric Vallabh Minikel bioRxiv, 2020 https://doi.org/10.1101/2020.06.18.159418AbstractPrion disease is a rapidly progressive neurodegenerative disorder caused by misfolding and aggregation of the prion protein (PrP), and there are currently no therapeutic options. PrP ligands could theoretically antagonize prion formation by protecting the native protein from misfolding or by targeting it for degradation, but no validated small-molecule binders have been discovered to date. We deployed a variety of screening methods in an effort to discover binders of PrP, including 19F-observed and saturation transfer difference (STD) nuclear magnetic resonance (NMR) spectroscopy, differential scanning fluorimetry (DSF), DNA-encoded library selection, and in silico screening. A single benzimidazole compound was confirmed in concentration-response, but affinity was very weak (Kd > 1 mM), and it could not be advanced further. The exceptionally low hit rate observed here suggests that PrP is a difficult target for small-molecule binders. While orthogonal binder discovery methods could yield high affinity compounds, non-small-molecule modalities may offer independent paths forward against prion disease.
Scott A Busby; Seth Carbonneau; John Concannon; Christoph E. Dumelin; YounKyoung Lee; Shin Numao; Nicole Renaud; Thomas M Smith; Douglas S. Auld ACS Chem. Biol., 2020, 15(10), 2636-2648 https://doi.org/10.1021/acschembio.0c00495AbstractAssays drive drug discovery from the exploratory phases to the clinical testing of drug candidates. As such, numerous assay technologies and methodologies have arisen to support drug discovery efforts. Robust identification and characteri-zation of tractable chemical matter requires biochemical, biophysical, and cellular approaches and often benefits from high-throughput methods. To increase throughput, efforts have been made to provide assays in miniaturized volumes which can be arrayed in microtiter plates to support the testing of as many as 100,000 samples/day. Alongside these ef-forts has been the growth of microtiter plate-free formats with encoded libraries that can support the screening of billions of compounds, a hunt for new drug modalities, as well as emphasis on more disease relevant formats using complex cell models of disease states. This review will focus on recent developments in high-throughput assay technologies applied to identify starting points for drug discovery. We also provide recommendations on strategies for implementing various as-say types to select high quality leads for drug development.
Yapeng Bao; Zongfa Deng; Jing Feng; Weiwei Zhu; Jin Li; Jinqiao Wan; Guansai Liu Org. Lett., 2020, 22(16), 6277-6282 https://doi.org/10.1021/acs.orglett.0c02032AbstractIndazolone cores are among the most common structural components in medicinal chemistry and can be found in many biologically active molecules. In this report, a mild and efficient approach to 2-substituted indazolones via B2(OH)4-mediated reductive N–N bond formation is developed. This strategy features mild conditions, no request for a metal catalyst, and a wide scope for both aliphatic and aromatic amines. Meanwhile, this method was further successfully applied on DNA to construct indazolone cores for a DNA-encoded library. This will enable the production of a very attractive indazolone-cored library from simple amines and scaffolds, which will provide considerable diversity.
Dirk Trauer; Belinda E. Hetzler; Andrej Shemet Synfacts, 2020, 16(07), 0850 https://doi.org/10.1055/s-0040-1707873AbstractDNA-barcoded chemical libraries are powerful tools in drug discovery and allow for the rapid screening of large compound numbers for biological target engagement. The authors showcased DNA-compatible cycloaddition methods that form sp3-rich, rigid, and diverse scaffolds that are ideally suited for library generation.
Philip Kocienski Synfacts, 2020, 16(07), 0758 https://doi.org/10.1055/s-0040-1707017AbstractTarget molecule is an indoleamine-2,3-dioxygenase-1 inhibitor that is of interest for the treatment of cancer. Crystalline I was poorly soluble with low bioavailability but the highly bioavailable phosphonooxymethyl prodrug L and its crystalline tris-salt N were rapidly converted into I in vivo.
Shorouk O. Badir; Gary A. Molander Chem, 2020, 6, 1327–1339 https://doi.org/10.1016/j.chempr.2020.05.013AbstractProgress in Ni/photoredox dual catalysis has enabled the construction of C(sp3)-hybridized centers under extremely mild reaction conditions in the presence of diverse functional groups. These strategies, however, are mainly restricted to the assembly of one C–C or C–heteroatom linkage because of the competitive two-component reactions and facile β-hydride elimination from alkylmetal complexes. Recently, photoinduced nickel-catalyzed 1,2-difunctionalizations of alkenes and alkynes have attracted extensive research efforts because they allow the construction of two sequential chemical bonds from inexpensive starting materials in one pot. Herein, we explore recent advances, state the current challenges, and discuss perspectives on the design of new catalytic systems.
Luka Stojanović; Miloš Popović; Nebojša Tijanić; Goran Rakočević; Marko Kalinić ChemRxiv, 2020 https://doi.org/10.26434/chemrxiv.12412694.v1AbstractDeep learning has demonstrated significant potential in advancing state of the art in many problem domains, especially those benefiting from automated feature extraction. Yet the methodology has seen limited adoption in the field of ligand-based virtual screening (LBVS), as traditional approaches typically require large, target-specific training sets, which limits their value in most prospective applications. Here, we report the development of a neural network architecture, and a learning framework designed to yield a generally applicable tool for LBVS. Our approach uses the molecular graph as input, and involves learning a representation that places compounds of similar biological profiles in close proximity within a hyperdimensional feature space; this is achieved by simultaneously leveraging historical screening data against a multitude of targets during training. Cosine distance between molecules in this space becomes a general similarity metric, and can readily be used to rank order database compounds in LBVS workflows. We demonstrate the resulting model generalizes exceptionally well to compounds and targets not used in its training. In three commonly employed LBVS benchmarks, our method outperforms popular fingerprinting algorithms without the need for any target-specific training. Moreover, we show the learned representation yields superior performance in scaffold hopping tasks, and is largely orthogonal to existing fingerprints. Summarily, we have developed and validated a framework for learning a molecular representation that is applicable to LBVS in a target-agnostic fashion, with as few as one query compound. Our approach can also enable organizations to generate additional value from large screening data repositories, and to this end we are making its implementation freely available at https://github.com/totient-bio/gatnn-vs
Xiangyu Liu; Jonas Kaindl; Magdalena Korczynska; Anne Stößel; Daniela Dengler; Markus Stanek; Harald Hübner; Mary J. Clark; Jake Mahoney; Rachel Ann Matt; Xinyu Xu; Kunio Hirata; Brian K. Shoichet; Roger K. Sunahara; Brian K. Kobilka; Peter Gmeiner Nat. Chem. Biol., 2020, 16(7), 749-755 https://doi.org/10.1038/s41589-020-0549-2AbstractMost drugs acting on G-protein-coupled receptors target the orthosteric binding pocket where the native hormone or neurotransmitter binds. There is much interest in finding allosteric ligands for these targets because they modulate physiologic signaling and promise to be more selective than orthosteric ligands. Here we describe a newly developed allosteric modulator of the β2-adrenergic receptor (β2AR), AS408, that binds to the membrane-facing surface of transmembrane segments 3 and 5, as revealed by X-ray crystallography. AS408 disrupts a water-mediated polar network involving E1223.41 and the backbone carbonyls of V2065.45 and S2075.46. The AS408 binding site is adjacent to a previously identified molecular switch for β2AR activation formed by I3.40, P5.50 and F6.44. The structure reveals how AS408 stabilizes the inactive conformation of this switch, thereby acting as a negative allosteric modulator for agonists and positive allosteric modulator for inverse agonists.
Xi Feng; Dongdong Liao; Dongyu Liu; An Ping; Zhiyu Li; Jinlei Bian J. Med. Chem., 2020, 63(24), 15115-15139 https://doi.org/10.1021/acs.jmedchem.0c00925AbstractIndoleamine 2,3-dioxygenase 1 (IDO1) has received increasing attention due to its immunosuppressive function in connection with various diseases, including cancer. A recent increase in the understanding of IDO1 has significantly contributed to the discovery of numerous novel inhibitors, but the latest clinical outcomes raised questions and have indicated a future direction of IDO1 inhibition for therapeutic approaches. Herein, we present a comprehensive review of IDO1, discussing the latest advances in understanding the IDO1 structure and mechanism, an overview of recent IDO1 inhibitor discoveries and potential therapeutic applications to provide helpful information for medicinal chemists investigating IDO1 inhibitors.
Xin Qian; Yan Tian; Xinxin Luo; Jingmiao Pan; Suya Deng; Yike Huang; Qifeng Fu; Zhining Xia Chin. J. Chromatogr., 2020, 38(10), 1170-1178 https://doi.org/10.3724/SP.J.1123.2020.05040AbstractCapillary electrophoresis(CE)shows enormous potential for application in new drug research and development.Because of the aqueous medium employed as the running buffer in CE,drug screening can be carried out in an environment similar to that in physiological testing media.Drug screening methods based on CE are different from other instrumental measurements in vitro.CE can not only sustain the biological activity of the screened molecules and ligands,but also help evaluate the interactions between the receptors and the ligands.Based on these interactions,some important pharmacological parameters related to drug screening,such as the association constant Kb,bonding rate constant Kon,and dissociation rate constant Koff,can be determined by CE.Thus,CE is an effective tool for simulating and predicting the entire interaction process between receptors and drugs in vivo.In this review,the history of CE for drug screening is revisited.The theories,common methods for drug screening by CE,and some application examples and related technologies are reviewed.The methods of drug screening by means of affinity CE and kinetic CE are introduced.Some selected studies on different ligands at the molecular and cellular level are reported,along with examples several types of drugs.Techniques based on a combination of CE with mass spectrometry and chemiluminescence are reviewed,with focus on the screening of candidate drugs and active compounds from traditional Chinese medicine.The application prospect of drug screening by CE combined with a DNA-encoded compound library is introduced.This paper discusses the core of the fraction collection step in CE and emphasizes the significance of combining CE with systematic evolution of ligands by exponential enrichment.In conclusion,various optional methods for CE drug screening would pave the way for new concepts related to drug screening and evaluation in the future.
Marco Catalano; Sebastian Oehler; Luca Prati; Nicholas Favalli; Gabriele Bassi; Joerg Scheuermann; Dario Neri Anal. Chem., 2020, 92(15), 10822-10829 https://doi.org/10.1021/acs.analchem.0c02304AbstractThe availability of reliable methods for the characterization of the binding of small molecule ligands to protein targets is crucially im-portant for Drug Discovery. We have adapted a method, routinely used for the characterization of monoclonal antibodies (Enzyme-linked immunosorbent assay, or “ELISA”), to small molecule ligands, using fluorescein conjugates and anti-fluorescein antibodies as de-tection reagents. The new small molecule-ELISA methodology was tested using a panel of binders specific to carbonic anhydrase II, with dissociation constants ranging between 6 μM and 14 nM. An excellent agreement was found between ELISA measurements and fluo-rescence polarization results. The methodology was also extended to BIAcore measurements and implemented for ligands coupled to oligonucleotides. Small molecule-ELISA procedures are particularly useful in the context of DNA-encoded libraries, for which hit valida-tion procedures need to be performed on dozens of candidate molecules and hit compounds can be conveniently resynthesized on DNA.
Marco Catalano; Sebastian Oehler; Luca Prati; Nicholas Favalli; Gabriele Bassi; Jörg Scheuermann; Dario Neri ChemRxiv, 2020 https://doi.org/10.26434/chemrxiv.12382580.v1AbstractThe availability of reliable methods for the characterization of the binding of small molecule ligands to protein targets is crucially important for Drug Discovery. We have adapted a method, routinely used for the characterization of monoclonal antibodies (Enzyme-linked immunosorbent assay, or “ELISA”), to small molecule ligands, using fluorescein conjugates and anti-fluorescein antibodies as detection reagents. The new small molecule-ELISA methodology was tested using a panel of binders specific to carbonic anhydrase II, with dissociation constants ranging between 6 uM and 14 nM. An excellent agreement was found between ELISA measurements and fluorescence polarization results. The methodology was also extended to BIAcore measurements and implemented for ligands coupled to oligonucleotides. Small molecule-ELISA procedures are particularly useful in the context of DNA-encoded libraries, for which hit validation procedures need to be performed on dozens of candidate molecules and hit compounds can be conveniently resynthesized on DNA.
Gabriele Bassi; Nicholas Favalli; Miriam Vuk; Marco Catalano; Adriano Martinelli; Anika Trenner; Antonio Porro; Su Yang; Chuin Lean Tham; Mustafa Moroglu; Wyatt W. Yue; Stuart J. Conway; Peter K. Vogt; Alessandro A. Sartori; Jörg Scheuermann; Dario Neri Adv. Sci., 2020, 2001970 https://doi.org/10.1002/advs.202001970AbstractA versatile and Lipinski‐compliant DNA‐encoded library (DEL), comprising 366 600 glutamic acid derivatives coupled to oligonucleotides serving as amplifiable identification barcodes is designed, constructed, and characterized. The GB‐DEL library, constructed in single‐stranded DNA format, allows de novo identification of specific binders against several pharmaceutically relevant proteins. Moreover, hybridization of the single‐stranded DEL with a set of known protein ligands of low to medium affinity coupled to a complementary DNA strand results in self‐assembled selectable chemical structures, leading to the identification of affinity‐matured compounds.
Alexander K. Price; Brian M. Paegel ACS Comb. Sci., 2020, 22(11), 649-655 https://doi.org/10.1021/acscombsci.0c00101AbstractDNA-encoded library (DEL) technology enables rapid, economical synthesis and exploration of novel chemical space. Reaction development for DEL synthesis has recently accelerated in pace with a specific emphasis on ensuring that the reaction does not compromise the integrity of the encoding DNA. However, the factors that contribute to a reaction’s “DNA compatibility” remain relatively unknown. We investigated several solid-phase reactions and encoding conditions and determined their impact on DNA compatibility. Conditions that minimized the accessibility of reactive groups on the DNA encoding tag (switching solvent, low temperature, double-stranded encoding tag) significantly improved compatibility. We showcased this approach in the multi-step synthesis of an acyldepsipeptide (ADEP1) fragment, which preserved 73% of DNA for a >100-fold improvement over canonical conditions. These results are particularly encouraging in the context of multi-step reaction sequences to access natural product-like scaffolds, and more broadly underscore the importance of reconciling the biophysical properties and reactivity of DNA with chemistry development to yield high-quality libraries of those scaffolds.
Reto Horst; Kathleen A. Farley; Bethany L. Kormos; Jane M. Withka J. Biomol. NMR, 2020, 74(10-11), 509-519 https://doi.org/10.1007/s10858-020-00330-0AbstractNuclear magnetic resonance (NMR) spectroscopy has evolved into a powerful tool within drug discovery over the last two decades. While traditionally being used by medicinal chemists for small molecule structure elucidation, it can also be a valuable tool for the identifcation of small molecules that bind to drug targets, for the characterization of target–ligand interactions and for hit-to-lead optimization. Here, we describe how NMR spectroscopy is integrated into the Pfzer drug discovery pipeline and how we utilize this approach to identify and validate initial hits and generate leads.
Yuqing Deng; Jianzhao Peng; Feng Xiong; Yinan Song; Yu Zhou; Jianfu Zhang; Fong Sang Lam; Chao Xie; Wenyin Shen; Yiran Huang; Ling Meng; Xiaoyu Li Angew. Chem. Int. Ed. Engl., 2020, 59 (35), 14965-14972 https://doi.org/10.1002/anie.202005070AbstractDynamic combinatorial library (DCL) is a powerful tool for ligand discovery in biomedical research; however, the development of DCL has been hampered by its low diversity. Recently, the concept of DNA encoding has been employed in DCL to create DNA-encoded dynamic libraries (DEDLs); however, all current DEDLs are limited to fragment identification, and a challenging process of fragment linking is required after selection. We report an anchor-directed DEDL approach that can identify full ligand structures from large-scale DEDLs. This method is also able to convert unbiased libraries to focused ones targeting specific protein classes. We demonstrated this method by selecting DEDLs against five proteins, and novel inhibitors have been identified for all targets. Notably, several selective BD1/BD2 inhibitors were identified from the selections against BRD4 (bromodomain 4), an important anti-cancer drug target. This work may provide a broadly applicable method for inhibitor discovery.
Sean B. Yeldell; Oliver Seitz Chem. Soc. Rev., 2020, 49(19), 6848-6865 https://doi.org/10.1039/D0CS00518EAbstractMultivalency is nature's way to establish firm and specific interactions when the binding sites of a protein receptor have only low affinity for monovalent ligands. Recently, researchers are increasingly using nucleic acid architectures for multivalent ligand presentation to unravel the mechanisms of multivalency-enhanced interactions and create high affinity binding agents. In contrast to other polymers, nucleic acid materials are capable of accessing a wide variety of rigid three-dimensional structures through the sequence-programed self-assembly of component strands. By controlling the number of ligands and their distances, researchers can construct tailor-made probes for interrogating multivalent interactions with Ångstrom precision. Nucleic acid assemblies have been used to address fundamental questions of multivalency in order to unravel how monovalent interaction strength, scaffold flexibility, distances between interacting sites and spatial arrangement influence the achievable affinity gains. In a slightly different approach, nucleic acid constructs have been applied as chemical dimerizers of protein receptors, to investigate the importance of receptor proximity or construct tools that provide control over biological signal transduction processes. In this review, we discuss multivalent nucleic acid–ligand conjugates in the context of the biological protein receptors they interrogate. We recount pioneering work and seminal studies performed within the last 10 years describing the in vitro interrogation of proteins recognizing carbohydrate ligands, small molecules, peptides and nucleic acid aptamers and we portray work performed with viruses, cell models, and whole organisms.
Xin Wen; Zhiqiang Duan; Jiaxiang Liu; Weiwei Lu; Xiaojie Lu Org. Lett., 2020, 22(15), 5721-5725 https://doi.org/10.1021/acs.orglett.0c01565AbstractTo expand the chemical space of DNA-encoded library (DEL) technology, we have developed the DNA-compatible cross-dehydrogenative coupling reaction between tetrahydroisoquinolines (THIQs) and a series of nucleophiles such as nitromethane, terminal alkynes, and so on. Both photo- and metal-promoted conditions have been successfully established for this C-1 derivation of THIQs, as exemplified by the addition of nitromethane. All of these new DNA-compatible transformations have paved the way for the divergent oriented synthesis of THIQ-focused DELs.
Herschel Mukherjee; J. Craig Blain; Lee E. Vandivier; Donovan N. Chin; Jessica E. Friedman; Fei Liu; Ashley Maillet; Chao Fang; Jenifer B. Kaplan; Jinxing Li; David M. Chenoweth; Allan Beck Christensen; Lars Kolster Petersen; Nils Jakob Vest Hansen; Luis Barrera; Neil Kubica; Gnanasambandam Kumaravel; Jennifer C. Petter ACS Chem. Biol., 2020, 15(9), 2374-2381 https://doi.org/10.1021/acschembio.0c00357AbstractRNA is emerging as a valuable target for the development of novel therapeutic agents. The rational design of RNA-targeting small molecules, however, has been hampered by the relative lack of methods for the analysis of small molecule–RNA interactions. Here, we present our efforts to develop such a platform using photoaffinity labeling. This technique, termed Photoaffinity Evaluation of RNA Ligation-Sequencing (PEARL-seq), enables the rapid identification of small molecule binding locations within their RNA targets and can provide information on ligand selectivity across multiple different RNAs. These data, when supplemented with small molecule SAR data and RNA probing data enable the construction of a computational model of the RNA–ligand structure, thereby enabling the rational design of novel RNA-targeted ligands.
Michael R. Green; Joseph Sambrook Cold Spring Harb. Protoc., 2020 https://doi.org/10.1101/pdb.top100743AbstractEscherichia coli DNA Pol I can carry out three enzymatic reactions: It possesses 5′ → 3′ DNA polymerase activity and 3′ → 5′ and 5′ → 3′ exonuclease activity. Pol I can be cleaved by mild treatment with subtilisin into two fragments; the larger fragment is known as the Klenow fragment. The Klenow fragment retains the polymerizing activity and the 3′ → 5′ exonuclease of the holo-enzyme but lacks its powerful 5′ → 3′ exonuclease activity. These enzymes and their applications in molecular cloning are introduced here.
Jinjian Liu; Lingxiang Lu; Devin Wood; Song Lin ACS Cent. Sci., 2020, 6(8), 1317-1340 https://doi.org/10.1021/acscentsci.0c00549AbstractAs the breadth of radical chemistry grows, new means to promote and regulate single-electron redox activities play increasingly important roles in driving modern synthetic innovation. In this regard, photochemistry and electrochemistry—both considered as niche fields for decades—have seen an explosive renewal of interest in recent years and gradually have become a cornerstone of organic chemistry. In this Outlook article, we examine the current state-of-the-art in the areas of electrochemistry and photochemistry, as well as the nascent area of electrophotochemistry. These techniques employ external stimuli to activate organic molecules and imbue privileged control of reaction progress and selectivity that is challenging to traditional chemical methods. Thus, they provide alternative entries to known and new reactive intermediates and enable distinct synthetic strategies that were previously unimaginable. Of the many hallmarks, electro- and photochemistry are often classified as “green” technologies, promoting organic reactions under mild conditions without the necessity for potent and wasteful oxidants and reductants. This Outlook reviews the most recent growth of these fields with special emphasis on conceptual advances that have given rise to enhanced accessibility to the tools of the modern chemical trade.
Verena B. K. Kunig; Marco Potowski; Mohammad Akbarzadeh; Mateja Klika Škopić; Denise Dos Santos Smith; Lukas Arendt; Ina Dormuth; Hélène Adihou; Blaž Andlovic; Hacer Karatas; Shabnam Shaabani; Tryfon Zarganes-Tzitzikas; Constantinos G. Neochoritis; Ran Zhang; Matthew Groves; Stéphanie M. Guéret; Christian Ottmann; Jörg Rahnenführer; Roland Fried; Alexander Dömling; Andreas Brunschweiger Angew. Chem. Int. Ed. Engl., 2020, 59(46), 20338-20342 https://doi.org/10.1002/anie.202006280AbstractDNA‐encoded combinatorial synthesis provides efficient and dense coverage of chemical space around privileged molecular structures. The indole side chain of tryptophan plays a prominent role in key, or “hot spot” regions of protein‐protein interactions. A DNA‐encoded combinatorial peptoid library was designed based on the Ugi four‐component reaction employing tryptophan‐mimetic indole side chains to probe target protein surface. Several peptoids were synthesized on a chemically stable hexathymidine adapter oligonucleotide “hexT”, encoded by DNA sequences and substituted by azide‐alkyne cycloaddition to yield a library of 8,112 molecules. Selection experiments on the tumor‐relevant proteins MDM2 and TEAD4 yielded MDM2 binders and a novel class of TEAD‐YAP interaction inhibitors that perturbed expression of a gene under the control of these Hippo pathway effectors.
Joseph Lau; Etienne Rousseau; Daniel Kwon; Kuo‐Shyan Lin; François Bénard; Xiaoyuan Chen Cancers, 2020, 12, 1312 https://doi.org/10.3390/cancers12051312AbstractWhile the development of positron emission tomography (PET) radiopharmaceuticals closely follows that of traditional drug development, there are several key considerations in the chemical and radiochemical synthesis, preclinical assessment, and clinical translation of PET radiotracers. As such, we outline the fundamentals of radiotracer design, with respect to the selection of an appropriate pharmacophore. These concepts will be reinforced by exemplary cases of PET radiotracer development, both with respect to their preclinical and clinical evaluation. We also provide a guideline for the proper selection of a radionuclide and the appropriate labeling strategy to access a tracer with optimal imaging qualities. Finally, we summarize the methodology of their evaluation in in vitro and animal models and the road to clinical translation. This review is intended to be a primer for newcomers to the field and give insight into the workflow of developing radiopharmaceuticals.
John W. Cuozzo; Matthew A. Clark; Anthony D. Keefe; Anna Kohlmann; Mark J Mulvihill; Haihong Ni; Louis Renzetti; Daniel I. Resnicow; Frank Ruebsam; Eric A. Sigel; Heather A. Thomson; Ce Wang; Zhifeng Xie; Ying Zhang J. Med. Chem., 2020, 63, 14, 7840-7856 https://doi.org/10.1021/acs.jmedchem.0c00688AbstractThe activity of the secreted phosphodiesterase autotaxin produces the inflammatory signaling molecule LPA and has been associated with a number of human diseases including idiopathic pulmonary fibrosis (IPF). We screened a single DNA-encoded chemical library (DECL) of 225 million compounds and identified a series of potent and selective inhibitors. Optimization of this series led to the discovery of compound 1 (X-165), a highly potent, selective and bioavailable small molecule. Co-crystallization of compound 1 with human autotaxin demonstrated that it has a novel binding mode occupying both the hydrophobic pocket and a channel near the autotaxin active site. Compound 1 inhibited the production of LPA in human and mouse plasma at nanomolar levels and showed efficacy in a mouse model of human lung fibrosis. After successfully completing IND-enabling studies, compound 1 was approved by the FDA for a Phase I clinical trial. These results demonstrate that DECL hits can be readily optimized into clinical candidates.
Chandravadivelu Gopi; Gudapati Krupamai; Chitikina Satya Sri; Magharla Dasaratha Dhanaraju Beni-Suef Univ. J. Basic Appl. Sci., 2020, 9, 57 https://doi.org/10.1186/s43088-020-00083-7AbstractBackground: In recent times, a powerful tool of combinatorial synthesis has been used for the preparation of large chemical entities through a small set up of reactions between different building blocks using solidphase and solution-phase techniques. This method reduced the time and cost of the drug discovery process substantially. Main text: Thousands of compounds are synthesised in a few reactions through combinatorial synthesis instead of getting a few compounds in the traditional method. This method also helps to identify chemical lead of the compounds and optimise them through the biological screening using a high-throughput method. There is no review concerning the recent research finding of combinatorial synthesis. Hence, an attempt had been made on the latest research findings (2002–2020) of newly synthesised compounds using combinatorial synthesis and their biological activities. Conclusion: To the best of our knowledge, the current review has completely analysed the importance of combinatorial synthesis and furnished an overview of solid-phase and solution-phase techniques as well as helped mankind by improving higher productivity at low cost, lead identification and optimization and preventing environmental pollution.
Gabriele Bassi; Nicholas Favalli; Sebastian Oehler; Adriano Martinelli; Marco Catalano; Jörg Scheuermann; Dario Neri Biochem. Biophys. Res. Commun., 2020, 533, 2, 223-229 https://doi.org/10.1016/j.bbrc.2020.04.035AbstractDNA-encoded chemical libraries (DEL) are increasingly being used for the discovery and optimization of small organic ligands to proteins of biological or pharmaceutical interest. The DNA fragments, that serve as amplifiable identification barcodes for individual compounds in the library, are typically used in double-stranded DNA format. To the best of our knowledge, a direct comparison of DEL selections featuring DNA in either single- or double-stranded DNA format has not yet been reported. In this article, we describe a comparative evaluation of selections with two DEL libraries (named GB-DEL and NF-DEL), based on different chemical designs and produced in both single- and double-stranded DNA format. The libraries were selected in identical conditions against multiple protein targets, revealing comparable and reproducible fingerprints for both types of DNA formats. Surprisingly, selections performed with single-stranded DNA barcodes exhibited improved enrichment factors compared to double-stranded DNA. Using high-affinity ligands to carbonic anhydrase IX as benchmarks for selection performance, we observed an improved selectivity for the NF-DEL library (on average 2-fold higher enrichment factors) in favor of single-stranded DNA. The enrichment factors were even higher for the GB-DEL selections (approximately 5-fold), compared to the same library in double-stranded DNA format. Collectively, these results indicate that DEL libraries can conveniently be synthesized and screened in both single- and double-stranded DNA format, but single-stranded DNA barcodes typically yield enhanced enrichment factors.
Richard A. Lerner; Dario Neri Biochem. Biophys. Res. Commun., 2020, 527(3), 757-759 https://doi.org/10.1016/j.bbrc.2020.04.080AbstractIn this short Editorial, we reflect on certain milestones that have led to the development of DNA-Encoded Chemical Libraries, while also providing a glimpse to future challenges and opportunities. We (RAL & DN) have been asked to write an introduction to the present volume. Since this is a collection of reports from prominent researchers in the field of DNA-encoded chemical libraries (DEL), merely summarizing its content would put us in the position of explaining accidents to eye witnesses. So instead, we have decided to focus on a more historical perspective, and specifically, to relate some of our personal experiences that have contributed to the origins and development of the DEL field. Our hope is that this may be both entertaining and useful - as sometimes history is a preface to the present, and even to the future. In an extraordinary event, the CRO Wuxi took up the challenge and built an 800 billion-member library, of which 80 billion members are freely available to academics under the general considerations mentioned above.
Patrick McEnaney; Madeline Blalzarini; HaJeung Park; Thomas Kodadek Chem. Commun., 2020, 56(72), 10560-10563 https://doi.org/10.1039/D0CC02588GAbstractA screen of a DNA-encoded library of PICCOs was screened against streptavidin (SA) resulting in the isolation of a high affinity ligand. The X-ray crystal structure of this complex is presented, providing the first detailed view of how a PICCO molecule interacts with a protein.
Dillon T. Flood; Kyle W. Knouse; Julien C. Vantourout; Brittany Sanchez; Emily J. Sturgell; Jason Chen; Phil Baran; Philip Dawson ChemRxiv, 2020 https://doi.org/10.26434/chemrxiv.12094053.v1AbstractThe controlled, site-specific ligation of molecules to native DNA remains an unanswered challenge. Herein, we report a simple solution to achieve this ligation through the tactical combination of two recently developed technologies: One for the manipulation of DNA in organic media, and another for the chemoselective labeling of alcohols. Reversible Adsorption of Solid Support (RASS) is employed to immobilize DNA and facilitate its transfer into dry acetonitrile. Subsequent ligation with P(V)-based Ψ reagents takes place in high yield with exquisite selectivity for the exposed 3’ or 5’ alcohols on DNA. This two-stage process, dubbed SENDR for Synthetic Elaboration of Native DNA by RASS, can be applied to a multitude of DNA conformations and sequences with a variety of functionalized Ψ reagents to generate useful constructs. Such entities can address numerous longstanding challenges, including the selective single coupling of DNA to proteins, ASOs, and functional small molecules, and also can allow the synthesis of doubly-labeled congeners for novel probe constructs including ones of potential interest to COVID-19 research. Finally, a prototype for the industrialization of SENDR in a kit format is presented.
Justin Hall; Timothy L. Foley; Qiuxia Chen; David I. Israel; Yanshan Xu; Kristen F. Ford; Ping Xie; Jing Fan; Jinqiao Wan Biochem. Biophys. Res. Commun., 2020, 527, 1, 250-256 https://doi.org/10.1016/j.bbrc.2020.04.024AbstractDNA-encoded libraries (DELs) can contain billions of unique chemical species; selecting against such large inputs, it is typical to find more candidate binders than is reasonable to pursue for follow-up synthesis and testing. Given this wealth of choices, common practice is to limit synthesis to only those compounds estimated to have the greatest chance of being high-affinity binders; of the many potential factors contributing to this estimation, the strength of the selection signal of a candidate binder is always important. We define here methods and equations which relate the theoretical selection signal of a compound to its affinity and chemical yield. Tests using known binders of BRD4 and ROCK2 support the theory backing these equations and suggest they should be of use for prospectively determining affinity and chemical yield from primary DEL selection data.
Meiying Cui; Yixin Zhang ChemBioChem, 2020, 21(17), 2503-2511 https://doi.org/10.1002/cbic.202000149AbstractDNA has become a promising candidate as future data storage medium, which makes DNA steganography indispensable in DNA data security. PCR primers are conventional secret keys in DNA steganography. Brutal force testing of different primers will be extremely time consuming, and practically unaffordable when high throughput sequencing is used. However, the information can be sequenced and read once the primers are intercepted. New steganography approach is needed to make the DNA‐encoded information safer. Herein, by mixing information DNA with partially degenerated DNA library containing single or multiple restriction sites, we build an additional protective layer, which can be removed by desired restriction enzymes as secondary secret keys. As PCR is inevitable for reading DNA‐encrypted information, heating will cause reshuffling and generate endonuclease‐resistant mismatched duplexes, especially for DNA with high sequence diversity. Consequently, with the incorporation of randomness, the DNA steganography possesses both quantum key distribution (QKD)‐like function for detecting PCR by an interceptor and self‐destructive property. It is noteworthy that the background noise generated through the protective layer is independent from any sequencing technology including Sanger and high throughput sequencing. With a DNA‐ink incorporating the steganography, we have shown that the authenticity of a writing can be confirmed only by authorized person with the knowledge of all keys.
Yi Qu; Huanan Wen; Rui Ge; Yanfen Xu; Hong Gao; Xiaodong Shi; Jiangong Wang; Weiren Cui; Wenji Su; Hongfang Yang; Letian Kuai; Alexander L. Satz; Xuanjia Peng Org. Lett., 2020, 22, 11, 4146-4150 https://doi.org/10.1021/acs.orglett.0c01219AbstractWe report a DNA-compatible copper-mediated efficient synthesis of 1,2,3-triazoles via a one-pot reaction of aryl borates with TMS-N3 followed by a click cycloaddition reaction. Employing the binuclear macrocyclic nanocatalyst Cu(II)-β-cyclodextrin, the reactions were performed under mild conditions with high conversions and wide functional group tolerance. We also demonstrate the reaction application toward a one-pot DNA-compatible intramolecular macrocyclization. Our optimized reaction protocol results in no significant DNA damage as judged by qPCR analysis and Sanger sequencing data.
Emory M. Payne; Daniel A. Holland-Moritz; Shuwen Sun; Robert T. Kennedy Lab Chip, 2020,20, 2247-2262 https://doi.org/10.1039/D0LC00347FAbstractIn two decades of development, impressive strides have been made for automating basic laboratory operations in droplet-based microfluidics, allowing the emergence of a new form of high-throughput screening and experimentation in nanoliter to femtoliter volumes. Despite advancements in droplet storage, manipulation, and analysis, the field has not yet been widely adapted for many high-throughput screening (HTS) applications. Broad adoption and commercial development of these techniques require robust implementation of strategies for the stable storage, chemical containment, generation of libraries, sample tracking, and chemical analysis of these small samples. We discuss these challenges for implementing droplet HTS and highlight key strategies that have begun to address these concerns. Recent advances in the field leave us optimistic about the future prospects of this rapidly developing technology.
Yulong An; Ke Li; Yanfang Shen; Zibing Hong; Linlin Chen; Yujing Hu; Lichun Zhou; Disha Wang; Xiaodong Shi; Shien Liu; Wenji Su; Weiren Cui; Letian Kuai; Hongfang Yang; Xuanjia Peng Org. Lett., 2020, 22, 10, 3931-3935 https://doi.org/10.1021/acs.orglett.0c01215AbstractA robust DNA-compatible Wittig reaction mediated by PPh2CH3 has been validated for DNA-conjugated α-chloroacetamides with aldehydes and, alternatively, DNA-conjugated aldehydes with α-halo acetamides or ketones. Further, 2-aminopyridines were acylated with α-chloroacetyl chloride and then reacted with DNA-conjugated aldehydes. Lastly, a pilot library employing our optimized Wittig reaction protocol was synthesized. The ability to generate α,β-unsaturated carbonyl compounds may be particularly useful for the design of DNA-encoded libraries capable of covalently interacting with protein targets.
Jens Bobers; Julia Gruehn; Stefan Hoeving; Tobias Pyka; Norbert Kockmann Org. Process Res. Dev., 2020, 24(10), 2094-2104 https://doi.org/10.1021/acs.oprd.0c00152AbstractThe transfer of batch processes to continuous flow is a major driver for the application of microreactors. Here, we present a methodology for the transfer of (bio-)chemical reactions in batch mode to two-phase continuous flow. For our purposes the coiled flow inverter (CFI) is a promising reactor design providing enhanced heat and mass transfer, narrow residence time distribution, and rapid mixing. First, this methodology is used for current development of a droplet-based reaction screening system, which was first tested with a Paal-Knorr pyrrole synthesis as model reaction. The reaction was successfully performed in the automated screening system. The yields compared to the batch mode revealed enhanced mass transfer of the product into the continuous phase. Second, we investigated the bio-catalyzed oxidation of ABTS by the enzyme laccase in a straight capillary for process development in a CFI. Because of its high flexibility regarding substrate specificity, laccase oxidizes many substrates with a colored product. Hence, an optical evaluation method for determination of reaction rate is used. We compare the Michaelis-Menten kinetic of the batch reaction and the continuous reaction in a capillary. The results show that the batch reaction can be mapped to the capillary setup. However, the capillary in continuous operation enables higher screening capacity of different reaction conditions and simple scale-up procedure.
Shukkoor Muhammed Kondengaden; Jiabin Zhang; Huajie Zhang; Aishwarya Parameswaran; Shameer M. Kondengadan; Shrikant Pawar; Akhila Puthengot; Rajshekhar Sunderraman; Jing Song; Samuel J. Polizzi; Liuqing Wen; Peng George Wang bioRxiv, 2020 https://doi.org/10.1101/2020.03.30.017012AbstractInteractions between glycans and glycan-binding proteins (GBPs) mediate diverse cellular functions, and therefore are of diagnostic and therapeutic significance. Current leading strategies for studying glycan-GBP interactions require specialized knowledge and instrumentation. In this study, we report a strategy for studying glycan-GBP interactions that uses PCR, qPCR and nextgeneration sequencing (NGS) technologies that are more routinely accessible. Our headpiece conjugation-code ligation (HCCL) strategy couples glycans with unique DNA codes that specify glycan sugar moieties and glycosidic linkages when sequenced. We demonstrate the technology by synthesizing a DNA encoded glycan library of 50 biologically relevant glycans (DEGL-50) and probing interactions against 25 target proteins including lectins and antibodies. Data show glycan-GPB interactions in solution that are consistent with lower content, lower throughput ELISA assays. Data further demonstrate how monovalent and multivalent headpieces can be used to increase glycan-GPB interactions and enrich signals while using smaller sample sizes. The flexibility of our modular HCCL strategy has potential for producing large glycan libraries, facilitating high content-high throughput glycan binding studies, and increasing access to lower cost glyco-analyses.
Emma. K. Grant; David J. Fallon; Michael M. Hann; Ken G. M. Fantom; Chad Quinn; Francesca Zappacosta; Roland S. Annan; Chun-wa Chung; Paul Bamborough; David P. Dixon; Peter Stacey; David House; Vipulkumar K. Patel; Nicholas C. O. Tomkinson; Jacob T. Bush ChemRxiv, 2020. 04.01 https://chemrxiv.org/articles/PhotoAffinity_Bits_A_Photoaffinity-Based_Fragment_Screening_Platform_for_Efficient_Identification_of_Protein_Ligands/12053445/1?file=22150278AbstractAdvances in genomic analyses enable the identification of new proteins that are associated with disease. To validate these targets, tool molecules are required to demonstrate that a ligand can have a disease-modifying effect. Currently, as tools are reported for only a fraction of the proteome, platforms for ligand discovery are essential to leverage insights from genomic analyses. Fragment screening offers an efficient approach to explore chemical space, however, it remains challenging to develop techniques that are both sufficiently high-throughput and sensitive. We present a fragment screening platform, termed PhABits (PhotoAffinity Bits), which utilises a library of photoreactive fragments to covalently capture fragment-protein interactions. Hits can be profiled to determine potency and site of crosslinking, and subsequently developed as reporters in a competitive displacement assay to identify novel hit matter. We envision that the PhABits will be widely applicable to novel protein targets, identifying starting points in the development of therapeutics.
Surendra Dawadi; Nicholas Simmons; Gabriella Miklossy; Kurt M. Bohren; John C. Faver; Melek Nihan Ucisik; Pranavanand Nyshadham; Zhifeng Yu; Martin M. Matzuk Proc. Natl. Acad. Sci. USA, 2020, 117(29), 16782-16789 https://doi.org/10.1073/pnas.2005447117AbstractDNA-encoded chemical libraries are collections of compounds individually coupled to unique DNA tags serving as amplifiable identification barcodes. By bridging split-and-pool combinatorial synthesis with the ligation of unique encoding DNA oligomers, million- to billion-member libraries can be synthesized for use in hundreds of healthcare target screens. Although structural diversity and desirable molecular property ranges generally guide DNA-encoded chemical library design, recent reports have highlighted the utility of focused DNA-encoded chemical libraries that are structurally biased for a class of protein targets. Herein, a protease-focused DNA-encoded chemical library was designed that utilizes chemotypes known to engage conserved catalytic protease residues. The three-cycle library features functional moieties such as guanidine, which interacts strongly with aspartate of the protease catalytic triad, as well as mild electrophiles such as sulfonamide, urea, and carbamate. We developed a DNA-compatible method for guanidinylation of amines and reduction of nitriles. Employing these optimized reactions, we constructed a 9.8-million-membered DNA-encoded chemical library. Affinity selection of the library with thrombin, a common protease, revealed a number of enriched features which ultimately led to the discovery of a 1 nM inhibitor of thrombin. Thus, structurally focused DNA-encoded chemical libraries have tremendous potential to find clinically useful high-affinity hits for the rapid discovery of drugs for targets (e.g., proteases) with essential functions in infectious diseases (e.g., severe acute respiratory syndrome coronavirus 2) and relevant healthcare conditions (e.g., male contraception).
Ki Tae Kim; Nicolas Winssinger Chem. Sci., 2020, 11, 4150-4157 https://doi.org/10.1039/D0SC00741BAbstractDNA or RNA templated reactions are attractive for nucleic acid sensing and imaging. As for any hybridization-based sensing, there is a tradeoff between sensitivity (detection threshold) and resolution (single nucleotide discrimination). Longer probes afford better sensitivity but compromise single nucleotide resolution due to the small thermodynamic penalty of a single mismatch. Herein we report a design that overcomes this tradeoff. The reaction is leveraged on the hybridization of a minimal substrate (covering 4 nucleotides) which is confined by two guide DNAs functionalized respectively with a ruthenium photocatalyst. The use of a catalytic reaction is essential to bypass the exchange of guide DNAs while achieving signal amplification through substrate turnover. The guide DNAs restrain the reaction to a unique site and enhance the hybridization of short substrates by providing two π-stacking interactions. The reaction was shown to enable the detection of SNPs and SNVs down to 50 pM with a discrimination factor ranging from 24 to 309 (median 82, 27 examples from 3 oncogenes). The clinical diagnostic potential of the technology was demonstrated with the analysis of RAS amplicons obtained directly from cell culture.
Wenting Wu; Zhen Sun; Xuan Wang; Xiaojie Lu; Dongcheng Dai Org. Lett., 2020, 22, 8, 3239-3244 https://doi.org/10.1021/acs.orglett.0c01016AbstractAn efficient and facile formal [4 + 2] cycloaddition reaction was developed to synthesize diverse thiazole-fused dihydropyrans (TFDP) on DNA. Mild reaction conditions, broad substrate scope, and compatibility with subsequent enzymatic ligation demonstrated the utility of this methodology in DNA-encoded library synthesis.
Fabrice Ndayisenga; Po-Han Lin; Runan Li; Abdul Hameed; Yue Zhang; Kamel Meguellati Int. J. Biosci., 2020, 16(3), 12-50 http://dx.doi.org/10.12692/ijb/16.3.12-50AbstractFunctionalization of proteins is a particular domain for the generation of modified proteins constructs which exhibit stimulating and diverse biological properties. The area has enormous potential for chemists and biologists to tune the proteins functions. The site-specific modification of proteins by installing new moieties or subunits provides an excellent opportunity to expand proteins functional abilities. In this review, we highlighted the most significant studies in protein functionalization which include chemical and enzymatic strategies. Exploration of novel methodologies is still on-going and deserves significant attention for future research in protein functionalization.
Tim J. Wigle; Danielle J. Blackwell; Laurie B. Schenkel; Yue Ren; W. David Church; Hetvi J. Desai; Kerren K. Swinger; Andrew G. Santospago; Christina R. Majer; Alvin Z. Lu; Mario Niepel; Nicholas R. Perl; Melissa M. Vasbinder; Heike Keilhack; Kevin W. Kuntz Cell Chem. Biol., 2020, 27(7), 877-887 https://doi.org/10.1016/j.chembiol.2020.06.009AbstractPoly(ADP-ribose) polymerase (PARP) enzymes use nicotinamide adenine dinucleotide (NAD+) to modify up to seven different amino acids with a single mono(ADP-ribose) unit (MARylation deposited by PARP monoenzymes) or branched poly(ADP-ribose) polymers (PARylation deposited by PARP polyenzymes). To enable the development of tool compounds for PARP monoenzymes and polyenzymes, we have developed active site probes for use in in vitro and cellular biophysical assays to characterize active site-directed inhibitors that compete for NAD+ binding. These assays are agnostic of the protein substrate for each PARP, overcoming a general lack of knowledge around the substrates for these enzymes. The in vitro assays use less enzyme than previously described activity assays, enabling discrimination of inhibitor potencies in the single-digit nanomolar range, and the cell-based assays can differentiate compounds with sub-nanomolar potencies and measure inhibitor residence time in live cells.
Kevin McCloskey; Eric A. Sigel; Steven Kearnes; Ling Xue; Xia Tian; Dennis Moccia; Diana Gikunju; Sana Bazzaz; Betty Chan; Matthew A. Clark; John W. Cuozzo; Marie-Aude Guié; John P. Guilinger; Christelle Huguet; Christopher D. Hupp; Anthony D. Keefe; Christopher J. Mulhern; Ying Zhang; Patrick Riley J. Med. Chem., 2020, 63(16), 8857-8866 https://doi.org/10.1021/acs.jmedchem.0c00452AbstractDNA-encoded small molecule libraries (DELs) have enabled discovery of novel inhibitors for many distinct protein targets of therapeutic value through screening of libraries with up to billions of unique small molecules. We demonstrate a new approach applying machine learning to DEL selection data by identifying active molecules from a large commercial collection and a virtual library of easily synthesizable compounds. We train models using only DEL selection data and apply automated or automatable filters with chemist review restricted to the removal of molecules with potential for instability or reactivity. We validate this approach with a large prospective study (nearly 2000 compounds tested) across three diverse protein targets: sEH (a hydrolase), ER{\alpha} (a nuclear receptor), and c-KIT (a kinase). The approach is effective, with an overall hit rate of {\sim}30% at 30 {\textmu}M and discovery of potent compounds (IC50 <10 nM) for every target. The model makes useful predictions even for molecules dissimilar to the original DEL and the compounds identified are diverse, predominantly drug-like, and different from known ligands. Collectively, the quality and quantity of DEL selection data; the power of modern machine learning methods; and access to large, inexpensive, commercially-available libraries creates a powerful new approach for hit finding.
William H. Press; John A. Hawkins; Stephen K. Jones; Jeffrey M. Schaub; Ilya J. Finkelstein Proc Natl Acad Sci USA. 2020, 117(31), 18489-18496 https://doi.org/10.1073/pnas.2004821117AbstractSynthetic DNA is rapidly emerging as a durable, high-density information storage platform. A major challenge for DNA-based information encoding strategies is the high rate of errors that arise during DNA synthesis and sequencing. Here, we describe the HEDGES (Hash Encoded, Decoded by Greedy Exhaustive Search) error-correcting code that repairs all three basic types of DNA errors: insertions, deletions, and substitutions. HEDGES also converts unresolved or compound errors into substitutions, restoring synchronization for correction via a standard Reed–Solomon outer code that is interleaved across strands. Moreover, HEDGES can incorporate a broad class of user-defined sequence constraints, such as avoiding excess repeats, or too high or too low windowed guanine–cytosine (GC) content. We test our code both via in silico simulations and with synthesized DNA. From its measured performance, we develop a statistical model applicable to much larger datasets. Predicted performance indicates the possibility of error-free recovery of petabyte- and exabyte-scale data from DNA degraded with as much as 10% errors. As the cost of DNA synthesis and sequencing continues to drop, we anticipate that HEDGES will find applications in large-scale error-free information encoding.
Kenzo Fujimoto; Kenta Ishida; Li Xue; Shigetaka Nakamura Photochem. Photobiol. Sci., 2020,19, 776-782 https://doi.org/10.1039/D0PP00098AAbstractUltrafast reversible DNA/RNA photo-cross-linking is a powerful tool for regulating the target strand in living cells. In particular, 3-cyanovinylcarbazole (CNVK) and 3-cyanovinylcarbazole modified by D-threoninol (CNVD) can photo-cross-link to pyrimidine bases within a few seconds of photoirradiation. However, these photo-cross-linkers can only cross-link to the counter base if it is adjacent to the 5′-side (−1 position). In this study, we synthesized novel photo-cross-linkers with varying linker lengths capable of photo-cross-linking with pyrimidine bases at locations other than the −1 position via click chemistry. The photo-cross-linking site was dependent on linker length.
Edward B. Miller; Robert B. Murphy; Daniel Sindhikara; Kenneth W. Borrelli; Matthew J. Grisewood; Fabio Ranalli; Steven L. Dixon; Steven Jerome; Nicholas A. Boyles; Tyler Day; Phani Ghanakota; Sayan Mondal; Salma B. Rafi; Dawn M. Troast; Robert Abel; Richard A. Friesner ChemRxiv, 2020,03 https://chemrxiv.org/articles/A_Reliable_and_Accurate_Solution_to_the_Induced_Fit_Docking_Problem_for_Protein-Ligand_Binding/11983845/1?file=22003809AbstractWe present a reliable and accurate solution to the induced fit docking problem for protein-ligand binding by combining ligand-based pharmacophore docking (Phase), rigid receptor docking (Glide), and protein structure prediction (Prime) with explicit solvent molecular dynamics simulations. We provide an in-depth description of our novel methodology and present results for 41 targets consisting of 415 cross-docking cases divided amongst a training and test set. For both the training and test-set, we compute binding modes with a ligand-heavy atom RMSD to within 2.5 Å or better in over 90% of cross-docking cases compared to less than 70% of cross-docking cases using our previously published induced-fit docking algorithm and less than 41% using rigid receptor docking. Applications of the predicted ligand-receptor structure in free energy perturbation calculations is demonstrated for both public data and in active drug discovery projects, both retrospectively and prospectively.
Jeishla Melendez Matos; Samantha Green; Yuge Chun; Vuong Q. Dang; Russell G. Dushin; Paul Richardson; Jason Chen; David W. Piotrowski; Brian Paegel; Ryan Shenvi ChemRxiv, 2020, 03, 13 https://chemrxiv.org/articles/Cycloisomerization_of_Olefins_in_Water/11977917AbstractPreparative chemical reactions that occur efficiently under dilute, buffered, aqueous conditions in the presence of biomolecules find application in ligation, peptide synthesis, polynucleotide synthesis and sequencing. However, the identification of functional groups or reagents that are mutually reactive with one another, but unreactive with biopolymers and water, is challenging. Here we show that cobalt catalysts will react with the alkenes of unsaturated tertiary amines under dilute, aqueous, buffered conditions and promote efficient cycloisomerization, in many cases mediating a formal Friedel-Crafts reaction. We find the constraining conditions of biorthogonal chemistry to be beneficial for reaction efficiency as we obtain superior conversion at low catalyst concentration and maintain competent rates in dilute conditions. The efficiency at high dilution in the presence of buffer and nucleobases suggests that these conditions may find use on or in the presence of biomolecules.
Qiuxia Chen; Justin Hall; Timothy L. Foley; Jinqiao Wan; You Li; David I. Israel Biochem. Biophys. Res. Commun., 2020, 533, 2, 249-255 https://doi.org/10.1016/j.bbrc.2020.04.029AbstractDEL selections are binding assays conducted with mixtures of chemically diverse DNA-tagged ligands and a screening target. DEL selections use DNA sequence counts to measure target binding, where ideally higher affinity ligands will have higher counts than weaker affinity ligands. However, there is not always a clear relationship between DNA sequence count (assay signal) and binding affinity. This disconnect may be due to the fidelity of library chemistry, where reactions often do not go to completion, and also to repetitive rounds of binding and elution that are standard practice in most DEL selection experiments. We describe here a strategy that addresses both of these issues and provides a means to calculate ligand affinity from primary selection data. The reaction yields of selected compounds during DEL library synthesis can also be predicted with this method.
Gregory D. Cuny; Alexei Degterev Semin. Cell Dev. Biol., 2020, S1084 https://doi.org/10.1016/j.semcdb.2020.06.014AbstractReceptor Interacting Protein Kinases (RIPKs) are a family of Ser/Thr/Tyr kinases whose functions, regulation and pathophysiologic roles have remained an enigma for a long time. In recent years, these proteins garnered significant interest due to their roles in regulating a variety of host defense functions including control of inflammatory gene expression, different forms of cell death, and cutaneous and intestinal barrier functions. In addition, there is accumulating evidence that while these kinases seemingly follow typical kinase blueprints, their functioning in cells can take forms that are atypical for protein kinases. Lastly, while these kinases generally belong to distinct areas of innate immune regulation, there are emerging overarching themes that may unify the functions of this kinase family. Our review seeks to discuss the biology of RIPKs, and how typical and atypical features of this family informs the activity of a rapidly growing repertoire of RIPK inhibitors.
Hongtao Xu; Yuang Gu; Shuning Zhang; Huan Xiong; Fei Ma; Fengping Lu; Qun Ji; Lili Liu; Peixiang Ma; Wei Hou; Guang Yang; Richard Alan Lerner Angew. Chem. Int. Ed. Engl., 2020, 59(32), 13273-13280 https://doi.org/10.1002/anie.202003595AbstractConventional direct C‐H selenylation suffers from simple selenation with limited atom economy and complicated reaction system. In this work, we designed benzoselenazolone as a novel bifunctional selenide reagent for both off‐ and on‐DNA C‐H selenylation under rhodium(III) catalysis. We showed that using benzoselenazolone allowed production of a series of selenylation products containing an adjacent aminoacyl group in a fast and efficient way, with high atom economy. The synthetic application of this method was demonstrated by taking advantage of the amide functionality as a nucleophile, directing group, and amide coupling partner. This work shows great potential in facilitating rapid construction of selenium‐containing DNA‐encoded chemical libraries (SeDELs), and lays the foundation for the development of selenium‐containing drugs.
Anna Fryszkowska; Paul N. Devine Curr. Opin. Chem. Biol., 2020, 55, 151-160 https://doi.org/10.1016/j.cbpa.2020.01.012AbstractEnzyme catalysis, enabled by advances in protein engineering and directed evolution, is beginning to transform chemical synthesis in the pharmaceutical industry. This review presents recent examples of the creative use of biocatalysis to enable drug discovery and development. We illustrate how increased access to novel biotransformations and the rise of cascade biocatalysis allowed fundamentally new syntheses of novel medicines, representing progress toward more sustainable pharmaceutical manufacturing. Finally, we describe the opportunities and challenges the industry must address to ensure the reduction to practice of biotechnological innovations to develop new therapies in a faster, more economical, and environmentally benign way.
Ada J. Kwong; Karl A. Scheidt Bioorg. Med. Chem. Lett., 2020, 127203 https://doi.org/10.1016/j.bmcl.2020.127203AbstractThe MAPK pathways are an enduring area of interest due to their essential roles in cell processes. Increased expression and activity can lead to a multitude of diseases, sparking research efforts in developing inhibitors against these kinases. Though great strides have been made in developing MEK1/2 inhibitors, there is a notable lack of chemical probes for MEK3-7, given their central role in stimuli response, cell growth, and development. This review summarizes the progress that has been made on developing small molecule probes for MEK3-7, the specific disease states in which they have been studied, and their potential to become novel therapeutics.
Arun Shivalingam; Lapatrada Taemaitree; Afaf H. El-Sagheer; Tom Brown Angew. Chem. Int. Ed. Engl., 2020, 59(28), 11416-11422 https://doi.org/10.1002/anie.202000209AbstractJoining oligonucleotides together (ligation) is a powerful means of retrieving information at the nanoscale. In order to recover this information, the linkages created must be compatible with polymerase enzymes. However, enzymatic ligation is restrictive and current chemical ligation methods lack flexibility. Here, a versatile ligation platform based on the formation of urea and squaramide artificial backbones from minimally modified 3'‐ and 5'‐amino oligonucleotides is described. One‐pot ligation gives a urea linkage with excellent read‐through speed, or a squaramide linkage that is read‐through under selective conditions. The squaramide linkage can be broken and reformed on demand, while stable pre‐activated precursor oligonucleotides expand the scope of the ligation reaction to reagent‐free, mild conditions. The utility of our system is demonstrated by replacing the enzymatically biased RNA‐to‐DNA reverse transcription step of RT‐qPCR with a rapid nucleic acid template‐dependent DNA chemical ligation system, an approach that allows direct RNA detection.
Pingdong Tao; Yuanyuan Kuang; Yu Li; Wenping Li; Zibei Gao; Lili Liu; Min Qiang; Zhao Zha; Kun Fan; Peixiang Ma; Jeffrey M. Friedman; Guang Yang; Richard A. Lerner Adv. Sci., 2020, 2000818 https://doi.org/10.1002/advs.202000818AbstractGrowth factor deficiency in adulthood constitutes a distinct clinical syndrome with significant morbidities including abnormal body composition, reduced energy, affective disturbances, dyslipidemia, and increased cardiovascular risk. Protein replacement therapies using recombinant proteins or enzymes represent the only approved treatment. Combinatorial antibodies have shown great promise as a new class of therapeutic molecules because they act as “mechanism‐based antibodies” with both agonist and antagonist activities. Using leptin, a key hormone in energy metabolism, as an example, a function‐guided approach is developed to select combinatorial antibodies with high potency and full agonist activity that substitute natural growth factors in vivo. The identified antibody shows identical biochemical properties and cellular profiles as leptin, and rescues leptin‐deficiency in ob/ob mice. Remarkably, the antibody activates leptin receptors that are otherwise nonfunctional because of mutations (L372A and A409E). Combinatorial antibodies have significant advantages over recombinant proteins for chronical usage in terms of immunological tolerance and biological stability.
Koki Shinbara; Wenyu Liu; Renier Herman Pieter van Neer; Takayuki Katoh; Hiroaki Suga Front. Chem.,2020, 8, 447 https://doi.org/10.3389/fchem.2020.00447AbstractBackbone macrocyclic structures are often found in diverse bioactive peptides and contribute to greater conformational rigidity, peptidase resistance, and potential membrane permeability compared to their linear counterparts. Therefore, such peptide scaffolds are an attractive platform for drug-discovery endeavors. Recent advances in synthetic methods for backbone macrocyclic peptides have enabled the discovery of novel peptide drug candidates against diverse targets. Here, we overview recent technical advancements in the synthetic methods including 1) enzymatic synthesis, 2) chemical synthesis, 3) split-intein circular ligation of peptides and proteins (SICLOPPS), and 4) in vitro translation system combined with genetic code reprogramming. We also discuss screening methodologies compatible with those synthetic methodologies, such as one-beads one-compound (OBOC) screening compatible with the synthetic method 2, cell-based assay compatible with 3, limiting-dilution PCR and mRNA display compatible with 4.
Huan Xiong; Yuang Gu; Shuning Zhang; Fengping Lu; Qun Ji; Lili Liu; Peixiang Ma; Guang Yang; Wei Hou; Hongtao Xu Chem. Commun., 2020, 56(34), 4692-4695 https://doi.org/10.1039/D0CC01647KAbstractAn efficient, selective and scalable C-H amidation of s-tetrazines under iridium(III) catalysis is reported. This reaction features broad substrate scope, high functional group tolerance and air and water tolerance. This reaction also shows great potential in the rapid preparation of tri- and tetra-functional building blocks, which can be applied either in bioconjugation or DNA-encoded library.
Jie Li; Yu Li; Fengping Lu; Lili Liu; Qun Ji; Ke Song; Qianqian Yin; Richard A. Lerner; Guang Yang; Hongtao Xu; Peixiang Ma Biochem. Biophys. Res. Commun., 2020, 533, 2, 241-248 https://doi.org/10.1016/j.bbrc.2020.04.022AbstractNatural products have been an invaluable source of drug discovery, but their targets remain largely unknown. Natural products enriched DNA-encoded chemical libraries (nDELs) empower the researchers to rapidly and economically screen numerous natural products against various protein targets, and therefore promote the elucidation of the molecular mechanisms. In this work, we used poly (ADP-ribose) polymerase 1 (PARP1), as an example to explore the usage of nDEL for the functional natural products selection. We used late-stage modification approach to label three positive binders with unique DNA barcodes, whose dissociation constants range from sub-micromolar to micromolar. The selection criterion was set up according to the enrichment of these controls. Five natural products selected by this criterion directly bind to PARP1 in SPR, among which luteolin exhibits the highest inhibitory activity against PARP1. Moreover, luteolin selectively induces accumulation of DNA double-strand breaks and G2/M phase arrest in BRCA-deficient cells. All the findings from these investigations on luteolin support that PARP1 inhibition is one of the mechanisms for its anti-cancer activity.
Tomas Brom; Francesco V. Reddavide; Stephan Heiden; Michael Thompson; Yixin Zhang Biochem. Biophys. Res. Commun., 2020, 533, 2, 230-234 https://doi.org/10.1016/j.bbrc.2020.04.025AbstractDNA-encoded chemical libraries (DECLs) are powerful tools for modern drug discovery. A DECL is a pooled mixture of small molecule compounds, each of which is tagged with a unique DNA sequence which functions as a barcode. After incubation with a drug target and washing to remove non-binders, the bound molecules are eluted and submitted for DNA sequencing to determine which molecules are binding the target. While the DECL technology itself is ultra-high throughput, the following re-synthesis of identified compounds for orthogonal validation experiments remains the bottleneck. Using existing DNA-small molecule conjugates directly for affinity measurements, as opposed to complete compound resynthesis, could accelerate the discovery process. To this end, we have tested various geometries of fluorescently-labelled DNA constructs for fluorescence anisotropy (FA) experiments. Minimizing the distance between the fluorescent moiety and ligand can maximize the correlation between ligand-protein interaction and corresponding change in fluorophore rotational freedom, thus leading to larger, easier to interpret changes in FA values. However, close proximity can also cause artifacts due to potentially promiscuous interactions between fluorophore and protein. By balancing these two opposite effects, we have identified applicable fluorescently labelled DNA constructs displaying either a single ligand or pairs of fragments for affinity measurement using a FA assay.
Nicholas G. Paciaroni; John Maina Ndungu; Thomas Kodadek Chem. Commun., 2020, 56(34), 4656-4659 https://doi.org/10.1039/D0CC01474EAbstractWe report chemistry suitable for the solid-phase synthesis of DNA-encoded libraries with an unusually high level of structural diversity. The strategy involves “exploding” an immobilized aldehyde into a plethora of different functional groups under DNA-compatible conditions.
Katharina Götte; Silvia Chines; Andreas Brunschweiger Tetrahedron Lett., 2020, 151889 https://doi.org/10.1016/j.tetlet.2020.151889AbstractDNA-encoded libraries of organic, synthesized molecules are an emerging technology for target-based compound screening. The design of first generation encoded libraries placed emphasis on ever increasing compound numbers that were synthesized from large sets of starting materials with a few well-established synthesis methods. In the last two years there has literally been an explosion of published research activities that have expanded the toolbox of reactions for designing DNA-encoded libraries. In this digest, we highlight latest developments that include modern photoredox chemistries as well as strategies to perform encoded compound synthesis in organic solvents or exploit nano-heterogeneous systems. The new chemistry developments carry the potential to revolutionize the technology as they enable access to unprecedented molecular diversity.
Karishma Patel; Louise J. Walport; James L. Walshe; Paul D. Solomon; Jason K. K. Low; Daniel H. Tran; Kevork S. Mouradian; Ana P. G. Silva; Lorna Wilkinson-White; Alexander Norman; Charlotte Franck; Jacqueline M. Matthews; J. Mitchell Guss; Richard J. Payne; Toby Passioura; Hiroaki Suga; Joel P. Mackay Proc. Natl. Acad. Sci. USA, 2020, 117(43), 26728-26738 https://doi.org/10.1073/pnas.2003086117AbstractLarge DNA-encoded libraries of cyclic peptides are emerging as powerful sources of molecules to tackle challenging drug targets. The structural and functional diversity contained within these libraries is, however, little explored. Here we demonstrate that one such library contains members that use unexpectedly diverse mechanisms to recognize the same surface on the same target proteins with high affinity and specificity. This range of binding modes is much larger than observed in natural ligands of the same proteins, demonstrating the power and versatility of the technology. Our data also reveal opportunities for the development of more sophisticated approaches to achieving specificity when trying to selectively target one member of a family of closely related proteins. Cyclic peptide library screening technologies show immense promise for identifying drug leads and chemical probes for challenging targets. However, the structural and functional diversity encoded within such libraries is largely undefined. We have systematically profiled the affinity, selectivity, and structural features of library-derived cyclic peptides selected to recognize three closely related targets: the acetyllysine-binding bromodomain proteins BRD2, -3, and -4. We report affinities as low as 100 pM and specificities of up to 106-fold. Crystal structures of 13 peptide–bromodomain complexes reveal remarkable diversity in both structure and binding mode, including both α-helical and β-sheet structures as well as bivalent binding modes. The peptides can also exhibit a high degree of structural preorganization. Our data demonstrate the enormous potential within these libraries to provide diverse binding modes against a single target, which underpins their capacity to yield highly potent and selective ligands.
Christopher G. Parker; Matthew R. Pratt Cell, 2020, 180(4), 605-632 https://doi.org/10.1016/j.cell.2020.01.025AbstractDespite advances in genetic and proteomic techniques, a complete portrait of the proteome and its complement of dynamic interactions and modifications remains a lofty, and as of yet, unrealized, objective. Specifically, traditional biological and analytical approaches have not been able to address key questions relating to the interactions of proteins with small molecules, including drugs, drug candidates, metabolites, or protein post-translational modifications (PTMs). Fortunately, chemists have bridged this experimental gap through the creation of bioorthogonal reactions. These reactions allow for the incorporation of chemical groups with highly selective reactivity into small molecules or protein modifications without perturbing their biological function, enabling the selective installation of an analysis tag for downstream investigations. The introduction of chemical strategies to parse and enrich subsets of the “functional” proteome has empowered mass spectrometry (MS)-based methods to delve more deeply and precisely into the biochemical state of cells and its perturbations by small molecules. In this Primer, we discuss how one of the most versatile bioorthogonal reactions, “click chemistry”, has been exploited to overcome limitations of biological approaches to enable the selective marking and functional investigation of critical protein-small-molecule interactions and PTMs in native biological environments.
Dominik K. Kölmel; Anokha S. Ratnayake; Mark E. Flanagan; Mei-Hsuan Tsai; Cong Duan; Chao Song Org. Lett., 2020, 22, 8, 2908-2913 https://doi.org/10.1021/acs.orglett.0c00574AbstractThe on-DNA synthesis of highly substituted cyclobutanes was achieved through a photocatalytic [2 + 2] cycloaddition reaction in aqueous solution. Readily available DNA-tagged styrene derivatives were reacted with structurally diverse cinnamates in the presence of an iridium-based photocatalyst, Ir(ppy)2(dtbbpy)PF6, to forge two new C(sp3)–C(sp3) bonds. This transformation was demonstrated to have excellent functional group tolerance and allowed for the facile installation of a variety of heteroaromatic substituents on a densely functionalized cyclobutane scaffold.
Yu Zhou; Jianzhao Peng; Wenyin Shen; Xiaoyu Li Biochem. Biophys. Res. Commun., 2020, 533, 2, 215-222 https://doi.org/10.1016/j.bbrc.2020.04.033AbstractDNA-encoded chemical library (DEL) has emerged as a powerful technology for ligand discovery in biomedical research. Recently, we have developed a DNA-encoded dynamic library (DEDL) approach by incorporating the concept of dynamic combinatorial library (DCL) with DELs. DEDL has shown excellent potential in ligand discovery towards a variety of protein targets. However, the requirement of having a pair of unnatural p-stilbazoles as the interstrand DNA crosslinker has limited the chemical diversity of DEDLs. Here, we replaced p-stilbazole with psoralen (PS) and tested the feasibility of psoralen as the crosslinker in DEDL selection. Since psoralen is commercially available and does not require any special crosslinking partner, existing DELs may be directly used to create high-diversity DEDLs. This study is expected to greatly facilitate the development of DEDLs as a versatile tool in drug discovery.
Alessandro Sannino; Adrián Gironda-Martínez; Emile M. D. Gorre; Luca Prati; Jacopo Piazzi; Joerg Scheuermann; Dario Neri; Etienne J. Donckele; Florent Samain ACS Comb. Sci., 2020, 22(4), 204-212 https://doi.org/10.1021/acscombsci.0c00023AbstractThe growing importance of DNA-encoded chemical libraries (DECLs) as tools for the discovery of protein binders has sparked an interest for the development of efficient screening methodologies, capable of discriminating between high- and medium-affinity ligands. Here, we present a systematic investigation of selection methodologies, featuring a library dis-played on single-stranded DNA, which could be hybridized to a complementary oligonucleotide carrying a diazirine photore-active group. Model experiments, performed using ligands of different affinity to carbonic anhydrase IX, revealed a recovery of preferential binders up to 10%, which was mainly limited by the highly-reactive nature of carbene intermediates generated during the photocrosslinking process. Ligands featuring acetazolamide or p-phenylsulfonamide exhibited a higher recovery compared to their counterparts based on 3-sulfamoyl benzoic acid, which had a lower affinity towards the target. A systemat-ic evaluation of experimental parameters revealed conditions that were ideally suited for library screening, which were used for the screening of a combinatorial DECL library, featuring 669240 combinations of two sets of building blocks. Compared to conventional affinity capture procedures on protein immobilized on solid supports, photocrosslinking provided a better discrimination of low-affinity CAIX ligands over the background signal and therefore can be used as a tandem methodology with the affinity capture procedures.
Xunan Zheng; Zhengning Ma; Dawei Zhang Pharmaceuticals, 2020, 13(3), 37 https://doi.org/10.3390/ph13030037AbstractImidazole and its derivatives are one of the most vital and universal heterocycles in medicinal chemistry. Owing to their special structural features, these compounds exhibit a widespread spectrum of significant pharmacological or biological activities, and are widely researched and applied by pharmaceutical companies for drug discovery. The van Leusen reaction based on tosylmethylisocyanides (TosMICs) is one of the most appropriate strategies to synthetize imidazole-based medicinal molecules, which has been increasingly developed on account of its advantages. In this review, we summarize the recent developments of the chemical synthesis and bioactivity of imidazole-containing medicinal small molecules, utilizing the van Leusen imidazole synthesis from 1977.
Jiayan Wang; Setayesh Yazdani; Ana Han; Matthieu Schapira bioRxiv, 2020 http://dx.doi.org/10.1101/2020.02.06.937730AbstractAlmost twenty years after the human genome was sequenced, the wealth of data produced by the international human genome project has not translated into a significantly improved drug discovery enterprise. This is in part because small molecule modulators that could be used to explore the cellular function of their target proteins and to discover new therapeutic opportunities are only available for a limited portion of the human proteome. International efforts are underway to develop such chemical tools for a few, specific protein families, and a Target 2035 call to enable, expand and federate these efforts towards a comprehensive chemical coverage of the druggable genome was recently announced. But what is the druggable genome? Here, we systematically review structures of human proteins bound to drug-like ligands available from the protein databank (PDB) and use ligand desolvation upon binding as a druggability metric to draw a landscape of the human druggable genome. We show that the vast majority of druggable protein families, including some highly populated and deeply associated with cancer according to genomic screens, are almost orphan of small molecule ligands, and propose a list of 46 druggable domains representing 3440 human proteins that could be the focus of large chemical probe discovery efforts.
Ke Li; Xiaohong Liu; Sixiu Liu; Yulong An; Yanfang Shen; Qingxia Sun; Xiaodong Shi; Wenji Su; Weiren Cui; Zhiqiang Duan; Letian Kuai; Hongfang Yang; Alexander L. Satz; Kaixian Chen; Hualiang Jiang; Mingyue Zheng; Xuanjia Peng; Xiaojie Lu iScience, 2020, 23(6), 101142 https://doi.org/10.1016/j.isci.2020.101142AbstractThe application of machine learning towards DNA encoded library (DEL) technology is lacking despite obvious synergy between these two advancing technologies. Herein, a machine learning algorithm has been developed that predicts the conversion rate for the DNA compatible reaction of a building block with a model DNA-conjugate. We exemplify the value of this technique with a challenging reaction, the Pictet-Spengler, where acidic conditions are normally required to achieve the desired cyclization between tryptophan and aldehydes to provide tryptolines. To avoid damaging the DNA our reaction conditions must be exceptionally mild, and therefore most building blocks fail to provide acceptable yields of desired product (<20% pass rate) in a test reaction employing our optimized protocol. In contrast, building blocks selected by our trained machine learning algorithm have a >78% pass rate. This is the first demonstration of using a machine learning algorithm to cull potential building blocks prior to their purchase and testing for DNA encoded library synthesis. Importantly, this allows for a challenging reaction, with an otherwise very low building block pass rate in the test reaction, to still be used in DEL synthesis. Furthermore, we discuss herein the rational design of DNA conjugated tryptophan substrates for our Pictet-Spengler reaction, and optimization of the reaction protocols. Lastly, because our protocol is solution-phase it is directly applicable to standard plate-based DEL synthesis.
Kenneth J. DiRico; Wenyi Hua; Chang Liu; Joseph W. Tucker; Anokha S. Ratnayake; Mark E. Flanagan; Matthew D. Troutman; Mark C. Noe; Hui Zhang ACS Med. Chem. Lett., 2020, 11, 6, 1101-1110 https://doi.org/10.1021/acsmedchemlett.0c00066AbstractHigh-throughput experimentation (HTE) has emerged as an important tool in drug discovery, providing a platform for preparing large compound libraries and enabling swift reaction screening over wide-ranging conditions. Recent advances in automated high-density, material-sparing HTE have necessitated the development of rapid analytics with sensitivity and resolution sufficient to identify products and/or assess reaction performance in a timely and data-rich manner. Combination of an ultrathroughput (UT) reader platform with Acoustic Droplet Ejection-Open Port Interface-Mass Spectrometry (ADE-OPI-MS) provides the requisite speed and sensitivity. Herein, we report the application of ADE-OPI-MS to HTE in the areas of parallel medicinal chemistry and reaction screening.
Sofie Martens; Sam Hofmans; Wim Declercq; Koen Augustyns; Peter Vandenabeele Trends Pharmacol. Sci., 2020, 41(3), 209-224 https://doi.org/10.1016/j.tips.2020.01.002AbstractThe scaffolding function of receptor-interacting protein kinase 1 (RIPK1) regulates prosurvival signaling and inflammatory gene expression, while its kinase activity mediates both apoptosis and necroptosis; the latter involving RIPK3 kinase activity. The mutual transition between the scaffold and kinase functions of RIPK1 is regulated by (de)ubiquitylation and (de)phosphorylation. RIPK1-mediated cell death leads to disruption of epithelial barriers and/or release of damage-associated molecular patterns (DAMPs), cytokines, and chemokines, propagating inflammatory and degenerative diseases. Many drug development programs have pursued targeting RIPK1, and to a lesser extent RIPK3 kinase activity. In this review, we classify existing and novel small-molecule drugs based on their pharmacodynamic (PD) type I, II, and III binding mode. Finally, we discuss their applicability and therapeutic potential in inflammatory and degenerative experimental disease models.
Animesh Roy; Thomas Kodadek Trends Pharmacol. Sci., 2020, 41(4), 225-227 https://doi.org/10.1016/j.tips.2020.01.007AbstractDNA-encoded libraries (DELs) comprise large numbers of small molecules, each of which is conjugated to an encoding DNA. Krusemark and colleagues recently described a method to introduce DELs into living cells and recover conjugates that bind to an intracellular target. This proof-of-principle study suggests that it may be feasible to screen DELs against protein targets in their native environment.
Yi Qu; Sixiu Liu; Huanan Wen; Yanfen Xu; Yulong An; Ke Li; Mengyuan Ni; Yanfang Shen; Xiaodong Shi; Wenji Su; Weiren Cui; Letian Kuai; Alexander L. Satz; Hongfang Yang; Xiaojie Lu; Xuanjia Peng Biochem. Biophys. Res. Commun., 2020, 533, 2, 209-214 https://doi.org/10.1016/j.bbrc.2020.04.027AbstractA mild reaction for DNA-compatible, palladium promoted Suzuki-Miyaura cross-coupling reaction of potassium Boc-protected aminomethyltrifluoroborate with DNA-conjugated aryl bromides has been developed efficiently. This novel DNA encoded chemistry reaction proceeded well with a wide range of functional group tolerance, including aryl bromides and heteroaryl bromides. Further, the utility our DNA conjugated aminomethylated arene products is demonstrated by reaction with various types of reagents (including amide formation with carboxylic acids, alkylation with aldehydes, and carbamoylation with amines) as would be desired for the production of a DNA encoded library.
Edward J. Griffen; Alexander G. Dossetter; Andrew G. Leach J. Med. Chem., 2020, 63(16), 8695-8704 https://doi.org/10.1021/acs.jmedchem.0c00163AbstractThe latest developments in artificial intelligence (AI) have arrived into an existing state of creative tension between computational and medicinal chemists. At their most productive, medicinal and computational chemists have made significant progress in delivering new therapeutic agents into the clinic. However, the relationship between these communities has the prospect of being weakened by application of oversimplistic AI methods that, if they fail to deliver, will reinforce unproductive prejudices. We review what can be learned from our history of integrating QSAR and structure-based methods into drug discovery. Now with synthesis and testing available as contract services, the environment for computational innovation has changed and we consider the impact this may have on the relationships in our disciplines. We discuss the current state of interdisciplinary communication and suggest approaches to bring the subdisciplines together in order to improve computational medicinal chemistry and, most importantly, deliver better medicines to the clinic faster.
Takuya Terai; Tomoyuki Koike; Naoto Nemoto Molecules, 2020, 25(6), 1472 https://doi.org/10.3390/molecules25061472AbstractBinding peptides for given target molecules are often selected in vitro during drug discovery and chemical biology research. Among several display technologies for this purpose, complementary DNA (cDNA) display (a covalent complex of a peptide and its encoding cDNA linked via a specially designed puromycin-conjugated DNA) is unique in terms of library size, chemical stability, and flexibility of modification. However, selection of cDNA display libraries often suffers from false positives derived from non-specific binding. Although rigorous washing is a straightforward solution, this also leads to the loss of specific binders with moderate affinity because the interaction is non-covalent. To address this issue, herein, we propose a method to covalently link cDNA display molecules with their target proteins using light irradiation. We designed a new puromycin DNA linker that contains a photocrosslinking nucleic acid and prepared cDNA display molecules using the linker. Target proteins were also labeled with a short single-stranded DNA that should transiently hybridize with the linker. Upon ultraviolet (UV) light irradiation, cDNA display molecules encoding correct peptide aptamers made stable crosslinked products with the target proteins in solution, while display molecules encoding control peptides did not. Although further optimization and improvement is necessary, the results pave the way for efficient selection of peptide aptamers in multimolecular crowding biosystems.
Peter Canning; Kristian Birchall; Catherine A. Kettleborough; Andy Merritt; Peter J. Coombs Drug Discov. Today, 2020, 25(11), 2030-2037 https://doi.org/10.1016/j.drudis.2020.09.003AbstractHere, we describe a novel workflow combining informatic and experimental approaches to enable evidence-based prioritising of targets from large sets in parallel. High-throughput protein production and biophysical fragment screening is used to identify those targets that are tractable and ligandable. As proof of concept we have applied this to a set of antibacterial targets comprising 146 essential genes. Of these targets, 51 were selected and 38 delivered results that allowed us to rank them by ligandability. The data obtained against these derisked targets have enabled rapid progression into structurally enabled drug discovery projects, demonstrating the practical value of the fragment-based target screening workflow.
Dirk Trauner; Andrej Shemet Synfacts, 2020, 16, 2, 0221 https://doi.org/10.1055/s-0039-1691634AbstractThe researchers reported a robust, mild, and efficient protocol for the diazo-transfer reaction that works under aqueous conditions in the presence of DNA. This methodology proved to be suitable for the synthesis of DNA-encoded libraries.
Pei-Pei Kung; Patrick Bingham; Benjamin J. Burke; Qiuxia Chen; Xuemin Cheng; Ya-Li Deng; Dengfeng Dou; Junli Feng; Gary M. Gallego; Michael R. Gehring; Stephan K. Grant; Samantha Greasley; Anthony R. Harris; Karen A. Maegley; Jordan Meier; Xiaoyun Meng; Jose L. Montano; Barry A. Morgan; Brigitte S. Naughton; Prakash B. Palde; Thomas A. Paul; Paul Richardson; Sylvie Sakata; Alex Shaginian; William K. Sonnenburg; Chakrapani Subramanyam; Sergei Timofeevski; Jinqiao Wan; Wen Yan; Albert E. Stewart ACS Med. Chem. Lett., 2020, 11(6), 1175-1184 https://doi.org/10.1021/acsmedchemlett.0c00029AbstractTwo novel compounds were identified as Naa50 binders/inhibitors using DNA-encoded technology screening. Biophysical and biochemical data as well as co-crystal structures were obtained for both compounds (3a and 4a) to understand their mechanism of action. These data were also used to rationalize the binding affinity differences observed between the two compounds and a MLGP peptide-containing substrate. Cellular target engagement experiments further confirm the Naa50 binding of 4a and demonstrate its selectivity toward related enzymes (Naa10 and Naa60). Additional analogs of inhibitor 4a were also evaluated to study the binding mode observed in the co-crystal structures.
Laurens Lindenburg; Tuomas Huovinen; Kayleigh van de Wiel; Michael Herger; Michael R Snaith; Florian Hollfelder Nucleic Acids Reaserch, 2020, 48(11), e63 https://doi.org/10.1093/nar/gkaa270AbstractSite-saturation libraries reduce protein screening effort in directed evolution campaigns by focusing on a limited number of rationally chosen residues. However, uneven library synthesis efficiency leads to amino acid bias, remedied at high cost by expensive custom synthesis of oligonucleotides, or through use of proprietary library synthesis platforms. To address these shortcomings, we have devised a method where DNA libraries are constructed on the surface of microbeads by ligating dsDNA fragments onto growing, surface-immobilised DNA, in iterative split-and-mix cycles. This method—termed SpliMLiB for Split-and-Mix Library on Beads—was applied towards the directed evolution of an anti-IgE Affibody (ZIgE), generating a 160,000-membered, 4-site, saturation library on the surface of 8 million monoclonal beads. Deep sequencing confirmed excellent library balance (5.1% ± 0.77 per amino acid) and coverage (99.3%). As SpliMLiB beads are monoclonal, they were amenable to direct functional screening in water-in-oil emulsion droplets with cell-free expression. A FACS-based sorting of the library beads allowed recovery of hits improved in Kd over wild-type ZIgE by up to 3.5-fold, while a consensus mutant of the best hits provided a 10-fold improvement. With SpliMLiB, directed evolution workflows are accelerated by integrating high-quality DNA library generation with an ultra-high throughput protein screening platform.
Dillon T. Flood; Cian Kingston; Julien C. Vantourout; Philip E. Dawson; Phil S. Baran Isr. J. Chem., 2020, 60(3-4), 268-280 https://doi.org/10.1002/ijch.201900133AbstractIn 1992, Brenner and Lerner hypothesized that individual chemical transformations could be encoded in DNA, allowing the rapid synthesis and screening of large collections of small molecules. Since their report, huge investments into the development of the DNA encoded library (DEL) technology have enabled the acceleration of the drug discovery process especially early phase discovery undertakings such as target validation and hit identification. As DEL lies at the nexus between chemistry and biology, there is an increasing need to expand the toolboxes of both organic transformations and biological methods. However, the myriad of techniques and reactions already reported can be difficult to digest for practitioners whose expertise resides outside the realm of DEL. This review therefore focuses on a stepwise presentation of DEL from the basic concepts to newest developments. The presentation includes the history, fundamentals, and successes of DEL, different methods for DEL synthesis and affinity selection, the conventional transformations, and finally the latest developments from a synthetic organic perspective.
Marco Potowski; Robin Esken; Andreas Brunschweiger Bioorg. Med. Chem., 2020, 115441 https://doi.org/10.1016/j.bmc.2020.115441AbstractThe Petasis three-component reaction gives rise to diverse substituted α-aryl glycines from readily available amines, boronic acids and glyoxalic acid. Thus, this reaction is highly attractive for DNA-encoded small molecule screening library synthesis. The Petasis reaction is for instance promoted by a potentially DNA damaging copper(I)/bipyridine reagent system in dry organic solvents. We found that solid phase-coupled DNA strands tolerated this reagent system at elevated temperature allowing for synthesis of diverse substituted DNA-tagged α-aryl glycines from DNA-conjugated secondary amines.
Peter Komar; Marko Kalinic ACS Comb. Sci., 2020, 22(8), 410-421 https://doi.org/10.1021/acscombsci.0c00007AbstractDNA-encoded libraries (DELs) are large, pooled collections of compounds in which every library member is attached to a stretch of DNA encoding its complete synthetic history. DEL-based hit discovery involves affinity selection of the library against a protein of interest, whereby compounds retained by the target are subsequently identified by next-generation sequencing of the corresponding DNA tags. When analyzing the resulting data, one typically assumes that sequencing output (i.e. read counts) is proportional to the binding affinity of a given compound, thus enabling hit prioritization and elucidation of any underlying structure-activity relationships (SAR). This assumption, though, tends to be severely confounded by a number of factors, including variable reaction yields, presence of incomplete products masquerading as their intended counterparts, and sequencing noise. In practice, these confounders are often ignored, potentially leading to false discoveries, and universally leading to loss of valuable information. To address this issue, we have developed a method for comprehensively denoising DEL selection outputs. Our method, dubbed ``deldenoiser'', is based on sparse learning and leverages inputs that are commonly available within a DEL generation and screening workflow. Using simulated and publicly available DEL affinity selection data, we show that ``deldenoiser'' is not only able to recover and rank true binders much more robustly than read count-based approaches, but also that it yields scores which accurately capture the underlying SAR. The proposed method can, thus, be of significant utility in hit prioritization following DEL screens.
Mathieu Surin; Sébastien Ulrich ChemistryOpen, 2020, 9(4), 480-498 https://doi.org/10.1002/open.202000013AbstractDNA-templated self-assembly represents a rich and growing subset of supramolecular chemistry where functional self-assemblies are programmed in a versatile manner using nucleic acids as readily-available and readily-tunable templates. In this review, we summarize the different DNA recognition modes and the basic supramolecular interactions at play in this context. We discuss the recent results that report the DNA-templated self-assembly of small molecules into complex yet precise nanoarrays, going from 1D to 3D architectures. Finally, we show their emerging functions as photonic/electronic nanowires, sensors, gene delivery vectors, and supramolecular catalysts, and their growing applications in a wide range of area from materials to biological sciences.
Luca Prati; Martina Bigatti; Etienne J. Donckele; Dario Neri; Florent Samain Biochem. Biophys. Res. Commun., 2020, 533, 2, 235-240 https://doi.org/10.1016/j.bbrc.2020.04.030AbstractDNA-encoded chemical libraries (DECLs) are large compound collections attached to DNA fragments, serving as amplifiable barcodes, which can be screened on target proteins of pharmaceutical interest. In DECL selections, ligands are identified by high-throughput DNA sequencing, by comparing their frequency before and after the affinity capture step. Hits identified using this procedure need to be validated by resynthesis and by performing affinity measurements. Here we report novel on-DNA hit validation strategies, which enable the facile confirmation of ligand-protein interaction as well as the determination of equilibrium and kinetic binding constants. The experimental procedures, which had been inspired by enzyme-linked immunosorbent assays (ELISA), were validated using ligands of different affinity to carbonic anhydrase II and IX.
Doris Mia Taylor; Justin Anglin; Suhyeorn Park; Melek N. Ucisik; John C. Faver; Nicholas Simmons; Zhuang Jin; Murugesan Palaniappan; Pranavanand Nyshadham; Feng Li; James Campbell; Liya Hu; Banumathi Sankaran; B. V. Venkataram Prasad; Hongbing Huang; Martin M. Matzuk; Timothy G. Palzkill ACS Infect. Dis., 2020, 6(5), 1214-1227 https://doi.org/10.1021/acsinfecdis.0c00015AbstractBacterial resistance to β-lactam antibiotics is largely mediated by β-lactamases, which catalyze the hydrolysis of these drugs and continue to emerge in response to antibiotic use. β-lactamases that hydrolyze the last resort carbapenem class of β-lactam antibiotics (carbapenemases) are a growing global health threat. Inhibitors have been developed to prevent β-lactamase-mediated hydrolysis and restore the efficacy of these antibiotics. However, there are few inhibitors available for problematic carbapenemases such as Oxacillinase-48 (OXA-48). A DNA-encoded chemical library approach was used to rapidly screen for compounds that bind and potentially inhibit OXA-48. Using this approach, a hit compound, CDD-97, was identified with sub-micromolar potency (Ki = 0.53 ± 0.08 µM) against OXA-48. X-ray crystallography showed that CDD-97 binds non-covalently in the active site of OXA-48. Synthesis and testing of derivatives of CDD-97 revealed structure-activity relationships and informed the design of a compound with a two-fold increase in potency. CDD-97, however, synergizes poorly with β-lactam antibiotics to inhibit the growth of bacteria expressing OXA-48 due to poor accumulation in E. coli. Despite the low in vivo activity, CDD-97 provides new insights into OXA-48 inhibition and demonstrates the potential of using DNA-encoded chemistry technology to rapidly identify β-lactamase binders and study β-lactamase inhibition, leading to clinically useful inhibitors.
Justin M. Rectenwald; Shiva Krishna Reddy Guduru; Zhao Dang; Leonard B. Collins; Yi-En Liao; Jacqueline L. Norris-Drouin; Stephanie H. Cholensky; Kyle W. Kaufmann; Scott M. Hammond; Dmitri B. Kireev; Stephen V. Frye; Kenneth H. Pearce Molecules, 2020, 25(4), 979 https://doi.org/10.3390/molecules25040979AbstractChromatin structure and function, and consequently cellular phenotype, is regulated in part by a network of chromatin-modifying enzymes that place post-translational modifications (PTMs) on histone tails. These marks serve as recruitment sites for other chromatin regulatory complexes that ‘read’ these PTMs. High-quality chemical probes that can block reader functions of proteins involved in chromatin regulation are important tools to improve our understanding of pathways involved in chromatin dynamics. Insight into the intricate system of chromatin PTMs and their context within the epigenome is also therapeutically important as misregulation of this complex system is implicated in numerous human diseases. Using computational methods, along with structure-based knowledge, we have designed and constructed a focused DNA-Encoded Library (DEL) containing approximately 60,000 compounds targeting bi-valent methyl-lysine (Kme) reader domains. Additionally, we have constructed DNA-barcoded control compounds to allow optimization of selection conditions using a model Kme reader domain. We anticipate that this target-class focused approach will serve as a new method for rapid discovery of inhibitors for multivalent chromatin reader domains.
Laurianne David; Amol Thakkar; Rocío Mercado; Ola Engkvist J. Cheminform., 2020, 12(1), 56 https://doi.org/10.1186/s13321-020-00460-5AbstractThe technological advances of the past century, marked by the computer revolution and the advent of high-throughput screening technologies in drug discovery, opened the path to the computational analysis and visualization of bioactive molecules. For this purpose, it became necessary to represent molecules in a syntax that would be readable by computers and understandable by scientists of various fields. A large number of chemical representations have been developed over the years, their numerosity being due to the fast development of computers and the complexity of producing a representation that encompasses all structural and chemical characteristics. We present here some of the most popular electronic molecular and macromolecular representations used in drug discovery, many of which are based on graph representations. Furthermore, we describe applications of these representations in AI-driven drug discovery. Our aim is to provide a brief guide on structural representations that are essential to the practice of AI in drug discovery. This review serves as a guide for researchers who have little experience with the handling of chemical representations and plan to work on applications at the interface of these fields.
Lindsay McMurray; Thomas M. McGuire; Rachel L. Howells Synthesis, 2020, 52(12), 1719-1737 https://doi.org/10.1055/s-0039-1690843AbstractThis review covers recent advances in decarboxylative photocatalysis applicable to the medicinal chemist. The review is not intended to be exhaustive, but instead is focussed on transformations that could be useful in the synthesis of drug-like compounds in order to highlight the utility of this methodology in the development of new pharmaceutical candidates. 1. Introduction; 2. C–C Bond Formation; 3. C–N and C–O Bond Formation; 4. Fluorination and Trifluoromethylation; 5. Hydrodecarboxylation; 6. Protein Functionalisation; 7. Conclusion
Tong Ye Wang; Liang Hu; Sergey N. Krylov Electrophoresis, 2020, 41(13-14), 1225-1229 https://doi.org/10.1002/elps.202000005AbstractIdeal‐filter capillary electrophoresis (IFCE) is a method for the selection of affinity binders for protein targets from oligonucleotide libraries, e.g., random‐sequence oligonucleotide libraries and DNA‐encoded libraries, in a single step of partitioning. In IFCE, protein–oligonucleotide complexes and unbound oligonucleotides move in the opposite directions facilitating very high efficiency of their partitioning. For any given protein target and oligonucleotide library, protein–oligonucleotide complexes and unbound oligonucleotides move in the opposite directions only for a limited range of electroosmotic flow (EOF) mobilities, which, in turn, corresponds to a limited range of pH and ionic strength values of the running buffer. Rational design of IFCE‐based partitioning requires a priori knowledge of this range of pH and ionic strength values, and here we introduce an approach to predict this range for a given type of the running buffer. The approach involves measuring EOF mobilities for a relatively wide range of pH and ionic strength (I) values and finding an empirical predictor function that related the EOF mobility with pH and ionic strength. In this work, we developed a predictor function for a running buffer (Tris‐HCl) that is commonly used in CE‐based partitioning of affinity binders for protein targets. This predictor function can be immediately used for the rational design of IFCE‐based partitioning in this running buffer, while the described approach will be used to develop predictor functions for other types of running buffer if needed.
Dillon T. Flood; Xuejing Zhang; Xiang Fu; Zhenxiang Zhao; Shota Asai; Brittany Sanchez; Emily J. Sturgell; Julien C. Vantourout; Paul Richardson; Mark E. Flanagan; David W. Piotrowski; Dominik K. Kölmel; Jinqiao Wan; Yong Chang; Zhao Wang; Jason Chen; Phil Baran; Philip Dawson Angew. Chem. Int. Ed. Engl., 2020, 59, 7377-7383 https://doi.org/10.1002/anie.201915493AbstractDNA Encoded Libraries have shown promise as a valuable technology for democratizing the hit discovery process. Although DEL provides relatively inexpensive access to libraries of unprecedented size, their production has been hampered by the idiosyncratic needs of the encoding DNA tag relegating DEL compatible chemistry to dilute aqueous environments. Recently Reversible Adsorption to Solid Support (RASS) has been demonstrated as a promising method to expand DEL reactivity using standard organic synthesis protocols. Here we demonstrate a suite of on-DNA chemistries to incorporate medicinally relevant and C–S, C–P and N–S linkages into DELs, which are underrepresented in the canonical methods.
Min Hao; Jianjun Qiao; Hao Qi Genes, 2020, 11(2), 116 https://doi.org/10.3390/genes11020116AbstractMethods for synthesizing arbitrary single-strand DNA (ssDNA) fragments are rapidly becoming fundamental tools for gene editing, DNA origami, DNA storage, and other applications. To meet the rising application requirements, numerous methods have been developed to produce ssDNA. Some approaches allow the synthesis of freely chosen user-defined ssDNA sequences to overcome the restrictions and limitations of different length, purity, and yield. In this perspective, we provide an overview of the representative ssDNA production strategies and their most significant challenges to enable the readers to make informed choices of synthesis methods and enhance the availability of increasingly inexpensive synthetic ssDNA. We also aim to stimulate a broader interest in the continued development of efficient ssDNA synthesis techniques and improve their applications in future research.
Zhoulong Fan; Shuai Zhao; Tao Liu; Peng-Xiang Shen; Zi-Ning Cui; Zhe Zhuang; Qian Shao; Jason Chen; Anokha S. Ratnayake; Mark E. Flanagan; Dominik K. Kölmel; David W. Piotrowski; Paul Richardson; Jinquan Yu ChemRxiv, 2020 https://doi.org/10.26434/chemrxiv.11514774.v1AbstractDNA-encoded library (DEL) technology has the potential to dramatically expedite hit identification in drug discovery owing to its ability to perform protein affinity selection with millions or billions of molecules in a single experiment. To expand the molecular diversity of DEL, it is critical to develop different types of DNA-encoded transformations that produces billions of molecules with distinct molecular scaffolds. Sequential functionalization of multiple C–H bonds provides a unique avenue for creating diversity and complexity from simple starting materials. However, the use of water as solvent, the presence of DNA, and the extremely low concentration of DNA-encoded coupling partners (0.001 M) have hampered the development DNA-encoded C(sp3)–H activation reactions. Herein, we report the realization of palladium-catalyzed C(sp3)–H arylation of aliphatic carboxylic acids, amides and ketones with DNA-encoded aryl iodides in water. Notably, the present method enables the use of alternative sets of bifunctional building blocks, and facilitates access to certain setups for DELs. Furthermore, sequential C–H arylation chemistry enabled the on-DNA synthesis of structurally-diverse scaffolds containing enriched C(sp3) character, chiral centers, cyclopropane, cyclobutane, and heterocycles.
Xiaoyun Meng; Lanjun Zhang; Hong Wei; Furong Li; Lihua Hu; Haiyun Ma; Qian Liu; Xiaoyu Li; Zhongchuan Liu BioTechniques, 2020, 69, 69(1), 427-433 https://doi.org/10.2144/btn-2019-0170AbstractRefolding of human interleukin 17A (IL-17A) has been reported; however, the key refolding protocol was not robust enough to deliver consistent results and to be easily scaled up for crystallization. Here we report an optimized refolding method for IL-17A. Although co-crystal structures of IL-17A with ligands have been obtained with a high-affinity peptide and an anti-IL-17A Fab as stabilizers, neither the production yield nor the characterization of the IL-17A/Fab complex was reported. To facilitate co-crystallization of IL-17A with small-molecule compounds derived from our DNA encoded library, we also describe the method for yield enhancement of anti-IL-17A Fab production and characterize the IL-17A/Fab complex for the first time, providing an essential prerequisite for structure-based drug discovery targeting IL-17A. METHOD SUMMARY: An Escherichia coli system was used for IL-17A expression. Two-way optimizations of the refolding conditions were implemented by exploring various ratios of oxidized and reduced forms of the oxido-shuffling reagents and the refolding duration. A detailed protocol for the optimized refolding has been submitted to protocols.io (doi 10.17504/protocols.io.bda3i2gn). Yield enhancement of anti-IL-17A Fab production was achieved after generating a stable HEK293-F cell line expressing the Fab fragment by G418 selection of transfected cells. The purified IL-17A/anti-IL-17A Fab complex was characterized by SDS-PAGE and western blotting. Single crystals of the quaternary complex, IL-17A/Fab/HAP (high affinity peptide) with small-molecule compounds derived from our DNA encoded library could be obtained using a batch method.
Yingchu Chen; John Faver; Angela F. Ku; Gabriella Miklossy; Kevin Riehle; Kurt M. Bohren; Melek N. Ucisik; Martin M. Matzuk; Zhifeng Yu; Nicholas Simmons Bioconjugate Chem., 2020, 31, 3, 770-780 https://doi.org/10.1021/acs.bioconjchem.9b00863AbstractDNA-encoded chemical library (DECL) screens are a rapid and economical tool to identify chemical starting points for drug discovery. As a robust transformation for drug discovery, palladium-catalyzed C–N coupling is a valuable synthetic method for the construction of DECL chemical matter—however currently disclosed methods have only been demonstrated on DNA-attached (hetero)aromatic iodide and bromide electrophiles. We developed conditions utilizing an N-heterocyclic carbene–palladium catalyst that extends this reaction to the coupling of DNA-conjugated (hetero)aromatic chlorides with (het-ero)aromatic and select aliphatic amine nucleophiles. In addition we evaluated steric and electronic effects within this catalyst series, carried out a large substrate scope study on two representative (hetero)aryl bromides, applied this newly-developed method within the construction of a 63 million-membered DECL and further validated reaction success through the identification of active hits from a screen of this DECL against the target polo-like kinase 1
Mohan Neetha; Salim Saranya; Nissy Ann Harry; Gopinathan Anilkumar ChemistrySelect, 2020, 5, 736-753 https://doi.org/10.1002/slct.201904436AbstractCoupling reactions form an important class of reactions in organic synthesis and these reactions are usually catalysed by metals and are of great significance. Copper due to its versatility and low toxicity has always gained much attention. This first review on copper‐catalysed amination of aryl and heteroaryl halides summarises the developments in this area and covers literature upto 2019.
Qiuxia Chen; Xuemin Cheng; Lifang Zhang; Xianyang Li; Purui Chen; Jian Liu; Lanjun Zhang; Hong Wei; Zhonghan Li; Dengfeng Dou SLAS Discov., 2020, 25(5), 523-529 https://doi.org/10.1177/2472555219893949AbstractDNA-encoded library (DEL) technology has been used as an ultra-high-throughput screening approach for hit identification of drug targets. This process is an affinity-based selection and requires incubation of DEL molecules with the target. Currently, in most reported cases, the input (i.e., the copy number) of individual DEL molecules varies from 10^5 to 10^7. With the ever-increasing DEL size and screening cost, lowering the input of DEL molecules while maintaining an appropriate signal-to-noise ratio in a selection is of paramount importance. In this article, we varied the input of DEL ranging from 10^3 to 10^5 in selections with two different protein targets to explore the lower limit of DEL molecule input. The results could facilitate the optimization of the DEL selection process and reduce costs related to library consumption.
Liqiang Su; Jing Feng; Ting Peng; Jinqiao Wan; Jing Fan; Jin Li; Jonathan O’Connell; David R. Lancia Jr.; G. Joseph Franklin; Guansai Liu Org. Lett., 2020, 22, 4, 1290-1294 https://doi.org/10.1021/acs.orglett.9b04578Abstract2-Aminobenzimidazole cores are among the most common structural components in medicinal chemistry and can be found in many biologically active molecules. Herein, we report a mild protocol for the synthesis of multifunctional 2-aminobenzimidazoles on-DNA with broad substrate scopes. The reaction conditions expand our ability to design and synthesize 2-aminobenzimidazole core-focused DNA-encoded libraries.
Lluc Farrera-Soler; Jean-Pierre Daguer; Patrick Raunft; Sofia Barluenga; Anne Imberty; Nicolas Winssinger Bioorg. Med. Chem., 2020, 115458 https://doi.org/10.1016/j.bmc.2020.115458AbstractSelections from dynamic combinatorial libraries (DCL) benefit from the dynamic nature of the library that can change constitution upon addition of a selection pressure, such as ligands binding to a protein. This technology has been predominantly used with small molecules interacting with each other through reversible covalent interaction. However, application of this technology in biomedical research and drug discovery has been limited by the reversibility of covalent exchange and the analytical deconvolution of small molecule fragments. Here we report a supramolecular approach based on the use of a constant short PNA tag to direct the combinatorial pairing of fragment. This PNA tag yields fast exchange kinetics, while still delivering the benefits of cooperativity, and provides favourable properties for analytical deconvolution by MALDI. A selection from > 6 000 assemblies of glycans (mono-, di-, tri-saccharides) targeting AFL, a lectin from pathogenic fungus, yielded a 95 nM assembly, nearly three orders of magnitude better in affinity than the corresponding glycan alone (41 µM).
Shorouk O. Badir; Jaehoon Sim; Katelyn Billings; Adam Csakai; Xuange Zhang; Weizhe Dong; Gary A. Molander Org. Lett., 2020, 22, 3, 1046-1051 https://doi.org/10.1021/acs.orglett.9b04568AbstractDNA-encoded library (DEL) technology has emerged as a novel interrogation modality for ligand discovery in the pharmaceutical industry. Given the increasing demand for a higher proportion of C(sp3)-hybridized centers in DEL platforms, a photoredox-mediated cross-coupling and defluorinative alkylation process is introduced using commercially available alkyl bromides and structurally diverse α-silylamines. Notably, no protecting group strategies for amines are necessary for the incorporation of a variety of amino-acid-based organosilanes, providing crucial branching points for further derivatization.
Shohei Hamada J. Synth. Org. Chem., 2020, 78(8), 813-815 https://doi.org/10.5059/yukigoseikyokaishi.78.813AbstractDNA encoded library （DEL） has emerged as a widely utilized chemical library, possessing DNA─tag on each compound, for discovering hit compounds in drug discovery. However, synthetic methods for DEL are not fully exploited due to the difficulty in chemical transformation of DNA encoded compounds. This mini─review focuses on the recent development on organic synthesis for diversity expansion of DEL.
Matthias V. Westphal; Liam Hudson; Jeremy W. Mason; Frédéric J. Zécri; Karin Briner; Stuart L. Schreiber J. Am. Chem. Soc., 2020, 142(17), 7776-7782 https://doi.org/10.1021/jacs.9b13186AbstractDNA-encoded libraries of small molecules are being explored extensively for the identification of binders in early drug-discovery efforts. Combinatorial syntheses of such libraries require water- and DNA-compatible reactions, and the paucity of these reactions currently limit the chemical features of resulting barcoded products. The present work introduces strain-promoted cycloadditions of cyclic allenes under mild conditions to DNA-encoded library synthesis. Owing to distinct cycloaddition modes of these reactive intermediates with activated olefins, 1,3-dipoles and dienes, the process generates diverse molecular architectures from a single precursor. The resulting DNA-barcoded compounds exhibit unprecedented ring and topographic features—related to elements found to be powerful in phenotypic screening.
Ainoa Figuerola-Conchas; Jacques Saarbach; Jean-Pierre Daguer; Adeline Cieren; Sofia Barluenga; Nicolas Winssinger; Monica Gotta ACS Chem. Biol., 2020, 15, 1, 243-253 https://doi.org/10.1021/acschembio.9b00832AbstractVCP/p97 belongs to the AAA+ ATPase family and has an essential role in several cellular processes ranging from cell division to protein homeostasis. Compounds targeting p97 inhibit the main ATPase domain and cause cell death. Here, using PNA-encoded chemical libraries, we have identified two small molecules that target the regulatory domain of p97, comprising the N-terminal and the D1 ATPase domains, and do not cause cell death. One molecule, NW1028, inhibits the degradation of a p97-dependent reporter, whereas the other, NW1030, increases it. ATPase assays show that NW1028 and NW1030 do not affect the main catalytic domain of p97. Mapping of the binding site using a photoaffinity conjugate points to a cleft at the interface of the N-terminal and the D1 ATPase domains. We have therefore discovered two new compounds that bind to the regulatory domain of p97 and modulate specific p97 cellular functions. Using these compounds, we have revealed a role for p97 in the regulation of mitotic spindle orientation in HeLa cells.
Seiya Kitamura; Qinheng Zheng; Jordan L. Woehl; Angelo Solan; Emily Chen; Nicholas Dillon; Mitchell Hull; Miyako Kotaniguchi; John R. Cappiello; Shinichi Kitamura; Victor Nizet; K. Barry Sharpless; Dennis Wolan J. Am. Chem. Soc., 2020, 142, 25, 10899-10904 https://doi.org/10.1021/jacs.9b13652AbstractOptimization of small-molecule probes or drugs is a lengthy, challenging and resource-intensive process. Lack of automation and reliance on skilled medicinal chemists is cumbersome in both academic and industrial settings. Here, we demonstrate a high-throughput hit-to-lead process based on the biocompatible SuFEx click chemistry. A modest high-throughput screening hit against a bacterial cysteine protease SpeB was modified with a SuFExable iminosulfur oxydifluoride [RN=S(O)F2] motif, rapidly diversified into 460 analogs in overnight reactions, and the products directly screened to yield drug-like inhibitors with 300-fold higher potency. We showed that the improved molecule is drug-like and biologically active in a bacteria-host coculture. Since these reactions can be performed on a picomole scale to conserve reagents, we anticipate our methodology can accelerate the development of robust biological probes and drug candidates.
Catrin Sohrabi; Andrew Foster; Ali Tavassoli Nat. Rev. Chem., 2020, 4, 90-101 https://doi.org/10.1038/s41570-019-0159-2AbstractDrug discovery has traditionally focused on using libraries of small molecules to identify therapeutic drugs, but new modalities, especially libraries of genetically encoded cyclic peptides, are increasingly used for this purpose. Several technologies now exist for the production of libraries of cyclic peptides, including phage display, mRNA display and split-intein circular ligation of peptides and proteins. These different approaches are each compatible with particular methods of screening libraries, such as functional or affinity-based screening, and screening in vitro or in cells. These techniques allow the rapid preparation of libraries of hundreds of millions of molecules without the need for chemical synthesis, and have therefore lowered the entry barrier to generating and screening for inhibitors of a given target. This ease of use combined with the inherent advantages of the cyclic-peptide scaffold has yielded inhibitors of targets that have proved difficult to drug with small molecules. Multiple reports demonstrate that cyclic peptides act as privileged scaffolds in drug discovery, particularly against ‘undruggable’ targets such as protein–protein interactions. Although substantial challenges remain in the clinical translation of hits from screens of cyclic-peptide libraries, progress continues to be made in this area, with an increasing number of cyclic peptides entering clinical trials. Here, we detail the various platforms for producing and screening libraries of genetically encoded cyclic peptides and discuss and evaluate the advantages and disadvantages of each approach when deployed for drug discovery.
James H. Hunter; Lisa Prendergast; Louis F. Valente; Andrew Madin; Garry Pairaudeau; Michael J. Waring Bioconjugate Chem., 2020, 31, 1, 149-155 https://doi.org/10.1021/acs.bioconjchem.9b00838AbstractDNA encoded chemical libraries provide a highly efficient means of screening vast numbers of small molecules against an immobilised protein target. Their potential is currently restricted by the constraints of carrying out library synthe-sis in the presence of attached DNA tags, for which a limited number of reactions and substrates can be used. Even established reactions, such as Suzuki-Miyaura couplings, do not give efficient coupling reactions across a wide range of substrates and can lead to significant DNA degradation. We developed an efficient protocol for carrying out Suzu-ki-Miyaura couplings on DNA tagged substrates that proceeds with unprecedented efficiency to the desired biaryl products (>98% on average with no detectable DNA degradation) across a wide range of drug-like substrates using a micellar promoted process with commercial TPGS-750-M surfactant. We have demonstrated the applicability of this method in DEL synthesis by preparing a prototypical 2-dimensional 36-membered library employing the Suzuki-Miyaura coupling methodology as the final library synthesis step. This work shows, for the first time, that standard micellar surfactants enable reactions for encoded library synthesis, leading to libraries of exceptional fidelity, and demonstrates the potential to expand the range of accessible DNA compatible chemistry.
Maja Sidstedt; Peter Rådström; Johannes Hedman Anal. Bioanal. Chem., 2020, 412, 2009-2023 https://doi.org/10.1007/s00216-020-02490-2AbstractDNA analysis has seen an incredible development in terms of instrumentation, assays and applications over the last years. Massively parallel sequencing (MPS) and digital PCR are now broadly applied in research and diagnostics, and quantitative PCR is used for more and more practises. All these techniques are based on in vitro DNA polymerization and fluorescence measurements. A major limitation for successful analysis is the various sample-related substances that interfere with the analysis, i.e. PCR inhibitors. PCR inhibition affects library preparation in MPS analysis and skews quantification in qPCR, and some inhibitors have been found to quench the fluorescence of the applied fluorophores. Here, we provide a deeper understanding of mechanisms of specific PCR inhibitors and how these impact specific analytical techniques. This background knowledge is necessary in order to take full advantage of modern DNA analysis techniques, specifically for analysis of samples with low amounts of template and high amounts of background material. The classical solution to handle PCR inhibition is to purify or dilute DNA extracts, which leads to DNA loss. Applying inhibitor-tolerant DNA polymerases, either single enzymes or blends, provides a more straightforward and powerful solution. This review includes mechanisms of specific PCR inhibitors as well as solutions to the inhibition problem in relation to cutting-edge DNA analysis.
Anthony J. Quartararo; Zachary P. Gates; Bente A. Somsen; Nina Hartrampf; Xiyun Ye; Arisa Shimada; Yasuhiro Kajihara; Christian Ottmann; Bradley L. Pentelute Nat. Commun., 2020, 11, 3183 https://doi.org/10.1038/s41467-020-16920-3AbstractHigh-diversity genetically-encoded combinatorial libraries (108−1013 members) are a rich source of peptide-based binding molecules, identified by affinity selection. Synthetic libraries can access broader chemical space, but typically examine only ~ 106 compounds by screening. Here we show that in-solution affinity selection can be interfaced with nano-liquid chromatography-tandem mass spectrometry peptide sequencing to identify binders from fully randomized synthetic libraries of 108 members—a 100-fold gain in diversity over standard practice. To validate this approach, we show that binders to a monoclonal antibody are identified in proportion to library diversity, as diversity is increased from 106–108. These results are then applied to the discovery of p53-like binders to MDM2, and to a family of 3–19 nM-affinity, α/β-peptide-based binders to 14-3-3. An X-ray structure of one of these binders in complex with 14-3-3σ is determined, illustrating the role of β-amino acids in facilitating a key binding contact.
Krištof Bozovičar; Tomaž Bratkovič Int. J. Mol. Sci., 2020, 21(1), 215 https://doi.org/10.3390/ijms21010215AbstractPeptides are widely used in pharmaceutical industry as active pharmaceutical ingredients, versatile tools in drug discovery, and for drug delivery. They find themselves at the crossroads of small molecules and proteins, possessing favorable tissue penetration and the capability to engage into specific and high-affinity interactions with endogenous receptors. One of the commonly employed approaches in peptide discovery and design is to screen combinatorial libraries, comprising a myriad of peptide variants of either chemical or biological origin. In this review, we focus mainly on recombinant peptide libraries, discussing different platforms for their display or expression, and various diversification strategies for library design. We take a look at well-established technologies as well as new developments and future directions
Jens Bobers; Mateja Klika Škopić; Robin Dinter; Piriyanth Sakthithasan; Laura Neukirch; Christian Gramse; Ralf Weberskirch; Andreas Brunschweiger; Norbert Kockmann ACS Comb. Sci., 2020, 22(3), 101-108 https://doi.org/10.1021/acscombsci.9b00207AbstractLab automation strategies have a vast potential for accelerating discovery processes. They enable higher efficiency and throughput for time-consuming screening procedures and reduce error-prone manual steps. Automating repetitive procedures can for instance support chemists in optimizing chemical reactions. Particularly, the technology of DNA-encoded libraries (DELs) may benefit from automation techniques, since translation of chemical reactions to DNA-tagged reactants often requires screening of multiple reaction parameters and evaluation of large numbers of reactants. Here, we describe a portable, automated system for reagent dispensing that was designed from open source materials. The system was validated performing amide coupling of carboxylic acids to DNA-linked amine and a micelle-mediated Povarov reaction to DNA-tagged hexahydropyrroloquinolines. The latter reaction required accurate pipetting of multiple components including different solvents and a surface-active reagent. Analysis of reactions demonstrated that the robotic system achieved high accuracy comparable to experimentation by an experienced chemist with the potential of higher throughput.
Wieslaw M. Kazmierski; Bing Xia; John Miller; Martha De La Rosa; David Favre; Richard M. Dunham; Yoshiaki Washio; Zhengrong Zhu; Feng Wang; Makda Mebrahtu; Hongfeng Deng; Jonathan Basilla; Liping Wang; Ghotas Evindar; Lijun Fan; Alison Olszewski; Ninad Prabhu; Christopher Davie; Jeffrey A. Messer; Vicente Samano J. Med. Chem., 2020, 63(7), 3552-3562 https://doi.org/10.1021/acs.jmedchem.9b01799AbstractWe report the discovery of a novel IDO1 inhibitor class through the affinity selection of a previously-unreported indole-based DNA-encoded library (DEL). The DEL exemplar, spiro-chromane 1 had moderate IDO1 potency but high in vivo clearance. Series optimization quickly afforded a potent, low in vivo clearance lead 11. Although amorphous 11 was highly bioavailable, crystalline 11 was poorly soluble and suffered disappointingly low bioavailability due to solubility-limited absorption. A prodrug approach was deployed and proved effective in discovering the highly-bioavailable phosphonooxymethyl 31, which rapidly converted to 11 in vivo. Obtaining crystalline 31 proved problematic, however, thus a salt screen was performed in an attempt to circumvent this obstacle and successfully delivered greatly soluble and bioavailable crystalline Tris-salt 32. IDO1 inhibitor 32 is characterized by low-calculated human dose, best-in-class potential, and unusual inhibition mode by binding the IDO1 heme-free (apo) form.
Dominik K. Kölmel; Anokha S. Ratnayake; Mark E. Flanagan Biochem. Biophys. Res. Commun., 2020, 533, 2, 201-208 https://doi.org/10.1016/j.bbrc.2020.04.028AbstractA catalytic manifold that enables photoredox cross-electrophile coupling of alkyl bromides with DNA-tagged aryl iodides in aqueous solution is presented. This metallaphotoredox transformation was aided by the identification of a new pyridyl bis(carboxamidine) ligand, which proved critical to the nickel catalytic cycle. The described C(sp2)-C(sp3) coupling tolerates a wide range of both DNA-tagged aryl iodides as well as alkyl bromides. Importantly, this reaction was optimized for parallel synthesis, which is a paramount prerequisite for the preparation of combinatorial libraries, by using a 96-well plate-compatible blue LED array as the light source. Therefore, this mild and DNA-compatible transformation is well positioned for the construction of DNA-encoded libraries.
Kelly A. McCarthy; G. Joseph Franklin; David R. Lancia Jr.; Martin Olbrot; Eneida Pardo; Jonathan C. O’Connell; Christopher S. Kollmann SLAS Discov., 2020, 25(5), 515-522 https://doi.org/10.1177/2472555220908240AbstractDNA-encoded library (DEL) technology has become a prominent screening platform in drug discovery owing to the capacity to screen billions or trillions of compounds in a single experiment. Although numerous successes with DEL technology have been reported, we are unaware of a rigorous examination of the many different variables that can influence a screen’s success. Herein, we explore the impact of variable sample sequencing depth on the detection of tool compounds with known affinities toward a given target while simultaneously probing the effect of initial compound input. Our sequencing data confirm reports that high-affinity compounds can be discovered directly from a DEL screen, but we demonstrate that a mismatch between selection output and sequencing quantity can obscure useful ligands. Our results highlight the importance of selection coverage in grasping the entire picture of a DEL screen where the signal of a weak or underrepresented ligand may be suppressed by the inherent noise of a selection. These potential missed ligands may be critical to the success or failure of a drug discovery program.
Antimo Gioiello; Alessandro Piccinno; Anna Maria Lozza; Bruno Cerra J. Med. Chem., 2020, 63, 13, 6624-6647 https://doi.org/10.1021/acs.jmedchem.9b01956AbstractMedicinal chemistry plays a fundamental and underlying role in chemical biology, pharmacology, and medicine to discover safe and efficacious drugs. Small molecule medicinal chemistry relies on iterative learning cycles composed of compound design, synthesis, testing, and data analysis to provide new chemical probes and lead compounds for novel and druggable targets. Using traditional approaches, the time from hypothesis to obtaining the results can be protracted, thus limiting the number of compounds that can be advanced into clinical studies. This challenge can be tackled with the recourse of enabling technologies that are showing great potential in improving the drug discovery process. In this Perspective, we highlight recent developments towards innovative medicinal chemistry strategies based on continuous flow systems coupled with automation and bioassays. After a discussion of the aims and concepts, we describe equipment and representative examples of automated flow systems and end-to-end prototypes realized to expedite medicinal chemistry discovery cycles.
Olivier B. C. Monty; Pranavanand Nyshadham; Kurt Bohren; Murugesan Palaniappan; Martin M. Matzuk; Damian W. Young; Nicholas Simmons ACS Comb. Sci., 2020, 22, 2, 80-88 https://doi.org/10.1021/acscombsci.9b00199AbstractReaction heterogeneity, poor pH control, and catalyst decomposition in the ring-closing metathesis (RCM) of DNA–chemical conjugates leads to poor yields of the cyclized products. Herein we address these issues with a RCM reaction sys-tem that includes a novel aqueous solvent combination to enable reaction homogeneity, an acidic buffer system which masks traditionally problematic functional groups and a decomposition-resistant catalyst which maximizes conversion to the cy-clized product. Additionally, we provide a systematic study of the substrate scope of the on-DNA RCM reaction, a demon-stration of its applicability to a single-substrate DNA-encoded chemical library that includes sequencing analysis, and the first successful stapling of an unprotected on-DNA [i, i+4] peptide.
Wei Hou; Bo Liu; Hongtao Xu Eur. J. Med. Chem., 2020, 189, 112081 https://doi.org/10.1016/j.ejmech.2020.112081AbstractCelastrol, a principal bioactive ingredient of Tripterygium wilfordii Hook F, has gained extensive exploration due to its unique structure features and multiple promising biological activities. This review will focus on the structural modifications, structure–activity relationships, pharmacology, and toxicology of celastrol in the past ten years. We hope this review would be helpful to get a better grasp of the progresses in the field and provide constructive suggestions for the further study of celastrol.
Jing Guo; Zhen Zhang; Ke Ding Expert Opin. Ther. Patents, 2020, 30(5), 341-350 https://doi.org/10.1080/13543776.2020.1732925AbstractIntroduction: Discoidin domain receptor 1 (DDR1) is a collagen-activated receptor tyrosine kinase. Upon collagen binding, DDR1 undergoes tyrosine autophosphorylation, which consequently triggers downstream genetic and cellular pathways and plays critical roles in regulation of cellular morphogenesis, differentiation, proliferation, adhesion, migration, and invasion. Increasing evidence suggests the potential roles of DDR1 in various human diseases including cancer, fibrosis, atherosclerosis, and other inflammatory disorders. Modulating the activity of DDR1 may be considered as a new therapeutic strategy for human cancer and inflammation-related diseases. Areas covered: This article summarizes current progress on the development of selective DDR1 inhibitors and their potential therapeutic application during the period from 2014 to 2019. Expert opinion: DDR1 is closely linked to a variety of human diseases, including fibrotic disorders, atherosclerosis, and cancer etc. Thus, DDR1 has been considered as a new potential target for drug discovery. A number of DDR1 inhibitors have been identified in the past 5 years, but most of them display relatively broad inhibition across the kinome. New generation DDR1 inhibitors targeting the allosteric sites outside of the canonical ATP-binding pocket or extracellular domain (allosteric inhibitors) may offer a new opportunity for selective DDR1 inhibition therapy development.
J. F. Hess; T. A. Kohl; M. Kotrová; K. Rönsch; T. Paprotka; V. Mohr; T. Hutzenlaub; M. Brüggemann; R. Zengerle; S. Niemann; N. Paust Biotechnol. Adv., 2020, 107537 https://doi.org/10.1016/j.biotechadv.2020.107537AbstractNext generation sequencing is in the process of evolving from a technology used for research purposes to one which is applied in clinical diagnostics. Recently introduced high throughput and benchtop instruments offer fully automated sequencing runs at a lower cost per base and faster assay times. In turn, the complex and cumbersome library preparation, starting with isolated nucleic acids and resulting in amplified and barcoded DNA with sequencing adapters, has been identified as a significant bottleneck. Library preparation protocols usually consist of a multistep process and require costly reagents and substantial hands-on-time. Considerable emphasis will need to be placed on standardisation to ensure robustness and reproducibility. This review presents an overview of the current state of automation of library preparation for next generation sequencing. Major challenges associated with library preparation are outlined and different automation strategies are classified according to their functional principle. Pipetting workstations allow high-throughput processing yet offer limited flexibility, whereas microfluidic solutions offer great potential due to miniaturisation and decreased investment costs. For the emerging field of single cell transcriptomics for example, microfluidics enable singularisation of tens of thousands of cells in nanolitre droplets and barcoding of the RNA to assign each nucleic acid sequence to its cell of origin. Finally, two applications, the characterisation of bacterial pathogens and the sequencing within human immunogenetics, are outlined and benefits of automation are discussed.
Julie S. Valastyan; Michael R. Tota; Isabelle R. Taylor; Vasiliki Stergioula; Graham A.B. Hone; Chari D. Smith; Brad R. Henke; Kenneth G. Carson; Bonnie L. Bassler ACS Chem. Biol., 2020, 15(2), 446-456 https://doi.org/10.1021/acschembio.9b00905AbstractPseudomonas aeruginosa is a leading cause of hospital-acquired infections in the United States. PqsE, a thioesterase enzyme, is vital for virulence of P. aeruginosa, making PqsE an attractive target for inhibition. Neither the substrate nor the product of PqsE catalysis has been identified. A library of 550 million DNA-encoded drug-like small molecules was screened for those that bind to the purified PqsE protein. The structures of the bound molecules were identified by high throughput sequencing of the attached DNA barcodes. Putative PqsE binders with the strongest affinity features were examined for inhibition of PqsE thioesterase activity in vitro. The most potent inhibitors were re-synthesized off DNA and examined for the ability to alter PqsE thermal melting and for PqsE thioesterase inhibition. Here, we report the synthesis, biological activity, mechanism of action, and early structure-activity relationships of a series of 2-(phenylcarbamoyl)benzoic acids that non-competitively inhibit PqsE. A small set of analogs designed to probe initial structure-activity relationships showed increases in potency relative to the original hits, the best of which has an IC50 = 5 uM. Compound refinement is required to assess their in vivo activities as the current compounds do not accumulate in the P. aeruginosa cytosol. Our strategy validates DNA-encoded compound library screening as a rapid and effective method to identify catalytic inhibitors of the PqsE protein, and more generally, for discovering binders to bacterial proteins revealed by genetic screening to have crucial in vivo activities but whose biological functions have not been well defined.
Samuel E. Dalton; Sebastien Campos ChemBioChem, 2020, 21(8), 1080-1100 https://doi.org/10.1002/cbic.201900674AbstractCovalent drugs have experienced significant renewed interest in drug discovery. This resurgence has been accompanied by a better understanding of the reactivity relationships required to engage selective covalent bonds between nucleophilic proteins and electrophilic small molecules. As a result, researchers have come to the realisation that covalent molecules could also represent useful and novel tools aimed at supporting medicinal chemistry programmes. This review surveys the increasing number of drug discovery platforms employing covalent chemistries, and highlights the utility of these techniques for identifying and characterising small molecules and biological targets.
Helena Andrade; Alvin K. Thomas; Weilin Lin; Francesco V. Reddavide; Yixin Zhang ChemBioChem, 2020, 21, 1144-1149 https://doi.org/10.1002/cbic.201900603AbstractThe study of populations of large size and high diversity is limited by the capability of collecting data. Moreover, for a pool of individuals, each associated with a unique characteristic feature, as the pool size grows, the possible interactions increase exponentially, quickly beyond the limit of computation and experimental studies. Herein, we present designs of DNA libraries with various diversity. Using a facile analytic method based on real time PCR, we can evaluate the diversity of a pool of DNA allowing extraordinarily high heterogenicity (e.g. > 1 trillion). We demonstrated that these DNA libraries can be used to model heterogeneous populations, exhibiting functions such as self-protection, suitable for biased expansion, and to evolve into amorphous structures. The method has shown the remarkable power of parallel computing using DNA, as it can resemble an analogue computer and be applied in selection-based biotechnology methods, such as DNA-encoded chemical libraries. As a chemical approach to solve problems traditionally for genetic and statistical analysis, the method provides a quick and cost-efficient evaluation of library diversity for the intermediate steps through a selection process.
Xuemei Xu; Pia Winterwerber; David Ng; Yuzhou Wu Top. Curr. Chem., 2020, 378, 31 https://doi.org/10.1007/s41061-020-0292-xAbstractDNA nanotechnology, based on sequence-specific DNA recognition, could allow programmed self-assembly of sophisticated nanostructures with molecular precision. Extension of this technique to the preparation of broader types of nanomaterials would significantly improve nanofabrication technique to lower nanometer scale and even achieve single molecule operation. Using such exquisite DNA nanostructures as templates, chemical synthesis of polymer and inorganic nanomaterials could also be programmed with unprecedented accuracy and flexibility. This review summarizes recent advances in the synthesis and assembly of polymer and inorganic nanomaterials using DNA nanostructures as templates, and discusses the current challenges and future outlook of DNA templated nanotechnology.
Petra Schneider; W. Patrick Walters; Alleyn T. Plowright; Norman Sieroka; Jennifer Listgarten; Robert A. Goodnow Jr.; Jasmin Fisher; Johanna M. Jansen; José S. Duca; Thomas S. Rush; Matthias Zentgraf; John Edward Hill; Elizabeth Krutoholow; Matthias Kohler; Jeff Blaney; Kimito Funatsu; Chris Luebkemann; Gisbert Schneider Nat. Rev. Drug Discov., 2020, 19(5), 353-364 https://doi.org/10.1038/s41573-019-0050-3AbstractArtificial intelligence (AI) tools are increasingly being applied in drug discovery. While some protagonists point to vast opportunities potentially offered by such tools, others remain sceptical, waiting for a clear impact to be shown in drug discovery projects. The reality is probably somewhere in-between these extremes, yet it is clear that AI is providing new challenges not only for the scientists involved but also for the biopharma industry and its established processes for discovering and developing new medicines. This article presents the views of a diverse group of international experts on the ‘grand challenges’ in small-molecule drug discovery with AI and the approaches to address them.
Minsoo Song; Gil Tae Hwang J. Med. Chem., 2020, 63, 13, 6578-6599 https://doi.org/10.1021/acs.jmedchem.9b01782AbstractDNA-encoded library technology (DELT) was introduced to our medicinal chemistry society more than 20 years ago. The application of DELT in the development of clinical candidates has been actively reported in the literature recently. A few representative examples include RIP1K inhibitors for inflammatory diseases and sEH inhibitors for endothelial dysfunction or abnormal tissue repair, among many others. Here, the authors would like to recall the recent developments in on-DNA synthetic methodologies for DEL construction and to analyze recent examples in the literature of DELT-based drug development efforts pursued in both the academic and industrial sectors. With this perspective, we hope to provide a useful summary of recent DELT-based drug discovery research and to discuss the future scope of DELT in medicinal chemistry.
Ayah Abdeldayem; Yasir S. Raouf; Stefan N. Constantinescu; Richard Moriggl; Patrick T. Gunning Chem. Soc. Rev., 2020, 49(9), 2617-2687 https://doi.org/10.1039/C9CS00720BAbstractOver the past decade, covalent kinase inhibitors (CKI) have seen a resurgence in drug discovery. Covalency affords a unique set of advantages as well as challenges relative to their non-covalent counterpart. After reversible protein target recognition and binding, covalent inhibitors irreversibly modify a proximal nucleophilic residue on the protein via reaction with an electrophile. To date, the acrylamide group remains the predominantly employed electrophile in CKI development, with its incorporation in the majority of clinical candidates and FDA approved covalent therapies. Nonetheless, in recent years considerable efforts have ensued to characterize alternative electrophiles that exhibit irreversible or reversibly covalent binding mechanisms towards cysteine thiols and other amino acids. This review article provides a comprehensive overview of CKIs reported in the literature over a decade period, 2007–2018. Emphasis is placed on the rationale behind warhead choice, optimization approach, and inhibitor design. Current FDA approved CKIs are also highlighted, in addition to a detailed analysis of the common trends and themes observed within the listed data set.
Chia-Te Kung; Hongyi Gao; Chia-Yen Lee; Yao-Nan Wang; Wenjun Dong; Chien-Hsuan Ko; Ge Wang; Lung-Ming Fu Chem. Eng. J., 2020, 399, 125748 https://doi.org/10.1016/j.cej.2020.125748AbstractNanomaterials have attracted significant attention in biomedical and environmental science in recent years due to their unique thermal, optical, electrical and magnetic properties. The properties of nanomaterials are highly correlated with their size and morphology. Consequently, efficient methods for synthesizing nanomaterials with a consistent size and shape are urgently required. Compared to conventional large-scale synthesis systems, microfluidic systems offer a far better control over the growth, nucleation and reaction conditions. As such, microfluidic synthesis has emerged as a key enabling technology for the rapid, low cost and reliable preparation of nanomaterials with complex properties and functions. This review provides a detailed overview of advances in the microfluidic synthesis technology field over the past five years. The review commences by describing the main microfluidic control mechanisms and microfluidic synthesis of particles. Some of the more common applications of microfluidic synthesized micro and nanomaterials are then introduced, including drug delivery, bioimaging, biosensing, environmental analysis, and cell analysis. The review concludes with a brief overview of the challenges facing the microfluidic synthesis field in the coming years, together with possible research directions aimed at overcoming these challenges.
Christopher J. Gerry; Stuart L. Schreiber Curr. Opin. Chem. Biol., 2020, 56, 1-9 https://doi.org/10.1016/j.cbpa.2019.08.008AbstractFor two decades, diversity-oriented synthesis (DOS) has facilitated the assembly of small-molecule libraries comprising a variety of complex molecular architectures. Here, we describe some of the recent achievements in this field, many of which promise to contribute to the development of new chemical probes and drug leads. In particular, we report progress along several avenues of bioactive discovery that leverage topographically complex compounds generated using DOS and other methods. We also discuss advances in DNA-compatible chemistry that enable syntheses of more three-dimensionally complex and diverse DNA-encoded libraries. Continual innovation in organic chemistry will be required to both expand and exploit our understanding of biological systems.
Huimin Lu; Heng Li; Chen Fan; Qing Qi; Yuxi Yan; Yanwei Wu; Chunlan Feng; Bing Wu; Yuanzhuo Gao; Jianping Zuo; Wei Tang Biochem. Pharmacol., 2020, 172, 113751 https://doi.org/10.1016/j.bcp.2019.113751AbstractBackground: The receptor-interacting protein kinase 1 (RIPK1) has emerged as a key upstream regulator that controls the inflammatory response via its kinase-dependent and independent functions, which makes it an attractive target for developing new drugs against inflammation-related diseases. Growing evidences illustrate that RIPK1 is certainly associated with pathogenesis of multiple tissue-damage diseases. However, what are intricate regulatory codes of RIPK1 inhibitor in diseases is still obscure. Methods: We used DSS-induced colitis model in vivo to study the therapeutic effects and the mechanisms of RIPK1 inhibitor. We next characterized the barrier function and the interaction between intestinal epithelial cells (IECs) and immunocytes both in vivo and in vitro. As a candidate in clinical study, GSK2982772 is the most well-developed drug of RIPK1 inhibitors, and we chose it as our study object. Results: We demonstrated that RIPK1 inhibitor could ameliorate the intestinal barrier injury by reducing tight junctions’ disruption and accompanying oxidative stress. Moreover, the release of chemokines and adhesion molecules from damaged IECs was suppressed by RIPK1 inhibitor treatment. And these protective effects were not only dependent on the suppression of necroptosis but also on the compromised activity of NF-κB. Taken together, RIPK1 inhibitor exerts suppressive function in intestinal inflammatory response possibly via protecting the intestinal epithelial barrier and maintaining the homeostasis of immune microenvironments. Eventually, the positive feedback immune response which triggered progressive epithelial cells injury could be restrained.
Christopher R. Wellaway; Dominique Amans; Paul Bamborough; Heather Barnett; Rino A. Bit; Jack A. Brown; Neil R. Carlson; Chun-wa Chung; Anthony W. J. Cooper; Peter D. Craggs; Robert P. Davis; Tony W. Dean; John P. Evans; Laurie Gordon; Isobel L. Harada; David J. Hirst; Philip G. Humphreys; Katherine L. Jones; Antonia J. Lewis; Matthew J. Lindon; Dave Lugo; Mahnoor Mahmood; Scott McCleary; Patricia Medeiros; Darren J. Mitchell; Michael O’Sullivan; Armelle Le Gall; Vipulkumar K. Patel; Chris Patten; Darren L. Poole; Rishi R. Shah; Jane E. Smith; Kayleigh A. J. Stafford; Pamela J. Thomas; Mythily Vimal; Ian D. Wall; Robert J. Watson; Natalie Wellaway; Gang Yao; Rab K. Prinjha J. Med. Chem. , 2020, 63, 2, 714-746 https://doi.org/10.1021/acs.jmedchem.9b01670AbstractThe bromodomain and extraterminal (BET) family of bromodomain-containing proteins are important regulators of the epigenome through their ability to recognize N-acetyl lysine (KAc) post-translational modifications on histone tails. These interactions have been implicated in various disease states and, consequently, disruption of BET–KAc binding has emerged as an attractive therapeutic strategy with a number of small molecule inhibitors now under investigation in the clinic. However, until the utility of these advanced candidates is fully assessed by these trials, there remains scope for the discovery of inhibitors from new chemotypes with alternative physicochemical, pharmacokinetic, and pharmacodynamic profiles. Herein, we describe the discovery of a candidate-quality dimethylpyridone benzimidazole compound which originated from the hybridization of a dimethylphenol benzimidazole series, identified using encoded library technology, with an N-methyl pyridone series identified through fragment screening. Optimization via structure- and property-based design led to I-BET469, which possesses favorable oral pharmacokinetic properties, displays activity in vivo, and is projected to have a low human efficacious dose.
Srikanth Gatadi; Srinivas Nanduri Bioorg. Chem., 2020, 95, 103534 https://doi.org/10.1016/j.bioorg.2019.103534AbstractMultidrug resistant Mycobacterium tuberculosis (MDR-MTB) infections have created a critical health problem globally. The appalling rise in drug resistance to all the current therapeutics has triggered the need for identifying new antimycobacterial agents effective against multidrug-resistant Mycobacterium tuberculosis. Structurally unique chemical entities with new mode of action will be required to combat this pressing issue. This review gives an overview of the structures and outlines on various aspects of in vitro pharmacological activities of new antimycobacterial agents, mechanism of action and brief structure activity relationships in the perspective of drug discovery and development. This review also summarizes on recent reports of new antimycobacterial agents.
Takashi Motoyaji Chem. Pharm. Bull., 2020, 68, 191-193 https://doi.org/10.1248/cpb.c19-00832AbstractAffinity selection (AS)-MS is a label-free binding assay technology for the analysis of interactions between targets and small drug molecules, which does not require modification of targets or compounds. AS-MS technology has been used in drug discovery research for more than 10 years, and is currently one of the most important affinity-based screening techniques. As such, it may be the driving force for novel small molecule drug discovery. This review introduces the principles of AS-MS technology and its use in high-throughput screening (HTS), then discusses strategies for its use in drug discovery and its application in target identification.
Connor W. Coley; Natalie S. Eyke; Klavs F. Jensen Angew. Chem. Int. Ed. Engl., 2020, 59(52), 23414-23436 https://doi.org/10.1002/anie.201909989AbstractThis two-part review examines how automation has contributed to different aspects of discovery in the chemical sciences. In this second part, we reflect on a selection of exemplary studies. It is increasingly important to articulate what the role of automation and computation has been in the scientific process and how that has or has not accelerated discovery. One can argue that even the best automated systems have yet to "discover" despite being incredibly useful as laboratory assistants. We must carefully consider how they have been and can be applied to future problems of chemical discovery in order to effectively design and interact with future autonomous platforms. The majority of this article defines a large set of open research directions, including improving our ability to work with complex data, build empirical models, automate both physical and computational experiments for validation, select experiments, and evaluate whether we are making progress toward the ultimate goal of autonomous discovery. Addressing these practical and methodological challenges will greatly advance the extent to which autonomous systems can make meaningful discoveries.
Yves Ruff; Roberto Martinez; Xavier Pellé; Pierre Nimsgern; Pascale Fille; Maxim Ratnikov; Frédéric Berst ACS Comb. Sci., 2020, 22, 3, 120-128 https://doi.org/10.1021/acscombsci.9b00164AbstractThe use of DNA-encoded libraries has emerged as a powerful hit generation technology. Combining the power of combinatorial chemistry to enumerate large compound collections with the efficiency of affinity selection in pools, the methodology makes it possible to interrogate vast chemical space against biological targets of pharmaceutical relevance. Thus, the chemical transformations employed for the synthesis of encoded libraries play a crucial role in the identification of diverse and drug-like starting points. Currently established transformations have mostly been limited to water-compatible reactions to accommodate the growing oligonucleotide tag. Herein, we describe the development of a practical catch-and-release methodology utilizing a cationic, amphiphilic PEG-based polymer to perform chemical transformations on immobilized DNA conjugates under anhydrous conditions. We demonstrate the usefulness of our APTAC (amphiphilic polymer-facilitated transformations under anhydrous conditions) approach by performing several challenging transformations on DNA-conjugated small molecules in pure organic solvents: the addition of a carbanion equivalent to a DNA-conjugated ketone in tetrahydrofuran, the synthesis of saturated heterocycles using the tin (Sn) amine protocol (SnAP) in dichloromethane, and the dual-catalytic (Ir/Ni) metallaphotoredox decarboxylative cross-coupling of carboxylic acids to DNA-conjugated aryl halides in DMSO. In addition, we demonstrate the feasibility of the latter in multititer-plate format.
Xiangyan Yi; Lian Xue; Tim Thomas; Jonathan B. Baell Future Med. Chem., 2020, 12(5), 423-437 https://doi.org/10.4155/fmc-2019-0212AbstractHere, we describe our action plan for hit identification (APHID) that guides the process of hit triage, with elimination of less tractable hits and retention of more tractable hits. We exemplify the process with reference to our high-throughput screening (HTS) campaign against the enzyme, KAT6A, that resulted in successful identification of a tractable hit. We hope that APHID could serve as a useful, concise and digestible guide for those involved in HTS and hit triage, especially those that are relatively new to this exciting and continually evolving technology.
Amber Hackler; Forrest G. FitzGerald; Vuong Q. Dang; Alexander L. Satz; Brian M. Paegel ACS Comb Sci., 2020, 22(1), 25-34 https://doi.org/10.1021/acscombsci.9b00153AbstractDNA-encoded library (DEL) technology is emerging as a key element of the small molecule discovery toolbox. Conventional DEL screens (i.e., on-DNA screening) interrogate large combinatorial libraries via affinity selection of DNA-tagged library members that are ligands of a purified and immobilized protein target. In these selections, the DNA tags can materially and undesirably influence target binding, and therefore the experiment outcome. Here, we use a solid-phase DEL and droplet-based microfluidic screening to separate the DEL member from its DNA tag (i.e. off-DNA screening), for subsequent in-droplet laser-induced fluorescence polarization (FP) detection of target binding, obviating DNA tag interference. Using the receptor tyrosine kinase (RTK) discoidin domain receptor 1 (DDR1) as a proof-of-concept target in a droplet-scale competition binding assay, we screened a 67,100-member solid-phase DEL of drug-like small molecules for competitive ligands of DDR1 and identified several known RTK inhibitor pharmacophores, including azaindole- and quinazolinone-containing monomers. Off-DNA affinity screening hit ligand families agreed with output from a conventional on-DNA DEL affinity binding screen using an analogous library. Off-DNA DEL affinity screening with FP detection is potentially amenable to a wide array of target classes, including nucleic acid binding proteins, proteins that are difficult to overexpress and purify, or targets with no known activity assay.
Jia Xie; Shuyue Wang; Peixiang Ma; Fei Ma; Jie Li; Wei Wang; Fengping Lu; Huan Xiong; Yuang Gu; Shuning Zhang; Hongtao Xu; Guang Yang; Richard A. Lerner iScience, 2020, 23(6), 101197 https://doi.org/10.1016/j.isci.2020.101197AbstractAlthough insulin is a life-saving medicine, administration by daily injection remains problematic. Our goal was to exploit the power of DNA-encoded libraries to identify molecules with insulin-like activity, but with the potential to be developed as oral drugs. Our strategy involved using a 104 member DNA-encoded library containing 160 Traditional Chinese Medicines (nDEL) to identify molecules that bind to and activate the insulin receptor. Importantly, we used the natural ligand, insulin, to liberate bound molecules. Using this selection method on our relatively small, but highly diverse, nDEL yielded a molecule capable of both binding to and activating the insulin receptor. Chemical analysis showed this molecule to be a polycyclic analog of the guanidine metformin, a known drug used to treat diabetes. By using our protocol with other, even larger, DELs we can expect to identify additional organic molecules capable of binding to and activating the insulin receptor.
Chunlin Zhuang; Fener Chen J. Med. Chem., 2020, 63(4), 1490-1510 https://doi.org/10.1021/acs.jmedchem.9b01317AbstractNecroptosis, an important form of programmed cell death (PCD), is a highly regulated caspase-independent type of cell death that plays a critical role in the pathophysiology of various inflammatory, infectious, and degenerative diseases. Currently, receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like protein (MLKL) have been widely recognized as critical therapeutic targets of the necroptotic machinery. Targeting RIPK1, RIPK3, and/or MLKL is a promising strategy for necroptosis-related diseases. Following the identification of the first RIPK1 inhibitor Nec-1 in 2005, the antinecroptosis field is attracting increasing research interest from multiple disciplines, including the biological and medicinal chemistry communities. Herein, we will review the functions of necroptosis in human diseases, as well as the related targets and representative small-molecule inhibitors, mainly focusing on research articles published during the past 10 years. Outlooks and perspectives on the associated challenges are also discussed.
Sijie Wang; Kyle E. Denton; Kathryn F. Hobbs; Tyler Weaver; James M.B. McFarlane; Katelyn E. Connelly; Michael C. Gignac; Natalia Milosevich; Fraser Hof; Irina Paci; Catherine A. Musselman; Emily C. Dykhuizen; Casey J. Krusemark ACS Chem. Biol., 2020, 15, 1, 112-131 https://doi.org/10.1021/acschembio.9b00654AbstractPolycomb repressive complex 1 (PRC1) is critical for mediating gene expression during development. Five chromobox (CBX) homolog proteins, CBX2,4,6,7,8, are incorporated into PRC1 complexes, where they mediate targeting to trimethylated lysine 27 of histone H3 (H3K27me3) via the N-terminal chromodomain (ChD). Individual CBX paralogs have been implicated as drug targets in cancer; however, high similarity in sequence and structure among the CBX ChDs provide a major obstacle in developing selective CBX ChD inhibitors. Here we report the selection of small, focused, DNA-encoded libraries (DELs) against multiple homologous ChDs to identify modifications to a parental ligand that confer both selectivity and potency for the ChD of CBX8. This on-DNA, medicinal chemistry approach enabled the development of SW2_110A, a selective, cell-permeable inhibitor of the CBX8 ChD. SW2_110A binds CBX8 ChD with a Kd of 800 nM, with minimal 5-fold selectivity for CBX8 ChD over all other CBX paralogs in vitro. SW2_110A specifically inhibits the association of CBX8 with chromatin in cells and inhibits the proliferation of THP1 leukemia cells driven by the MLL-AF9 translocation. In THP1 cells, SW2_110A treatment results in the significant decrease in the expression of MLL-AF9 target genes, including HOXA9, validating the previously established role for CBX8 in MLL-AF9 transcriptional activation, and defining the ChD as necessary for this function. The success of SW2_110A provides great promise for the development of highly selective and cell permeable probes for the full CBX family. In addition, the approach taken provides a proof-of-principle demonstration of how DELs can be used iteratively for optimization of both ligand potency and selectivity.
Amanda J. Kennedy; Linda Sundström; Stefan Geschwindner; Eunice K. Y. Poon; Yuhong Jiang; Rongfeng Chen; Rob Cooke; Shawn Johnstone; Andrew Madin; Junxian Lim; Qingqi Liu; Rink-Jan Lohman; Anneli Nordqvist; Maria Fridén-Saxin; Wenzhen Yang; Dean G. Brown; David P. Fairlie; Niek Dekker Commun. Biol., 2020, 3, 782 https://doi.org/10.1038/s42003-020-01504-0AbstractProtease-activated receptor-2 (PAR2) has been implicated in multiple pathophysiologies but drug discovery is challenging due to low small molecule tractability and a complex activation mechanism. Here we report the pharmacological profiling of a potent new agonist, suggested by molecular modelling to bind in the putative orthosteric site, and two novel PAR2 antagonists with distinctly different mechanisms of inhibition. We identify coupling between different PAR2 binding sites. One antagonist is a competitive inhibitor that binds to the orthosteric site, while a second antagonist is a negative allosteric modulator that binds at a remote site. The allosteric modulator shows probe dependence, more effectively inhibiting peptide than protease activation of PAR2 signalling. Importantly, both antagonists are active in vivo, inhibiting PAR2 agonist-induced acute paw inflammation in rats and preventing activation of mast cells and neutrophils. These results highlight two distinct mechanisms of inhibition that potentially could be targeted for future development of drugs that modulate PAR2.
Manh V. Pham; Milan Bergeron-Brlek; Christian Heinis ChemBioChem, 2020, 21(4), 543-549 https://doi.org/10.1002/cbic.201900390AbstractDNA-encoded chemical library technologies enable the screening of large combinatorial libraries of chemically and structurally diverse molecules, including short cyclic peptides. A challenge in the combinatorial synthesis of cyclic peptides is the final step, the cyclization of linear peptides that typically suffers from incomplete reactions and large variability between substrates. Several efficient peptide cyclization strategies rely on the modification of thiol groups, such as the formation of disulfide or thioether bonds between cysteines. In this work, we established a strategy and reaction conditions for the efficient chemical synthesis of cyclic peptide-DNA conjugates based on linking the side chains of cysteines. We tested two different thiol-protecting groups and found that tert-butylthio (S-tBu) works best for incorporating a pair of cysteines, and we show that the DNA-linked peptides can be efficiently cyclized through disulfide and thioether bond formation. In combination with established procedures for DNA encoding, the strategy for incorporation of cysteines may be readily applied for the generation and screening of disulfide- and thioether-cyclized peptide libraries.
Oleksandr O. Grygorenko; Dmitriy M. Volochnyuk; Sergey V. Ryabukhin; Duncan B. Judd Chem. Eur. J., 2020, 26, 6, 1196-1237 https://doi.org/10.1002/chem.201903232AbstractAll pharmaceutical products contain organic molecules; the source may be a natural product or a fully synthetic molecule, or a combination of both. Thus, it follows that organic chemistry underpins both existing and upcoming pharmaceutical products. The reverse relationship has also affected organic synthesis, changing its landscape towards increasingly complex targets. This Review article sets out to give a concise appraisal of this symbiotic relationship between organic chemistry and drug discovery, along with a discussion of the design concepts and highlighting key milestones along the journey. In particular, criteria for a high‐quality compound library design enabling efficient virtual navigation of chemical space, as well as rise and fall of concepts for its synthetic exploration (such as combinatorial chemistry; diversity‐, biology‐, lead‐, or fragment‐oriented syntheses; and DNA‐encoded libraries) are critically surveyed.
Bruno David; Antonio Grondin; Philippe Schambel; Marc Vitorino; Denis Zeyer Phytochem Rev., 2020, 19, 1141-1156 https://doi.org/10.1007/s11101-019-09612-4AbstractPlant natural products (PNP) (e.g., secondary vegetal metabolites and their derivatives) have been a productive source of active ingredients for the pharmaceutical industry. The High Throughput Screening of Plant Natural Products (PNP-HTS) with extracts or isolated compounds has shown to be time consuming, expensive, and not as successful as expected. Recently building upon the innovative fragment-based drug discovery (FBDD) a disruptive approach was developed based on PNP. The fragment approach involves elaboration and/or isolation of weakly binding small molecules with molecular weights between 150 and 250 Da. This method is fundamentally different from HTS in almost every aspect (i.e., size of the compound library, screening methods, and optimization steps from hit to lead). Due to their nature, vegetal natural fragments have unique three-dimensional (3D) properties, high Fsp3, low aromaticity, and large chemo-diversities which represent potential opportunities for developing novel drugs. Preliminary results using vegetal natural fragments appear to be a promising and emerging field which offers valuable prospects for developing new drugs.
Peyton Shieh ChemBioChem,2020, 22(3), 481-482https://doi.org/10.1002/cbic.202000529 Abstract Research at the biological‐material interface often has translation in mind, with applications in medical implants, drug delivery, and regenerative medicine. While the clinical impact of this research is undeniable, a clearer picture of the in vivo behavior of materials is needed to address longstanding limitations in performance and function. Advances in chemical biology and biotechnology have propelled our understanding of how small molecules and biologics behave in living systems. Adapting these techniques to the study of synthetic materials, enabled by modern polymer chemistry, will bring molecular resolution to biological‐material interactions and guide the development of next‐generation biomaterials for therapeutic and diagnostic applications.
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FEXhttps://pdbj.org/mine/summary/6fex Abstract Epithelial discoidin domain-containing receptor 1, 2-[4-bromanyl-2-oxidanylidene-1'-(1~{H}-pyrazolo[4,3-b]pyridin-5-ylcarbonyl)spiro[indole-3,4'-piperidine]-1-yl]-~{N}-[2,2,2-tris(fluoranyl)ethyl]ethanamide, IODIDE ION
Steven Kearnes arXiv:2009.00707https://arxiv.org/abs/2009.00707 Abstract Retrospective testing of predictive models does not consider the real-world context in which models are deployed. Prospective validation, on the other hand, enables meaningful comparisons between data generation processes by incorporating trained models and considering the subjective decisions that affect reproducibility. Prospective experiments are essential for consistent progress in modeling.
Marianne Schimpl; J. Willem M. Nissink; Carolyn Blackett; Kristin Goldberg; Edward J. Hennessy; Elizabeth Hardaker; William McCoull; Lindsay McMurray; Olga Collingwood; Ross Overman; Alexander Pflug; Marian Preston; Philip B. Rawlins; Emma Rivers; Paul Smith; Elizabeth Underwood; Caroline Truman; Juli Warwicker; Jon Winter-Holt; Simon Woodcock PDB - 7AW2https://www.rcsb.org/structure/7AW2 Abstract 5-(2'-chloro-[1,1'-biphenyl]-4-yl)-N-(imidazo[1,2-a]pyridin-6-ylmethyl)-N-methyl-1,3,4-oxadiazol-2-amine, Classification: SIGNALING PROTEIN Organism(s): Homo sapiens Expression System: Escherichia coli BL21(DE3) Mutation(s): Yes
John W. Cuozzo PDB Entry - 6W35https://doi.org/10.2210/pdb6W35/pdb Abstract Classification: LIPID BINDING PROTEIN Organism(s): Pan troglodytes Expression System: Homo sapiens Mutation(s): Yes
Doris Mia Taylor; Liya Hu; B. V. Venkataram Prasad; Timothy Palzkill PDB Entry - 6UVKhttps://doi.org/10.2210/pdb6UVK/pdb Abstract Classification: HYDROLASE/HYDROLASE INHIBITOR Organism(s): Klebsiella pneumoniae Expression System: Escherichia coli BL21(DE3)
Samantha Greasley; Junli Feng; Ya-Li Deng; Albert E. Stewart PDB Entry - 6WFNhttps://doi.org/10.2210/pdb6WFN/pdb Abstract Classification: TRANSFERASE/INHIBITOR Organism(s): Homo sapiens Expression System: Escherichia coli BL21(DE3) Mutation(s): No
Francesco V. Reddavide; Michael Thompson; Luca Mannocci; Yixin Zhang Aldrichimica Acta, 2019, 52, 3, 63 https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma-Aldrich/General_Information/2/acta-52-3-mk.pdf#page=63AbstractAn urgent challenge in chemistry and biotechnology is to develop a routine, robust, and cost-effective method for the identification of molecules that specifically bind to a large variety of protein targets. In recent years, an elegant selection method for small-molecule drug discovery, DNA-encoded chemical library (DECL) technology, has been receiving much attention from the pharmaceutical and biotechnology industries. Here, we review the major recent developments in DECL technology, with a focus on the self-assembling dual-display format, which aims to combine the bio-inspired selection process with a fragmentbased approach to discover potent binders to protein targets.
Ying Huang; Olena Savych; Yuii Moroz; Yiyun Chen; Robert A. Goodnow Aldrichimica Acta, 2019, 52, 3, 75 https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma-Aldrich/General_Information/2/acta-52-3-mk.pdf#page=63AbstractThe use of DNA for encoding and decoding in smallmolecule synthesis for lead identification continues to gain widespread attention and application—more than a quarter century after its first disclosure. Successful execution of a diverse, drug-like library usually requires hundreds to thousands of commonly functionalized building blocks of relatively similar reactivity profiles. Aqueous and DNA-compatible organic reactions that utilize a large number of functionalized building blocks are perhaps among the most obvious and often discussed aspects of the successful application of this chemistry. This review highlights recent (since ~2015), relevant, new, and potentially highly useful such chemical transformations. Thereafter follows a discussion of the properties, requirements, costs, and diversity of building blocks that are currently available and may be useful for the construction of DNA-encoded libraries.
Asher Mullard Nat. Rev. Drug Discov., 2019, 18(12), 887-891 https://doi.org/10.1038/d41573-019-00195-5AbstractFive anti-cancer KRAS inhibitors, with three different modes of action, are in the clinic. More are on the way.
Wei Liu; Wei Deng; Saisai Sun; Chunyan Yu; Xubo Su; Aliang Wu; Youlang Yuan; Zhonglin Ma; Ke Li; Hongfang Yang; Xuanjia Peng; Justin Dietrich Org. Lett., 2019, 21, 24, 9909-9913 https://doi.org/10.1021/acs.orglett.9b03843AbstractAn efficient method is reported to synthesize sulfonamides on DNA from sulfinic acids or sodium sulfinates and amines in the presence of iodine under mild conditions. This method demonstrates a major expansion of scope of sulfonamide formation on DNA through the utilization of a novel sodium carbonate–sodium sulfinate bifunctional reagent class.
Adrián Gironda-Martínez; Dario Neri; Florent Samain; Etienne J. Donckele Org. Lett., 2019, 21, 23, 9555-9558 https://doi.org/10.1021/acs.orglett.9b03726AbstractDNA-encoded chemical libraries (DECLs) are increasingly employed in hit discovery toward proteins of pharmaceutical interest. Protected amino acids are the most commonly used building blocks for the construction of DECLs; therefore, the expansion of reaction scope with the subsequent free amine is highly desired. Here, we developed a robust DNA-compatible diazo-transfer reaction using imidazole-1-sulfonyl azide tetrafluoroborate salt converting a wide range of primary amines into their corresponding azides in good to excellent yields.
Tinghu Zhang; John M. Hatcher; Mingxing Teng; Nathanael S. Gray; Milka Kostic Cell Chem. Biol., 2019, 26,11, 1486-1500 https://doi.org/10.1016/j.chembiol.2019.09.012AbstractSome of the most widely used drugs, such as aspirin and penicillin, are covalent drugs. Covalent binding can improve potency, selectivity, and duration of the effects, but the intrinsic reactivity represents a potential liability and may result in idiosyncratic toxicity. For decades, the cons were believed to outweigh the pros, and covalent targeting was deprioritized in drug discovery. Recently, several covalent inhibitors have been approved for cancer treatment, thus rebooting the field. In this review, we briefly reflect on the history of selective covalent targeting, and provide a comprehensive overview of emerging developments from a chemical biology stand-point. Our discussion will reflect on efforts to validate irreversible covalent ligands, expand the scope of targets, and discover new ligands and warheads. We conclude with a brief commentary of remaining limitations and emerging opportunities in selective covalent targeting.
Claudio Zambaldo; Stefanie N. Geigle; Alexander L. Satz Org. Lett., 2019, 21, 23, 9353-9357 https://doi.org/10.1021/acs.orglett.9b03553AbstractHerein we provide a generalizable method for the cost-effective synthesis of thousands of building blocks (BBs) employing DNA-incompatible chemistries. The ability to produce large numbers of crude products via solid-phase synthesis has existed for decades; however, our work demonstrates a practical use of such crude reaction mixtures and employs DNA-conjugation to simultaneously encode, purify, and rapidly analyze the desired products. This workflow generated sp3-rich BBs that could be encoded by DNA in a high-throughput manner.
Geoffrey Holdgate; Kevin Embrey; Alexander Milbradt; Gareth Davies1 ADMET and DMPK, 2019, 7(4), 222-241 http://dx.doi.org/10.5599/admet.733AbstractBiophysical methods such as mass spectrometry, surface plasmon resonance, nuclear magnetic resonance, and both differential scanning isothermal titration calorimetry are now well established as key components of the early drug discovery process. These approaches are used successfully for a range of activities, including assay development, primary screening, hit confirmation and detailed mechanistic characterisation of compound binding. Matching the speed, sensitivity and information content of the various techniques to the generation of critical data and information required at each phase of the drug discovery process has been key. This review describes the framework by which these methods have been applied in the drug discovery process and provides case studies to exemplify the impact.
Hongtao Xu; Fei Ma; Nan Wang; Wei Hou; Huan Xiong; Fengping Lu; Jie Li; Shuyue Wang; Peixiang Ma; Guang Yang; Richard A. Lerner Adv. Sci., 2019, 6(23), 1901551 https://doi.org/10.1002/advs.201901551AbstractUsing (hetero)aryl fluorosulfonates as versatile electrophiles, facile on‐DNA cross‐coupling reactions of Suzuki, Sonogashira, and Buchwald are reported here. Notably, all of these reactions show excellent functional group tolerance, mild reaction conditions (relative low temperature and open to air), rich heterocyclic coupling partners, and more importantly, DNA‐compatibility. Thus, these new reactions based on efficient formation of C(sp2)‐C(sp2), C(sp2)‐C(sp), and C(sp2)‐N bonds are highly amenable to synthesis of DNA‐encoded libraries with great molecular diversity.
Stefanie N. Geigle; Ann C. Petersen; Alexander L. Satz Org. Lett., 2019, 21, 22, 9001-9004 https://doi.org/10.1021/acs.orglett.9b03406AbstractDNA-encoded libraries (DELs) have generated recent interest due to their ability to provide new small molecule ligands for pharmaceutically important proteins. The chemical diversity of DELs determines their ability to provide potent, novel, and drug-like chemical matter, and DEL chemical diversity is limited by the scope of DNA-compatible chemical reactions. Herein, the one-pot three-component Van Leusen chemistry is applied to DEL synthesis, providing the first reported DNA-compatible method to generate novel highly functionalized imidazoles.
Klavs F. Jensen; Connor W. Coley; Natalie S. Eyke Angew. Chem. Int. Ed. Engl., 2019 https://doi.org/10.1002/anie.201909987AbstractThis two‐part review examines how automation has contributed to different aspects of discovery in the chemical sciences. In this first part, we describe a classification for discoveries of physical matter (molecules, materials, devices), processes, and models and how they are unified as search problems. We then introduce a set of questions and considerations relevant to assessing the extent of autonomy. Finally, we describe many case studies of discoveries accelerated by or resulting from computer assistance and automation from the domains of synthetic chemistry, drug discovery, inorganic chemistry, and materials science. These illustrate how rapid advancements in hardware automation and machine learning continue to transform the nature of experimentation and modelling. Part two reflects on these case studies and identifies a set of open challenges for the field.
Marco Potowski; Florian Losch; Elena Wunnemann; Janina K. Dahmen; Silvia Chines; Andreas Brunschweiger Chem. Sci., 2019, 10, 10481-10492 https://doi.org/10.1039/C9SC04708EAbstractDNA-encoded compound libraries are a widely used technology for target-based small molecule screening. Generally, these libraries are synthesized by solution phase combinatorial chemistry requiring aqueous solvent mixtures and reactions that are orthogonal to DNA reactivity. Initiating library synthesis with readily available controlled pore glass-coupled DNA barcodes benefits from enhanced DNA stability due to nucleobase protection and choice of dry organic solvents for encoded compound synthesis. We screened the compatibility of solid-phase coupled DNA sequences with 53 metal salts and organic reagents. This screening experiment suggests design of encoded library synthesis. Here, we show the reaction optimization and scope of three sp3-bond containing heterocyclic scaffolds synthesized on controlled pore glass-connected DNA sequences. A ZnCl2-promoted aza-Diels–Alder reaction with Danishefsky's diene furnished diverse substituted DNA-tagged pyridones, and a phosphoric acid organocatalyst allowed for synthesis of tetrahydroquinolines by the Povarov reaction and pyrimidinones by the Biginelli reaction, respectively. These three reactions caused low levels of DNA depurination and cover broad and only partially overlapping chemical space though using one set of DNA-coupled starting materials.
Min Hyeon Shin; Kang Ju Lee; Hyun-Suk Lim Bioconjugate Chem., 2019, 30, 11, 2931-2938 https://doi.org/10.1021/acs.bioconjchem.9b00628AbstractWe report the design and synthesis of a DNA-encoded one-bead one-compound library of cyclic peptoids composed of more than 11 million molecules. We show that affinity-based screening of this large library can identify cyclic peptoid ligands for a target protein. In this work, we developed a simple method for amplifying PCR product from DNA tags on a single bead, thereby enabling to determine the structures of hit cyclic peptoids with no need for high-throughput sequencing and complicated data analysis. We also developed a sub-library screening strategy to minimize false positives caused by the interference of coding DNA tags before starting laborious and impractical hit confirmation. Given the simplicity and robustness of the synthesis and screening, along with the desirable features of macrocyclic peptoids including improved conformational rigidity, our method will be highly useful for discovering biologically active molecules modulating challenging targets such as protein-protein interactions that are not easily targeted by typical peptidomimetics and small-molecules.
Jacques Saarbach; Pramod M. Sabale; Nicolas Winssinger Curr. Opin. Chem. Biol., 2019, 52, 112-124 https://doi.org/10.1016/j.cbpa.2019.06.006AbstractPeptide nucleic acid (PNA) stands as one of the most successful artificial oligonucleotide mimetics. Salient features include the stability of hybridization complexes (either as duplexes or triplexes), metabolic stability, and ease of chemical modifications. These features have enabled important applications such as antisense agents, gene editing, nucleic acid sensing and as a platform to program the assembly of PNA-tagged molecules. Here, we review recent advances in these areas.
Xiaojian Huang; Bowei Ni; Yang Xi; Xiangyu Chu; Rui Zhang; Hongbo You Aging, 2019, 11, 24, 12532-12545 https://doi.org/10.18632/aging.102586AbstractOsteoarthritis (OA) is a highly prevalent joint disorder blamed for pain and disability in older individuals. It’s commonly accepted that inflammation, apoptosis, autophagy and cellular senescence participate in the progress of OA. Protease activated receptor 2 (PAR2), a member of the G-protein coupled receptors, is involved in the regulation of various inflammation diseases. Previous studies have identified PAR2 as a potential therapeutic target for the treatment of OA. Here, we investigated the role of PAR2 antagonist AZ3451 in inflammation response, apoptosis, autophagy and cellular senescence during OA. We confirmed that PAR2 expression was significantly up-regulated in OA articular cartilage tissues as well as in interleukin 1β (IL-1β) stimulated chondrocytes. We demonstrated AZ3451 could prevent the IL-1β-induced inflammation response, cartilage degradation and premature senescence in chondrocytes. Further study showed that AZ3451 attenuated chondrocytes apoptosis by activating autophagy in vitro. The P38/MAPK, NF-κB and PI3K/AKT/mTOR pathways were involved in the protective effect of AZ3451. In vivo, we found that intra-articular injection of AZ3451 could ameliorate the surgery induced cartilage degradation in rat OA model. Our work provided a better understanding of the mechanism of PAR2 in OA, and indicated that PAR2 antagonist AZ3451 might serve as a promising strategy for OA treatment.
Eachan O. Johnson; Deborah T. Hung ACS Chem. Biol., 2019, 14, 12, 2497-2511 https://doi.org/10.1021/acschembio.9b00714AbstractFor the past several decades, chemical biologists have been leveraging chemical principles for understanding biology, tackling disease, and biomanufacturing, while systems biologists have holistically applied computation and genome-scale experimental tools to the same problems. About a decade ago, the benefit of combining the philosophies of chemical biology with systems biology into systems chemical biology was advocated, with the potential to systematically understand the way small molecules affect biological systems. Recently, there has been an explosion in new technologies that permit massive expansion in the scale of biological experimentation, increase access to more diverse chemical space, and enable powerful computational interpretation of large datasets. Fueled by these rapidly increasing capabilities, systems chemical biology is now at an inflection point, poised to enter a new era of more holistic and integrated scientific discovery. Systems chemical biology is primed to reveal an integrated understanding of fundamental biology and to discover new chemical probes to comprehensively dissect and systematically understand that biology, thereby providing a path to novel strategies for discovering therapeutics, designing drug combinations, avoiding toxicity, and harnessing beneficial polypharmacology. In this Review, we examine the emergence of new capabilities driving us to this inflection point in systems chemical biology, and highlight holistic approaches and opportunities that are arising from integrating chemical biology with a systems-level understanding of the intersection of biology and chemistry.
Hongyao Zhu; Mark E. Flanagan; Robert V. Stanton J. Chem. Inf. Model., 2019, 59, 11, 4645-4653 https://doi.org/10.1021/acs.jcim.9b00729AbstractDNA encoded libraries (DEL) are being used as a complement or alternative to traditional high throughput screening (HTS). To maximize the chances of finding chemically attractive lead material that is appropriate for medicinal chemistry optimization, for example, in the Rule of Five compliant chemistry space, it is important to design DEL library compounds such that they are highly diverse and fall within a desired property space. Currently available library design methods can be classified as either monomer-based or product-based. As monomers may undergo significant structural changes when participating in a reaction, monomer based design can provide a poor representation of the properties of resultant DEL products. However, product-based design introduces a technical obstacle due to the enormous chemical design space for many DELs. Here a new method for monomer based selections is described using representative sublibraries as surrogates for fully enumerated DEL property-based optimization. Through a series of rational and systematic library enumerations and property calculations, building-block representatives are identified and representative sublibraries are defined to drive the optimization process. A published data set for a triazine library was used to demonstrate the effectiveness of the multiple objective optimization for six properties. All of the evaluated properties for the designed library are shown to consistently shift toward the desired property distribution as driven by the design criteria.
Meiying Cui; Francesco Reddavide; Stephan Heiden; Luca Mannocci; Michael Thompson; Yixin Zhang bioRxiv, 2019, 08, 24 http://dx.doi.org/10.1101/744680AbstractInformation processing functions are essential for biological organisms to perceive and react to their complex enviornment, as well as for human to analyze and rationalize them. While our brain has an extraordinary power to process complex information, winner(s)-take-all computation is one of the simplest models of lateral inhibition and competition among biological neurons. It has been implemented as DNA-based neural networks, for example, to mimic pattern recognition. However, the utility of DNA-based computation in information processing for real biotechnological applications remains to be demonstrated. In this paper, we developed a winner(s)-take-all method for non-linear amplification of mixtures of DNA sequences. Unlike conventional biological experiments, selected species were not directly subjected to analysis. Instead, parallel computation among myriad of different DNA sequences was carried out with a neural network-inspired winner-take-all function, to reduce the information complexity. The method could be used for various oligonucleotide-encoded libraries, as we have demonstrated its application in decoding and data analysis for selection experiment with DNA-encoded chemical library against protein target.
Thomas Kodadek; Nicholas G. Paciaroni; Madeline Balzarini; Paige Dickson Chem. Commun., 2019, 55, 13330-13341 https://doi.org/10.1039/C9CC06256DAbstractDNA-encoded library (DEL) screening has emerged as an important method for early stage drug and probe molecule discovery. The vast majority of screens using DELs have been relatively simple binding assays. The library is incubated with a target molecule, which is almost always a protein, and the DNAs that remain associated with the target after thorough washing are amplified and deep sequenced to reveal the chemical structures of the ligands they encode. Recently however, a number of different screening formats have been introduced that demand more than simple binding. These include a format that demands hits exhibit high selectivity for target vs. off-targets, a protocol to screen for enzyme inhibitors and another to identify organocatalysts in a DEL. These and other novel assay formats are reviewed in this article. We also consider some of the most significant remaining challenges in DEL assay development.
Sheila Dakhel; Tiziano Ongaro; Baptiste Gouyou; Mattia Matasci; Alessandra Villa; Dario Neri; Samuele Cazzamalli Oncotarget, 2019, 10, 62, 6678-6690 https://doi.org/10.18632/oncotarget.27320AbstractIntroduction: Cytokine-based products are gaining importance for cancer immunotherapy. L19-TNF is a clinical-stage antibody-cytokine fusion protein that selectively accumulates to tumors and displays potent anticancer activity in preclinical models. Here, we describe an innovative approach to transiently inhibit off-target toxicity of L19-TNF, while maintaining antitumor activity. Methods: GSK’963, a potent small molecule inhibitor of RIPK1, was tested in tumor-bearing mice for its ability to reduce acute toxicity associated with TNF signaling. The biological effects of L19-TNF on tumor cells, lymphocytes and tumor vessels were investigated with the aim to enable the administration of TNF doses, which would otherwise be lethal. Results: Transient inhibition of RIPK1 allowed to increase the maximal tolerated dose of L19-TNF. The protective effect of GSK’963 did not affect the selective localization of the immunocytokine to tumors as evidenced by quantitative biodistribution analysis and allowed to reach high local TNF concentrations around tumor blood vessels, causing diffused vascular shutdown and hemorrhagic necrosis within the neoplastic mass. Conclusions: The selective inhibition of RIPK1 with small molecule inhibitors can be used as a pharmaceutical tool to transiently mask TNF activity and improve the therapeutic window of TNF-based biopharmaceuticals. Similar approaches may be applicable to other pro-inflammatory cytokines.
Bo Cai; Dongwook Kim; Saeed Akhand; Yixing Sun; Robert Cassell; Aktan Alpsoy; Emily C. Dykhuizen; Richard M. van Rijn; Michael K. Wendt; Casey J. Krusemark J. Am. Chem. Soc., 2019, 141, 43, 17057-17061 https://doi.org/10.1021/jacs.9b08085AbstractWe report the selection of DNA-encoded small molecule libraries against protein targets within the cytosol and on the surface of live cells. The approach relies on generation of a covalent linkage of the DNA to protein targets by affinity labeling. This crosslinking event enables subsequent co-purification by a tag on the recombinant protein. To access targets within cells, a cyclic cell-penetrating peptide is appended to DNA-encoded libraries for delivery across the cell membrane. As this approach assesses binding of DELs to targets in live cells, it provides a strategy for selection of DELs against challenging targets that cannot be expressed and purified as active.
Sandra Röhm; Benedict-Tilman Berger; Martin Schröder; Apirat Chaikuad; Rob Winkel; Koen F. W. Hekking; Jorg J. C. Benningshof; Gerhard Müller; Roberta Tesch; Mark Kudolo; Michael Forster; Stefan Laufer; Stefan Knapp J. Med. Chem., 2019, 62, 23, 10757-10782 https://doi.org/10.1021/acs.jmedchem.9b01227Abstractp38 mitogen-activated protein kinases are key mediators of environmental stress response and are promising targets for treatment of inflammatory diseases and cancer. Numerous efforts have led to the discovery of several potent inhibitors; however, so far no highly selective type-II inhibitors have been reported. We previously identified VPC-00628 as a potent and selective type-II inhibitor of p38α/β with few off-targets. Here we analyzed the chemical building blocks of VPC-00628 that played a key role in achieving potency and selectivity through targeting an inactive state of the kinases induced by a unique folded P-loop conformation. Using a rapid, systematic combinatorial synthetic approach, we identified compound 93 (SR-318) with excellent potency and selectivity for p38α/β, which potently inhibited the TNF-α release in whole blood. SR-318 therefore presents a potent and selective type-II inhibitor of p38α/β that can be used as a chemical probe for targeting this particular inactive state of these two p38 isoforms.
Misael A. Romero-Reyes; Jennifer M. Heemstra ACS Materials Lett., 2019, 1, 568−572 https://doi.org/10.1021/acsmaterialslett.9b00288AbstractSequestration of small molecules from aqueous solutions poses a significant and important challenge in environmental science and human health. Current methods focus on broadly sequestering all small molecules but are unable to address specific small molecules of interest. Additionally, these procedures require large amounts of resources, such as electricity and pressure. We propose to address this challenge through the use of DNA aptamer-functionalized ultrafiltration membranes. To demonstrate this approach, we developed an aptamer-functionalized membrane that sequesters and removes the small-molecule contaminant bisphenol A (BPA) from water. We show that BPA is depleted and that the membranes can be regenerated for multiple uses, which can allow for recovery of the small molecule when desired. Aptamers can be selected for a wide variety of target small molecules, making this approach highly generalizable beyond our initial demonstration. Together, this research offers a promising solution to improving the efficacy of small molecule removal and recovery from aqueous matrices.
Verena B. K. Kunig; Christiane Ehrt; Alexander Dömling; Andreas Brunschweiger Org. Lett., 2019, 21, 18, 7238-7243 https://doi.org/10.1021/acs.orglett.9b02448AbstractIsocyanide multicomponent reactions play a prominent role in drug discovery. This chemistry has hardly been investigated for compatibility with DNA-encoded combinatorial synthesis. The Ugi, Ugi-azide, and Groebke–Blackburn–Bienaymé reactions are well-tolerated by DNA on the solid phase and show a broad scope. However, an oxadiazole-forming variant of the Ugi reaction caused DNA depurination, requiring a more stable hexathymidine DNA for encoded library synthesis. Cheminformatic analysis revealed that isocyanide multicomponent-reaction-based encoded libraries cover a diverse chemical space.
Jian-Yuan Li; Gabriella Miklossy; Ram K. Modukuri; Kurt M. Bohren; Zhifeng Yu; Murugesan Palaniappan; John C. Faver; Kevin Riehle; Martin M. Matzuk; Nicholas Simmons Bioconjugate Chem., 2019, 30, 8, 2209-2215 https://doi.org/10.1021/acs.bioconjchem.9b00447AbstractA strategy for DNA-compatible, palladium-catalyzed hydroxycarbonylation of (hetero)aryl halides on DNA-chemical conjugates has been developed. This method generally provided the corresponding carboxylic acids in moderate to very good conversions for (hetero)aryl iodides and bromides, and in poor to moderate conversions for (hetero)aryl chlorides. These conditions were further validated by application within a DNA-encoded chemical library synthesis and subsequent discovery of enriched features from the library in selection experiments against two protein targets
Dongwook Kim; Yixing Sun; Dan Xie; Kyle E. Denton; Hao Chen; Hang Lin; Michael K. Wendt; Carol Beth Post; Casey J. Krusemark Molecules, 2019, 24(15), 2764 https://doi.org/10.3390/molecules24152764AbstractAs aberrant activity of protein kinases is observed in many disease states, these enzymes are common targets for therapeutics and detection of activity levels. The development of non-natural protein kinase substrates offers an approach to protein substrate competitive inhibitors, a class of kinase inhibitors with promise for improved specificity. Also, kinase activity detection approaches would benefit from substrates with improved activity and specificity. Here, we apply a substrate-mediated selection to a peptidomimetic DNA-encoded chemical library for enrichment of molecules that can be phosphorylated by the protein tyrosine kinase, c-Src. Several substrates were identified and characterized for activity. A lead compound (SrcDEL10) showed both the ability to serve as a substrate and to promote ATP hydrolysis by the kinase. In inhibition assays, compounds displayed IC50′s ranging from of 8–100 µM. NMR analysis of SrcDEL10 bound to the c-Src:ATP complex was conducted to characterize the binding mode. An ester derivative of the lead compound demonstrated cellular activity with inhibition of Src-dependent signaling in cell culture. Together, the results show the potential for substrate-mediated selections of DNA-encoded libraries to discover molecules with functions other than simple protein binding and offer a new discovery method for development of synthetic tyrosine kinase substrates.
Nannan Sun; Huimin Guo; Yonghui Wang Expert Opin. Ther. Patents, 2019, 29, 9, 663-674 https://doi.org/10.1080/13543776.2019.1655541AbstractIntroduction: RORγt is critical for the differentiation of Th17 cells and the production of IL-17. Inhibition of RORγt is considered as a promising strategy to treat Th17-mediated autoimmune diseases. Quite a number of RORγt inhibitors have been progressed into clinical trials, besides much biological interests in this attractive target. Areas covered: This article reviews the progress of RORγt inhibitors (antagonists and inverse agonists) that are active in clinical development based on an analysis of the related patents published by the corresponding companies in the period of January 2016 through May 2019. Expert opinion: The development of RORγt inhibitors has gone through a boom period in the past three years. However, with a little bit frustration, some of the frontrunner clinical compounds were either discontinued or suspended for further development possibly due to some safety concerns or lack of efficacy in humans. There is a need to probe deeply into these concerns in the on-going pre-clinical and clinical studies. Given the effectiveness of a few recently FDA-approved anti-IL-17(R) antibodies on psoriasis, the discovery of RORγt inhibitors continues to be an exciting field for the development of novel treatment approaches.
Pinwen Cai; Guanyu Yang; Lanzhou Zhao; Jinqiao Wan; Jin Li; Guansai Liu Org. Lett., 2019, 21, 17, 6633-6637 https://doi.org/10.1021/acs.orglett.9b02132Abstract3-Alkylated indole cores have been found in countless natural products and many biologically active compounds, including pharmaceuticals. In this report, a highly efficient approach to C3-alkylated indole derivatives on DNA via indolyl alcohol formation followed by metal-free transfer hydrogenation is developed. This on-DNA C3 alkylation approach is attractive because library compounds can be constructed from simple aldehydes or acid functionalized aldehydes, which are widely commercially available.
Anokha S. Ratnayake; Mark E. Flanagan; Timothy L. Foley; Justin D. Smith; Jillian G. Johnson; Justin Bellenger; Justin I. Montgomery; Brian M. Paegel ACS Comb. Sci., 2019, 21, 10, 650-655 https://doi.org/10.1021/acscombsci.9b00113AbstractDNA-Encoded Chemical Library (DECL) synthesis must occur in aqueous media under conditions that preserve the integrity of the DNA encoding tag. While the identification of "DNA-compatible" reaction conditions is critical for the development of DECL designs that explore previously inaccessible chemical space, reports measuring such compatibility have been largely restricted to methods that do not faithfully capture the impact of reaction conditions on DNA fidelity in solution phase. Here we report a comprehensive methodology that uses soluble DNA substrates that exactly recapitulate DNA’s exposure to the chemically reactive species of DECL synthesis. This approach includes the assessment of chemical fidelity (reaction yield and purity), encoding fidelity (ligation efficiency), and readability (DNA compatibility), revealing the fate of the DNA tag during DECL chemistry from a single platform.
Cedric J. Stress; Basilius Sauter; Lukas A. Schneider; Timothy Sharpe; Dennis Gillingham Angew. Chem. Int. Ed. Engl., 2019, 58(28), 9570-9574 https://doi.org/10.1002/anie.201902513AbstractHerein we perform a seven-step chemical synthesis of a DNA-encoded macrocycle library (DEML) on DNA. Inspired by polyketide and mixed peptide-polyketide natural products, the library was designed to incorporate rich backbone diversity. Achieving this diversity, however, comes at the cost of custom synthesis of bifunctional building block libraries. Our work outlines the importance of careful retrosynthetic design in DNA-encoded libraries, while revealing areas where new DNA synthetic methods are needed.
Mateja Klika Škopi; Katharina Götte; Christian Gramse; Marvin Dieter; Sabrina Pospich; Stefan Raunser; Ralf Weberskirch; Andreas Brunschweiger J. Am. Chem. Soc., 2019, 141, 26, 10546-10555 https://doi.org/10.1021/jacs.9b05696AbstractThe translation of well-established molecular biology methods such as genetic coding, selection and DNA sequencing to combinatorial organic chemistry and compound identification has made extremely large compound collections, termed DNA-encoded libraries, accessible for drug screening. However, the reactivity of the DNA imposes limitations on the choice of chemical methods for encoded library synthesis. For example, strongly acidic reaction conditions must be avoided because they damage the DNA by depurination, i. e. the cleavage of purine bases from the oligomer. Application of micellar catalysis holds much promise for encoded chemistry. Aqueous micellar dispersions enabled compound synthesis under often appealingly mild conditions. Amphiphilic block copolymers covalently functionalized with sulfonic acid moieties in the lipophilic portion assemble in water and locate the Brønsted catalyst in micelles. These acid nanoreactors enabled the reaction of DNA-conjugated aldehydes to diverse substituted tetrahydroquinolines and aminoimidazopyridines by Povarov, and Groebke-Blackburn-Bienaymé reactions, respectively, and the cleavage of tBoc protective groups from amines. The polymer micelle design was successfully translated to the Cu/Bipyridine/TEMPO system mediating the oxidation of DNA-coupled alcohols to the corresponding aldehydes. These results suggest a potentially broad applicability of polymer micelles for encoded chemistry.
Deirdre Jill Cohen; Shubham Pant; Bert O'Neil; Jill Marinis; James Winnberg; Christoph Matthias Ahlers; Justin Callaway; Chetan Rathi; Andre Acusta; Adeline Verticelli; John Bertin; James F. Smothers J. Clin. Oncol., 2019, 37, no. 15_suppl https://doi.org/10.1200/JCO.2019.37.15_suppl.TPS4165AbstractBackground: The immunosuppressive myeloid infiltrate characteristic of the tumor microenvironment in pancreatic cancer represents a major therapeutic barrier in this disease. Modulation of this infiltrate may increase sensitivity to immune checkpoint blockade in this and other tumors with a similar phenotype. The receptor interacting protein 1 (RIP1) is a serine/threonine kinase that becomes active upon homeostatic disruptions. Bound to RIP3 and mixed lineage kinase domain-like protein (MLKL), RIP1 kinase activity drives necroptosis. However, RIP1 also signals in response to inflammatory stimuli independently of its association with RIP3. A correlation between increased RIP1 protein expression and a worse prognosis has been reported in a variety of solid tumors. Furthermore, in an unbiased screen RIP1 was identified as a top gene contributing to resistance to immunotherapy (Manguso 2017). In murine models, RIP1 kinase activity has been reported to drive pancreatic oncogenesis. Inhibition of RIP1 in the pancreatic TME leads to the replacement of tumor-permissive myeloid infiltrates with innate cells promoting an anti-tumor response by the adaptive immune system (Seifert 2016; Wang 2018) and synergized with anti-PD-1 treatment. These data suggest that the small molecule RIP1 inhibitor GSK3145095 may have therapeutic potential in multiple tumor types. Methods: This is a four-part phase 1/2 study designed to evaluate the safety, PK, PD, and preliminary activity of GSK3145095 given orally to participants with selected advanced or recurrent solid tumors. Part 1 will be conducted in approximately 30 adults with pancreatic cancer with escalating doses of GSK3145095. Part 2 will combine escalating doses of GSK3145095 with 200 mg pembrolizumab and may be conducted in a broader population of selected solid tumors. Part 3 represents a cohort expansion of Part 2. Part 4 may investigate the combination of additional anticancer agent(s) with one or more doses of GSK3145095 identified as safe in Part 1. References: Manguso RT. Nature. 2017;547(7664):413-418. Seifert L. Nature. 2016;532(7598):245-249. Wang W. Cancer Cell 2018; 34: 757-774. Clinical trial information: NCT03681951.
Dehua Pei; George Appiah Kubi Expert Opin. Drug Discov., 2019, 14(11), 1097-1102 https://doi.org/10.1080/17460441.2019.1647164AbstractIntroduction: Combinatorial chemistry provides a cost-effective method for rapid discovery of drug hits/leads. The one-bead-one-compound (OBOC) library method is in principle ideally suited for this application, because it permits a large number of structurally diverse compounds to be rapidly synthesized and simultaneously screened for binding to a target of interest. However, application of OBOC libraries in drug discovery has encountered significant technical challenges. Areas covered: This Special Report covers the challenges associated with first-generation OBOC libraries (difficulty in structural identification of non-peptidic hits, screening biases and high false positive rates, and poor scalability). It also covers the many strategies developed over the past two decades to overcome these challenges. Expert opinion: With most of the technical challenges now overcome and the advent of powerful intracellular delivery technologies, OBOC libraries of metabolically stable and conformationally rigidified molecules (macrocyclic peptides and peptidomimetics, rigidified acyclic oligomers, and D-peptides) can be routinely synthesized and screened to discover initial hits against previously undruggable targets such as intracellular protein-protein interactions. On the other hand, further developments are still needed to expand the utility of the OBOC method to non-peptidic chemical scaffolds.
Aarti Kawatkar; Michelle Schefter; Nils-Olov Hermansson; Arjan Snijder; Niek Dekker; Dean G. Brown; Thomas Lundbäck; Andrew X. Zhang; M. Paola Castaldi ACS Chem. Biol., 2019, 14, 9, 1913-1920 https://doi.org/10.1021/acschembio.9b00399AbstractDemonstration of target binding is a key requirement for understanding the mode of action of new therapeutics. The cellular thermal shift assay (CETSA) has been introduced as a powerful label-free method to assess target engagement in physiological environments. Here, we present the application of live-cell CETSA to different classes of integral multipass transmembrane proteins using three case studies, the first showing a large and robust stabilization of the outer mitochondrial five-pass transmembrane protein TSPO, the second being a modest stabilization of SERCA2, and the last describing an atypical compound-driven stabilization of the GPCR PAR2. Our data demonstrated that using modified protocols with detergent extraction after the heating step, CETSA can reliably be applied to several membrane proteins of different complexity. By showing examples with distinct CETSA behaviors, we aim to provide the scientific community with an overview of different scenarios to expect during CETSA experiments, especially for challenging, membrane bound targets.
Oliver Seitz J. Pep. Sci., 2019, 25, e3198. https://doi.org/10.1002/psc.3198AbstractIn light of the 2018 Max Bergmann Medal, this review discusses advancements on chemical biology–driven templated chemistry developed in the author's laboratories. The focused review introduces the template categories applied to orient functional units such as functional groups, chromophores, biomolecules, or ligands in space. Unimolecular templates applied in protein synthesis facilitate fragment coupling of unprotected peptides. Templating via bimolecular assemblies provides control over proximity relationships between functional units of two molecules. As an instructive example, the coiled coil peptide–templated labelling of receptor proteins on live cells will be shown. Termolecular assemblies provide the opportunity to put the proximity of functional units on two (bio)molecules under the control of a third party molecule. This allows the design of conditional bimolecular reactions. A notable example is DNA/RNA–triggered peptide synthesis. The last section shows how termolecular and multimolecular assemblies can be used to better characterize and understand multivalent protein‐ligand interactions.
An T. H. Le; Svetlana M. Krylova; Sergey N. Krylov Electrophoresis, 2019, 40, 2553-2564 https://doi.org/10.1002/elps.201900028AbstractSelection of affinity ligands for protein targets from oligonucleotide libraries currently involves multiple rounds of alternating steps of partitioning of protein‐bound oligonucleotides (binders) from protein‐unbound oligonucleotides (nonbinders). We have recently introduced Ideal‐Filter Capillary Electrophoresis (IFCE) for binder selection in a single step of partitioning. In IFCE, protein‐binder complexes and nonbinders move inside the capillary in the opposite directions, and the efficiency of their partitioning reaches 10^9; i.e., only one of a billion molecules of nonbinders leaks through IFCE while all binders pass through. The condition of IFCE can be satisfied when the magnitude of the mobility of electroosmotic flow is smaller than that of protein–binder complexes and larger than that of nonbinders. The efficiency of partitioning in IFCE is ten million times higher than those of solid‐phase‐based methods of partitioning typically used in selection of affinity ligands for protein targets from oligonucleotide libraries. Here, we provide additional details on our justification for IFCE development. We elaborate on electrophoretic aspects of the method and define the theoretical range of electroosmotic‐flow mobilities which support IFCE. Based on these theoretical results, we identity an experimental range of background electrolyte's ionic strength which supports IFCE. We also extend our interpretation of the results and discuss in‐depth IFCE's prospective in practical applications and fundamental studies.
Christopher J. Helal; Mark Bundesmann; Susan Hammond; Melissa Holmstrom; Jacquelyn Klug-McLeod; Bruce A. Lefker; Dale McLeod; Chakrapani Subramanyam; Oleg Zakaryants; Sylvie Sakata ACS Med. Chem. Lett., 2019, 10, 8, 1104-1109 https://doi.org/10.1021/acsmedchemlett.9b00205AbstractMany pharmaceutical companies have invested millions of dollars in establishing internal chemical stores to provide reliable access to large numbers of building blocks (BB) for the synthesis of new molecules, especially for the timely design and execution of parallel (library) synthesis. Recognizing budget and logistical limitations, we required a more economically scalable process to provide diverse BB. We disclose a novel business partnership that achieves the goals of just-in-time, economical access to com-mercial BB that increases chemical space coverage and accelerates the synthesis of new drug candidates. We believe that this model can be of benefit to companies of all sizes that are engaged in drug discovery by reducing cost, increasing diversity of ana-log molecules in a time-conscious manner, and reducing BB inventory. More efficient use of BB by customers may allow com-mercial vendors to devote a greater portion of their resources to preparing novel BB that increase chemical diversity as opposed to resynthesizing out-of-stock compounds that are inaccessible within company compound collections.
Amber Y. S. Balazs; Rodrigo J. Carbajo; Nichola L. Davies; Yu Dong; Alexander W. Hird; Jeffrey W. Johannes; Michelle L. Lamb; William McCoull; Piotr Raubo; Graeme R. Robb; Martin J. Packer; Elisabetta Chiarparin J. Med. Chem., 2019, 62, 21, 9418-9437 https://doi.org/10.1021/acs.jmedchem.9b00716AbstractThe three-dimensional conformations adopted by a free ligand in solution impact bioactivity and physicochemical properties. Solution 1D NMR spectra inherently contain information on ligand conformational flexibility and three-dimensional shape, as well as the propensity of the free ligand to fully preorganize into the bioactive conformation. Herein we discuss some key learnings, distilled from our experience developing potent and selective synthetic macrocyclic inhibitors, including Mcl-1 clinical candidate AZD5991. Case studies have been selected from recent oncology research projects, demonstrating how 1D NMR conformational signatures can complement X-ray protein–ligand structural information to guide medicinal chemistry optimization. Learning to extract free ligand conformational information from routinely available 1D NMR signatures has proven to be fast enough to guide medicinal chemistry decisions within design cycles for compound optimization.
Alexander Litovchick; Xia Tian; Michael I. Monteiro; Kaitlyn M. Kennedy; Marie-Aude Guié; Paolo A. Centrella; Ying Zhang; Matthew A. Clark; Anthony D. Keefe Molecules, 2019, 24(10), E2026 https://doi.org/10.3390/molecules24102026AbstractInspired by the many reported successful applications of DNA-encoded chemical libraries in drug discovery projects with protein targets, we decided to apply this platform to nucleic acid targets. We used a 120-billion-compound set of 33 distinct DNA-encoded chemical libraries and affinity-mediated selection to discover binders to a panel of DNA targets. Here, we report the successful discovery of small molecules that specifically interacted with DNA G-quartets, which are stable structural motifs found in G-rich regions of genomic DNA, including in the promoter regions of oncogenes. For this study, we chose the G-quartet sequence found in the c-myc promoter as a primary target. Compounds enriched using affinity-mediated selection against this target demonstrated high-affinity binding and high specificity over DNA sequences not containing G-quartet motifs. These compounds demonstrated a moderate ability to discriminate between different G-quartet motifs and also demonstrated activity in a cell-based assay, suggesting direct target engagement in the cell. DNA-encoded chemical libraries and affinity-mediated selection are uniquely suited to discover binders to targets that have no inherent activity outside of a cellular context, and they may also be of utility in other nucleic acid structural motifs
Maomao Yan; Yuyang Zhu; Xueyun Liu; Yi Lasanajak; Jinglin Xiong; Jingqiao Lu; Xi Lin; David Ashline; Vernon Reinhold; David F. Smith; Xuezheng Song Anal. Chem., 2019, 91, 9221−9228 https://doi.org/10.1021/acs.analchem.9b01988AbstractInteractions of glycans with proteins, cells, and microorganisms play important roles in cell–cell adhesion and host–pathogen interaction. Glycan microarray technology, in which multiple glycan structures are immobilized on a single glass slide and interrogated with glycan-binding proteins (GBPs), has become an indispensable tool in the study of protein–glycan interactions. Despite its great success, the current format of the glycan microarray requires expensive, specialized instrumentation and labor-intensive assay and image processing procedures, which limit automation and possibilities for high-throughput analyses. Furthermore, the current microarray is not suitable for assaying interaction with intact cells due to their large size compared to the two-dimensional microarray surface. To address these limitations, we developed the next-generation glycan microarray (NGGM) based on artificial DNA coding of glycan structures. In this novel approach, a glycan library is presented as a mixture of glycans and glycoconjugates, each of which is coded with a unique oligonucleotide sequence (code). The glycan mixture is interrogated by GBPs followed by the separation of unbound coded glycans. The DNA sequences that identify individual bound glycans are quantitatively sequenced (decoded) by powerful next-generation sequencing (NGS) technology, and copied numbers of the DNA codes represent relative binding specificities of corresponding glycan structures to GBPs. We demonstrate that NGGM generates glycan–GBP binding data that are consistent with that generated in a slide-based glycan microarray. More importantly, the solution phase binding assay is directly applicable to identifying glycan binding to intact cells, which is often challenging using glass slide-based glycan microarrays.
Peixiang Ma; Hongtao Xu; Jie Li; Fengping Lu; Fei Ma; Shuyue Wang; Huan Xiong; Wei Wang; Damiano Buratto; Francesco Zonta; Nan Wang; Kaiwen Liu; Tian Hua; Zhi-Jie Liu; Guang Yang; Richard A. Lerner Angew. Chem. Int. Ed. Engl., 2019, 58(27), 9254-9261 https://doi.org/10.1002/anie.201901485AbstractDNA encoded chemical libraries (DELs) link the powers of genetics and chemical synthesis via combinatorial optimization. Through combinatorial chemistry DELs can grow to the unprecedented size of billions to trillions, providing a rich chemical diversity for biological and pharmaceutical research. While in most cases at the molecular level, the diversity is confined to available building blocks of DNA compatible chemical reactions, modern chemical methods are now being used to increase the diversity. To take full advantage of the DEL approach, linking the power of genetics directly to chemical structures would offer even greater diversity in a finite chemical world. Natural products have evolved an incredible structural diversity along with their biological evolution. Here we use traditional Chinese medicine (TCM) as an example, because there are many of them, they are functionally diverse, and they can be obtained in high purity. We describe a late stage modification toolbox approach to annotate these complex organic compounds with amplifiable DNA barcodes, which could be easily incorporated into a DEL. The method of end‐products labeling also generates a cluster of isomers with a single DNA tag at different sites. These isomers provide an additional spatial diversity for multiple accessible pockets of targeted proteins. Notably, a novel PARP‐1 inhibitor with TCM activity has been identified from the natural products enriched DEL (nDEL).
Kevin S. Currie; Leena Patel; Kassandra F. Sedillo Bioorg. Med. Chem. Lett., 2019, 29, 2034-2041 https://doi.org/10.1016/j.bmcl.2019.06.042AbstractInflammatory bowel disease (IBD) is a chronic, debilitating condition with a significant impact on quality of life. In spite of recent advances with antibody therapies, there remains a significant unmet medical need in IBD. Ongoing research and development efforts aim to identify new therapies that will increase remission rates beyond those achieved with current standard-of-care, while maintaining a high safety margin. This review will provide an overview of the small-molecule agents that are being explored in this regard.
Dominik K. Kölmel; Jiang Meng; Mei-Hsuan Tsai; Jiamin Que; Richard P. Loach; Thomas Knauber; Jinqiao Wan; Mark E. Flanagan ACS Comb. Sci., 2019, 21, 8, 588-597 https://doi.org/10.1021/acscombsci.9b00076AbstractA new catalytic manifold that merges photoredox with nickel catalysis in aqueous solution is presented. Specifically, the combination of a highly active, yet air-stable, nickel pre-catalyst with a new electron-deficient pyridyl carboxamidine ligand was key to the development of a water-compatible nickel catalysis platform, which is a crucial requirement for the preparation of DNA-encoded libraries (DELs). Together with an iridium-based photocatalyst and a powerful light source, this dual catalysis approach enabled the efficient decarboxylative arylation of α-amino acids with DNA-tagged aryl halides. This C(sp2)-C(sp3) coupling tolerates a wide variety of functional groups on both the amino acid and the aryl halide substrates. Due to the mild and DNA-compatible reaction conditions, the presented transformation holds great potential for the construction of DELs. This was further evidenced by showing that well plate-compatible LED arrays can serve as competent light sources to facilitate parallel synthesis. Lastly, we demonstrate that this procedure can serve as a blueprint toward the adaptation of other established nickel metallaphotoredox transformations to the idiosyncratic requirements of DEL.
Dirk Trauner; Andrej Shemet Synfacts, 2019, 15(05), 0565 https://doi.org/10.1055/s-0037-1612508AbstractDNA-encoded library (DEL) technology has emerged as a powerful tool for the rapid identification of pharmaceutical lead compounds. New reactions compatible with DELs can significantly expand the potential chemical space available to this technology. However, they must work under aqueous conditions in the presence of DNA.
An T. H. Le; Svetlana M. Krylova; Sergey N. Krylov Anal. Chem., 2019, 91, 8532−8539 https://doi.org/10.1021/acs.analchem.9b01801AbstractIdeal-filter capillary electrophoresis (IFCE) allows selection of protein binders from oligonucleotide libraries in a single step of partitioning in which protein-bound and unbound oligonucleotides move in the opposite directions. In IFCE, the unbound oligonucleotide does not reach the detector, imposing a problem for finding the equilibrium constant (Kd) and rate constant (koff) of protein–oligonucleotide complex dissociation. We report a double-passage approach that allows finding Kd and koff under the IFCE conditions, i.e. near-physiological pH and ionic strength. First, a plug of the protein–oligonucleotide equilibrium mixture passes to the detector in a pressure-driven flow, allowing for both the complex and free oligonucleotide to be detected as a single first peak. Second, the pressure is turned off and the voltage is applied to reverse the migration of only the complex which is detected as the second peak. The experiment is repeated with a lower voltage consequently resulting in longer travel time of the complex to the detector, greater extent of complex dissociation, and the decreased area of the second peak. Finally, the peak areas are used to calculate the values of Kd and koff. Here we explain theoretical and practical aspects of the double-passage approach, prove its validity quantitatively, and, demonstrate its application to determine Kd and koff for an affinity complex between a protein and its DNA aptamer. The double-passage approach for finding Kd and koff of protein–oligonucleotide complexes under the IFCE conditions is a perfect complement for IFCE-based selection of protein binders from oligonucleotide libraries.
Dillon T. Flood; Shota Asai; Xuejing Zhang; Jie Wang; Leonard Yoon; Zoë C. Adams; Blythe C. Dillingham; Brittany Sanchez; Julien C. Vantourout; Mark E. Flanagan; David W. Piotrowski; Paul Richardson; Samantha Green; Ryan Shenvi; Jason Chen; Phil Baran; Philip Dawson J. Am. Chem. Soc., 2019, 141, 25, 9998-10006 https://doi.org/10.1021/jacs.9b03774AbstractHerein, we present the adaptation of reversible adsorption to solid support (RASS) for a DEL setting, which allows reactions to be performed in organic solvents at near anhydrous conditions opening previously inaccessible chemical reactivities to DEL. The RASS approach enabled the rapid development of C(sp2)-C(sp3) decarboxylative cross-couplings with broad substrate scope, an electrochemical amination (the first electrochemical synthetic transformation performed in a DEL context), and improved reductive amination conditions. We believe that RASS will offer expedient access to new DEL reactivities, expanded chemical space, and ultimately more drug-like libraries.
Dafni C. Delivoria; Sean Chia; Johnny Habchi; Michele Perni; Ilias Matis; Nikoletta Papaevgeniou; Martin Reczko; Niki Chondrogianni; Christopher M. Dobson; Michele Vendruscolo; Georgios Skretas Sci. Adv, 2019, 5(10), eaax5108 https://doi.org/10.1126/sciadv.aax5108AbstractProtein misfolding and aggregation are associated with a many human disorders, including Alzheimer's and Parkinson's diseases. Toward increasing the effectiveness of early-stage drug discovery for these conditions, we report a bacterial platform that enables the biosynthesis of molecular libraries with expanded diversities and their direct functional screening for discovering protein aggregation inhibitors. We illustrate this approach by performing, what is to our knowledge, the largest functional screen of small-size molecular entities described to date. We generated a combinatorial library of ~200 million drug-like, cyclic peptides and rapidly screened it for aggregation inhibitors against the amyloid-β peptide (Aβ42), linked to Alzheimer's disease. Through this procedure, we identified more than 400 macrocyclic compounds that efficiently reduce Aβ42 aggregation and toxicity in vitro and in vivo. Finally, we applied a combination of deep sequencing and mutagenesis analyses to demonstrate how this system can rapidly determine structure-activity relationships and define consensus motifs required for bioactivity.
Johannes Ottl; Lukas Leder; Jonas V. Schaefer; Christoph E. Dumelin Molecules, 2019, 24(8), 1629 https://doi.org/10.3390/molecules24081629AbstractThe scope of targets investigated in pharmaceutical research is continuously moving into uncharted territory. Consequently, finding suitable chemical matter with current compound collections is proving increasingly difficult. Encoded library technologies enable the rapid exploration of large chemical space for the identification of ligands for such targets. These binders facilitate drug discovery projects both as tools for target validation, structural elucidation and assay development as well as starting points for medicinal chemistry. Novartis internalized two complementing encoded library platforms to accelerate the initiation of its drug discovery programs. For the identification of low-molecular weight ligands, we apply DNA-encoded libraries. In addition, encoded peptide libraries are employed to identify cyclic peptides. This review discusses how we apply these two platforms in our research and why we consider it beneficial to run both pipelines in-house
Deepak Bandyopadhyay; Constantine Kreatsoulas; Pat G. Brady; Joseph Boyer; Zangdong He; Genaro Scavello Jr.; Tyler Peryea; Ajit Jadhav; Dac-Trung Nguyen; Rajarshi Guha J. Chem. Inf. Model., 2019, 59, 11, 4880-4892 https://doi.org/10.1021/acs.jcim.9b00243AbstractWe present a method for visualizing and navigating large screening datasets while also taking into account their activities and properties. Our approach is to annotate the data with all possible scaffolds contained within each molecule. We have developed a Spotfire visualization, coupled to a fuzzy clustering approach based on the scaffold decomposition of the screening deck, used to drive the hit triage process. Progression decisions can be made using aggregate scaffold parameters and data from multiple datasets merged at the scaffold level. This visualization reveals overlaps that help prioritize hits, highlight tractable series, and posit ways to combine aspects of multiple hits. The structure–activity relationship of a large and complex hit is automatically mapped onto all constituent scaffolds making it possible to navigate, via any shared scaffold, to all related hits. This scaffold “walking” helps address bias toward a handful of potent and ligand-efficient molecules at the expense of coverage of chemical space. We consider two scaffold generation methods and explored their similarities and differences both qualitatively and quantitatively. The workflow of a Spotfire visualization used in combination with fuzzy clustering and structure annotation provides an intuitive view of large and diverse screening datasets. This allows teams to effortlessly navigate between structurally related molecules and enriches the population of leads considered and progressed in a manner complementary to established approaches.
Kit S. Lam Nat. Chem. Biol., 2019, 15, 320-321 https://doi.org/10.1038/s41589-019-0253-2AbstractPowerful combinatorial peptide library methods allow the discovery of peptide leads from diverse libraries. A new platform based on tandem mass spectrometry peptide sequencing coupled with high-performance size-exclusion chromatography enables identification of high-affinity peptidic ligands from focused libraries.
Philip A. Harris; Jill M. Marinis; John D Lich; Scott B. Berger; Anirudh Chirala; Julie A Cox; Patrick M Eidam; Joshua N. Finger; Peter J. Gough; Jae U Jeong; James Kang; Viera Kasparcova; Lara K Leister; Mukesh K. Mahajan; George Miller; Rakesh Nagilla; Michael T. Ouellette; Michael A. Reilly; Alan R. Rendina; Elizabeth J. Rivera; Helen H. Sun; James H. Thorpe; Rachel D Totoritis; Wei Wang; Dongling Wu; Daohua Zhang; John Bertin; Robert W Marquis ACS Med. Chem. Lett., 2019, 10, 6, 857-862 https://doi.org/10.1021/acsmedchemlett.9b00108AbstractRIP1 regulates cell death and inflammation and is believed to play an important role in contributing to a variety of hu-man pathologies, including immune-mediated inflammatory diseases and cancer. While small-molecule inhibitors of RIP1 kinase have been advanced to the clinic for inflammatory diseases and CNS indications, RIP1 inhibitors for oncol-ogy indications have yet to be described. Herein we report on the discovery and profile of GSK3145095 (compound 6). Compound 6 potently binds to RIP1 with exquisite kinase specificity and has excellent activity in blocking RIP1 kinase-dependent cellular responses. Highlighting its potential as a novel cancer therapy, the inhibitor was also able to pro-mote a tumor suppressive T cell phenotype in pancreatic adenocarcinoma organ cultures. Compound 6 is currently in phase 1 clinical studies for pancreatic adenocarcinoma and other selected solid tumors.
Huang-Chi Du; Madison C. Bangs; Nicholas Simmons; Martin M. Matzuk Bioconjugate Chem., 2019, 30, 5, 1304-1308 https://doi.org/10.1021/acs.bioconjchem.9b00188AbstractA multistep protocol for the synthesis of 3,5-disubstituted 1,2,4-oxadiazoles on DNA-chemical conjugates has been developed. A set of six DNA-connected aryl nitriles were converted to corresponding amidoximes with hydroxylamine followed by the O-acylation with a series of aryl and aliphatic carboxylic acids. After cyclodehydration of the O-acyl amidoximes by heating at 90 °C in pH 9.5 borate buffer for 2 h, the desired oxadiazole products were observed in 51–92% conversion with the cleavage of O-acylamidoximes as the major side-product. The reported protocol paves the way for the synthesis of oxadiazole core-focused DNA-encoded chemical libraries.
Paige Dickson; Thomas Kodadek Org. Biomol. Chem., 2019, 17, 4676-4688 https://doi.org/10.1039/C9OB00581AAbstractDNA-encoded libraries represent an exciting and powerful modality for high-throughput screening. In this article, we highlight recent important advances in this field and also suggest some important directions that would make the technology even more powerful.
Gianluca Veggiani; María Carla Rosales Gerpe; Sachdev S. Sidhu; Wei Zhang Pharmacol. Therapeut., 2019, 199, 139–154 https://doi.org/10.1016/j.pharmthera.2019.03.003AbstractDevelopment of effective cancer therapeutic strategies relies on our ability to interfere with cellular processes that are dysregulated in tumors. Given the essential role of the ubiquitin proteasome system (UPS) in regulating a myriad of cellular processes, it is not surprising that malfunction of UPS components is implicated in numerous human diseases, including many types of cancer. The clinical success of proteasome inhibitors in treating multiple myeloma has further stimulated enthusiasm for targeting UPS proteins for pharmacological intervention in cancer treatment, particularly in the precision medicine era. Unfortunately, despite tremendous efforts, the paucity of potent and selective UPS inhibitors has severely hampered attempts to exploit the UPS for therapeutic benefits. To tackle this problem, many groups have been working on technology advancement to rapidly and effectively screen for potent and specific UPS modulators as intracellular probes or early-phase therapeutic agents. Here, we review several emerging technologies for developing chemical- and protein-based molecules to manipulate UPS enzymatic activity, with the aim of providing an overview of strategies available to target ubiquitination for cancer therapy.
Torsten Hoffmann; Marcus Gastreich Drug Discov. Today, 2019, 24, 1148-1156 https://doi.org/10.1016/j.drudis.2019.02.013AbstractRecent innovations have brought pharmacophore-driven methods for navigating virtual chemical spaces, the size of which can reach into the billions of molecules, to the fingertips of every chemist. There has been a paradigm shift in the underlying computational chemistry that drives chemical space search applications, incorporating intelligent reaction knowledge into their core so that they can readily deliver commercially available molecules as nearest neighbor hits from within giant virtual spaces. These vast resources enable medicinal chemists to execute rapid scaffold-hopping experiments, rapid hit expansion, and structure–activity relationship (SAR) exploitation in largely intellectual property (IP)-free territory and at unparalleled low cost.
Alwin M. Hartman; Robin M. Gierse; Anna K. H. Hirsch Eur. J. Org. Chem., 2019, 3581–3590 https://doi.org/10.1002/ejoc.201900327AbstractDynamic combinatorial chemistry (DCC) is a powerful tool to identify bioactive compounds. This efficient technique allows the target to select its own binders and circumvents the need for synthesis and biochemical evaluation of all individual derivatives. An ever‐increasing number of publications report the use of DCC on biologically relevant target proteins. This minireview complements previous reviews by focusing on the experimental protocol and giving detailed examples of essential steps and factors that need to be considered, such as protein stability, buffer composition and cosolvents.
Wesley Cochrane; Marie Lynne Malone; Vuong Q Dang; Valerie J. Cavett; Alexander L. Satz; Brian M Paegel ACS Comb. Sci., 2019, 21, 5, 425-435 https://doi.org/10.1021/acscombsci.9b00037AbstractRobotic high-throughput compound screening (HTS) and, increasingly, DNA-encoded library (DEL) screening are driving bioactive chemical matter discovery in the post-genome era. HTS enables activity-based investigation of highly complex targets using static compound libraries. Conversely, DEL grants efficient access to novel chemical diversity, although screening is limited to affinity-based selections. Here, we describe an integrated droplet-based microfluidic circuit that directly screens solid-phase DELs for activity. An example screen of a 67,100-member library for inhibitors of the phosphodiesterase autotaxin yielded 35 high-priority structures for nanomole-scale synthesis and validation (20 active), guiding candidate selection for synthesis at scale (5/5 compounds with IC50s 4–10 µM). We further compared activity-based hits with those of an analogous affinity-based DEL selection. This miniaturized screening platform paves the way toward applying DELs to more complex targets (signaling pathways, cellular response), and represents a distributable approach to small molecule discovery.
Feng Liu; Hua Wang; Suhua Li; Grant A. L. Bare; Xuemin Chen; Chu Wang; John E. Moses; Peng Wu; K. Barry Sharpless Angew. Chem. Int. Ed. Engl., 2019, 58(24), 8029-8033 https://doi.org/10.1002/anie.201902489AbstractWe report here the development of a suite of biocompatible SuFEx transformations from the SOF4-derived iminosulfur oxydifluoride-hub in aqueous buffer conditions. Thes biocompatible SuFEx reactions of iminosulfur oxydifluorides (R-N=SOF2) with primary amines give sulfamides (8 examples, up to 98%), while the reaction with secondary amines furnish sulfuramidimidoyl fluoride products (8 examples, up to 97%). Likewise, under mild buffered conditions, phenols react with the iminosulfur oxydifluorides (Ar-N=SOF2) to produce sulfurofluoridoimidates (13 examples, up to 99%), which can themselves be further modified by nucleophiles. These transformations open the potential for asymmetric and trisubstituted linkages projecting from the sulfur(VI) center, including versatile S-N and S-O connectivity (9 examples, up to 94%). Finally, the SuFEx bioconjugation of iminosulfur oxydifluorides to amine-tagged single-stranded DNA and to BSA protein demonstrate the potential of SOF4 derived SuFEx click chemistry in biological applications.
Guixian Zhao; Yiran Huang; Yu Zhou; Yizhou Li; Xiaoyu Li Expert Opin. Drug Discov., 2019, 14(8), 735-753 https://doi.org/10.1080/17460441.2019.1614559AbstractIntroduction: DNA-encoded chemical libraries (DELs) have come of age and emerged to become a powerful technology platform for ligand discovery in biomedical research and drug discovery. Today, DELs have been widely adopted in the pharmaceutical industry and employed in drug discovery programs worldwide. DELs are capable of interrogating drug targets with an extremely large number of compounds highly efficiently. Area covered: In this review, the authors introduce the history of DELs and provide an overview of the major technological components, including encoding methods, library synthesis, chemistry, selection methods, hit deconvolution strategy, and post-selection data analysis. A brief update on the hit compounds recently discovered from DEL selections against drug targets is also provided. Finally, the authors discuss their views on the present challenges and future directions for the development and application of DELs in drug discovery. Expert opinion: DELs have provided great opportunities for lead compound discovery at an unprecedented scale and efficiency in drug discovery. The key to the future success of DELs as true discovery modalities, rather than just ‘a way to make many compounds,’ is to go beyond physical binding to functional or even phenotypic assays with the capability to probe the biological system.
Francesco Vincenzo Reddavide; Meiying Cui; Weilin Lin; Naiqiang Fu; Stephan Heiden; Helena Andrade; Michael Sion Thompson; Yixin Zhang Chem. Commun., 2019, 55, 3753-3756 https://doi.org/10.1039/C9CC01429BAbstractWe present a DNA-encoded chemical library, which allows dynamic selection followed by ligation of the encoding strands. As a chemical approach to mimic the genetic recombination process of adaptive immunity, the technology led to enhanced enrichment factor and signal-to-noise ratio as compared to static libraries.
Philip A. Harris; Nicolas Faucher; Nicolas George; Patrick M. Eidam; Bryan W. King; Gemma V. White; Niall A. Anderson; Deepak Bandyopadhyay; Allison M. Beal; Veronique Beneton; Scott B. Berger; Nino Campobasso; Sebastien Campos; Carol A. Capriotti; Julie A. Cox; Alain Daugan; Frederic Donche; Marie-Helé ne Fouchet; Joshua N. Finger; Brad Geddes; Peter J. Gough; Pascal Grondin; Bonnie L. Hoffman; Sandra J. Hoffman; Susan E. Hutchinson; Jae U. Jeong; Emilie Jigorel; Pauline Lamoureux; Lara K. Leister; John D. Lich; Mukesh K. Mahajan; Jamel Meslamani; Julie E. Mosley; Rakesh Nagilla; Pamela M. Nassau; Sze-Ling Ng; Michael T. Ouellette; Kishore K. Pasikanti; Florent Potvain; Michael A. Reilly; Elizabeth J. Rivera; Stephane Sautet; Michelle C. Schaeffer; Clark A. Sehon; Helen Sun; James H. Thorpe; Rachel D. Totoritis; Paris Ward; Natalie Wellaway; David D. Wisnoski; James M. Woolven; John Bertin; Robert W. Marquis J. Med. Chem., 2019, 62, 5096−5110 https://doi.org/10.1021/acs.jmedchem.9b00318AbstractRIP1 kinase regulates necroptosis and inflammation and may play an important role in contributing to a variety of human pathologies, including inflammatory and neurological diseases. Currently, RIP1 kinase inhibitors have advanced into early clinical trials for evaluation in inflammatory diseases such as psoriasis, rheumatoid arthritis, and ulcerative colitis and neurological diseases such as amyotrophic lateral sclerosis and Alzheimer’s disease. In this paper, we report on the design of potent and highly selective dihydropyrazole (DHP) RIP1 kinase inhibitors starting from a high-throughput screen and the lead-optimization of this series from a lead with minimal rat oral exposure to the identification of dihydropyrazole 77 with good pharmacokinetic profiles in multiple species. Additionally, we identified a potent murine RIP1 kinase inhibitor 76 as a valuable in vivo tool molecule suitable for evaluating the role of RIP1 kinase in chronic models of disease. DHP 76 showed efficacy in mouse models of both multiple sclerosis and human retinitis pigmentosa.
Christopher J. Gerry; Mathias J. Wawer; Paul A. Clemons; Stuart L. Schreiber J. Am. Chem. Soc., 2019, 141, 26, 10225-10235 https://doi.org/10.1021/jacs.9b01203AbstractIt is challenging to incorporate stereochemical diversity and topographic complexity into DNA-encoded libraries (DELs) because DEL syntheses cannot fully exploit the capabilities of modern synthetic organic chemistry. Here, we describe the design, construction, and validation of DOS-DEL-1, a library of 107 616 DNA-barcoded chiral 2,3-disubsituted azetidines and pyrrolidines. We used stereospecific C–H arylation chemistry to furnish complex scaffolds primed for DEL synthesis, and we developed an improved on-DNA Suzuki reaction to maximize library quality. We then studied both the structural diversity of the library and the physicochemical properties of individual compounds using Tanimoto multifusion similarity analysis, among other techniques. These analyses revealed not only that most DOS-DEL-1 members have “drug-like” properties, but also that the library more closely resembles compound collections derived from diversity synthesis than those from other sources (e.g., commercial vendors). Finally, we performed validation screens against horseradish peroxidase and carbonic anhydrase IX, and we developed a novel, Poisson-based statistical framework to analyze the results. A set of assay positives were successfully translated into potent carbonic anhydrase inhibitors (IC50 = 20.1–68.7 nM), which confirmed the success of the synthesis and screening procedures. These results establish a strategy to synthesize DELs with scaffold-based stereochemical diversity and complexity that does not require the development of novel DNA-compatible chemistry.
Huang-Chi Du; Nicholas Simmons; John C. Faver; Zhifeng Yu; Murugesan Palaniappan; Kevin Riehle; Martin M. Matzuk Org. Lett., 2019, 21, 7, 2194-2199 https://doi.org/10.1021/acs.orglett.9b00497AbstractA hypodiboric acid system for the reduction of nitro groups on DNA–chemical conjugates has been developed. This transformation provided good to excellent yields of the reduced amine product for a variety of functionalized aromatic, heterocyclic, and aliphatic nitro compounds. DNA tolerance to reaction conditions, extension to decigram scale reductions, successful use in a DNA-encoded chemical library synthesis, and subsequent target selection are also described.
Zachary Wu; S. B. Jennifer Kan; Russell D. Lewis; Bruce J. Wittmann; Frances H. Arnold Proc. Natl. Acad. Sci. USA, 2019, 116(18), 8852-8858 https://doi.org/10.1073/pnas.1901979116AbstractTo reduce experimental effort associated with directed protein evolution and to explore the sequence space encoded by mutating multiple positions simultaneously, we incorporate machine learning into the directed evolution workflow. Combinatorial sequence space can be quite expensive to sample experimentally, but machine-learning models trained on tested variants provide a fast method for testing sequence space computationally. We validated this approach on a large published empirical fitness landscape for human GB1 binding protein, demonstrating that machine learning-guided directed evolution finds variants with higher fitness than those found by other directed evolution approaches. We then provide an example application in evolving an enzyme to produce each of the two possible product enantiomers (i.e., stereodivergence) of a new-to-nature carbene Si-H insertion reaction. The approach predicted libraries enriched in functional enzymes and fixed seven mutations in two rounds of evolution to identify variants for selective catalysis with 93% and 79% ee (enantiomeric excess). By greatly increasing throughput with in silico modeling, machine learning enhances the quality and diversity of sequence solutions for a protein engineering problem.
Xiangyu Liu; Ali Masoudi; Alem W. Kahsai; Li-Yin Huang; Biswaranjan Pani; Dean P. Staus; Paul J. Shim; Kunio Hirata; Rishabh K. Simhal; Allison M. Schwalb; Paula K. Rambarat; Seungkirl Ahn; Robert J. Lefkowitz; Brian Kobilka Science, 2019, 364, 6447, 1283-1287 https://doi.org/10.1126/science.aaw8981AbstractDrugs targeting the orthosteric, primary binding site of G protein-coupled receptors are the most common therapeutics. Allosteric binding sites, elsewhere on the receptors, are less well-defined, and so less exploited clinically. We report the crystal structure of the prototypic β2-adrenergic receptor in complex with an orthosteric agonist and compound-6FA, a positive allosteric modulator of this receptor. It binds on the receptor's inner surface in a pocket created by intracellular loop 2 and transmembrane segments 3 and 4, stabilizing the loop in an a-helical conformation required to engage the G protein. Structural comparison explains the selectivity of the compound for β2- over the β1- adrenergic receptor. Diversity in location, mechanism, and selectivity of allosteric ligands provides potential to expand the range of receptor drugs.
Kim T. Mortensen; Thomas J. Osberger; Thomas A. King; Hannah F. Sore; David R. Spring Chem. Rev., 2019, 119, 17, 10288-10317 https://doi.org/10.1021/acs.chemrev.9b00084AbstractMacrocycles have long been recognized as useful chemical entities for medicine, with naturally occurring and synthetic macrocycles clinically approved for use as prescription drugs. Despite this promise, the synthesis of collections of macrocycles has been historically challenging due to difficulties in the formation of large rings. Diversity-Oriented Synthesis (DOS) emerged in the early 2000s as a powerful strategic solution to the construction of diverse molecular libraries. This review details the various strategies developed within the field of DOS for the synthesis of macrocycle libraries, utilizing modern synthetic methodology to deliver structurally diverse collections of macrocyclic molecules, and the exploration of their therapeutic potential. Section 1 of this work details the use of algorithmic strategies and is divided into Build/Couple/Pair, Advanced Build/Couple/Pair, Initiate/Propagate/Terminate, Fragment-Based Domain Shuffling, Two-Directional Synthesis, and Successive Ring Expansion. Section 2 covers strategies based on ring distortion reactions, including Sequential Cycloaddition/Fragmentation, Ring Expansions, and Miscellaneous.
Nicholas Favalli; Gabriele Bassi; Tania Zanetti; Jörg Scheuermann; Dario Neri Helv. Chim. Acta, 2019, 102, e1900033 https://doi.org/10.1002/hlca.201900033AbstractThe construction of DNA‐encoded chemical libraries (DECLs) crucially relies on the availability of chemical reactions, which are DNA‐compatible and which exhibit high conversion rates for a large number of diverse substrates. In this work, we present our optimization and validation procedures for three copper and palladium‐catalyzed reactions (Suzuki cross‐coupling, Sonogashira cross‐coupling, and copper(I)‐catalyzed alkyne‐azide cycloaddition (CuAAC)), which have been successfully used by our group for the construction of large encoded libraries.
Yan Lu; Shanshan Qin; Bingjie Zhang; Antao Dai; Xiaoqing Cai; Mengna Ma; Zhan-Guo Gao; Dehua Yang; Raymond C. Stevens; Kenneth A. Jacobson; Ming-Wei Wang; Wenqing Shui Anal. Chem., 2019, 91, 13, 8162-8169 https://doi.org/10.1021/acs.analchem.9b00477AbstractAffinity mass spectrometry (MS) enables rapid screening of compound mixtures for ligands bound to a specific protein target, yet its current throughput is limited to individually assay pools of 400-2000 compounds. Typical affinity MS screens implemented in pharmaceutical industry labs identify putative ligands based on qualitative analysis of compound binding to the target whereas no quantitative information is acquired to discriminate high- and low-affinity ligands in the screening phase. Furthermore, these screens require purification of a stabilized form of the protein target, which poses a great challenge for membrane receptor targets. Here, we describe a new, potentially general affinity MS strategy that allows screening of 20,000 compounds in one pool for highly efficient ligand discovery towards a G protein-coupled receptor (GPCR) target. Quantitative measurement of compound binding to the receptor enables high-affinity ligand selection using both the purified receptor and receptor-embedded cell membranes. This high-throughput, label-free and quantitative affinity MS screen resulted in discovery of three new antagonists of the A2A adenosine receptor.
Marco Potowski; Verena Kunig; Florian Losch; Andreas Brunschweiger Med. Chem. Commun., 2019, 10, 1082-1093 https://doi.org/10.1039/C9MD00042AAbstractDNA-encoded libraries of chemically synthesized compounds are an important small molecule screening technology. The synthesis of encoded compounds in solution is currently restricted to a few DNA-compatible and water-tolerant reactions. Encoded compound synthesis on short DNA-barcodes covalently connected to solid support benefits from a broad choice of organic solvents. Here we show that this encoded chemistry approach allows for synthesis of DNA-coupled isoquinolones by Yb(III)-mediated Castagnoli-Cushman reaction under anhydrous reaction conditions and for the synthesis of highly substituted pyrrolidines by Ag(I)-mediated 1,3-dipolar azomethine ylide cycloaddition. An encoding scheme for these DNA-barcoded compounds based on a DNA hairpin is demonstrated.
James P. Phelan; Simon B. Lang; Jaehoon Sim; Simon Berritt; Andrew J. Peat; Katelyn Billings; Lijun Fan; Gary A. Molander J. Am. Chem. Soc., 2019, 141, 8, 3723-3732 https://doi.org/10.1021/jacs.9b00669AbstractDNA-encoded library (DEL) technology is a powerful tool commonly used by the pharmaceutical industry for the identification of compounds with affinity to biomolecular targets. Success in this endeavor lies in sampling diverse chemical libraries. However, current DELs tend to be deficient in C(sp3) carbon counts. We report unique solutions to the challenge of increasing both the chemical diversity of these libraries and their C(sp3) carbon counts by merging Ni/photoredox dual catalytic C(sp2)–C(sp3) cross-coupling as well as photoredox-catalyzed radical/polar crossover alkylation protocols with DELs. The successful integration of multiple classes of radical sources enables the rapid incorporation of a diverse set of alkyl fragments.
Oliver Plettenburg Isr. J. Chem., 2019, 59, 29-36 https://doi.org/10.1002/ijch.201900007AbstractIn early times, the drug discovery process was mainly based on profiling of new medications in in vivo systems. In the absence of knowledge on enzymatic structures, biochemical signaling or precise cellular processes, researchers were dependent on knowledgeable readout of phenotypic effects in living animals.
Bingbing Shi; Yuqing Deng; Xiaoyu Li ACS Comb. Sci., 2019, 21, 5, 345-349 https://doi.org/10.1021/acscombsci.9b00011AbstractDNA-encoded chemical libraries (DELs) have become an important ligand discovery technology in biomedical research and drug discovery. DELs can be comprised of hundreds of millions to billions of candidate molecules and provide outstanding chemical diversity for discovering novel ligands and inhibitors for a large variety of biological targets. However, in most cases, DELs are selected against purified and immobilized proteins based on binding affinity. The development and application of DELs to more complex biological targets requires selection methods compatible with nonimmobilized and unpurified proteins. Here, we describe an approach using polymerase-based extension and target-directed photo-cross-linking and its application to the interrogation of a solution-phase protein target, carbonic anhydrase II.
Santanu Mondal; Paul R. Thompson Acc. Chem. Res., 2019, 52, 818−832 https://doi.org/10.1021/acs.accounts.9b00024AbstractProteins are well-known to undergo a variety of post-translational modifications (PTMs). One such PTM is citrullination, an arginine modification that is catalyzed by a group of hydrolases called protein arginine deiminases (PADs). Hundreds of proteins are known to be citrullinated and hypercitrullination is associated with autoimmune diseases including rheumatoid arthritis (RA), lupus, ulcerative colitis (UC), Alzheimer’s disease, multiple sclerosis (MS), and certain cancers. In this Account, we summarize our efforts to understand the structure and mechanism of the PADs and to develop small molecule chemical probes of protein citrullination. PAD activity is highly regulated by calcium. Structural studies with PAD2 revealed that calcium-binding occurs in a stepwise fashion and induces a series of dramatic conformational changes to form a catalytically competent active site. These studies also identified the presence of a calcium-switch that controls the overall calcium-dependence and a gatekeeper residue that shields the active site in the absence of calcium. Using biochemical and site-directed mutagenesis studies, we identified the key residues (two aspartates, a cysteine, and a histidine) responsible for catalysis and proposed a general mechanism of citrullination. Although all PADs follow this mechanism, substrate binding to the thiolate or thiol form of the enzyme varies for different isozymes. Substrate-specificity studies revealed that PADs 1–4 prefer peptidyl-arginine over free arginine and certain citrullination sites on a peptide substrate. Using high-throughput screening and activity-based protein profiling (ABPP), we identified several reversible (streptomycin, minocycline, and chlorotetracycline) and irreversible (streptonigrin, NSC 95397) PAD-inhibitors. Screening of a DNA-encoded library and lead-optimization led to the development of GSK199 and GSK484 as highly potent PAD4-selective inhibitors. Furthermore, use of an electrophilic, cysteine-targeted haloacetamidine warhead to mimic the guanidinium group in arginine afforded several mechanism-based pan-PAD-inhibitors including Cl-amidine and BB-Cl-amidine. These compounds are highly efficacious in various animal models, including those mimicking RA, UC, and lupus. Structure–activity relationships identified numerous covalent PAD-inhibitors with different bioavailability, in vivo stability, and isozyme-selectivity (PAD1-selective: D-Cl-amidine; PAD2-selective: compounds 16–20; PAD3-selective: Cl4-amidine; and PAD4-selective: TDFA). Finally, this Account describes the development of PAD-targeted and citrulline-specific chemical probes. While PAD-targeted probes were utilized for identifying off-targets and developing high-throughput inhibitor screening platforms, citrulline-specific probes enabled the proteomic identification of novel diagnostic biomarkers of hypercitrullination-related autoimmune diseases.
William C. Reisdorf; Qing Xie; Xin Zeng; Wensheng Xie; Neetu Rajpal; Bao Hoang; Mark E. Burgert; Vinod Kumar; Mark R. Hurle; Deepak K. Rajpal; Sarah O’Donnell; Thomas T. MacDonald; Anna Vossenkämper; Lin Wang; Mike Reilly; Bart J. Votta; Yolanda Sanchez; Pankaj Agarwal PLoS ONE, 2019, 14(4), e0215033 https://doi.org/10.1371/journal.pone.0215033AbstractEpoxyeicosatrienoic acids (EETs) are signaling lipids produced by cytochrome P450 epoxygenation of arachidonic acid, which are metabolized by EPHX2 (epoxide hydrolase 2, alias soluble epoxide hydrolase or sEH). EETs have pleiotropic effects, including anti-inflammatory activity. Using a Connectivity Map (CMAP) approach, we identified an inverse-correlation between an exemplar EPHX2 inhibitor (EPHX2i) compound response and an inflammatory bowel disease patient-derived signature. To validate the gene-disease link, we tested a pre-clinical tool EPHX2i (GSK1910364) in a mouse disease model, where it showed improved outcomes comparable to or better than the positive control Cyclosporin A. Up-regulation of cytoprotective genes and down-regulation of proinflammatory cytokine production were observed in colon samples obtained from EPHX2i-treated mice. Follow-up immunohistochemistry analysis verified the presence of EPHX2 protein in infiltrated immune cells from Crohn’s patient tissue biopsies. We further demonstrated that GSK2256294, a clinical EPHX2i, reduced the production of IL2, IL12p70, IL10 and TNFα in both ulcerative colitis and Crohn's disease patient-derived explant cultures. Interestingly, GSK2256294 reduced IL4 and IFNγ in ulcerative colitis, and IL1β in Crohn's disease specifically, suggesting potential differential effects of GSK2256294 in these two diseases. Taken together, these findings suggest a novel therapeutic use of EPHX2 inhibition for IBD.
Christopher J. Gerry; Zhenhua Yang; Michele Stasi; Stuart L. Schreiber Org. Lett., 2019, 21, 5, 1325-1330 https://doi.org/10.1021/acs.orglett.9b00017AbstractThe limited scope of DNA-compatible chemistry restricts the types of chemical features that can be incorporated into DNA-encoded libraries (DELs). Here, a method to synthesize DNA-conjugated polycyclic isoxazolidines via a [3+2] nitrone–olefin cycloaddition is described. The reaction is compatible with many olefin-containing substrates and diverse N-alkylhydroxylamines. The ability to perform subsequent DNA ligation and PCR amplification was also confirmed. This methodology facilitates the synthesis of DELs containing topographically complex compounds with under-explored chemical features.
An T. H. Le; Svetlana M. Krylova; Mirzo Kanoatov; Shrey Desai; Sergey N. Krylov Angew. Chem. Int. Ed. Engl., 2019, 58(9), 2739-2743 https://doi.org/10.1002/anie.201812974AbstractSelection of aptamers from oligonucleotide libraries currently requires multiple rounds of alternating steps of partitioning of binders from nonbinders and enzymatic amplification of all collected oligonucleotides. Here we report a highly‐practical solution for reliable one‐step selection of aptamers. We introduce partitioning by Ideal‐Filter Capillary Electrophoresis (IFCE) in which binders and nonbinders move in the opposite directions. The efficiency of IFCE‐based partitioning reaches 10^9, which is ten million times higher than that of typical solid‐phase partitioning methods. One step of IFCE‐based partitioning is sufficient for selection of a high‐affinity aptamer pool for a protein target. Partitioning by IFCE promises to become an indispensable tool for fast and robust selection of binders from different types of oligonucleotide libraries.
Markus Follmann; Hans Briem; Andreas Steinmeyer; Alexander Hillisch; Monika H. Schmitt; Helmut Haning; Heinrich Meier Drug Discov. Today, 2019, 24(3), 668-672 https://doi.org/10.1016/j.drudis.2018.12.003AbstractPharmaceutical companies often refer to ‘screening their library’ when performing high-throughput screening (HTS) on a corporate compound collection to identify lead structures for small-molecule drug discovery programs. Characteristics of such a library, including the size, chemical space covered, and physicochemical properties, often determine the success of a screening campaign. Therefore, strategies to maintain and enhance the overall quality of screening collections are crucial to stay competitive and to cope with the ‘novelty erosion’ that is observed gradually. The Next Generation Library Initiative (NGLI), the enhancement of Bayer’s HTS collection by 500 000 newly designed compounds within 5 years, is addressing exactly this challenge. Here, we describe this collaborative project, which involves all internal medicinal chemists in a crowd-sourcing approach, as well as selected external partners, to reach this ambitious goal.
Xuan Wang; Hui Sun; Jiaxiang Liu; Wenge Zhong; Mingqiang Zhang; Hu Zhou; Dongcheng Dai; Xiaojie Lu Org. Lett., 2019, 21, 3, 719-723 https://doi.org/10.1021/acs.orglett.8b03926AbstractOptimal conditions for palladium-promoted Heck reaction on DNA were developed with good to excellent conversions. Versatility with either DNA-conjugated styrene/acrylamide or aryl iodide and a broad substrate scope of the corresponding coupling partners were established. Furthermore, robustness of the Heck reaction conditions on single-strand DNA and feasibility for DNA-encoded library production were demonstrated.
Alessandro Sannino; Elena Gabriele; Martina Bigatti; Sara Mulatto; Jacopo Piazzi; Jörg Scheuermann; Dario Neri; Etienne J. Donckele; Florent Samain ChemBioChem, 2019, 20(7), 955-962 https://doi.org/10.1002/cbic.201800766AbstractDNA‐Encoded chemical libraries are often used for the discovery of ligands against protein targets of interest. These large collections of DNA‐barcoded chemical compounds are typically screened by using affinity capture methodologies followed by PCR amplification and DNA sequencing procedures. However, the performance of individual steps in the selection procedures has been scarcely investigated so far. Here, we describe a quantitative analysis of selection experiments, using three ligands with different affinity to carbonic anhydrase IX as model compounds. In a first set of experiments, we used quantitative PCR (qPCR) procedures, in order to evaluate the recovery and selectivity for affinity capture procedures performed on different solid phase supports, which are commonly used for library screening. In a second step, we used both qPCR and analysis of DNA sequencing results, in order to assess the recovery and selectivity of individual carbonic anhydrase IX ligands in a library, containing 360’000 compounds. Collectively, our study reveals that selection procedures can be efficient for ligands with submicromolar dissociation constant to the target protein of interest, but also that selection performance dramatically drops when 10^4 copies per library member are used as input.
Joerg Holenz; Patrick Stoy Bioorg. Med. Chem. Lett., 2019, 29(4), 517-524 https://doi.org/10.1016/j.bmcl.2018.12.001AbstractLead Generation represents a critical drug discovery phase where chemical starting points and their respective mechanism of action, quality, and potential liabilities are largely predefined. Recent advances such as DNA-encoded libraries or fragment-, chemical biology-, and virtual screening-based approaches are today as common as traditional High Throughput Screening. Innovations in characterizing lead quality have allowed more informed decision-making by discovery teams. The key challenge today is to individually tailor the right mix of methods for each project to facilitate data integration with the purpose of creating multiple high-quality lead series, ultimately translating to reduced chemistry-related pipeline attrition.
Stefanie Geigle; Dnyaneshwar B. Rasale; Dennis Gillingham Helv. Chim. Acta., 2019, 102(1), e1800183 https://doi.org/10.1002/hlca.201800183AbstractWe report here the synthesis and catalytic evaluation in DNA alkylation of a series of water‐soluble copper complexes bearing N‐heterocyclic carbene (NHC) ligands. The NHC ligands were chosen to cover the gamut of commonly used scaffold variations, but in many cases, copper complexes could not be obtained or were unstable. Nevertheless, we identified several complexes that were both stable and catalytically active. Our studies provide guidance and starting scaffolds for any researchers interested in aqueous copper(I) catalysis. A key practical aspect of our findings is that azide‐bearing copper‐NHC complexes are excellent substrates for the azide‐alkyne cycloaddition, which allows late‐stage tailoring of the copper complexes.
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto ACS Chem. Biol., 2019, 14, 1, 37-49 https://doi.org/10.1021/acschembio.8b00866AbstractThe importance of DDR1 in renal fibrosis has been shown via gene knockout and use of antisense oligonucleotides; however, these techniques act via a reduction of DDR1 protein while we prove the therapeutic potential of inhibiting DDR1 phosphorylation with a small molecule. To date, efforts to generate a selective small-molecule to specifically modulate the activity of DDR1 in an in vivo model have been unsuccessful. We performed parallel DNA encoded library screens against DDR1 and DDR2, and discovered a chemical series that is highly selective for DDR1 over DDR2. Structure-guided optimization efforts yielded the potent DDR1 inhibitor 2.45, which possesses excellent kinome selectivity (including 64-fold selectivity over DDR2 in a biochemical assay), a clean in vitro safety profile, and favorable pharmacokinetic and physicochemical properties. As desired, compound 2.45 modulates DDR1 phosphorylation in vitro as well as prevents collagen-induced activation of renal epithelial cells expressing DDR1. Compound 2.45 preserves renal function and reduces tissue damage in Col4a3−/− mice (the preclinical mouse model of Alport syndrome) when employing a therapeutic dosing regime, indicating the real therapeutic value of selectively inhibiting DDR1 phosphorylation in vivo. Our results may have wider significance as Col4a3−/− mice also represent a model for chronic kidney disease, a disease which affects 10% of the global population.
Eduardo de Pedro Beato; Julián Priego; Adrián Gironda-Martínez; Fernando González; Jesús Benavides; Jesús Blas; María Dolores Martín-Ortega; Miguel Ángel Toledo; Jesús Ezquerra; Alicia Torrado ACS Comb. Sci., 2019, 21, 2, 69-74 https://doi.org/10.1021/acscombsci.8b00142AbstractDNA-encoded library technology (ELT) has emerged in the pharmaceutical industry as a powerful tool for hit and lead generation. Over the last 10 years, a number of DNA-compatible chemical reactions have been published and used to synthesize libraries. Among the most commonly used reactions in medicinal chemistry is the C–N bond formation, and its application to DNA-encoded library technology affords an alternative approach to identify high-affinity binders for biologically relevant protein targets. Herein we report a newly developed Pd-promoted C–N cross coupling reaction between DNA-conjugated aryl bromides and a wide scope of arylamines in good to excellent yields. The mild reaction conditions should facilitate the synthesis of novel DNA-encoded combinatorial libraries.
Mikael Madsen; Kurt V. Gothelf Chem. Rev., 2019, 119, 6384−6458 https://doi.org/10.1021/acs.chemrev.8b00570AbstractThe predictable nature of DNA interactions enables the programmable assembly of highly advanced 2D and 3D DNA structures of nanoscale dimensions. The access to ever larger and more complex structures has been achieved through decades of work on developing structural design principles. Concurrently, an increased focus has emerged on the applications of DNA nanostructures. In its nature, DNA is chemically inert and nanostructures based on unmodified DNA mostly lack function. However, functionality can be obtained through chemical modification of DNA nanostructures and the opportunities are endless. In this review, we discuss methodology for chemical functionalization of DNA nanostructures and provide examples of how this is being used to create functional nanodevices and make DNA nanostructures more applicable. We aim to encourage researchers to adopt chemical modifications as part of their work in DNA nanotechnology and inspire chemists to address current challenges and opportunities within the field.
Linlin Tang; Jialian Tan; Yi Xiang; Zhubo Li; Chengde Mao; Hua Zuo Chinese Sci. Bull., 2019, 64, 10, 1076-1083 https://doi.org/10.1360/N972018-00892AbstractDNA has the unique chemical and physical characteristics as a carrier of genetic information. With the development of structural DNA nanotechnology, it has been recognized that DNA can perform versatile functions with promising applications in nanomachine, computing, sensing and nanoarchitectures. In addition, the chemical incorporation with various functional groups brings in a diverse range of additional properties to DNA molecules for broadened applications. Therefore, post-synthetic modification of DNA has been widely used to introduce additional functionalities to DNA molecules, for example, design and synthesis of DNA-encoded library, preparation of DNA-based probes for biosensing, improving binding activities of DNA aptamers, etc. The modification efficiency is determined by both the intrinsic factors of the molecules involved (DNA and reagents) and the external factors (experimental conditions). However, previous studies on the modification focused on the optimization of reaction conditions. To determine the important intrinsic factors that affect modification efficiency, we have conducted a systematic study on the reaction with a model system— acylation of amino-DNA. Nine amino-modified DNAs with two different DNA conformations (a–i) were reacted with two carboxylic acids with different molecule weights (12-oxo-2,5,8,11-tetraoxapentadecan-15-oic acid, MW 264.1209, and 30-oxo-2,5,8,11,14,17,20,23,26,29-decaoxatritriacontan-33-oic acid, MW 528.2782), respectively, and characterized by polyacrylamide gel electrophoresis (PAGE). The bands were analyzed and calculated by Image J. In the present study, three factors are considered for the design. First, acylation is one of the most widely methods used for chemical modifications of DNA. The acylation reactions afford amides and these amides binding to DNA form more structurally complex and diverse DNA building blocks. Therefore, we explored the acylation reaction between amino-modified DNA with carboxylic acids. Second, two DNA conformations (single-stranded random coils, ssDNAs, and rigid, double-stranded DNA duplexes, dsDNA) were included. Single-stranded DNAs are more flexible than rigid DNA duplexes and thus can provide more variability in terms of modulating orientation of the reaction group to facilitate reactions. Third, the two small organic molecules with a varying number of polyethylene glycol (PEG) units were used, because PEG has been widely used in the field of chemical modification of biomacromolecules due to its excellent properties including a wide range of solubility, predominant bio-compatibility, good stability, low toxicity and no irritation, etc. We, therefore, systematically studied the DNA modification reaction to investigate the influences of two intrinsic factors (MW and conformation) on the reaction. The primary factor for DNA-NH2 acylation reaction is molecular weight (MW). The higher the MW of the regents (both small molecules and DNAs) are, the slower the reactions are. DNA conformation plays a much minor role in the reaction. Compared with rigid dsDNA duplexes, flexible ssDNA chains facilitate reactions. Based on the current study, we would suggest for post-synthetic modification of DNA: (1) Keep the molecular weights of all reagents (both small molecules and DNA molecules) to the minimal in the design to allow fast diffusion. (2) Keep the DNA strands near the modification location to be single stranded to facilitate orientation adjustment. If high molecular weight and rigid conformation (duplex) are not avoidable, reaction time and/or reagent concentration should be elongated to ensure the modification efficiency.
Marcello Anzola; Nicolas Winssinger Chem. Eur. J., 2019, 24, 2, 334-342 https://doi.org/10.1002/chem.201804283AbstractHere, the synthesis of a RuII photocatalyst by light‐directed oligonucleotide‐templated ligation reaction is described. The photocatalyst was found to have tremendous potential for signal amplification with >15000 turnovers measured in 9 hours. A templated reaction was used to turn on the activity of this ruthenium(II) photocatalyst in response to a specific DNA sequence. The photocatalysis of the ruthenium(II) complex was harnessed to uncage a new precipitating dye that is highly fluorescent and photostable in the solid state. This reaction was used to discriminate between different DNA analytes based on localization of the precipitate as well as for in cellulo miRNA detection. Finally, a bipyridine ligand functionalized with two different peptide nucleic acid (PNA) sequences was shown to enable template‐mediated ligation (turn on of the ruthenium(II) photocatalysis) and recruitment of substrate for templated photocatalysis.
Stuart L. Schreiber Isr. J. Chem., 2019, 59, 52-59 https://doi.org/10.1002/ijch.201800113AbstractA simplistic view of drug discovery is that it begins, most often using “model organisms”, with biological inferences of a disease that suggest the need to interfere with some activity, function or process. An enzyme should be inhibited or a pathogen should be killed. Chemical experimentation yields the desired inhibitor, and clinical investigations then test the underlying hypothesis in humans. If the stars align, an effective drug emerges
John C. Faver; Kevin Riehle; David R. Lancia; Jr.; Jared B. J. Milbank; Christopher S. Kollmann; Nicholas Simmons; Zhifeng Yu; Martin M. Matzuk ACS Comb. Sci., 2019, 21, 2, 75-82 https://doi.org/10.1021/acscombsci.8b00116AbstractDNA-encoded chemical libraries (DELs) provide a high-throughput and cost-effective route for screening billions of unique molecules for binding affinity for diverse protein targets. Identifying candidate compounds from these libraries involves affinity selection, DNA sequencing, and measuring enrichment in a sample pool of DNA barcodes. Successful detection of potent binders is affected by many factors, including selection parameters, chemical yields, library amplification, sequencing depth, sequencing errors, library sizes, and the chosen enrichment metric. To date, there has not been a clear consensus about how enrichment from DEL selections should be measured or reported. We propose a normalized z-score enrichment metric using a binomial distribution model that satisfies important criteria that are relevant for analysis of DEL selection data. The introduced metric is robust with respect to library diversity and sampling and allows for quantitative comparisons of enrichment of n-synthons from parallel DEL selections. These features enable a comparative enrichment analysis strategy that can provide valuable information about hit compounds in early stage drug discovery.
Glen Liszczak; Tom W. Muir Angew. Chem. Int. Ed. Engl., 2019, 58(13), 4144-4162 https://doi.org/10.1002/anie.201808956AbstractThe emergence of high‐throughput DNA sequencing technologies sparked an immediate revolution in the field of genomics that has rippled into many branches of the life and physical sciences. The remarkable sensitivity, specificity, throughput, and multiplexing capacity that are inherent to massively parallel DNA sequencing have since motivated its use as a broad‐spectrum molecular counter in small molecule and peptide‐based inhibitor discovery, high‐throughput biochemistry, protein and cellular detection and diagnostics, and even materials science. A key aspect of extrapolating DNA sequencing to 'non‐traditional' applications is the underlying need to append nucleic acid barcodes to entities of interest. In this review, we describe the chemical and biochemical approaches that have enabled facile nucleic acid barcoding of proteinaceous and non‐proteinaceous materials, and provide exciting examples of downstream technologies that have been made possible by DNA‐encoded molecules. Considering that commercially available high‐throughput sequencers were first released less than 15 years ago, we believe related applications will continue to mature for years to come, and close by proposing potential new frontiers to support this assertion.
Lik Hang Yuen; Srikanta Dana; Yu Liu; Samuel I Bloom; Ann-Gerd Thorsell; Dario Neri; Anthony J. Donato; Dmitri B. Kireev; Herwig Schüler; Raphael M. Franzini J. Am. Chem. Soc., 2019, 141, 13, 5169-5181 https://doi.org/10.1021/jacs.8b08039AbstractDNA-encoded chemical libraries are increasingly used in pharmaceutical research because they enable the rapid discovery of synthetic protein ligands. Here we explored whether target-class focused DNA-encoded chemical libraries can be cost-effective tools to achieve robust screening productivity for a series of proteins. The study revealed that a DNA-encoded library designed for NAD+-binding pockets (NADEL) effectively sampled the chemical binder space of enzymes with ADP-ribosyltransferase activity. The extracted information directed the synthesis of inhibitors for several enzymes including PARP15 and SIRT6. The high dissimilarity of NADEL screening fingerprints for different proteins translated into inhibitors that showed selectivity for their target. The discovery of patterns of enriched structures for six out of eight tested proteins is remarkable for a library of 58,302 DNA-tagged structures and illustrates the prospect of focused DNA-encoded libraries as economic alternatives to large library platforms.
Matthias Gehringer; Stefan A. Laufer J. Med. Chem., 2019, 62, 12, 5673-5724 https://doi.org/10.1021/acs.jmedchem.8b01153AbstractTargeted covalent inhibitors (TCIs) are designed to bind poorly conserved amino acids by means of reactive groups, the so-called warheads. Currently, targeting noncatalytic cysteine residues with acrylamides and other α,β-unsaturated carbonyl compounds is the predominant strategy in TCI development. The recent ascent of covalent drugs has stimulated considerable efforts to characterize alternative warheads for the covalent-reversible and irreversible engagement of noncatalytic cysteine residues as well as other amino acids. This Perspective article provides an overview of warheads—beyond α,β-unsaturated amides—recently used in the design of targeted covalent ligands. Promising reactive groups that have not yet demonstrated their utility in TCI development are also highlighted. Special emphasis is placed on the discussion of reactivity and of case studies illustrating applications in medicinal chemistry and chemical biology.
Nienke Moret; Nicholas A. Clark; Marc Hafner; Yuan Wang; Eugen Lounkine; Mario Medvedovic; Jinhua Wang; Nathanael Gray; Jeremy Jenkins; Peter K. Sorger Cell Chem. Biol., 2019, 26, 5, 765-777 https://doi.org/10.1016/j.chembiol.2019.02.018AbstractLibraries of well-annotated small molecules have many uses in chemical genetics, drug discovery, and therapeutic repurposing. Multiple libraries are available, but few data-driven approaches exist to compare them and design new libraries. We describe an approach to scoring and creating libraries based on binding selectivity, target coverage, and induced cellular phenotypes as well as chemical structure, stage of clinical development, and user preference. The approach, available via the online tool http://www.smallmoleculesuite.org, assembles sets of compounds with the lowest possible off-target overlap. Analysis of six kinase inhibitor libraries using our approach reveals dramatic differences among them and led us to design a new LSP-OptimalKinase library that outperforms existing collections in target coverage and compact size. We also describe a mechanism of action library that optimally covers 1,852 targets in the liganded genome. Our tools facilitate creation, analysis, and updates of both private and public compound collections.
Yichao Huang; Mareike Margarete Wiedmann; Hiroaki Suga Chem. Rev., 2019, 119, 17, 10360-10391 https://doi.org/10.1021/acs.chemrev.8b00430AbstractThe past two decades have witnessed the emergence of macrocycles, including macrocyclic peptides, as a promising yet underexploited class of de novo drug candidates. Both rational/computational design and in vitro display systems have contributed tremendously to the development of cyclic peptide binders of either traditional targets such as cell-surface receptors and enzymes or challenging targets such as protein–protein interaction surfaces. mRNA display, a key platform technology for the discovery of cyclic peptide ligands, has become one of the leading strategies that can generate natural-product-like macrocyclic peptide binders with antibody-like affinities. On the basis of the original cell-free transcription/translation system, mRNA display is highly evolvable to realize its full potential by applying genetic reprogramming and chemical/enzymatic modifications. In addition, mRNA display also allows the follow-up hit-to-lead development using high-throughput focused affinity maturation. Finally, mRNA-displayed peptides can be readily engineered to create chemical conjugates based on known small molecules or biologics. This review covers the birth and growth of mRNA display and discusses the above features of mRNA display with success stories and future perspectives and is up to date as of August 2018.
Jiankun Lyu; Sheng Wang; Trent E. Balius; Isha Singh; Anat Levit; Yurii S. Moroz; Matthew J. O’Meara; Tao Che; Enkhjargal Algaa; Kateryna Tolmachova; Andrey A. Tolmachev; Brian K. Shoichet; Bryan L. Roth; John J. Irwin Nature, 2019, 566, 224-229 https://doi.org/10.1038/s41586-019-0917-9AbstractDespite intense interest in expanding chemical space, libraries containing hundreds-of-millions to billions of diverse molecules have remained inaccessible. Here we investigate structure-based docking of 170 million make-on-demand compounds from 130 well-characterized reactions. The resulting library is diverse, representing over 10.7 million scaffolds that are otherwise unavailable. For each compound in the library, docking against AmpC β-lactamase (AmpC) and the D4 dopamine receptor were simulated. From the top-ranking molecules, 44 and 549 compounds were synthesized and tested for interactions with AmpC and the D4 dopamine receptor, respectively. We found a phenolate inhibitor of AmpC, which revealed a group of inhibitors without known precedent. This molecule was optimized to 77 nM, which places it among the most potent non-covalent AmpC inhibitors known. Crystal structures of this and other AmpC inhibitors confirmed the docking predictions. Against the D4 dopamine receptor, hit rates fell almost monotonically with docking score, and a hit-rate versus score curve predicted that the library contained 453,000 ligands for the D4 dopamine receptor. Of 81 new chemotypes discovered, 30 showed submicromolar activity, including a 180-pM subtype-selective agonist of the D4 dopamine receptor.
Zhengrong Zhu; LaShadric C. Grady; Yun Ding; Kenneth E. Lind; Christopher P. Davie; Christopher B. Phelps; Ghotas Evindar SLAS Discov., 2019, 24(2), 169-174 https://doi.org/10.1177/2472555218808454AbstractDNA-encoded libraries (DELs) have been broadly applied to identify chemical probes for target validation and lead discovery. To date, the main application of the DEL platform has been the identification of reversible ligands using multiple rounds of affinity selection. Irreversible (covalent) inhibition offers a unique mechanism of action for drug discovery research. In this study, we report a developing method of identifying irreversible (covalent) ligands from DELs. The new method was validated by using 3C protease (3CP) and on-DNA irreversible tool compounds (rupintrivir derivatives) spiked into a library at the same concentration as individual members of that library. After affinity selections against 3CP, the irreversible tool compounds were specifically enriched compared with the library members. In addition, we compared two immobilization methods and concluded that microscale columns packed with the appropriate affinity resin gave higher tool compound recovery than magnetic beads.
Michael D. Shultz J. Med. Chem., 2019, 62, 4, 1701–1714 https://doi.org/10.1021/acs.jmedchem.8b00686AbstractTwo decades have passed since the rule of five ushered in the concept of “drug-like” properties. Attempts to quantify, correlate, and categorize molecules based on Ro5 parameters evolved into the introduction of efficiency metrics with far reaching consequences in decision making by industry leaders and scientists seeking to discover new medicines. Examination of oral drug parameters approved before and after the original Ro5 analysis demonstrates that some parameters such as clogP and HBD remained constant while the cutoffs for parameters such as molecular weight and HBA have increased substantially over the past 20 years. The time dependent increase in the molecular weight of oral drugs during the past 20 years provides compelling evidence to disprove the hypothesis that molecular weight is a “drug-like” property. This analysis does not validate parameters that have not changed as being “drug-like” but instead calls into question the entire hypothesis that “drug-like” properties exist.
Solange Moll; Alexis Desmoulièrec; Marcus J. Moeller; Jean-Claude Pache; Laura Badi; Filippo Arcadu; Hans Richter; Alexander Satz; Sabine Uhles; Andrea Cavalli; Faye Drawnel; Leonardo Scapozza; Marco Prunotto BBA-Mol. Cell. Res., 2019, 1866, 11, 118474 https://doi.org/10.1016/j.bbamcr.2019.04.004AbstractDiscoidin domain receptor1 (DDR1) is a collagen activated receptor tyrosine kinase and an attractive anti-fibrotic target. Its expression is mainly limited to epithelial cells located in several organs including skin, kidney, liver and lung. DDR1's biology is elusive, with unknown downstream activation pathways; however, it may act as a mediator of the stromal-epithelial interaction, potentially controlling the activation state of the resident quiescent fibroblasts. Increased expression of DDR1 has been documented in several types of cancer and fibrotic conditions including skin hypertrophic scars, idiopathic pulmonary fibrosis, cirrhotic liver and renal fibrosis. The present review article focuses on: a) detailing the evidence for a role of DDR1 as an anti-fibrotic target in different organs, b) clarifying DDR1 tissue distribution in healthy and diseased tissues as well as c) exploring DDR1 protective mode of action based on literature evidence and co-authors experience; d) detailing pharmacological efforts attempted to drug this subtle anti-fibrotic target to date.
Christopher R. Wellaway; Dominique Amans; Paul Bamborough; Heather Barnett; Rino A. Bit; Jack A. Brown; Neil R. Carlson; Chun-wa Chung; Anthony W. J. Cooper; Peter D. Craggs; Robert P. Davis; Tony W. Dean; John P. Evans; Laurie Gordon; Isobel L. Harada; David J. Hirst; Philip G. Humphreys; Katherine L. Jones; Antonia J. Lewis; Matthew J. Lindon; Dave Lugo; Mahnoor Mahmood; Scott McCleary; Patricia Medeiros; Darren J. Mitchell; Michael O’Sullivan; Armelle Le Gall; Vipulkumar K. Patel; Chris Patten; Darren L. Poole; Rishi R. Shah; Jane E. Smith; Kayleigh A. J. Stafford; Pamela J. Thomas; Mythily Vimal; Ian D. Wall; Robert J. Watson; Natalie Wellaway; Gang Yao; Rab K. Prinjha PDBJ - 6TPZhttps://pdbj.org/mine/summary/6tpz Abstract Bromodomain-containing protein 4, 1,2-ETHANEDIOL, 5-[1-(1,3-dimethoxypropan-2-yl)-5-morpholin-4-yl-benzimidazol-2-yl]-1,3-dimethyl-pyridin-2-one
Christopher R. Wellaway; Dominique Amans; Paul Bamborough; Heather Barnett; Rino A. Bit; Jack A. Brown; Neil R. Carlson; Chun-wa Chung; Anthony W. J. Cooper; Peter D. Craggs; Robert P. Davis; Tony W. Dean; John P. Evans; Laurie Gordon; Isobel L. Harada; David J. Hirst; Philip G. Humphreys; Katherine L. Jones; Antonia J. Lewis; Matthew J. Lindon; Dave Lugo; Mahnoor Mahmood; Scott McCleary; Patricia Medeiros; Darren J. Mitchell; Michael O’Sullivan; Armelle Le Gall; Vipulkumar K. Patel; Chris Patten; Darren L. Poole; Rishi R. Shah; Jane E. Smith; Kayleigh A. J. Stafford; Pamela J. Thomas; Mythily Vimal; Ian D. Wall; Robert J. Watson; Natalie Wellaway; Gang Yao; Rab K. Prinjha PDBJ - 6TPYhttps://pdbj.org/mine/summary/6tpy Abstract Bromodomain-containing protein 4, 1,2-ETHANEDIOL, 1,3-dimethyl-5-[1-(oxan-4-ylmethyl)benzimidazol-2-yl]pyridin-2-one
Christopher R. Wellaway; Dominique Amans; Paul Bamborough; Heather Barnett; Rino A. Bit; Jack A. Brown; Neil R. Carlson; Chun-wa Chung; Anthony W. J. Cooper; Peter D. Craggs; Robert P. Davis; Tony W. Dean; John P. Evans; Laurie Gordon; Isobel L. Harada; David J. Hirst; Philip G. Humphreys; Katherine L. Jones; Antonia J. Lewis; Matthew J. Lindon; Dave Lugo; Mahnoor Mahmood; Scott McCleary; Patricia Medeiros; Darren J. Mitchell; Michael O’Sullivan; Armelle Le Gall; Vipulkumar K. Patel; Chris Patten; Darren L. Poole; Rishi R. Shah; Jane E. Smith; Kayleigh A. J. Stafford; Pamela J. Thomas; Mythily Vimal; Ian D. Wall; Robert J. Watson; Natalie Wellaway; Gang Yao; Rab K. Prinjha PDBJ - 6TPXhttps://pdbj.org/mine/summary/6tpx Abstract Bromodomain-containing protein 4, 2-(3,5-dimethyl-4-oxidanyl-phenyl)-1-[(1-ethanoylpiperidin-4-yl)methyl]-~{N}-methyl-benzimidazole-5-carboxamide, 1,2-ETHANEDIOL
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FILhttps://pdbj.org/mine/summary/6fil Abstract Epithelial discoidin domain-containing receptor 1, 2-[8-(2~{H}-indazol-5-ylcarbonyl)-4-oxidanylidene-1-phenyl-1,3,8-triazaspiro[4.5]decan-3-yl]-~{N}-methyl-ethanamide, IODIDE ION
Hailong Li; Zhen Sun; Wenting Wu; Xuan Wang; Mingqiang Zhang; Xiaojie Lu; Wenge Zhong; Dongcheng Dai Org. Lett., 2018, 20, 22, 7186-7191 https://doi.org/10.1021/acs.orglett.8b03114AbstractThe synthesis of pyridazines on DNA has been developed on the basis of inverse-electron-demand Diels-Alder (IEDDA) reactions of 1,2,4,5-tetrazines. The broad substrate scope is explored. Functionalized pyridazine products are selected for subsequent DNA-compatible Suzuki-Miyaura coupling, acylation, and SNAr substitution reactions, demonstrating the feasibility and versatility of IEDDA reactions for DNA-encoded library synthesis
Jian-Yuan Li; Hongbing Huang Bioconjugate Chem., 2018, 29, 11, 3841-3846 https://doi.org/10.1021/acs.bioconjchem.8b00676AbstractDNA-encoded chemical libraries (DELs) are a cost-effective technology for the discovery of novel chemical probes and drug candidates. A major limiting factor in assembling productive DELs is the availability of DNA-compatible chemical reactions in aqueous media. In an effort to increase the chemical accessibility and structural diversity of small molecules displayed by DELs, we developed a robust SuzukiMiyaura reaction protocol that is compatible with the DNA structures. By employing a water soluble Pdprecatalyst, we developed conditions that allow efficient coupling of DNA-linked aryl halides with a wide variety of boronic acids/esters including heteroaryl boronates
Yu Zhou; Chen Li; Jianzhao Peng; Liangxu Xie; Ling Meng; Qingrong Li; Jianfu Zhang; Xiang David Li; Xin Li; Xuhui Huang; Xiaoyu Li J. Am. Chem. Soc., 2018, 140, 46, 15859-15867 https://doi.org/10.1021/jacs.8b09277AbstractDynamic combinatorial library (DCL) has emerged as an efficient tool for ligand discovery and become an important discovery modality in biomedical research. However, the applications of DCLs have been significantly hampered by low library diversity and limited analytical methods capable of processing large libraries. Here, we report a strategy that has addressed this limitation and can select cooperatively binding small-molecule pairs from large-scale dynamic libraries. Our approach is based on DNA-mediated dynamic hybridization, DNA-encoding, and a photo-cross-linking-based decoding scheme. To demonstrate the generality and performance of this approach, a 10 000-member DNA-encoded dynamic library has been prepared and selected against six protein targets. Specific binders have been identified for each target, and we have validated the biological activities of selected ligands for the targets that are implicated in important cellular functions including protein deacetylation and sumoylation. Notably, a series of novel and selective sirtuin-3 inhibitors have been developed. Our study has circumvented a major obstacle in DCL and may provide a broadly applicable method for ligand discovery against biological targets.
Jonas Boström; Dean G. Brown; Robert J. Young; György M. Keserü Nat. Rev. Drug Discov., 2018, 17(10), 709-727 https://doi.org/10.1038/nrd.2018.116AbstractThe key objectives of medicinal chemistry are to efficiently design and synthesize bioactive compounds that have the potential to become safe and efficacious drugs. Most medicinal chemistry programmes rely on screening compound collections populated by a range of molecules derived from a set of known and robust chemistry reactions. Analysis of the role of synthetic organic chemistry in subsequent hit and lead optimization efforts suggests that only a few reactions dominate. Thus, the uptake of new synthetic methodologies in drug discovery is limited. Starting from the known limitations of reaction parameters, synthesis design tools, synthetic strategies and innovative chemistries, here we highlight opportunities for the expansion of the medicinal chemists' synthetic toolbox. More intense crosstalk between synthetic and medicinal chemists in industry and academia should enable enhanced impact of new methodologies in future drug discovery.
Derek Lowe Med. Chem. Commun., 2018, 9, 1397-1398 https://doi.org/10.1039/c8md90041hAbstractWe're going to need a bigger compound deck…
X.Zhou; P.Pathak; J.Jayawickramarajah Chem. Commun., 2018, 54, 11668-11680 https://doi.org/10.1039/C8CC06716CAbstractWith this Feature Article we review, for the first time, the development of DNA–host conjugates—a nascent yet rapidly growing research focus within the ambit of DNA supramolecular chemistry. Synthetic hosts (such as cyclodextrins, cucurbiturils, and calixarenes) are well-suited to be partnered with DNA, since DNA assembly and host–guest binding both thrive in aqueous media, are largely orthogonal, and exhibit controllable and input-responsive properties. The covalent braiding of these two supramolecular synthons thus leads to advanced self-assemblies and nanostructures with exciting function that range from drug delivery agents to input-triggered switches. The latter class of DNA–host conjugates have been demonstrated to precisely control protein activity, and have also been used as modulable catalysts and versatile biosensors.
Likun Xu; Dongna Zhang; Yuanyuan Dou; Yabin Song; Baogang Wang; Hongquan Wang J. Int. Pharm. Res., 2018, 45, 10, 736-742 http://202.38.153.236:81/Jweb_jipr/CN/10.13220/j.cnki.jipr.2018.10.002AbstractIf lead compounds are obtained by high-throughput screening (HTS) of traditional compound libraries, a process that requires the storage and processing of thousands of compounds and the identification of molecules bound specifically to target proteins will be a formidable challenge. DNA Encoding chemical library (DEL) is a group of small organic molecules that bind to DNA tags in a covalent manner and encode molecules with unique DNA markers. This technique allows the construction and screening of combinatorial libraries on an unprecedented scale, through protein target affinity screening and HTS decoding, to efficiently and cheaply discover many unusual affinity ligands. This paper reviews and discusses the characteristics of DEL technology, its construction and screening methods, as well as its application in academic and industrial fields, and the development of DEL and the prospect of the future.
Bjarke N. Hansen; Kim S. Larsen; Daniel Merkle; Alexei Mihalchuk Natural Computing, 2018, 17, 693-707 https://doi.org/10.1007/s11047-018-9697-7AbstractIn chemistry, synthesis is the process in which a target compound is produced in a step-wise manner from given base compounds. A recent, promising approach for carrying out these reactions is DNA-templated synthesis, since, as opposed to more traditional methods, this approach leads to a much higher effective molarity and makes much desired (sequential) one-pot synthesis possible. With this method, compounds are tagged with DNA sequences and reactions can be controlled by bringing two compounds together via their tags. This leads to new cost optimization problems of minimizing the number of different tags or strands to be used under various conditions. We identify relevant optimization criteria, provide the first computational approach to automatically inferring DNA-templated programs, and obtain efficient optimal and near-optimal results, and also provide a brute-force integer linear programming approach for complete solutions to smaller instances.
Dominik K. Kölmel; Richard P. Loach; Thomas Knauber; Mark E. Flanagan ChemMedChem, 2018, 13, 2159-2165 https://doi.org/10.1002/cmdc.201800492AbstractA new procedure for the photoredox‐mediated conjugate addition of radicals that can be conveniently generated from a‐amino acids, to DNA‐tagged Michael acceptors and styrenes is presented. This C(sp3)‐C(sp3) coupling tolerates a broad array of structurally diverse radical precursors, including all of the 20 proteinogenic amino acids. Importantly, this reaction proceeds under mild conditions and in DNA‐compatible aqueous media. Furthermore, the presented reaction conditions are compatible with DNA, making this reaction platform well suited for the construction of DNA‐encoded libraries. The scope and limitations of the chemistry are discussed herein along with proposals for how this methodology might be utilized to construct DNA‐encoded libraries.
Dennis G. Gillingham; Dnyaneshwar Rasale Chimia, 2018, 72, 777–781 https://doi.org/10.2533/chimia.2018.777AbstractWe present the state-of-the-art in direct RNA modification as well as the challenges that hold back further development of RNA mechanistic probes and medicines. Solid-phase synthesis has revolutionized the synthesis of short DNAs and RNAs. Many open questions in RNA biology are with large long-non-coding RNAs or mRNAs and there is also interest in developing these big RNAs as medicines. Techniques for direct modification will become more important in the coming years and we give a current snapshot of the field here, with a bias towards our own contributions.
Diana I. S. P. Resende; Amalia M. Estévez; André M. Alker; Rainer E. Martin; Hans Peter Wessel Med. J. Chem., 2018, 7(2), 135-144 https://doi.org/10.13171/mjc72/01809051415-wesselAbstractFor the generation of compound libraries for drug discovery a central scaffold containing three exit vectors with defined chirality was devised starting from commercially available tri-O-acetyl-glucal. Surprisingly, the reaction of a 4-O-mesylate with sodium azide did not lead to the expected 4-azido-4-deoxy derivative but to a 3-azido-3-deoxy regioisomer via intermediate epoxide formation. The absolute stereochemical configuration of the final tetrahydropyran building block was proven by X-ray crystallography. This scaffold endowed with a carboxylic acid, a secondary alcohol, and an azide functionality may be connected to a DNA tag at any of the three distinct exit vectors, thus providing ready access to several different compound libraries.
Sven Kochmann; An T. H. Le; Ryan Hili; Sergey N. Krylov Electrophoresis, 2018, 39(23), 2991-2996 https://doi.org/10.1002/elps.201800270AbstractNonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM) is an affinity method for separating binder‐target complexes from nonbinders by gel‐free CE. NECEEM is a promising high‐efficiency method for partitioning protein binders from nonbinders in DNA‐encoded libraries (DEL), such binders are used as “hits” in drug development. It is important to be able to predict the efficiency of NECEEM‐based partitioning, which is the efficiency of collecting binders while removing nonbinders for a specific protein and a specific DEL with a minimum of empirical information. Here, we derive and study the dependence of efficiency of NECEEM‐based partitioning on electrophoretic mobilities of the protein and the DNA moiety in DEL compounds. Our derivation is based upon a previously found relation between the electrophoretic mobility of protein‐binder complex and measured electrophoretic mobilities of the protein and unbound DEL and their estimated sizes. The derivation utilizes the assumption of Gaussian shapes of electrophoretic peaks and the approximation of the efficiency of partitioning by the background of nonbinders – a fraction of nonbinders, which elutes along with protein‐binder complexes. Our results will serve as a guiding tool for planning the NECEEM‐based partitioning of protein binders from non‐binders in DELs. In particular, it can be used to estimate a minimum number of rounds of partitioning required for the desired level of DEL enrichment.
Ghotas Evindar Nat. Chem., 2018, 10(7), 690-691 https://doi.org/10.1038/s41557-018-0094-8AbstractGhotas Evindar, Chemistry Group Leader at GlaxoSmithKline, talks with Nature Chemistry about the advantages of using encoded libraries in drug discovery and the challenges these technologies present.
N/A Nat. Chem., 2018, 10(7), 687 https://doi.org/10.1038/s41557-018-0103-yAbstractEncoded chemical libraries can be used to screen a vast array of compounds against a protein target to identify potent binders. A collection of articles in this issue discuss different methods to increase the chemical space sampled by encoded macrocycle libraries and the advantages that such libraries offer for discovering new drug leads.
Xuan Wang; Hui Sun; Jiaxiang Liu; Dongcheng Dai; Mingqiang Zhang; Hu Zhou; Wenge Zhong; Xiaojie Lu Org. Lett., 2018, 20, 16, 4764-4768 https://doi.org/10.1021/acs.orglett.8b01837AbstractThe first example of DNA-compatible C-H activation reaction between DNA-conjugated acrylamides and aromatic acids has been developed. This new transformation enables aromatic acid, previously considered as monofunctional building block, to act like a bifunctional building block for the DNA encoded library synthesis. The general scope of aromatic acid was established for this new on-DNA C-H activation, which paved the way for its application in combinatorial library preparation.
Charlotte Kielar; Francesco V. Reddavide; Stefan Tubbenhauer; Meiying Cui; Xiaodan Xu; Guido Grundmeier; Yixin Zhang; Adrian Keller Angew. Chem. Int. Ed. Engl., 2018, 57(45), 14873-14877 https://doi.org/10.1002/anie.201806778AbstractThe rational combination of techniques from the fields of nanotechnology, single molecule detection, and lead discovery could provide elegant solutions to enhance the throughput of drug screening. We have synthesized nanoarrays of small pharmacophores on DNA origami substrates that are displayed either as individual ligands or as fragment pairs and thereby reduced the feature size by several orders of magnitude, as compared with standard microarray techniques. Atomic force microscopy‐based single‐molecule detection allowed us to distinguish potent protein‐ligand interactions from weak binders. Several independent binding events, i.e., strong binding, weak binding, symmetric bidentate binding, and asymmetric bidentate binding are directly visualized and evaluated. We apply this method to the discovery of bidentate trypsin binders based on benzamidine paired with aromatic fragments. Pairing of benzamidine with the dye TAMRA results in tenfold enhancement of the trypsin binding yield.
Verena Kunig; Marco Potowski; Anne Gohla; Andreas Brunschweiger Biol. Chem., 2018, 399(7), 691-710 https://doi.org/10.1515/hsz-2018-0119AbstractDNA-encoded compound libraries are a highly attractive technology for discovery of small molecule protein ligands. These compound collections consist of small molecules covalently connected to individual DNA sequences carrying readable information about the compound structure. DNA-tagging allows for efficient synthesis, handling, and interrogation of vast numbers of chemically synthesized, drug-like compounds. They are screened on proteins by an efficient, generic assay based on Darwinian principles of selection. To date, selection of DNA-encoded libraries allowed for identification of numerous bioactive compounds. Some of these compounds uncovered hitherto unknown allosteric binding sites on target proteins; several compounds proved their value as chemical biology probes unravelling complex biology; and first examples of clinical candidates that trace their ancestry to a DNA-encoded library were reported. Thus, DNA-encoded libraries proved their value for the biomedical sciences as a generic technology for the identification of bioactive drug-like molecules numerous times. However, large scale experiments showed that even selection of billions of compounds failed to deliver bioactive compounds for the majority of proteins in an unbiased panel of target proteins. This raises the question of compound library design.
Robert A. Goodnow SLAS Discov., 2018, 23(5), 385-386 https://doi.org/10.1177/2472555218766250AbstractDrug discovery scientists have made great efforts over the years to explore a maximum possible number of chemical structures against a biological target or targets of interest. The advent of automation and miniaturization has enabled significant progress in the high-throughput screening, delivery, and analysis of hundreds of thousands to millions of compounds. This journal has published numerous manuscripts that report on the efficiency of such technologies applied to screening and laboratory sciences. With increased efficiency in any process, one must consider its inputs and outputs. This special collection of SLAS Discovery focuses on DNA-encoded library technology (DELT) to shed light on the new opportunity to make available high numbers of small-molecule structures in the hit-finding process, something that has been a limiting factor in drug discovery. DELT specifically addresses the problem of exploring the interface of chemistry and biology with hundreds of millions of small molecules in an efficient manner that has been neither possible nor feasible before.
Peng Zhao; Sirsendu Bhowmick; Jianchao Yu; Jun Wang Adv. Sci., 2018, 5(9), 1800672 https://doi.org/10.1002/advs.201800672AbstractHighly multiplexed detection of proteins secreted by single cells is always challenging. Herein, a multiplexed in situ tagging technique based on single-stranded DNA encoded microbead arrays and multicolor successive imaging for assaying single-cell secreted proteins with high throughput and high sensitivity is presented. This technology is demonstrated to be capable of increasing the multiplexity exponentially. Upon integration with polydimethylsiloxane microwells, this platform is applied to detect ten immune effector proteins from differentiated single macrophages stimulated with lipopolysaccharide. Significant heterogeneity is observed when the derived human primary macrophages are analyzed. This versatile technology is expected to open new opportunities in systems biology, immune regulation studies, signaling analysis, and molecular diagnostics.
Ryosuke Ueki; Shinsuke Sando MedChem. News, 2018, 28(2), 93-97 https://doi.org/10.14894/medchem.28.2_93AbstractDNA-encoded library (DEL) has attracted much attention as a promising approach for efficient ligand generation. Taking advantage of unique characteristics of DNA, DEL has realized drug discovery using a small molecule library with an unprecedented diversity. This review article summarizes the construction methods for DEL with a particular focus on chemical properties of DNA
Dean G. Brown; Jonas Boström J. Med. Chem., 2018, 61, 21, 9442-9468 https://doi.org/10.1021/acs.jmedchem.8b00675AbstractAn analysis of 66 published clinical candidates from Journal of Medicinal Chemistry has been conducted to shed light on which lead generation strategies are most frequently employed in identifying drug candidates. The most frequent lead generation strategy (producing a drug candidate) was based on starting points derived from previously known compounds (43%) followed by random high throughput screening (29%). The remainder of approaches included focused screening, structure-based drug design (SBDD), fragment-based lead generation (FBLG) and DNA-encoded library screening (DEL). An analysis of physical chemical properties on the hit-to-clinical pairs show an average increase in molecular weight (ΔMW +85) but no change in lipophilicity (ΔclogP -0.2), although exceptions are noted. The majority (>50%) of clinical candidates were found to be structurally very different from their starting point, as well as being more complex. Finally, several reports of non-covalent scaffolds modified by a covalent warhead using SBDD approaches are discussed.
Marian C. Bryan; Naomi S. Rajapaksa J. Med. Chem., 2018, 61, 20, 9030-9058 https://doi.org/10.1021/acs.jmedchem.8b00667AbstractSmall molecule inhibitors targeting autoimmune and inflammatory processes have been an area of intense focus within academia and industry. Much of this work has been aimed at key kinases operating as central nodes in inflammatory signaling pathways. While this focus has led to over 30 FDA-approved small molecule kinase inhibitors, only one is currently approved for autoimmune and inflammatory diseases. Despite this lack of success, there remains tremendous reason for excitement. Our growing understanding of the biology involved in the inflammatory response, the factors that lead to safer small molecule kinase inhibitors, and the availability of selective tool molecules for interrogating specific nodes and pathways are all pushing the field forward. This article focuses on recent developments requiring novel approaches to create safe and effective small molecule kinase inhibitors and where further work is needed to realize the promise of small molecule kinase inhibitors for patient benefit.
Jie Wang; Helena Lundberg; Shota Asai; Pedro Martín-Acosta; Jason S. Chen; Stephen Brown; William Farrell; Russell G. Dushin; Christopher J. O’Donnell; Anokha S. Ratnayake; Paul Richardson; Zhiqing Liu; Tian Qin; Donna G. Blackmond; Phil S. Baran Proc. Natl. Acad. Sci. USA, 2018, 115(28), E6404-E6410 https://doi.org/10.1073/pnas.1806900115AbstractDNA-encoded libraries (DEL)-based discovery platforms have recently been widely adopted in pharmaceutical industry, mainly due to its powerful diversity and incredible number of molecules. In the past two decades since its disclosure, great strides have been made to expand the toolbox of reaction modes that are compatible with the idiosyncratic aqueous, dilute, and DNA-sensitive parameters of this system. However, construction of highly important C(sp3)-C(sp3) linkages on DNA through cross-coupling remains unexplored. In this article, we describe a systematic approach to translating standard organic reactions to a DEL-setting through the tactical combination of kinetic analysis and empirical screening with information captured from data mining. To exemplify this model, implementation of the Giese addition to forge high value C–C bonds on DNA was studied, which represents the first radical-based synthesis in DEL.
Yves Ruff; Frédéric Berst Med. Chem. Commun., 2018, 9, 1188-1193 https://doi.org/10.1039/C8MD00185EAbstractHerein, we describe the development of a copper-catalyzed amination of DNA-conjugated aryl iodides with aliphatic amines. This protocol leverages a novel ligand, 2-((2,6-dimethoxyphenyl)amino)-2-oxoacetic acid, to effect the transformation in aqueous DMSO, under mild conditions, in air, making it an ideal candidate for the synthesis of DNA-encoded libraries.
Hongming Chen; Thierry Kogej; Ola Engkvist Mol. Inform., 2018, 37(9-10), e1800041 https://doi.org/10.1002/minf.201800041AbstractCheminformatics has established itself as a core discipline within large scale drug discovery operations. It would be impossible to handle the amount of data generated today in a small molecule drug discovery project without persons skilled in cheminformatics. In addition, due to increased emphasis on “Big Data”, machine learning and artificial intelligence, not only in the society in general, but also in drug discovery, it is expected that the cheminformatics field will be even more important in the future. Traditional areas like virtual screening, library design and high‐throughput screening analysis are highlighted in this review. Applying machine learning in drug discovery is an area that has become very important. Applications of machine learning in early drug discovery has been extended from predicting ADME properties and target activity to tasks like de novo molecular design and prediction of chemical reactions.
Katrine Qvortrup; Thomas Eiland Nielsen; Vitaly Komnatnyy Chem. Commun., 2018, 54, 6759-6771 https://doi.org/10.1039/C8CC02486CAbstractHigh-throughput screening is an important component of the drug discovery process. The screening of libraries containing hundreds of thousands of compounds requires assays amenable to miniaturisation and automization. Combinatorial chemistry holds a unique promise to deliver structurally diverse libraries for early drug discovery. Among the various library forms, the one-bead-one-compound (OBOC) library, where each bead carries many copies of a single compound, holds the greatest potential for the rapid identification of novel hits against emerging drug targets. However, this potential has not yet been fully realized due to a number of technical obstacles. In this feature article, we review the progress that has been made in bead-based library screening and its application to the discovery of bioactive compounds. We identify the key challenges of this approach and highlight key steps needed for making a greater impact in the field.
Gerald Kolodny; Xiaoyu Li; Steven Balk BioEssays, 2018, 40, e1800057 https://doi.org/10.1002/bies.201800057AbstractMajor problems in cancer chemotherapy are toxicity, resistance, and cancer heterogeneity. A new theranostic paradigm has been proposed by the authors. Many million small molecules (SM) are bound to the proteins extracted from a patient's cancer. SM that also bind proteins extracted from normal human tissues are subtracted from the cancer protein bound SM leaving a large array of SM targeting many sites on each of the cancer biomarkers. Targeting many more than the conventional 1 – 4 cancer biomarkers will reduce development of tumor resistance. After several cycles of selection and counter selection, DNA codes appended to the SM will be PCR amplified to provide templates for restricted libraries of the SM to improve selectivity and sensitivity. The large array of selected and counter selected SM assures that many of the compounds in the array will penetrate the cell membrane and bind to intracellular targets, low tumor resistance, low background for imaging, low therapeutic toxicity, and targeting of the diverse biomarkers present in the heterogeneous mixture of cells in primary and metastatic cancer. Theranostic use of radiolabeled SM binding many sites on many, not necessarily critical, biomarkers provides high cancer cell killing. Experiments to provide proof of principle of this novel concept suggested by the authors.
Nicholas Favalli; Stefan Biendl; Marco Hartmann; Jacopo Piazzi; Filippo Sladojevich; Susanne Gräslund; Peter J. Brown; Katja Näreoja; Herwig Schüler; Jörg Scheuermann; Raphael Franzini; Dario Neri ChemMedChem, 2018, 13, 1303-1307 https://doi.org/10.1002/cmdc.201800193AbstractA DNA‐encoded chemical library (DECL) with 1.2 million compounds was synthesized by combinatorial reaction of seven central scaffolds with two sets of 343×492 building blocks. Library screening by affinity capture revealed that for some target proteins, the chemical nature of building blocks dominated the selection results, whereas for other proteins, the central scaffold also crucially contributed to ligand affinity. Molecules based on a 3,5‐bis(aminomethyl)benzoic acid core structure were found to bind human serum albumin with a Kd value of 6 nm, while compounds with the same substituents on an equidistant but flexible l‐lysine scaffold showed 140‐fold lower affinity. A 18 nm tankyrase‐1 binder featured l‐lysine as linking moiety, while molecules based on d‐Lysine or (2S,4S)‐amino‐l‐proline showed no detectable binding to the target. This work suggests that central scaffolds which predispose the orientation of chemical building blocks toward the protein target may enhance the screening productivity of encoded libraries.
Nicholas Favalli; Gabriele Bassi; Jörg Scheuermann; Dario Neri FEBS Lett., 2018, 592(12), 2168-2180 https://doi.org/10.1002/1873-3468.13068AbstractDNA-encoded chemical libraries (DECLs) are collections of compounds, individually coupled to DNA tags serving as amplifiable identification barcodes. Since individual compounds can be identified by the associated DNA tag, they can be stored as a mixture, allowing the synthesis and screening of combinatorial libraries of unprecedented size, facilitated by the implementation of split&pool synthetic procedures or other experimental methodologies. In this review, we briefly present relevant concepts and technologies, which are required for the implementation and interpretation of screening procedures with DNA-encoded chemical libraries. Moreover, we illustrate some success stories, detailing how novel ligands were discovered from encoded libraries. Finally, we critically review what can realistically be achieved with the technology at the present time, highlighting challenges and opportunities for the future.
Alexander L. Satz ACS Med. Chem. Lett., 2018, 9, 5, 408-410 https://doi.org/10.1021/acsmedchemlett.8b00128AbstractUse of DNA-encoded libraries (DELs) in the pharmaceutical industry has rapidly increased. We discuss what to expect when you run a DEL screen and contemplate guidelines for library design. Additionally, we consider some visionary work and extrapolate to the future
Eric Valeur; Patrick Jimonet J. Med. Chem., 2018, 61, 20, 9004-9029 https://doi.org/10.1021/acs.jmedchem.8b00378AbstractWith the rise of novel biology and high potential target identification technologies originating from advances in genomics, medicinal chemists are progressively facing targets of increasing complexity and often unprecedented. Novel hit finding technologies, combined with a wider choice of drug modalities, has resulted in a unique repertoire of options to address these challenging targets and to identify suitable starting points for optimization. Furthermore, innovative solutions originating from a range of academic groups and biotech companies require new types of collaborative models to leverage and integrate them in the drug discovery process. This perspective provides a guide for medicinal chemists covering contemporary probe and lead generation approaches and discusses the strengths and limitations of each strategy. Moreover, the expansion of strategies to modulate proteins creates the opportunity of a modality-agnostic and mode-of-action centric hit finding paradigm.
Melanie Leveridge; Chun-Wa Chung; Jeffrey W. Gross; Christopher B. Phelps; Darren Green SLAS Discov., 2018, 23(9), 881-897 https://doi.org/10.1177/2472555218778503AbstractThere has been much debate around the success rates of various screening strategies to identify starting points for drug discovery. Although high-throughput target-based and phenotypic screening has been the focus of this debate, techniques such as fragment screening, virtual screening, and DNA-encoded library screening are also increasingly reported as a source of new chemical equity. Here, we provide examples in which integration of more than one screening approach has improved the campaign outcome and discuss how strengths and weaknesses of various methods can be used to build a complementary toolbox of approaches, giving researchers the greatest probability of successfully identifying leads. Among others, we highlight case studies for receptor-interacting serine/threonine-protein kinase 1 and the bromo- and extra-terminal domain family of bromodomains. In each example, the unique insight or chemistries individual approaches provided are described, emphasizing the synergy of information obtained from the various tactics employed and the particular question each tactic was employed to answer. We conclude with a short prospective discussing how screening strategies are evolving, what this screening toolbox might look like in the future, how to maximize success through integration of multiple tactics, and scenarios that drive selection of one combination of tactics over another
John A Hawkins; Stephen K Jones Jr.; Ilya J Finkelstein; William H Press Proc. Natl. Acad. Sci. USA, 2018, 115(27), E6217-E6226 https://doi.org/10.1073/pnas.1802640115AbstractMany large-scale, high-throughput experiments use DNA barcodes, short DNA sequences prepended to DNA libraries, for identification of individuals in pooled biomolecule populations. However, DNA synthesis and sequencing errors confound the correct interpretation of observed barcodes and can lead to significant data loss or spurious results. Widely used error-correcting codes borrowed from computer science (e.g., Hamming, Levenshtein codes) do not properly account for insertions and deletions (indels) in DNA barcodes, even though deletions are the most common type of synthesis error. Here, we present and experimentally validate filled/truncated right end edit (FREE) barcodes, which correct substitution, insertion, and deletion errors, even when these errors alter the barcode length. FREE barcodes are designed with experimental considerations in mind, including balanced guanine-cytosine (GC) content, minimal homopolymer runs, and reduced internal hairpin propensity. We generate and include lists of barcodes with different lengths and error correction levels that may be useful in diverse high-throughput applications, including >10^6 single-error-correcting 16-mers that strike a balance between decoding accuracy, barcode length, and library size. Moreover, concatenating two or more FREE codes into a single barcode increases the available barcode space combinatorially, generating lists with >10^15 error-correcting barcodes. The included software for creating barcode libraries and decoding sequenced barcodes is efficient and designed to be user-friendly for the general biology community.
Kaicheng Meng; Paul Shim; Qingtin Wang; Shuai Zhao; Ting Gu; Alem W. Kahsai; Seungkirl Ahn; Xin Chen Bioorg. Med. Chem., 2018, 26, 9, 2320-2330 https://doi.org/10.1016/j.bmc.2018.03.023AbstractThe β2-adrenergic receptor (β2AR), a G protein-coupled receptor, is an important therapeutic target. We recently described Cmpd-15, the first small molecule negative allosteric modulator (NAM) for the β2AR. Herein we report in details the design, synthesis and structure-activity relationships (SAR) of seven Cmpd-15 derivatives. Furthermore, we provide in a dose-response paradigm, the details of the effects of these derivatives in modulating agonist-induced β2AR activities (G-protein-mediated cAMP production and β-arrestin recruitment to the receptor) as well as the binding affinity of an orthosteric agonist in radio-ligand competition binding assay. Our results show that some modifications, including removal of the formamide group in the para-formamido phenylalanine region and bromine in the meta-bromobenzyl methylbenzamide region caused dramatic reduction in the functional activity of Cmpd-15. These SAR results provide valuable insights into the mechanism of action of the NAM Cmpd-15 as well as the basis for future development of more potent and selective modulators for the β2AR based on the chemical scaffold of Cmpd-15.
Robert J. Young; Paul D. Leeson J. Med. Chem., 2018, 61, 6421−6467 https://doi.org/10.1021/acs.jmedchem.8b00180AbstractThe practices and tactics employed in successful optimizations are examined, judged from the trajectories of ligand efficiency and property evolution. A wide range of targets is analyzed, encompassing a variety of hit finding methods (HTS, fragments, encoded library technology) and types of molecules, including those beyond the rule of five. The wider employment of efficiency metrics and lipophilicity control are evident in contemporary practice and the impact on quality demonstrable. What is clear is that while targets are different, successful molecules are almost invariably amongst the most efficient for their target, even at the extremes. Trajectory mapping, based on principles rather than rules, is useful in assessing quality and progress in optimizations, whilst benchmarking against competitors and assessing property-dependent risks.
Seungkirl Ahn; Biswaranjan Pani; Alem W. Kahsai; Eva K. Olsen; Gitte Husemoen; Mikkel Vestrgaard; Lei Jin; Shuai Zhao; Laura M. Wingler; Paula K. Rambarat; Rishabh K. Simhal; Thomas T. Xu; Lillian D. Sun; Paul J. Shim; Dean P. Staus; Li-Yin Huang; Thomas Franch; Xin Chen; Robert J. Lefkowitz Mol. Pharmacol., 2018, 94(2), 850-861 https://doi.org/10.1124/mol.118.111948AbstractConventional drug discovery efforts at the β2-adrenoceptor (β2AR) have led to the development of ligands that bind almost exclusively to the receptor's hormone-binding orthosteric site. However, targeting the largely unexplored and evolutionarily unique allosteric sites has potential for developing more specific drugs with fewer side effects than orthosteric ligands. Using our recently developed approach for screening G protein-coupled receptors (GPCRs) with DNA-encoded small molecule libraries, we have discovered and characterized the first β2AR small molecule positive allosteric modulators (PAMs) - compound-6 [(R)-N-(4-amino-1-(4-(tert-butyl)phenyl)-4-oxobutan-2-yl)-5-(N-isopropyl-N-methylsulfamoyl)-2-((4-methoxyphenyl) thio)benzamide] and its analogs. We utilized purified human β2ARs, occupied by a high affinity agonist, for the affinity-based screening of over 500 million distinct library compounds, which yielded compound-6. It exhibits a low micro-molar affinity for the agonist-occupied β2AR, and displays positive cooperativity with orthosteric agonists, thereby enhancing their binding to the receptor and ability to stabilize its active state. Compound-6 is cooperative with G protein and β-arrestin1 (a.k.a. arrestin2) to stabilize high-affinity, agonist-bound active states of the β2AR, and potentiates downstream cAMP production and receptor-recruitment of β-arrestin2 (a.k.a. arrestin3). Compound-6 is specific for the β2AR compared to the closely related β1AR. Structure-activity studies of select compound-6 analogs defined the chemical groups that are critical for its biological activity. We thus introduce the first small molecule PAMs for the β2AR, which may serve as a lead molecule for the development of novel therapeutics. The approach described here establishes a broadly applicable proof-of-concept strategy for affinity-based discovery of small molecule allosteric compounds targeting unique conformational states of GPCRs.
Jacques Saarbach; Eric Lindberg; Nicolas Winssinger Chimia, 2018, 72, 207-211 https://doi.org/10.2533/chimia.2018.207AbstractTemplated reactions proceed by bringing reagents in close proximity through their interaction with a template thus raising their effective concentrations. Templated reactions empower chemists to perform reactions at low concentrations in complex environments. Herein, we discuss our work on templated reactions leveraged on ruthenium photocatalysis. Over the past five years, we have used this reaction to uncage reporter molecules and sense or image nucleic acids or proteins of interest. The ruthenium photocatalysis chemistry has proven to be extremely robust and compatible with complex biological environments.
Yun Ding; Jing Chai; Paolo A. Centrella; Chenaimwoyo Gondo; Jennifer L. DeLorey; Matthew A. Clark ACS Comb. Sci., 2018, 20, 5, 251-255 https://doi.org/10.1021/acscombsci.8b00009AbstractEncoded library technology (ELT) is an effective approach to the discovery of novel small-molecule ligands for biologi-cal targets. A key factor for the success of the technology is the chemical diversity of the libraries. Here we report the development of DNA-conjugated benzimidazoles. Using 4-fluoro-3-nitrobenzoic acid as a key synthon, we synthesized a 320 million-member DNA-encoded benzimidazole library using Fmoc-protected amino acids, amines and aldehydes as diversity elements. Affinity selection of the library led to the discovery of a novel, potent and specific antagonist of the NK3 receptor.
Dario Neri; Richard A. Lerner Annu. Rev. Biochem., 2018, 87, 479-502 https://doi.org/10.1146/annurev-biochem-062917-012550AbstractThe discovery of organic ligands that bind specifically to proteins is a central problem in chemistry, biology, and the biomedical sciences. The encoding of individual organic molecules with distinctive DNA tags, serving as amplifiable identification bar codes, allows the construction and screening of combinatorial libraries of unprecedented size, thus facilitating the discovery of ligands to many different protein targets. Fundamentally, one links powers of genetics and chemical synthesis. After the initial description of DNA-encoded chemical libraries in 1992, several experimental embodiments of the technology have been reduced to practice. This review provides a historical account of important milestones in the development of DNA-encoded chemical libraries, a survey of relevant ongoing research activities, and a glimpse into the future.
Jae Eun Cheong; Anil Ekkati; Lijun Sun Expert Opin. Ther. Patents, 2018, 28, 4, 317-330 https://doi.org/10.1080/13543776.2018.1441290AbstractIntroduction: Indoleamine 2,3-dioxygenase 1 (IDO1) is overexpressed by cancer cells and the antigen presenting dendritic cells in the tumor microenvironment (TME). Activation of IDO1 depletes tryptophan and produces kynurenine, which induces T cell anergy and suppresses tumor control by the immune system. When combined with an immune checkpoint inhibitor, IDO1 inhibitors have shown promising anticancer activity in preclinical tumor models as well as in early stage clinical trials. Areas covered: IDO1 inhibitors disclosed in the patent literature from 2013-2017 are categorized, when applicable, according to their structural similarity to the clinical development candidates indoximod and PF-06840003, navoximod, epacadostat, KHK2455 and aryl-1,2-diamines, and BMS-986205 among others, respectively. Representative structures and their IDO1 inhibitory activity are presented to highlight the novelty and activity. Finally, the reported cocrystal structures were analyzed to provide insights for inhibitor-enzyme interactions and guidance for the design and discovery of next generation inhibitors. Expert opinion: This review demonstrates that the structural diversity of new IDO1 inhibitors could be expanded via a number of approaches.
Tommi Österlund; Heidi Korhonen; Pasi Virta Org. Lett., 2018, 20(6), 1496-1499 https://doi.org/10.1021/acs.orglett.8b00113AbstractThe potential of N(Me)-alkoxyamine glycosylation as a DNA-templated ligation has been studied. On a hairpin stem-template model, a notable rate enhancement and an increased equilibrium yield are observed compared to the corresponding reaction without a DNA catalyst. The N-glycosidic connection is dynamic at pH 5, whereas it becomes irreversible at pH 7. The N(Me)-alkoxyamine glycosylation may hence be an attractive pH controlled reaction for the assembly of DNA-based dynamic products.
Giulia Opassi; Alessandro Gesù; Alberto Massarotti Drug Discov. Today, 2018, 3(23), 565-574 https://doi.org/10.1016/j.drudis.2018.01.007AbstractWe are used to considering chemical and biological spaces as two different entities; although they represent a more-interconnected world, in fact they represent a Yin-Yang concept in drug discovery. Chemical-biological space is as vast as the universe and, as Douglas Adams famously said, ‘Space is big. You just won’t believe how vastly, hugely, mind-bogglingly big it is’. However, many researchers are convinced that it is not so infinite, and are designing computational and experimental tools to help identify and explore all possible chemical-biological space. Here, we provide an analysis of their approaches and discuss possible future research studies.
Keitou Shu; Thomas Kodadek ACS Comb. Sci., 2018, 20(5), 277-281 https://doi.org/10.1021/acscombsci.8b00001AbstractOne-bead-one-compound (OBOC) libraries constructed by solid-phase split-and-pool synthesis are a valuable source of protein ligands. Most OBOC libraries are comprised of oligoamides, particularly peptides, peptoids and peptoid-inspired molecules. Further diversification of the chemical space covered by OBOC libraries is desirable. Towards this end, we report here the efficient proline-catalyzed asymmetric aldol reaction between immobilized aldehydes and soluble ketones. These reaction conditions do not compromise the amplification of DNA by the polymerase chain reaction. Thus, this chemistry should be useful for the construction of novel DNA-encoded OBOC libraries by solid-phase synthesis
Xiaofang Zuo; Zhipeng Huo; Dongwei Kang; Gaochan Wu; Zhongxia Zhou; Xinyong Liu; Peng Zhan Expert Opin. Ther. Patents, 2018, 28, 4, 299-316 https://doi.org/10.1080/13543776.2018.1438410AbstractIntroduction: To deal with the rapid emergence of drug resistance challenges, together with the difficulty to eradicate the virus, off-target effects and significant cumulative drug toxicities, it is still imperative to develop next-generation anti-HIV agents with novel chemical classes or new mechanisms of action. Areas covered: We primarily focused on current strategies to discover novel anti-HIV agents. Moreover, examples of anti-HIV lead compounds were mainly selected from recently patented publications (reported between 2014 and 2017). In particular, ‘privileged structure’-focused substituents decorating approach, scaffold hopping, natural-product diversification and prodrug are focused on. Furthermore, exploitation of new compounds with unexplored mechanisms of action and medicinal chemistry strategies to deplete the HIV reservoir were also described. Perspectives that could inspire future anti-HIV drug discovery are delineated. Expert opinion: Even if a large number of patents have been disclosed recently, additional HIV inhibitors are still required, especially novel chemical skeletons displaying a unexploited mechanism of action. Current medicinal chemistry strategies are inadequate, and appropriate and new methodologies and technologies should be exploited to identify novel anti-HIV drug candidates in a time- and cost- effective manner.
Ki Tae Kim; Dalu Chang; Nicolas Winssinger Helv. Chim. Acta, 2018, 101, e170095 https://doi.org/10.1002/hlca.201700295AbstractRNA, originally perceived as a simple information transfer biopolymer, is emerging as an important regulator in cellular processes. A number of non-coding RNAs are double-stranded and there is a need for technologies to reliably detect and image such RNAs for biological and biomedical research. Herein we report double-stranded RNA-specific templated reaction resulting from PNA-reagent conjugates that are brought within reactive distance through the formation of sequence-specific triplexes onto double-stranded RNA. The reaction makes use of a ruthenium-based photocatalyst that reduces a pyridinium-based immolative linker, unmasking a profluorophore. The reaction was shown to proceed with signal amplification and to be selective for double-stranded RNA over DNA as well as single-stranded RNA. The generality of the triplex formation was enabled by non-canonical nucleobases that extend the Hoogsteen base-pairing repertoire. The technology was applied to a templated reaction using pre-microRNA 31
Kristin K. Brown; Michael M. Hann; Ami S. Lakdawala; Rita Santos; Pamela J. Thomas; Kieran Todd Med. Chem. Commun., 2018, 9, 606-613 https://doi.org/10.1039/C7MD00633KAbstractAbstract. The assessment of the suitability of novel targets to intervention by different modalities, e.g. small molecules or antibodies, is increasingly seen as important in helping to select the most progressable targets at the outset of a drug discovery project. This perspective considers differing aspects of tractability and how it can be assessed using in silico and experimental approaches. We also share some of our experiences in using these approaches.
Nicolas Probst; Rémy Lartia; Océane Théry; Mouâd Alami; Eric Defrancq; Samir Messaoudi Chem. Eur. J., 2018, 24(8), 1795-1800 https://doi.org/10.1002/chem.201705371AbstractAn efficient method for the thioglycoconjugation of iodinated oligonucleotides by Buchwald-Hartwig-Migita cross-coupling under mild conditions is reported. The method enables divergent synthesis of many different functionalized thioglycosylated ODNs in good yields, without affecting the integrity of the other A, C, and G nucleobases
Ruben Ragg ChemBioChem, 2018, 19, 4-6 https://doi.org/10.1002/cbic.201700640AbstractChemBioChem had a successful year in 2017 receiving its 10 000 th submission. By broadening the journal scope through the addition of biomaterials, ChemBioChem will help to intertwine chemistry and biology even further. Also up next in 2018, a young chemists Special Issue, so gather up ChemBioTalents!
Yiran Huang; Wenlu Zheng; Xiaoyu Li Anal. Biochem., 2018, 545, 84-90 https://doi.org/10.1016/j.ab.2018.01.010AbstractDNA-based probes are powerful analytical tools for protein detection and analysis. Target-induced DNA assembly is a widely used strategy to transduce target-ligand binding to detectable signals. However, most of the existing methods based on DNA assembly require two or more binding sites on the target protein. Here we report a novel detection method suitable for protein targets with just a single binding site. This method is based on target-induced probe assembly, DNA-templated photo-crosslinking, and DNA-mediated toehold strand displacement to form a tri-probe complex that is specific for target protein.
Letian Kuai; Thomas O’Keeffe; Christopher C. Arico-Muendel SLAS Discov., 2018, 23(5), 405-416 https://doi.org/10.1177/2472555218757718AbstractDNA Encoded Libraries (DELs) use unique DNA sequences to tag each chemical warhead within a library mixture to enable deconvolution following affinity selection against a target protein. With next-generation sequencing, millions to billions of sequences can be read and counted to report binding events. This unprecedented capability has enabled researchers to synthesize and analyze numerically large chemical libraries. Despite the common perception that each library member undergoes a miniaturized affinity assay, selections with higher complexity libraries often produce results that are difficult to rank order. In this study, we aimed to understand the robustness of DEL selection by examining the sequencing readouts of warheads and chemotype families among a large number of experimentally repeated selections. The results revealed that (1) the output of DEL selection is intrinsically noisy but can be reliably modeled by the Poisson distribution, and (2) Poisson noise is the dominating noise at low copy counts and can be estimated even from a single experiment. We also discuss the shortcomings of data analyses based on directly using copy counts and their linear transformations, and propose a framework that incorporates proper normalization and confidence interval calculation to help researchers better understand DEL data.
Masato Yoshikawa; Morihisa Saitoh; Taisuke Katoh; Tomohiro Seki; Simone V. Bigi; Yuji Shimizu; Tsuyoshi Ishii; Takuro Okai; Masako Kuno; Harumi Hattori; Etsuro Watanabe; Kumar S. Saikatendu; Hua Zou; Masanori Nakakariya; Takayuki Tatamiya; Yoshihisa Nakada; Takatoshi Yogo J. Med. Chem., 2018, 61, 6, 2384-2409 https://doi.org/10.1021/acs.jmedchem.7b01647AbstractWe report the discovery of 7-oxo-2,4,5,7-tetrahydro-6H-pyrazolo[3,4-c]pyridine derivatives as a novel class of receptor interacting protein 1 (RIP1) kinase inhibitors. On the basis of the overlay study between HTS hit 10 and GSK2982772 (6) in RIP1 kinase, we designed and synthesized a novel class of RIP1 kinase inhibitor 11 possessing moderate RIP1 kinase inhibitory activity and P-gp mediated efflux. The optimization of the core structure and the exploration of appropriate substituents utilizing SBDD approach led to the discovery of 22, a highly potent, orally available, and brain-penetrating RIP1 kinase inhibitor with excellent PK profiles. Compound 22 significantly suppressed necroptotic cell death both in mouse and human cells. Oral administration of 22 (10 mg/kg, bid) attenuated disease progression in the mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Moreover, analysis of structure–kinetic relationship (SKR) for our novel chemical series was also discussed.
Kevin Pels; Paige Dickson; Hongchan An; Thomas Kodadek ACS Comb. Sci., 2018, 20, 2, 61-69 https://doi.org/10.1021/acscombsci.7b00169AbstractWe demonstrate that the Knoevenagel condensation can be exploited in combinatorial synthesis on the solid phase. Condensation products from such reactions were structurally characterized, and their Michael reactivity with thiol and phosphine nucleophiles is described. Cyanoacrylamides were previously reported to react reversibly with thiols, and notably, we show that dilution into low pH buffer can trap covalent adducts which are isolable via chromatography. Finally, we synthesized both traditional and DNA-encoded one-bead, one-compound libraries containing cyanoacrylamides as a source of cysteine-reactive, reversibly covalent protein ligands.
Dalu Chang; Ki Tae Kim; Eric Lindberg; Nicolas Winssinger Bioconjugate Chem., 2018, 29, 1, 158-163 https://doi.org/10.1021/acs.bioconjchem.7b00663AbstractNucleic acid templated reactions have attracted attention as an important technology to sense oligonucleotides and to translate nucleic acid-based instructions into diverse outputs. Great progress has been made in accelerating the reaction in order to improve signal amplification, reaching rates where substrate turnover rather than chemical reaction is rate limiting. Herein we explore the utility of architectures inspired by three-way junction that yield a cleavage of a strand thus accelerating substrate turnover. We demonstrate that such design can overcome product inhibition in templated reactions and operate close to the rate of hybridization.
Chad Townsend; Akihiro Furukawa; Joshua Schwochert; Cameron Pye; Quinn Edmondson; R. Scott Lokey Bioorg. Med. Chem., 2018, 26(6), 1232-1238 https://doi.org/10.1016/j.bmc.2018.01.027AbstractCyclic peptides are of great interest as therapeutic compounds due to their potential for specificity and intracellular activity, but specific compounds can be difficult to identify from large libraries without resorting to molecular encoding techniques. Large libraries of cyclic peptides are often DNA-encoded or linearized before sequencing, but both of those deconvolution strategies constrain the chemistry, assays, and quantification methods which can be used. We developed an automated sequencing program, CycLS, to identify cyclic peptides contained within large synthetic libraries. CycLS facilitates quick and easy identification of all library-members via tandem mass spectrometry data without requiring any specific chemical moieties or modifications within the library. Validation of CycLS against a library of 400 cyclic hexa-peptide peptoid hybrids (peptomers) of unique mass yielded a result of 95% accuracy when compared against a simulated library size of 234,256 compounds. CycLS was also evaluated by resynthesizing pure compounds from a separate 1800-member library of cyclic hexapeptides and hexapeptomers with high mass redundancy. Of 22 peptides resynthesized, 17 recapitulated the retention times assigned to them from the whole-library bulk assay results. Implementing a database-matching approach, CycLS is fast and provides a robust method for sequencing cyclic peptides that is particularly applicable to the deconvolution of synthetic libraries.
Nam Tran Hoang; Thomas Kodadek ACS Comb. Sci., 2018, 20, 2, 55-60 https://doi.org/10.1021/acscombsci.7b00151AbstractOne-bead-one-compound (OBOC) libraries constructed by solid-phase split-and-pool synthesis are a valuable source of protein ligands. Most OBOC libraries are comprised of oligoamides, particularly peptides, peptoids and peptoid-inspired molecules. Further diversification of the chemical space covered by OBOC libraries is desirable. Towards this end, we report here the efficient proline-catalyzed asymmetric Mannich reaction between immobilized aldehydes and soluble ketones and anilines. The reaction conditions do not compromise the amplification of DNA by the PCR. Thus, this chemistry will be useful for the construction of novel DNA-encoded libraries by solid-phase synthesis.
Dmitry L. Usanov; Alix I. Chan; Juan Pablo Maianti; David R. Liu Nat. Chem., 2018, 10(7), 704-714 https://doi.org/10.1038/s41557-018-0033-8AbstractDNA-encoded libraries have emerged as a widely used resource for the discovery of bioactive small molecules, and offer substantial advantages compared with conventional small-molecule libraries. Here, we have developed and streamlined multiple fundamental aspects of DNA-encoded and DNA-templated library synthesis methodology, including computational identification and experimental validation of a 20 × 20 × 20 × 80 set of orthogonal codons, chemical and computational tools for enhancing the structural diversity and drug-likeness of library members, a highly efficient polymerase-mediated template library assembly strategy, and library isolation and purification methods. We have integrated these improved methods to produce a second-generation DNA-templated library of 256,000 small-molecule macrocycles with improved drug-like physical properties. In vitro selection of this library for insulin-degrading enzyme affinity resulted in novel insulin-degrading enzyme inhibitors, including one of unusual potency and novel macrocycle stereochemistry (IC50 = 40 nM). Collectively, these developments enable DNA-templated small-molecule libraries to serve as more powerful, accessible, streamlined and cost-effective tools for bioactive small-molecule discovery.
Kyle E. Denton; Sijie Wang; Michael C. Gignac; Natalia Milosevich; Fraser Hof; Emily C. Dykhuizen; Casey J. Krusemark SLAS Discov., 2018, 23(5), 417-428 https://doi.org/10.1177/2472555217750871AbstractThe identification of protein ligands from a DNA-encoded library is commonly conducted by an affinity selection assay. These assays are often not validated for robustness, raising questions about selections that fail to identify ligands and the utility of enrichment values for ranking ligand potencies. Here, we report a method for optimizing and utilizing affinity selection assays to identify potent and selective peptidic ligands to the highly related chromodomains of CBX proteins. To optimize affinity selection parameters, statistical analyses (Z' factors) were used to define the ability of selection assay conditions to identify and differentiate ligands of varying affinity. A DNA-encoded positional scanning library of peptidomimetics was constructed around a trimethyllysine-containing parent peptide, and parallel selections against the chromodomains from CBX8 and CBX7 were conducted over three protein concentrations. Relative potencies of off-DNA hit molecules were determined through a fluorescence polarization assay and were consistent with enrichments observed by DNA sequencing of the affinity selection assays. In addition, novel peptide-based ligands were discovered with increased potency and selectivity to the chromodomain of CBX8. The results indicate low DNA tag bias and show that affinity-based in vitro selection assays are sufficiently robust for both ligand discovery and determination of quantitative structure-activity relationships.
Osayemwenre Erharuyia; Scott Simanski; Patrick J. McEnaney; Thomas Kodadek Bioorg. Med. Chem. Lett., 2018, 28(16), 2773-2778 https://doi.org/10.1016/j.bmcl.2018.01.033AbstractOne bead one compound (OBOC) libraries can be screened against serum samples to identify ligands to antibodies in this mixture. In this protocol, hit beads are identified by staining with a fluorescent labeled secondary antibody. When screens are conducted against two different sets of serum, antibodies, and ligands to them, can be discovered that distinguish the two populations. The application of DNA-encoding technology to OBOC libraries has allowed the use of 10 µm beads for library preparation and screening, which pass through a standard flow cytometer, allowing the fluorescent hit beads to be separated from beads displaying non-ligands easily. An important issue in using this approach for the discovery of antibody biomarkers is its analytical sensitivity. In other words, how abundant must an IgG be to allow it to be pulled out of serum in an unbiased screen using a flow cytometer? We report here a model study in which monoclonal antibodies with known ligands of varying affinities are doped into serum. We find that for antibody ligands typical of what one isolates from an unbiased combinatorial library, the target antibody must be present at 10–50 nM. True antigens, which bind with significantly higher affinity, can detect much less abundant serum antibodies.
Zhengrong Zhu; Alex Shaginian; LaShadric C. Grady; Thomas O'Keeffe; Xiangguo E. Shi; Christopher P. Davie; Graham L Simpson; Jeffrey A. Messer; Ghotas Evindar; Robert N. Bream; Praew P. Thansandote; Naomi R Prentice; Andrew M. Mason; Sandeep Pal ACS Chem. Biol., 2018, 13, 1, 53-59 https://doi.org/10.1021/acschembio.7b00852AbstractA DNA-encoded macrocyclic peptide library was designed and synthesized with 2.4×(10^12) members composed of 4-20 natural and non-natural amino acids. Affinity-based selection was performed against two therapeutic targets, VHL and RSV N protein. Based on selection data some peptides were selected for resynthesis without DNA tag and their activity was confirmed
Jorge Amigo; Ramón Rama-Garda; Xabier Bello; Beatriz Sobrino; Jesús de Blas; María Martín-Ortega; Theodore C. Jessop; Ángel Carracedo; María Isabel García Loza; Eduardo Domínguez SLAS Discov., 2018, 23(5), 397-404 https://doi.org/10.1177/2472555217753840AbstractAvailable tools to analyze sequencing data coming from DNA-encoded chemical libraries (DELs) are often limited to in-house methods, which usually rely on strictly looking for the particular DEL structure used. Current methods do not take into account technological errors, such as library codification and sequencing errors, when detecting the sequences. The vast amount of data produced by next-generation sequencing of DEL screens is usually enough to extract the minimum information needed for compound identification. Here, we report a methodology to deconvolute encoding oligonucleotides, thus optimizing the sequencing power regardless of the library size, design complexity, or sequencing technology chosen. tagFinder is a highly flexible tool for fast tag detection and thorough DEL results characterization, which requires minimal hardware resources, scales linearly, and does not introduce any analytical error. The methodology can even deal with sequencing errors and PCR duplicates on single- or double-stranded DNA, enhancing the analytical detection and quantification of molecules and the informativeness of the entire process. Source code is available at https://github.com/jamigo/tagFinder
Dean G. Brown; Giles A. Brown; Paolo A. Centrella; Kaan Certel; Robert M. Cooke; John W. Cuozzo; Niek Dekker; Christoph E. Dumelin; Andrew Ferguson; Cédric Fiez-Vandal; Stefan Geschwindner; Marie-Aude Guié; Sevan Habeshian; Anthony D. Keefe; Oliver Schlenker; Eric A. Sigel; Arjan Snijder; Holly T. Soutter; Linda Sundström; Dawn M. Troast; Giselle Wiggin; Jing Zhang; Ying Zhang; Matthew A. Clark SLAS Discov., 2018, 23(5), 429-436 https://doi.org/10.1177/2472555217749847AbstractThe discovery of ligands via affinity-mediated selection of DNA-encoded chemical libraries is driven by the quality and concentration of the protein target. G-protein-coupled receptors (GPCRs) and other membrane-bound targets can be difficult to isolate in their functional state and at high concentrations, and therefore have been challenging for affinity-mediated selection. Here, we report a successful selection campaign against protease-activated receptor 2 (PAR2). Using a thermo-stabilized mutant of PAR2, we conducted affinity selection using our >100-billion-compound DNA-encoded library. We observed a number of putative ligands enriched upon selection, and subsequent cellular profiling revealed these ligands to comprise both agonists and antagonists. The agonist series shared structural similarity with known agonists. The antagonists were shown to bind in a novel allosteric binding site on the PAR2 protein. This report serves to demonstrate that cell-free affinity selection against GPCRs can be achieved with mutant stabilized protein targets.
Jesús Castañón; José Pablo Román; Theodore C. Jessop; Jesús de Blas; Rubén Haro SLAS Discov., 2018, 23(5), 387-396 https://doi.org/10.1177/2472555217752091AbstractDNA-encoded libraries (DELs) have emerged as an efficient and cost-effective drug discovery tool for the exploration and screening of very large chemical space using small-molecule collections of unprecedented size. Herein, we report an integrated automation and informatics system designed to enhance the quality, efficiency, and throughput of the production and affinity selection of these libraries. The platform is governed by software developed according to a database-centric architecture to ensure data consistency, integrity, and availability. Through its versatile protocol management functionalities, this application captures the wide diversity of experimental processes involved with DEL technology, keeps track of working protocols in the database, and uses them to command robotic liquid handlers for the synthesis of libraries. This approach provides full traceability of building-blocks and DNA tags in each split-and-pool cycle. Affinity selection experiments and high-throughput sequencing reads are also captured in the database, and the results are automatically deconvoluted and visualized in customizable representations. Researchers can compare results of different experiments and use machine learning methods to discover patterns in data. As of this writing, the platform has been validated through the generation and affinity selection of various libraries, and it has become the cornerstone of the DEL production effort at Lilly.
Stefan Matysiak; Klaus Hellmuth; Afaf H. El-Sagheer; Arun Shivalingam; Yavuz Ariyurek; Marco de Jong; Martine J. Hollestelle; Ruud Out; Tom Brown Org. Biomol. Chem., 2018, 16, 48-52 https://doi.org/10.1039/C7OB02119DAbstractDNA encoded ligands are self-assembled into bivalent complexes and chemically ligated to link their identities. To demonstrate their potential as a combinatorial screening platform for avidity interactions, the optimal bivalent aptamer design (examplar ligands) for human alpha-thrombin is determined in a single round of selection and the DNA scaffold replaced with minimal impact on the final design.
Hiroshi Abe; Yasuaki Kimura Chem. Pharm. Bull., 2018, 66, 117-122 https://doi.org/10.1248/cpb.c17-00615AbstractChemical ligation of oligonucleotides (ONs) is the key reaction for various ON-based technologies. We have tried to solve the problems of RNA interference (RNAi) technology by applying ON chemical ligation to RNAi. We designed a new RNAi system, called intracellular buildup RNAi (IBR-RNAi), where the RNA fragments are built up into active small-interference RNA (siRNA) in cells through a chemical ligation reaction. Using the phosphorothioate and iodoacetyl groups as reactive functional groups for the ligation, we achieved RNAi effects without inducing immune responses. Additionally, we developed a new chemical ligation for IBR-RNAi, which affords a more native-like structure in the ligated product. The new ligation method should be useful not only for IBR-RNAi but also for the chemical synthesis of biofunctional ONs.
Jessica Sayers; Richard J. Payne; Nicolas Winssinger Chem. Sci., 2018, 9, 896-903 https://doi.org/10.1039/C7SC02736BAbstractThe development of a rapid and chemoselective selenocystine–selenoester peptide ligation that operates at nanomolar reactant concentrations has been developed by utilising PNA templation. Kinetic analysis of the templated peptide ligation revealed that the selenocystine–selenoester reaction was 10 times faster than traditional native chemical ligation at cysteine and to our knowledge is the fastest templated ligation reaction reported to date. The efficiency and operational simplicity of this technology is highlighted through the formation of hairpin molecular architectures and in a novel paper-based lateral flow assay for the rapid and sequence specific detection of oligonucleotides, including miRNA in cell lysates.
Yizhou Li; Roberto De Luca; Samuele Cazzamalli; Francesca Pretto; Davor Bajic; Jörg Scheuermann; Dario Neri Nat. Chem., 2018, 10(4), 441-448 https://doi.org/10.1038/s41557-018-0017-8AbstractIn nature, specific antibodies can be generated as a result of an adaptive selection and expansion of lymphocytes with suitable protein binding properties. We attempted to mimic antibody–antigen recognition by displaying multiple chemical diversity elements on a defined macrocyclic scaffold. Encoding of the displayed combinations was achieved using distinctive DNA tags, resulting in a library size of 35,393,112. Specific binders could be isolated against a variety of proteins, including carbonic anhydrase IX, horseradish peroxidase, tankyrase 1, human serum albumin, alpha-1 acid glycoprotein, calmodulin, prostate-specific antigen and tumour necrosis factor. Similar to antibodies, the encoded display of multiple chemical elements on a constant scaffold enabled practical applications, such as fluorescence microscopy procedures or the selective in vivo delivery of payloads to tumours. Furthermore, the versatile structure of the scaffold facilitated the generation of protein-specific chemical probes, as illustrated by photo-crosslinking.
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FINhttps://pdbj.org/mine/summary/6fin Abstract Epithelial discoidin domain-containing receptor 1, IODIDE ION, 3-[(3-cyclopropyl-1,2,4-oxadiazol-5-yl)methyl]-8-(1~{H}-indazol-5-ylcarbonyl)-1-phenyl-1,3,8-triazaspiro[4.5]decan-4-one
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FEWhttps://pdbj.org/mine/history/6few Abstract Epithelial discoidin domain-containing receptor 1, 2-[8-(2~{H}-indazol-5-ylcarbonyl)-4-oxidanylidene-1-phenyl-1,3,8-triazaspiro[4.5]decan-3-yl]-~{N}-methyl-ethanamide
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FIQhttps://pdbj.org/mine/summary/6fiq Abstract Epithelial discoidin domain-containing receptor 1, 1-(1~{H}-indazol-5-ylcarbonyl)-5'-methoxy-1'-[2-oxidanylidene-2-[(2~{S})-2-(trifluoromethyl)pyrrolidin-1-yl]ethyl]spiro[piperidine-4,3'-pyrrolo[3,2-b]pyridine]-2'-one, CHLORIDE ION
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FERhttps://pdbj.org/mine/summary/6fer Abstract Discoidin domain-containing receptor 2, 2-[4,5-bis(fluoranyl)-2-oxidanylidene-1'-(1~{H}-pyrazolo[3,4-b]pyridin-5-ylcarbonyl)spiro[indole-3,4'-piperidine]-1-yl]-~{N}-[2,2,2-tris(fluoranyl)ethyl]ethanamide
Hans Richter; Alexander L. Satz; Marc Bedoucha; Bernd Buettelmann; Ann C. Petersen; Anja Harmeier; Ricardo Hermosilla; Remo Hochstrasser; Dominique Burger; Bernard Gsell; Rodolfo Gasser; Sylwia Huber; Melanie N. Hug; Buelent Kocer; Bernd Kuhn; Martin Ritter; Markus G. Rudolph; Franziska Weibel; Judith Molina-David; Jin-Ju Kim; Javier Varona Santos; Martine Stihle; Guy J. Georges; R. Daniel Bonfil; Rafael Fridman; Sabine Uhles; Solange Moll; Christian Faul; Alessia Fornoni; Marco Prunotto PDBJ - 6FIOhttps://pdbj.org/mine/summary/6fio Abstract Epithelial discoidin domain-containing receptor 1, 2-[1'-(1~{H}-indazol-5-ylcarbonyl)-4-methyl-2-oxidanylidene-spiro[indole-3,4'-piperidine]-1-yl]-~{N}-[2,2,2-tris(fluoranyl)ethyl]ethanamide
Amalia M. Estévez; Felix Gruber; Alexander L. Satz; Rainer E. Martin; Hans Peter Wessel Tetrahedron Asymm.，2017, 28, 6, 837-842https://doi.org/10.1016/j.tetasy.2017.04.007 Abstract For the generation of DNA-encoded libraries, a central scaffold containing three exit vectors with defined chirality was devised starting from commercially available tri-O-acetyl-glucal. This scaffold may be connected to the DNA barcode at any of the three different exit vectors, thus making it a versatile approach as this readily leads to three different libraries. For one of the three possibilities, the compatibility with the DNA tag was demonstrated. The reactions include amide bond formation, Boc deprotection, urea formation, azide reduction and reductive alkylation
P. McEnaney; P. Dickson; V. Dang; A. MacConnell; V. Cavett; S. Reza; H. Park; B. Paegel; T. Kodadek PDBJ - 6AVKhttps://pdbj.org/mine/summary/6avk Abstract Streptavidin, N-[(2H-1,3-benzodioxol-5-yl)methyl]-2-({[(2H-1,3-benzodioxol-5-yl)methyl][2-(chloromethyl)-1,3-oxazole-4-carbonyl]amino}methyl)-N-[(4-carbamoyl-1,3-oxazol-2-yl)methyl]-1,3-oxazole-4-carboxamide, 1-hydroxydodecan-4-one
Robert A. Goodnow; Christopher P. Davie Annu. Rep. Med. Chem., 2017, 50, 1-15 https://doi.org/10.1016/bs.armc.2017.09.002AbstractIn the 25 years since the first publication of the concept of encoded combinatorial library synthesis with sequences of DNA, DNA-encoded library technology (DELT) has become a widely accepted method for hit finding along with high-throughput screening. This method has been shown to be useful by the numerous, valuable hits that have been published as well as by the announcement of several drug development candidates that originated from DELT hits. Multiple methods exist for directing and encoding library synthesis, demonstrating the robust nature of this informational and functional platform. In its current, commonly practiced format, this method is in theory readily applicable to any hit finding problem for soluble protein targets. The robust nature of DNA binding and recognition properties is part of the continuous evolution of DNA-encoded and DNA-directed methods
Kathleen Weisel; Nicola E. Scott; Debra J. Tompson; Bartholomew J. Votta; Sujith Madhavan; Kat Povey; Allen Wolstenholme; Monica Simeoni; Todd Rudo; Lauren Richards‐Peterson; Tarjinder Sahota; J. Gene Wang; John Lich; Joshua Finger; Adeline Verticelli; Michael Reilly; Peter J. Gough; Philip A. Harris; John Bertin; Mei‐Lun Wang Pharmacol. Res. Perspect., 2017, e00365 https://doi.org/10.1002/prp2.365AbstractGSK2982772 is a highly selective inhibitor of receptor‐interacting protein kinase 1 (RIPK1) being developed to treat chronic inflammatory diseases. This first‐in‐human study evaluated safety, tolerability, pharmacokinetics (PK), and exploratory pharmacodynamics (PD) of GSK2982772 administered orally to healthy male volunteers. This was a Phase I, randomized, placebo‐controlled, double‐blind study. In Part A, subjects received single ascending doses of GSK2982772 (0.1‐120 mg) or placebo in a crossover design during each of 4 treatment periods. In Part B, subjects received repeat doses of GSK2982772 (20 mg once daily [QD] to up to 120 mg twice daily [BID]) or placebo for 14 days. Part C was an open‐label relative bioavailability study comparing 20‐mg tablets vs capsules. Safety, tolerability, pharmacokinetics (PK), RIPK1 target engagement (TE), and pharmacodynamics (PD) were assessed. The most common adverse events (AEs) were contact dermatitis and headache. Most AEs were mild in intensity, and there were no deaths or serious AEs. The PK of GSK2982772 was approximately linear over the dose range studied (up to 120 mg BID). There was no evidence of drug accumulation upon repeat dosing. Greater than 90% RIPK1 TE was achieved over a 24‐hour period for the 60‐mg and 120‐mg BID dosing regimens. Single and repeat doses of GSK2982772 were safe and well tolerated. PK profiles showed dose linearity. The high levels of RIPK1 TE support progression into Phase II clinical trials for further clinical development.
Samuele Cazzamalli; Alberto Dal Corso; Dario Neri Chimia, 2017, 71, 712-715 https://doi.org/10.2533/chimia.2017.712AbstractCytotoxic drugs, which are commonly used for the pharmacotherapy of many forms of cancer, often cause substantial toxicity to the patient without being able to induce long-lasting remissions. Ligands specific to accessible tumor-associated targets, capable of selective localization at the neoplastic site, may facilitate the preferential delivery of anti-cancer drugs, boosting activity and helping spare normal organs. In this article, we present a critical analysis of the limitation of conventional anti-cancer drugs and we contrast monoclonal antibodies and small organic ligands, as vehicles for pharmacodelivery applications.
Joshua N. Finger; Jean-Marie Brusq; Nino Campobasso; Michael N. Cook; Jennifer Deutsch; Heather Haag; Philip A. Harris; Earl L. Jenkins; Devika Joglekar; John D. Lich; Sean Maguire; Rakesh Nagilla; Elizabeth J. Rivera; Helen Sun; Bartholomew J. Votta; John Bertin; Peter J. Gough Pharmacol. Res. Perspect., 2017, e00377 https://doi.org/10.1002/prp2.377AbstractTherapies that suppress RIPK1 kinase activity are emerging as promising therapeutic agents for the treatment of multiple inflammatory disorders. The ability to directly measure drug binding of a RIPK1 inhibitor to its target is critical for providing insight into pharmacokinetics, pharmacodynamics, safety and clinical efficacy, especially for a first-in-class small-molecule inhibitor where the mechanism has yet to be explored. Here, we report a novel method for measuring drug binding to RIPK1 protein in cells and tissues. This TEAR1 (Target Engagement Assessment for RIPK1) assay is a pair of immunoassays developed on the principle of competition, whereby a first molecule (ie, drug) prevents the binding of a second molecule (ie, antibody) to the target protein. Using the TEAR1 assay, we have validated the direct binding of specific RIPK1 inhibitors in cells, blood and tissues following treatment with benzoxazepinone (BOAz) RIPK1 inhibitors. The TEAR1 assay is a valuable tool for facilitating the clinical development of the lead RIPK1 clinical candidate compound, GSK2982772, as a first-in-class RIPK1 inhibitor for the treatment of inflammatory disease.
Martina Bigatti; Alberto Dal Corso; Sara Vanetti; Samuele Cazzamalli; Ulrike Rieder; Jörg Scheuermann; Dario Neri; Filippo Sladojevich ChemMedChem, 2017, 12(21), 1748-1752 https://doi.org/10.1002/cmdc.201700569AbstractThe screening of encoded self-assembling chemical libraries allows the identification of fragment pairs that bind to adjacent pockets on target proteins of interest. For practical applications, it is necessary to link these ligand pairs into discrete organic molecules, devoid of any nucleic acid component. Here we describe the discovery of a synergistic binding pair for acid alpha-1 glycoprotein and a chemical strategy for the identification of optimal linkers, connecting the two fragments. The procedure yielded a set of small organic ligands, the best of which exhibited a dissociation constant of 9.9 nM, as measured in solution by fluorescence polarization
Andrew M. Davis; Alleyn T. Plowright; Eric Valeur Nat. Rev. Drug Discov., 2017, 16(10), 681-698 https://doi.org/10.1038/nrd.2017.146AbstractThe strong biological rationale to pursue challenging drug targets such as protein–protein interactions has stimulated the development of novel screening strategies, such as DNA-encoded libraries, to allow broader areas of chemical space to be searched. There has also been renewed interest in screening natural products, which are the result of evolutionary selection for a function, such as interference with a key signalling pathway of a competing organism. However, recent advances in several areas, such as understanding of the biosynthetic pathways for natural products, synthetic biology and the development of biosensors to detect target molecules, are now providing new opportunities to directly harness evolutionary pressure to identify and optimize compounds with desired bioactivities. Here, we describe innovations in the key components of such strategies and highlight pioneering examples that indicate the potential of the directed-evolution concept. We also discuss the scientific gaps and challenges that remain to be addressed to realize this potential more broadly in drug discovery
Ulrike Reisacher; Linda Antusch; Robert Hofsäß; Christian Schwechheimer; Benjamin Lehmann; Hans-Achim Wagenknecht Curr. Opin. Chem. Biol., 2017, 40, 119-126 https://doi.org/10.1016/j.cbpa.2017.07.011AbstractThe chemical toolbox for synthetic modification by nucleotide building blocks and postsynthetic methods delivers light-induced functions to DNA in great variety and allows not only to initiate photoinduced processes but additionally the temporal and spatial control of these artificial functions. Herein, selected light-induced artificial functions in DNA are briefly summarized. This includes the postsynthetic ‘photoclick’ labeling strategy, benzophenone and acetophenone nucleosides as photosensitizers to induce [2+2] cycloadditions, molecular switches and energy transfer based fluorophore pairs, called “DNA traffic lights”.
Mateja Klika Škopić; Suzanne Willems; Bernd Wagner; Justin Schieven; Norbert Krause; Andreas Brunschweiger Org. Biomol. Chem., 2017, 15, 8648-8654 https://doi.org/10.1039/c7ob02347bAbstractWe demonstrate a Au(I)-mediated three-component reaction to DNA-tagged highly substituted 6-oxa-1,2-diazaspiro[4.4]nonanes from either DNA-coupled aldehydes, hydrazides, or alkynols. The choice of the starting material coupled to the DNA tag was critial for the purity of the product as the DNA-aldehyde conjugate yielded the purest products, whereas the alkynol- and hydrazide conjugates returned complex product mixtures. The reaction was compatible with thymine-, cytosine-, and, surprisingly, with adenine-DNA, while guanine-containing DNA strands were degraded under the reaction conditions.
Victor Bandlow; Susanne Liese; OrcidDaniel Lauster; Kai Ludwig; Roland R. Netz; Andreas Herrmann; Oliver Seitz J. Am. Chem. Soc., 2017, 139, 45, 16389-16397 https://doi.org/10.1021/jacs.7b09967AbstractAttachment of the Influenza A virus onto host cells involves multivalent interactions between virus surface hemagglutinin (HA) and sialoside-containing glyco ligands. Despite the development of extremely powerful multivalent binders of the Influenza virus and other viruses, comparably little is known about the optimal spacing of HA ligands, which ought to bridge binding sites within or across the trimeric HA molecules. To explore the criteria for ligand economical high affinity binding, we systematically probed distance–affinity relationships by means of two differently behaving scaffold types based on (i) flexible polyethylene glycol (PEG) conjugates and (ii) rigid self-assembled DNA·PNA complexes. The bivalent scaffolds presented two sialyl-LacNAc ligands in 23–101 Å distance. A combined analysis of binding by means of microscale thermophoresis measurements and statistical mechanics models exposed the inherent limitations of PEG-based spacers. Given the distance requirements of HA, the flexibility of PEG scaffolds is too high to raise the effective concentration of glyco ligands above a value that allows interactions with the low affinity binding site. By contrast, spatial screening with less flexible, self-assembled peptide nucleic acid (PNA)·DNA complexes uncovered a well-defined and, surprisingly, bimodal distance–affinity relationship for interactions of the Influenza A virus HA with bivalent displays of the natural sialyl-LacNAc ligand. Optimal constructs conferred 103-fold binding enhancements with only two ligands. We discuss the existence of secondary binding sites and shine light on the preference for intramolecular rather than intermolecular recognition of HA trimers on the virus surface.
Stephen Middel; Cornelia H. Panse; Swantje Nawratil; Ulf Diederichsen ChemBioChem, 2017, 18, 23, 2328-2332 https://doi.org/10.1002/cbic.201700487AbstractA novel peptide–peptide ligation strategy is introduced that has the potential to provide peptide libraries of linearly or branched coupled fragments and will be suited to introduce simultaneous protein modifications at different ligation sites. Ligation is assisted by templating peptide nucleic acid (PNA) strands, and therefore, ligation specificity is solely encoded by the PNA sequence. PNA templating, in general, allows for various kinds of covalent ligation reactions. As a proof of principle, a native chemical ligation strategy was elaborated. This PNA‐templated ligation includes easy on‐resin procedures to couple linkers and PNA to the respective peptides, and a traceless photocleavage of the linker/PNA oligomer after the ligation step. A 4,5‐dimethoxy‐2‐nitrobenzaldehyde‐based linker that allowed the photocleavable linkage of two bio‐oligomers was developed.
Carlo Baggio; Parima Udompholkul; Elisa Barile; Maurizio Pellecchia ACS Chem. Biol., 2017, 12, 12, 2981-2989 https://doi.org/10.1021/acschembio.7b00717AbstractIn modern drug discovery, the ability of biophysical methods, including nuclear magnetic resonance spectroscopy or surface plasmon resonance, to detect and characterize ligand–protein interactions accurately and unambiguously makes these approaches preferred versus conventional biochemical high-throughput screening of large collections of compounds. Nonetheless, ligand screening strategies that address simultaneously potency and selectivity have not yet been fully developed. In this work, we propose a novel method for screening large collections of combinatorial libraries using enthalpy measurements as a primary screening technique. We demonstrate that selecting binders that are driven by enthalpy (ΔH) results in agents that are not only potent but also more selective for a given target. This general and novel approach, we termed ΔH screening of fPOS (enthalpy screening of focused positional scanning library), combines the principles of focused combinatorial chemistry with rapid calorimetry measurements to efficiently identify potent and selective inhibitors.
Amaury E. Fernández-Montalván; Markus Berger; Benno Kuropka; Seong Joo Koo; Volker Badock; Joerg Weiske; Vera Puetter; Simon J. Holton; Detlef Stöckigt; Antonius ter Laak; Paolo A. Centrella; Matthew A. Clark; Christoph E Dumelin; Eric A Sigel; Holly H Soutter; Dawn M. Troast; Ying Zhang; John W. Cuozzo; Anthony D. Keefe; Didier Roche; Vincent RODESCHINI; Apirat Chaikuad; Laura Díaz-Sáez; James M. Bennett; Oleg Fedorov; Kilian V. M. Huber; Jan Huebner; Hilmar Weinmann; Ingo V Hartung; Matyas Gorjanacz ACS Chem. Biol., 2017, 12, 11, 2730-2736 https://doi.org/10.1021/acschembio.7b00708AbstractATAD2 (ANCCA) is an epigenetic regulator and transcriptional co-factor, whose over-expression has been linked to the progress of various cancer types. Here we report a DNA-encoded library screen leading to the discovery of BAY-850, a potent and isoform selective inhibitor that specifically induces ATAD2 bromodomain dimerization, and prevents interactions with acetylated histones in vitro, as well as with chromatin in cells. These features qualify BAY-850 as chemical probe to explore ATAD2 biology
Wesley G. Cochrane; Amber L. Hackler; Valerie J. Cavett; Alexander K. Price; Brian M. Paegel Anal. Chem., 2017, 89, 24, 13227-13234 https://doi.org/10.1021/acs.analchem.7b03070AbstractReaction incubation is ubiquitous in high-throughput screening workflows, including those at the microfluidic droplet scale. Fully integrated microfluidic processors that generate, incubate, and sort droplets for continuous droplet screening must passively handle a flowing emulsion such that droplet-droplet incubation time variation is minimized. Here, we disclose an integrated microfluidic emulsion creamer that close packs assay droplets by draining away excess oil through microfabricated drain channels. The drained oil co-flows with creamed emulsion and then reintroduces the oil to disperse the droplets at the circuit terminus for analysis. Creamed droplet emulsion assay incubation time dispersion was 1.7%, 3-fold less than other reported incubators. The integrated, continuous emulsion creamer (ICEcreamer) was used to miniaturize and optimize measurements of various enzymatic activities (phosphodiesterase, kinase, bacterial translation) under multiple- and single-turnover conditions. Combining the ICEcreamer with current integrated microfluidic DNA-encoded library bead processors eliminates potentially cumbersome instrumentation engineering challenges and provides a rich array of target classes and assay formats for distributed small molecule discovery.
Huang-Chi Du; Hongbing Huang Bioconjugate Chem., 2017, 28, 10, 2575-2580 https://doi.org/10.1021/acs.bioconjchem.7b00416AbstractDNA-encoded chemical libraries have emerged as a cost-effective alternative to high-throughput scree (HTS) for hit identification in drug discovery. A key factor for productive DNA-encoded libraries is chemical diversity of small molecule moiety that is attached to an encoding DNA oligomer. The library structure diversity is often limited to DNA compatible chemical reactions in aqueous media. Herein, we describe a facile process of reducing aryl nitro groups to aryl amines. The new protocol offers easy operation and circumvents pyrophoric potential of the conventional method (Raney Ni). The reaction is performed in aqueous solution and does not compromise DNA structure integrity. The utility of this method is demonstrated in versatile synthesis of benzimidazoles on DNA
Jörn Saupe; Oliver Kunz; Lars Ole Haustedt; Sven Jakupovic; Christian Mang Chem. Eur. J., 2017, 23, 11784-11791 https://doi.org/10.1002/chem.201703209AbstractMacrocycles are a structural class bearing great promise for future challenges in medicinal chemistry. Nevertheless there are few flexible approaches for the rapid generation of structurally diverse macrocyclic compound collections. We have developed an efficient method for the generation of novel macrocyclic peptidebased scaffolds. The process, which we called MacroEvoLution, is based on a cyclization screening approach which gives reliable access to novel macrocyclic architectures. Classification of building blocks into specific pools ensures that scaffolds with orthogonally addressable functionalities are generated which can easily be used for the generation of structurally diverse compound libraries. The method grants rapid access to novel scaffolds with scalable synthesis (multi gram scale) and the introduction of further diversity on a late stage. Despite being developed for peptidic systems the approach can easily be extended for the synthesis of systems with a decreased peptidic character
Dongwook Kim; Rachael R. Jetson; Casey J. Krusemark Chem. Commun., 2017, 53, 9474-9477 https://doi.org/10.1039/c7cc05236gAbstractHere, we describe an immunoassay approach for the detection of enzyme activity by quantitative PCR (qPCR) or parallel DNA sequencing which relies on activity-based probes linked to barcoding DNAs. We demonstrate this technique in the detection of serine hydrolase activities using a fluorophosphonate-oligonucleotide conjugate.
Christine C. Genick; S. Kirk Wright Expert Opin. Drug Discov., 2017, 12, 897-907 https://doi.org/10.1080/17460441.2017.1349096AbstractIntroduction: There are many challenges to the drug discovery process, including the complexity of the target, its interactions, and how these factors play a role in causing the disease. Traditionally, biophysics has been used for hit validation and chemical lead optimization. With its increased throughput and sensitivity, biophysics is now being applied earlier in this process to empower target characterization and hit finding. Areas covered: In this article, the authors provide an overview of how biophysics can be utilized to assess the quality of the reagents used in screening assays, to validate potential tool compounds, to test the integrity of screening assays, and to create follow-up strategies for compound characterization. They also briefly discuss the utilization of different biophysical methods in hit validation to help avoid the resource consuming pitfalls caused by the lack of hit overlap between biophysical methods. Expert opinion: The use of biophysics early on in the drug discovery process has proven crucial to identifying and characterizing targets of complex nature. It also has enabled the identification and classification of small molecules which interact in an allosteric or covalent manner with the target. By applying biophysics in this manner and at the early stages of this process, the chances of finding chemical leads with novel mechanisms of action are increased. In the future, focused screens with biophysics as a primary readout will become increasingly common.
K. Delaney Hook; John T. Chambers; Ryan Hili Chem. Sci., 2017,8, 7072-7076 https://doi.org/10.1039/C7SC02779FAbstractWe have developed a novel high-throughput screening platform for the discovery of small-molecules catalysts for bond-forming reactions. The method employs an in vitro selection for bond-formation using amphiphilic DNA-encoded small molecules charged with reaction substrate, which enables selections to be conducted in a variety of organic or aqueous solvents. Using the amine-catalysed aldol reaction as a catalytic model and high-throughput DNA sequencing as a selection read-out, we demonstrate the 1200-fold enrichment of a known aldol catalyst from a library of 16.7-million uncompetitive library members.
Bingbing Shi; Yuqing Deng; Peng Zhao; Xiaoyu Li Bioconjugate Chem., 2017, 28, 9, 2293-2301 https://doi.org/10.1021/acs.bioconjchem.7b00343AbstractDNA-encoded chemical library (DEL) has recently emerged and become an important technology platform in biomedical research and drug discovery. DELs containing large numbers of compounds can be prepared and selected against biological targets to discover novel ligands and inhibitors. In practice, DELs are usually selected against purified and immobilized proteins using the typical “bind-wash-elute” protocol; however, selection methods compatible with non-immobilized proteins would be able to greatly expand the target scope of DELs beyond purified proteins to more complex and biologically relevant targets. Using a novel “ligate-crosslink-purify” strategy, here we report a method capable of selecting DELs against unmodified and non-immobilized protein targets. In addition, this method has shown excel-lent capability in identify binders with moderate and weak affinities.
Roman Lagoutte; Remi Patouret; Nicolas Winssinger Curr. Opin. Chem. Biol., 2017, 39, 54-63 https://doi.org/10.1016/j.cbpa.2017.05.008AbstractThere is a resurging interest in compounds that engage their target through covalent interactions. Cysteine's thiol is endowed with enhanced reactivity, making it the nucleophile of choice for covalent engagement with a ligand aligning an electrophilic trap with a cysteine residue in a target of interest. The paucity of cysteine in the proteome coupled to the fact that closely related proteins do not necessarily share a given cysteine residue enable a level of unprecedented rational target selectivity. The recent demonstration that a lysine's amine can also be engaged covalently with a mild electrophile extends the potential of covalent inhibitors. The growing database of protein structures facilitates the discovery of covalent inhibitors while the advent of proteomic technologies enables a finer resolution in the selectivity of covalently engaged proteins. Here, we discuss recent examples of discovery and design of covalent inhibitors
Xiaojie Lu; Sarah Roberts; George Joseph Franklin; Christopher P. Davie Med. Chem. Commun., 2017, 8, 1614-1617 https://doi.org/10.1039/C7MD00289KAbstractEncoded Library Technology (ELT) is a novel hit identification platform synergistic with HTS, Fragment Hit ID and Focused Screening. It provides both an ultra high-throughput and a cost-efficient tool for the discovery of small molecules that bind to protein targets of pharmaceutical interest. The success of ELT relies heavily on the chemical diversity accessed through DNA-Encoded Library (DEL) synthesis. We developed unprecedented on-DNA cross-coupling reactions, including on-DNA Pd and Cu promoted C-N cross-coupling reactions. These reported reactions have strong potential for application in DNA-Encoded Library (DEL) synthesis
Rachel K. O’Reilly; Andrew J. Turberfield; Thomas R. Wilks Acc Chem. Res., 2017, 50, 10, 2496-2509 https://doi.org/10.1021/acs.accounts.7b00280AbstractPrecise control over reactivity and molecular structure is a fundamental goal of the chemical sciences. Billions of years of evolution by natural selection have resulted in chemical systems capable of information storage, self-replication, catalysis, capture and production of light, and even cognition. In all these cases, control over molecular structure is required to achieve a particular function: without structural control, function may be impaired, unpredictable, or impossible. The search for molecules with a desired function is often achieved by synthesizing a combinatorial library, which contains many or all possible combinations of a set of chemical building blocks (BBs), and then screening this library to identify “successful” structures. The largest libraries made by conventional synthesis are currently of the order of 10^8 distinct molecules. To put this in context, there are 10^13 ways of arranging the 21 proteinogenic amino acids in chains up to 10 units long. Given that we know that a number of these compounds have potent biological activity, it would be highly desirable to be able to search them all to identify leads for new drug molecules. Large libraries of oligonucleotides can be synthesized combinatorially and translated into peptides using systems based on biological replication such as mRNA display, with selected molecules identified by DNA sequencing; but these methods are limited to BBs that are compatible with cellular machinery. In order to search the vast tracts of chemical space beyond nucleic acids and natural peptides, an alternative approach is required. DNA-templated synthesis (DTS) could enable us to meet this challenge. DTS controls chemical product formation by using the specificity of DNA hybridization to bring selected reactants into close proximity, and is capable of the programmed synthesis of many distinct products in the same reaction vessel. By making use of dynamic, programmable DNA processes, it is possible to engineer a system that can translate instructions coded as a sequence of DNA bases into a chemical structure—a process analogous to the action of the ribosome in living organisms but with the potential to create a much more chemically diverse set of products. It is also possible to ensure that each product molecule is tagged with its identifying DNA sequence. Compound libraries synthesized in this way can be exposed to selection against suitable targets, enriching successful molecules. The encoding DNA can then be amplified using the polymerase chain reaction and decoded by DNA sequencing. More importantly, the DNA instruction sequences can be mutated and reused during multiple rounds of amplification, translation, and selection. In other words, DTS could be used as the foundation for a system of synthetic molecular evolution, which could allow us to efficiently search a vast chemical space. This has huge potential to revolutionize materials discovery—imagine being able to evolve molecules for light harvesting, or catalysts for CO2 fixation. The field of DTS has developed to the point where a wide variety of reactions can be performed on a DNA template. Complex architectures and autonomous “DNA robots” have been implemented for the controlled assembly of BBs, and these mechanisms have in turn enabled the one-pot synthesis of large combinatorial libraries. Indeed, DTS libraries are being exploited by pharmaceutical companies and have already found their way into drug lead discovery programs. This Account explores the processes involved in DTS and highlights the challenges that remain in creating a general system for molecular discovery by evolution.
Christoph Boss; Julien Hazemann; Thierry Kimmerlin; Modest von Korff; Urs Lüthi; Oliver Peter; Thomas Sander; Romain Siegrist Chimia, 2017, 71, 667-677 https://doi.org/10.2533/chimia.2017.667AbstractIn this case study on an essential instrument of modern drug discovery, we summarize our successful efforts in the last four years toward enhancing the Actelion screening compound collection. A key organizational step was the establishment of the Compound Library Committee (CLC) in September 2013. This cross-functional team consisting of computational scientists, medicinal chemists and a biologist was endowed with a significant annual budget for regular new compound purchases. Based on an initial library analysis performed in 2013, the CLC developed a New Library Strategy. The established continuous library turn-over mode, and the screening library size of 300'000 compounds were maintained, while the structural library quality was increased. This was achieved by shifting the selection criteria from 'druglike' to 'leadlike' structures, enriching for non-flat structures, aiming for compound novelty, and increasing the ratio of higher cost 'Premium Compounds'. Novel chemical space was gained by adding natural compounds, macrocycles, designed and focused libraries to the collection, and through mutual exchanges of proprietary compounds with agrochemical companies. A comparative analysis in 2016 provided evidence for the positive impact of these measures. Screening the improved library has provided several highly promising hits, including a macrocyclic compound, that are currently followed up in different Hit-to-Lead and Lead Optimization programs. It is important to state that the goal of the CLC was not to achieve higher HTS hit rates, but to increase the chances of identified hits to serve as the basis of successful early drug discovery programs. The experience gathered so far legitimates the New Library Strategy.
Xiaojie Lu; Lijun Fan; Christopher B. Phelps; Christopher P. Davie; Christine P. Donahue Bioconjugate Chem., 2017, 28, 6, 1625-1629 https://doi.org/10.1021/acs.bioconjchem.7b00292AbstractDNA-encoded library technology (ELT) is now widely used in pharmaceutical, biotech and academic research for hit identification and target validation. New on-DNA reactions are keys to exploring greater chemical space and accessing challenging chemotypes such as configurationally constrained macrocycles. Herein, we describe the first on-DNA ring-closing metathesis (RCM) and cross-metathesis (CM) reactions promoted by fast initiating Grubbs Ru reagents. Under the optimized conditions, MgCl2 was used to protect the DNA from Ru-induced decomposition. The substrate scope for on-DNA RCM was established and the same conditions were applied to a CM reaction with good conversions.
Alix I. Chan; Lynn M. McGregor; Tara Jain; David R. Liu J. Am. Chem. Soc., 2017, 139, 30, 10192-10195 https://doi.org/10.1021/jacs.7b04880AbstractWe previously reported interaction determination using unpurified proteins (IDUP), a method to selectively amplify DNA sequences encoding ligand:target pairs from a mixture of DNA-linked small molecules and unpurified protein targets in cell lysates. In this study we applied IDUP to libraries of DNA-encoded bioactive compounds and DNA-tagged human kinases to identify ligand:protein binding partners out of 32,096 possible combinations in a single solution-phase library × library experiment. The results recapitulated known small molecule:protein interactions and also revealed that ethacrynic acid is a novel ligand and inhibitor of MAP2K6 kinase. Ethacrynic acid inhibits MAP2K6 in part through alkylation of a non-conserved cysteine residue. This work validates the ability of IDUP to discover ligands for proteins of biomedical relevance.
Ruiwu Liu; Xiaocen Li; Kit S. Lam Curr. Opin. Chem. Biol., 2017, 38, 117-126 https://doi.org/10.1016/j.cbpa.2017.03.017AbstractSeveral combinatorial methods have been developed to create focused or diverse chemical libraries with a wide range of linear or macrocyclic chemical molecules: peptides, non-peptide oligomers, peptidomimetics, small-molecules, and natural product-like organic molecules. Each combinatorial approach has its own unique high-throughput screening and encoding strategy. In this article, we provide a brief overview of combinatorial chemistry in drug discovery with emphasis on recently developed new technologies for design, synthesis, screening and decoding of combinatorial library. Examples of successful application of combinatorial chemistry in hit discovery and lead optimization are given. The limitations and strengths of combinatorial chemistry are also briefly discussed. We are now in a better position to truly leverage the power of combinatorial technologies for the discovery and development of next-generation drugs.
Dong-Yao Wang; Yan Cao; Le-Yi Zheng; Lang-Dong Chen; Xiao-Fei Chen; Zhan-Ying Hong; Zhen‐Yu Zhu; Xiaoyu Li; Yi-Feng Chai Chem. Eur. J., 2017, 23, 10906-10914 https://doi.org/10.1002/chem.201702033AbstractAccurate identification of the molecular targets of bioactive small molecules is a highly important yet challenging task in biomedical research. Previously we have developed a method named DPAL (DNA-Programmed Affinity Labelling) for labelling and identifying the cellular targets of small molecules and nucleic acids. Here we applied DPAL for the target identification of Alisertib (MLN8237), a highly specific Aurora kinase A (AKA) inhibitor and a drug candidate being tested in clinical trials for cancer treatment. Besides its well-established target of AKA, several potential new targets of MLN8237 were identified. Among them, p38 mitogen-activated protein kinase (p38) and laminin receptor (LAMR) were validated to be implicated in the anti-cancer activities of MLN8237. Interestingly, these new targets were not identified with non-DNA-based affinity probes either in our experiments or in previous studies. This work may facilitate the understanding the molecular basis of the efficacy and side-effects of MLN8237 as a clinical drug candidate. On the other hand, this work has also demonstrated that the method of DPAL could be a useful tool for target identification of bioactive small molecules.
Nadine Schneider; Nikolas Fechner; Gregory A. Landrum; Nikolaus Stiefl J. Chem. Inf. Model., 2017, 57(8), 1816-1831 https://doi.org/10.1021/acs.jcim.7b00249AbstractBig data is one of the key transformative factors which are increasingly influencing all aspects of modern life. Although this transformation brings vast opportunities it also generates novel challenges, not the least of which is organizing and searching this data deluge. The field of medicinal chemistry is not different: more and more data are being generated, for instance by technologies such as DNA encoded libraries, peptide libraries, text mining of large literature corpora, and new in silico enumeration methods. Handling those huge sets of molecules effectively is quite challenging and requires compromises that often come at the expense of the interpretability of the results. In order to find an intuitive and meaningful approach to organizing large molecular datasets, we adopted a probabilistic framework called "topic modeling" from the text-mining field. Here we present the first chemistry-related implementation of this method, which allows large molecule sets to be assigned to "chemical topics" and investigating the relationships between those. In this first study, we thoroughly evaluate this novel method in different experiments and discuss both its disadvantages and advantages. We show very promising results in reproducing human-assigned concepts by using the approach to identify and retrieve chemical series from sets of molecules. We have also created an intuitive visualization of the chemical topics output by the algorithm. This is a huge benefit compared to other unsupervised machine-learning methods, like clustering, which are commonly used to group sets of molecules. Finally, we applied the new method to the 1.6 million molecules of the ChEMBL22 dataset to test its robustness and efficiency. In about 1h we built a 100-topic model of this large dataset in which we could identify interesting topics like "proteins", "DNA" or "steroids". Along with this publication we provide our datasets and an open-source implementation of the new method (CheTo) which will be part of an upcoming version of the open-source cheminformatics toolkit RDKit.
Gunther Zimmermann; Yizhou Li; Ulrike Rieder; Martin Mattarella; Dario Neri; Jörg Scheuermann ChemBioChem, 2017, 18(9), 853-857 https://doi.org/10.1002/cbic.201600637AbstractA multiplexing approach: DNA‐encoded chemical libraries can be screened on target proteins of interest. Hits need to be confirmed by resynthesis and by performing affinity measurements. We present new methods for hit validation, based on hybridization of oligonucleotide conjugates with fluorescently labeled complementary oligonucleotides and on fluorescence anisotropy, Alphascreen, and microscale thermophoresis technology. Abstract DNA‐encoded chemical libraries (DECLs) are large collections of compounds linked to DNA fragments, serving as amplifiable barcodes, which can be screened on target proteins of interest. In typical DECL selections, preferential binders are identified by high‐throughput DNA sequencing, by comparing their frequency before and after the affinity capture step. Hits identified in this procedure need to be confirmed, by resynthesis and by performing affinity measurements. In this article we present new methods based on hybridization of oligonucleotide conjugates with fluorescently labeled complementary oligonucleotides; these facilitate the determination of affinity constants and kinetic dissociation constants. The experimental procedures were demonstrated with acetazolamide, a binder to carbonic anhydrase IX with a dissociation constant in the nanomolar range. The detection of binding events was compatible not only with fluorescence polarization methodologies, but also with Alphascreen technology and with microscale thermophoresis.
Margherita Di Pisa; Oliver Seitz ChemMedChem, 2017, 12, 872-882 https://doi.org/10.1002/cmdc.201700266AbstractNucleic acid directed bioorthogonal reactions offer the fascinating opportunity to unveil and redirect a plethora of intracellular mechanisms. Nano‐ to picomolar amounts of specific RNA molecules serve as templates and catalyze the selective formation of molecules that 1) exert biological effects, or 2) provide measurable signals for RNA detection. Turnover of reactants on the template is a valuable asset when concentrations of RNA templates are low. The idea is to use RNA‐templated reactions to fully control the biodistribution of drugs and to push the detection limits of DNA or RNA analytes to extraordinary sensitivities. Herein we review recent and instructive examples of conditional synthesis or release of compounds for in cellulo protein interference and intracellular nucleic acid imaging.
Gunther Zimmermann; Ulrike Rieder; Davor Bajic; Sara Vanetti; Apirat Chaikuad; Stefan Knapp; Jörg Scheuermann; Martin Mattarella; Dario Neri Chem. Eur. J., 2017, 23, 8152-8155 https://doi.org/10.1002/chem.201701644AbstractWe describe the construction of a DNA-encoded chemical library comprising 148'135 members, generated through the self-assembly of two sub-libraries, containing 265 and 559 members, respectively. The library was designed to contain building blocks potentially capable of forming covalent interactions with target proteins. Selections performed against JNK1, a kinase containing a conserved cysteine residue close to the ATP binding site, revealed the preferential enrichment of a 2-phenoxynicotinic acid moiety (building block A82) and a 4-(3,4-difluorophenyl)-4-oxobut-2-enoic acid moiety (building block B272). When the two compounds were joined by a short PEG linker, the resulting bidentate binder (A82-L-B272) was able to covalently modify JNK1 in the presence of a large molar excess of glutathione (0.5 mM), used to simulate intracellular reducing conditions. By contrast, derivatives of the individual building blocks were not able to covalently modify JNK1 in the same experimental conditions. The A82-L-B272 ligand was selective over related kinases (BTK and GAK), which also contain targetable cysteine residues in the vicinity of the active site.
Andrew B MacConnell; Brian M Paegel ACS Comb. Sci., 2017, 19, 8, 524-532 https://doi.org/10.1021/acscombsci.7b00061AbstractMicrofluidic droplet-based screening of DNA-encoded one-bead-one-compound combinatorial libraries is a miniaturized, potentially widely distributable approach to small molecule discovery. In these screens, a microfluidic circuit distributes library beads into droplets of activity assay reagent, photochemically cleaves the compound from the bead, then incubates and sorts the droplets based on assay result for subsequent DNA sequencing-based hit compound structure elucidation. Pilot experimental studies revealed that Poisson statistics describe nearly all aspects of such screens, prompting the development of simulations to understand system behavior. Monte Carlo screening simulation data showed that increasing mean library sampling (ε), mean droplet occupancy, or library hit rate all increase the false discovery rate (FDR). Compounds identified as hits on k > 1 beads (the replicate k class) were much more likely to be authentic hits than singletons (k = 1), in agreement with previous findings. Here, we explain this observation by deriving an equation for authenticity, which reduces to the product of a library sampling bias term (exponential in k) and a sampling saturation term (exponential in ε) setting a threshold that the k-dependent bias must overcome. The equation thus quantitatively describes why each hit structure’s FDR is based on its k class, and further predicts the feasibility of intentionally populating droplets with multiple library beads, assaying the micro-mixtures for function, and identifying the active members by statistical deconvolution.
Jae Uk Jeong; Philip A. Harris; James Kang; Lara Leister; Yunfeng Lan; Joseph Romano; Xiaoyang Dong; Robert W. Marquis Tetrahedron Lett., 2017, 58, 23, 2306-2308 https://doi.org/10.1016/j.tetlet.2017.05.001AbstractTwo new synthetic routes were developed to prepare the RIP1 kinase inhibitor clinical candidate GSK2982772 involving a key (S)-3-amino-benzo[b][1,4]oxazepin-4-one intermediate prepared via Mitsunobu and SNAr cyclization reactions. Both routes are practical and cost effective compared to the initial medicinal chemistry route and are also applicable to kilogram scale-up to support on-going clinical studies.
Jacques Saarbach; Daniela Masi; Claudio Zambaldo; Nicolas Winssinger Bioorg. Med. Chem., 2017, 25(19), 5171-5177 https://doi.org/10.1016/j.bmc.2017.05.064AbstractPeptide nucleic acids (PNAs) derivatized with functional molecules are increasingly used in diverse biosupramolecular applications. PNAs have proven to be highly tolerant to modifications and different applications benefit from the use of modified PNAs, in particular modifications at the γ position. Herein we report simple protocols to access modified PNAs from iterative Ugi couplings which allow modular modifications at the α, β or γ position of the PNA backbone from simple starting materials. We demonstrate the utility of the method with the synthesis of several bioactive small molecules (a peptide ligand, a kinase inhibitor and a glycan)-PNA conjugates
Markus G. Rudolph BIOspektrum, 2017, 23, 2, 142-145 https://doi.org/10.1007/s12268-017-0778-4AbstractStructure-based drug design requires lead molecules, which traditionally are found by tedious and expensive screening campaigns. By contrast,screening of DNA-encoded small molecule libraries can be much more efficient. Here, the principle and application of the recently developed DNA-encoded library technology (DELT) are discussed.
V. C. Willis; N. K. Banda; K. N. Cordova; P. E. Chandra; W. H. Robinson; D. C. Cooper; D. Lugo; G. Mehta; S. Taylor; P. P. Tak; R. K. Prinjha; H. D. Lewis; V. M. Holers Clin. Exp. Immunol., 2017, 188(2), 263–274 https://doi.org/10.1111/cei.12932AbstractCitrullination of joint proteins by the protein arginine deiminase (PAD) family of enzymes is recognized increasingly as a key process in the pathogenesis of rheumatoid arthritis. This present study was undertaken to explore the efficacy of a novel PAD4‐selective inhibitor, GSK199, in the murine collagen‐induced arthritis model of rheumatoid arthritis. Mice were dosed daily from the time of collagen immunization with GSK199. Efficacy was assessed against a wide range of end‐points, including clinical disease scores, joint histology and immunohistochemistry, serum and joint citrulline levels and quantification of synovial autoantibodies using a proteomic array containing joint peptides. Administration of GSK199 at 30 mg/kg led to significant effects on arthritis, assessed both by global clinical disease activity and by histological analyses of synovial inflammation, pannus formation and damage to cartilage and bone. In addition, significant decreases in complement C3 deposition in both synovium and cartilage were observed robustly with GSK199 at 10 mg/kg. Neither the total levels of citrulline measurable in joint and serum, nor levels of circulating collagen antibodies, were affected significantly by treatment with GSK199 at any dose level. In contrast, a subset of serum antibodies reactive against citrullinated and non‐citrullinated joint peptides were reduced with GSK199 treatment. These data extend our previous demonstration of efficacy with the pan‐PAD inhibitor Cl‐amidine and demonstrate robustly that PAD4 inhibition alone is sufficient to block murine arthritis clinical and histopathological end‐points.
Dario Neri ChemBioChem, 2017, 18(9), 827-828 https://doi.org/10.1002/cbic.201700130AbstractThe availability of DNA-encoded chemical libraries containing billions of compounds facilitates the discovery of binding molecules for pharmaceutical applications and for investigating biological processes. This Special Issue highlights the use of this library technology and some of the latest developments in the field.
Takatoshi Yogo Pharmacia, 2017, 3, 261 https://doi.org/10.14894/faruawpsj.53.3_261AbstractIn the research and development of small molecule drugs, it is important to obtain good hit compounds that can expand the structure. DNA-encoded compound library technology (ELT) has a DNA tag. This is a technique in which a library of hundreds of millions of compounds is subjected to a binding test with a target protein in a mixture, and the binding compounds are identified through affinity selection, amplification of DNA tags by PCR, and DNA sequencing (Figure 1A). Has attracted attention as a hit compound identification technology in recent years because it enables high-speed screening of huge and diverse low-molecular weight compounds against target proteins. The case of GlaxoSmithKline (GSK) is introduced, and this paper introduces the research results based on the following documents.
S. Grünzner; F. V. Reddavide; C. Steinfelder; M. Cui; M. Busek; U. Klotzbach; Y. Zhang; F. Sonntag Microfluidics, BioMEMS, and Medical Microsystems XV, 2017, 1006111 https://doi.org/10.1117/12.2253840AbstractThe fast development of DNA-encoded chemical libraries (DECL) in the past 10 years has received great attention from pharmaceutical industries. It applies the selection approach for small molecular drug discovery. Because of the limited choices of DNA-compatible chemical reactions, most DNA-encoded chemical libraries have a narrow structural diversity and low synthetic yield. There is also a poor correlation between the ranking of compounds resulted from analyzing the sequencing data and the affinity measured through biochemical assays. By combining DECL with dynamical chemical library, the resulting DNA-encoded dynamic library (EDCCL) explores the thermodynamic equilibrium of reversible reactions as well as the advantages of DNA encoded compounds for manipulation/detection, thus leads to enhanced signal-to-noise ratio of the selection process and higher library quality. However, the library dynamics are caused by the weak interactions between the DNA strands, which also result in relatively low affinity of the bidentate interaction, as compared to a stable DNA duplex. To take advantage of both stably assembled dual-pharmacophore libraries and EDCCLs, we extended the concept of EDCCLs to heat-induced EDCCLs (hi-EDCCLs), in which the heat-induced recombination process of stable DNA duplexes and affinity capture are carried out separately. To replace the extremely laborious and repetitive manual process, a fully automated device will facilitate the use of DECL in drug discovery. Herein we describe a novel lab-on-a-chip platform for high throughput drug discovery with hi-EDCCL. A microfluidic system with integrated actuation was designed which is able to provide a continuous sample circulation by reducing the volume to a minimum. It consists of a cooled and a heated chamber for constant circulation. The system is capable to generate stable temperatures above 75 °C in the heated chamber to melt the double strands of the DNA and less than 15 °C in the cooled chamber, to reanneal the reshuffled library. In the binding chamber (the cooled chamber) specific retaining structures are integrated. These hold back beads functionalized with the target protein, while the chamber is continuously flushed with library molecules. Afterwards the whole system can be flushed with buffer to wash out unspecific bound molecules. Finally the protein-loaded beads with attached molecules can be eluted for further investigation.
Alessandro Zorzi; Kaycie Deyle; Christian Heinis Curr. Opin. Chem. Biol., 2017, 38, 24-29 https://doi.org/10.1016/j.cbpa.2017.02.006AbstractCyclic peptides combine several favorable properties such as good binding affinity, target selectivity and low toxicity that make them an attractive modality for the development of therapeutics. Over 40 cyclic peptide drugs are currently in clinical use and around one new cyclic peptide drug enters the market every year on average. The vast majority of clinically approved cyclic peptides are derived from natural products, such as antimicrobials or human peptide hormones. New powerful techniques based on rational design and in vitro evolution have enabled the de novo development of cyclic peptide ligands to targets for which nature does not offer solutions. A look at the cyclic peptides currently under clinical evaluation shows that several have been developed using such techniques. This new source for cyclic peptide ligands introduces a freshness to the field, and it is likely that de novo developed cyclic peptides will be in clinical use in the near future.
Zhen Gong; Guoping Hu; Qiang Li; Zhiguo Liu; Fei Wang; Xuejin Zhang; Jian Xiong; Peng Li; Yan Xu; Rujian Ma; Shuhui Chen; Jian Li Curr. Drug Discov. Technol., 2017, 14, 216-228 https://doi.org/10.2174/1570163814666170425155154AbstractHit identification is the starting point of small-molecule drug discovery and is therefore very important to the pharmaceutical industry. One of the most important approaches to identify a new hit is to screen a compound library using an in vitro assay. High-throughput screening has made great contributions to drug discovery since the 1990s but requires expensive equipment and facilities, and its success depends on the size of the compound library. Recent progress in the development of compound libraries has provided more efficient ways to identify new hits for novel drug targets, thereby helping to promote the development of the pharmaceutical industry, especially for first-in-class drugs. In this review, the sources and classification of compound libraries are summarized. The progress made in combinatorial libraries and DNA-encoded libraries is reviewed. Library design methods, especially for focused libraries, are introduced in detail. In the final part, the status of the compound libraries at WuXi is reported
Xiangyu Liu; Seungkirl Ahn; Alem W. Kahsai; Kai-Cheng Meng; Naomi R. Latorraca; Biswaranjan Pani; A. J. Venkatakrishnan; Ali Masoudi; William I. Weis; Ron O. Dror; Xin Chen; Robert J. Lefkowitz; Brian K. Kobilka Nature, 2017, 548(7668), 480-484 https://doi.org/10.1038/nature23652AbstractG-protein-coupled receptors (GPCRs) pose challenges for drug discovery efforts because of the high degree of structural homology in the orthosteric pocket, particularly for GPCRs within a single subfamily, such as the nine adrenergic receptors. Allosteric ligands may bind to less-conserved regions of these receptors and therefore are more likely to be selective. Unlike orthosteric ligands, which tonically activate or inhibit signalling, allosteric ligands modulate physiologic responses to hormones and neurotransmitters, and may therefore have fewer adverse effects. The majority of GPCR crystal structures published to date were obtained with receptors bound to orthosteric antagonists, and only a few structures bound to allosteric ligands have been reported. Compound 15 (Cmpd-15) is an allosteric modulator of the β2 adrenergic receptor (β2AR) that was recently isolated from a DNA-encoded small-molecule library. Orthosteric β-adrenergic receptor antagonists, known as beta-blockers, are amongst the most prescribed drugs in the world and Cmpd-15 is the first allosteric beta-blocker. Cmpd-15 exhibits negative cooperativity with agonists and positive cooperativity with inverse agonists. Here we present the structure of the β2AR bound to a polyethylene glycol-carboxylic acid derivative (Cmpd-15PA) of this modulator. Cmpd-15PA binds to a pocket formed primarily by the cytoplasmic ends of transmembrane segments 1, 2, 6 and 7 as well as intracellular loop 1 and helix 8. A comparison of this structure with inactive- and active-state structures of the β2AR reveals the mechanism by which Cmpd-15 modulates agonist binding affinity and signalling
Taavi K. Neklesa; James D. Winkler; Craig M. Crews Pharmacol. Ther., 2017, 174, 138-144 https://doi.org/10.1016/j.pharmthera.2017.02.027AbstractTargeted protein degradation using the PROTAC technology is emerging as a novel therapeutic method to address diseases driven by the aberrant expression of a disease-causing protein. PROTAC molecules are bifunctional small molecules that simultaneously bind a target protein and an E3-ubiquitin ligase, thus causing ubiquitination and degradation of the target protein by the proteasome. Like small molecules, PROTAC molecules possess good tissue distribution and the ability to target intracellular proteins. Herein, we highlight the advantages of protein degradation using PROTACs, and provide specific examples where degradation offers therapeutic benefit over classical enzyme inhibition. Foremost, PROTACs can degrade proteins regardless of their function. This includes the currently “undruggable” proteome, which comprises approximately 85% of all human proteins. Other beneficial aspects of protein degradation include the ability to target overexpressed and mutated proteins, as well as the potential to demonstrate prolonged pharmacodynamics effect beyond drug exposure. Lastly, due to their catalytic nature and the pre-requisite ubiquitination step, an exquisitely potent molecules with a high degree of degradation selectivity can be designed. Impressive preclinical in vitro and in vivo PROTAC data have been published, and these data have propelled the development of clinically viable PROTACs. With the molecular weight falling in the 700–1000 Da range, the delivery and bioavailability of PROTACs remain the largest hurdles on the way to the clinic. Solving these issues and demonstrating proof of concept clinical data will be the focus of many labs over the next few years.
Afaf H. El-Sagheer; Tom Brown Chem. Commun., 2017, 53, 10700-10702 https://doi.org/10.1039/C7CC00858AAbstractTemplated chemical ligation of 5′-amino and 3′-phosphate oligonucleotides was used to synthesise a 762 base pair gene for green fluorescent protein. The phosphoramidate linkage can be read by DNA polymerase and transcribed to make RNA. We also show that phosphoramidate ligation and orthogonal CuAAC-mediated DNA ligation can be used simultaneously.
Mateja Klika Škopić; Hazem Salamon; Olivia Bugain; Kathrin Jung; Anne Gohla; Lara J. Doetsch; Denise dos Santos; Avinash Bhat; Bernd Wagner; Andreas Brunschweiger Chem. Sci., 2017, 8, 3356-3361 https://doi.org/10.1039/C7SC00455AAbstractLibraries of DNA-tagged compounds are a validated screening technology for drug discovery. They are synthesized through combinatorial iterations of alternated coding and preparative synthesis steps. Thus,large chemical space can be accessed for target-based screening. However, the need to preserve the functionality of the DNA tag severely restricts the choice of chemical methods for library synthesis. Acidic organocatalysts, transition metals, and oxidants furnish diverse drug-like structures from simple starting materials, but cause loss of genetic information by depurination. A hexathymidine oligonucleotide, called “hexT” allows the chemist utilizing these classes of catalysts to access a potentially broad variety of structures in the initial step of library synthesis. We exploited its catalyst tolerance to efficiently synthesize diverse substituted b-carbolines, pyrazolines, and pyrazoles from readily available starting materials as hexT conjugates by acid- and Au(I)-catalysis, respectively. The hexT conjugates were ligated to coding DNA sequences yielding encoded screening libraries inspired by drug structures.
Alexander L. Satz; Remo Hochstrasser; Ann C. Peters ACS Comb. Sci., 2017, 19, 4, 234-238 https://doi.org/10.1021/acscombsci.7b00023AbstractTo optimize future DNA-encoded library design, we have attempted to quantify the library size at which the signal becomes undetectable. To accomplish this we (i) have calculated that percent yields of individual library members following a screen range from 0.002 to 1%, (ii) extrapolated that ∼1 million copies per library member are required at the outset of a screen, and (iii) from this extrapolation predict that false negative rates will begin to outweigh the benefit of increased diversity at library sizes >108. The above analysis is based upon a large internal data set comprising multiple screens, targets, and libraries; we also augmented our internal data with all currently available literature data. In theory, high false negative rates may be overcome by employing larger amounts of library; however, we argue that using more than currently reported amounts of library (≫10 nmoles) is impractical. The above conclusions may be generally applicable to other DNA encoded library platforms, particularly those platforms that do not allow for library amplification.
Margherita Di Pisa; Anett Hauser; Oliver Seitz ChemBioChem, 2017, 18, 9, 872-879 https://doi.org/10.1002/cbic.201600687AbstractA chemical reaction that is triggered by a specific RNA molecule might provide opportunities for the design of artificial feedback loops. We envision a peptidyl transfer reaction in which mRNA encoding an antiapoptotic protein would instruct the synthesis of apoptosis‐inducing peptides. In this study, we used the RNA‐programmed synthesis of a 16‐mer peptide derived from the BH3 domain of the protein Bak, which inhibits the antiapoptotic protein Bcl‐xL. The reaction involves the transfer of a thioester‐linked donor peptide fragment from one PNA conjugate to an acceptor peptide–PNA conjugate. We asked two key questions. What are the chemical requirements that allow RNA‐templated synthesis of a 16‐mer peptide to proceed at lower (nanomolar) concentrations of RNA, that is, the concentration range found in cancer cells? Will such reactions provide sufficient amounts of peptide product and sufficient affinity to interfere with the targeted protein–protein interaction? Perhaps surprisingly, the lengths of the peptides involved in peptidyl transfer chemistry have little effect on the achievable rate enhancements. However, the nature of the thioester C terminus, the distance between the targeted template annealing sites, and template affinity play important roles. The investigation revealed guidelines for the reaction design for peptidyl transfer with low amounts (1–10 nm) of RNA, yet still provide sufficient product to antagonize a protein–protein interaction.
Richard A. Lerner; Sydney Brenner Angew. Chem. Int. Ed. Engl., 2017, 56, 1164-1165 https://doi.org/10.1002/anie.201612143Abstract“… We envisioned an iterative system where a unique DNA tag identifier that encoded the event was appended to each newly formed molecule. These vast collections of molecules are known today as DNA‐ encoded chemical libraries (DECLs), and allow scientists to do selections on the benchtop that previously required access to large and complex high‐throughput screening centers …” Read more in the Guest Editorial by Richard A. Lerner and Sydney Brenner.
Svetlana L. Belyanskaya; Yun Ding; James Callahan; Aili Lazaar; David I. Israel ChemBioChem, 2017, 18(9), 837-842 https://doi.org/10.1002/cbic.201700014AbstractWe describe the development of a DNA‐encoded small‐molecule technology platform, and its use in discovering the soluble epoxide hydrolase (sEH) inhibitor GSK2256294. This molecule progressed through preclinical development and entered a human clinical trial where it was found to be orally bioavailable and well tolerated, demonstrating potent and dose‐dependent inhibition of sEH. Abstract DNA-encoded chemical library technology was developed with the vision of its becoming a transformational platform for drug discovery. The hope was that a new paradigm for the discovery of low-molecular-weight drugs would be enabled by combining the vast molecular diversity achievable with combinatorial chemistry, the information-encoding attributes of DNA, the power of molecular biology, and a streamlined selection-based discovery process. Here, we describe the discovery and early clinical development of GSK2256294, an inhibitor of soluble epoxide hydrolase (sEH, EPHX2), by using encoded-library technology (ELT). GSK2256294 is an orally bioavailable, potent and selective inhibitor of sEH that has a long half life and produced no serious adverse events in a first-time-in-human clinical study. To our knowledge, GSK2256294 is the first molecule discovered from this technology to enter human clinical testing and represents a realization of the vision that DNA-encoded chemical library technology can efficiently yield molecules with favorable properties that can be readily progressed into high-quality drugs.
Aleksejs Kontijevskis J. Chem. Inf. Model., 2017, 57, 4, 680-699 https://doi.org/10.1021/acs.jcim.7b00006AbstractThe emergence of DNA-encoded chemical libraries (DEL) field in past decade has attracted attention of pharmaceutical industry as a powerful mechanism for the discovery of novel drug-like hits for various biological targets. Nuevolution Chemetics technology enables DNA encoded synthesis of billions of chemically diverse drug-like small molecule compounds, and the efficient screening and optimization of these, facilitating effective identification of drug candidates at an unprecedented speed and scale. Although many approaches have been developed by the cheminformatics community for the analysis and visualization of drug-like chemical space, most of them are restricted to the analysis of maximum few millions of compounds and cannot handle collections of 10^8-10^12 compounds typical for DELs. To address this big chemical data challenge, we developed Reduced Complexity Molecular (RCM) frameworks methodology as an abstract and very general way of representing chemical structures. By further introducing RCM framework descriptors we constructed a global framework map of drug-like chemical space and demonstrate how chemical space occupied by multi-million-member drug-like Chemetics DNA-encoded libraries and virtual combinatorial libraries with >10^12 members could be analysed and mapped without a need for library enumeration. We further validate the approach by performing RCM framework-based searches in drug-like chemical universe and mapping Chemetics library selection outputs for LSD1 target on a global framework chemical space map.
Lik Hang Yuen; Raphael M. Franzini Bioconjugate Chem., 2017, 28, 4, 1076-1083 https://doi.org/10.1021/acs.bioconjchem.7b00005AbstractOligonucleotide conjugates of small molecules are widely used in chemical biology and have found increasing interest in the context of DNA-encoded chemical libraries for drug discovery. Attachment of molecules to DNA bound to the solid support is an attractive small-molecule conjugation method that permits the use of organic solvents, rigorous reaction conditions, and simple workup. However, the conjugated structures must be resistant to the harsh DNA deprotection/cleavage conditions and the stabilities of building blocks under various deprotection conditions are mostly unexplored. In the present study, we analyzed the stability of 131 structurally diverse fragments that contain amides and amide-like elements during DNA deprotection protocols. Structural features susceptible to decomposition in DNA deprotection conditions were identified and a protocol that enabled the synthesis of DNA conjugates with labile fragments on solid support was identified.
Andrew B. MacConnell; Alexander K. Price; Brian M. Paegel ACS Comb. Sci., 2017, 19, 181−192 https://doi.org/10.1021/acscombsci.6b00192AbstractDNA-encoded synthesis is rekindling interest in combinatorial compound libraries for drug discovery and in technology for automated and quantitative library screening. Here, we disclose a microfluidic circuit that enables functional screens of DNA-encoded compound beads. The device carries out library bead distribution into picoliter-scale assay reagent droplets, photochemical cleavage of compound from the bead,assay incubation, laser-induced fluorescence-based assay detection, and fluorescence-activated droplet sorting to isolate hits. DNA-encoded compound beads (10-μm diameter) displaying a photocleavable positive control inhibitor pepstatin A were mixed (1920 beads, 729 encoding sequences) with negative control beads (58 000 beads, 1728 encoding sequences) and screened for cathepsin D inhibition using a biochemical enzyme activity assay. The circuit sorted 1518 hit droplets for collection following 18 min incubation over a 240 min analysis.Visual inspection of a subset of droplets (1188 droplets) yielded a 24% false discovery rate (1166 pepstatin A beads; 366 negative control beads). Using template barcoding strategies, it was possible to count hit collection beads (1863) using next-generation sequencing data. Bead-specific barcodes enabled replicate counting, and the false discovery rate was reduced to 2.6% by only considering hit-encoding sequences that were observed on >2 beads. This work represents a complete distributable small molecule discovery platform, from microfluidic miniaturized automation to ultrahigh-throughput hit deconvolution by sequencing.
Eric Defrancq; Samir Messaoudi ChemBioChem, 2017, 18, 426-431 https://doi.org/10.1002/cbic.201600599AbstractDNA probes: New applications of Pd‐catalyzed coupling reactions for post‐conjugation of nucleic acids have been developed recently. Breakthroughs in this area might now pave the way for the development of sophisticated DNA probes potentially of great interest in chemical biology, nanotechnology, and bioanalysis, as well as in diagnostic domains. Abstract New applications of Pd-catalyzed coupling reactions (Suzuki-Miyaura, Sonogashira, and Stille–Migita coupling) for post-conjugation of nucleic acids have been developed recently. Breakthroughs in this area might now pave the way for the development of sophisticated DNA probes, which might be of great interest in chemical biology, nanotechnology, and bioanalysis, as well as in diagnostic domains.
Baptiste Thomas; Xiaojie Lu; William R. Birmingham; Kun Huang; Peter Both; Juana Elizabeth; Reyes Martinez; Robert J. Young; Christopher P. Davie; Sabine L. Flitsch ChemBioChem, 2017, 18(9), 858-863 https://doi.org/10.1002/cbic.201600678AbstractCracking the codes: Biocatalysis is particularly beneficial for DNA‐encoded libraries, as it is highly selective and can be performed in aqueous environments. In this work, we demonstrated the application of biocatalysis for the on‐DNA synthesis of carbohydrate‐based libraries by using enzymatic oxidation and glycosylation in combination with traditional organic chemistry. Abstract DNA-encoded libraries are increasingly used for the discovery of bioactive lead compounds in high-throughput screening programs against specific biological targets. Although a number of libraries are now available, they cover limited chemical space due to bias in ease of synthesis and the lack of chemical reactions that are compatible with DNA tagging. For example,compound libraries rarely contain complex biomolecules such as carbohydrates with high levels of functionality, tereochemistry,and hydrophilicity. By using biocatalysis in combination with chemical methods, we aimed to significantly expand chemical space and generate generic libraries with potentially better biocompatibility. For DNA-encoded libraries, biocatalysis is particularly advantageous, as it is highly selective and can be performed in aqueous environments, which is an essential feature for this split-and-mix library technology. In this work, we demonstrated the application of biocatalysis for the on-DNA synthesis of carbohydrate-based libraries by using enzymatic oxidation and glycosylation in combination with traditional organic chemistry.
Dalu Chang; Eric Lindberg; Nicolas Winssinger J. Am. Chem. Soc., 2017, 139, 4, 1444-1447 https://doi.org/10.1021/jacs.6b12764AbstractNucleic acid-templated reactions have attracted significant attention for nucleic acid sensing and imaging. The level of signal amplification obtained from templated reactions is a function of the template turnover, wherein the template acts as the catalyst. Herein, we report the application of a pyridinium linker that immolates upon photocatalytic reduction with a ruthenium complex to yield the fastest nucleic acid templated reaction reported to date. We show that the templated reaction turnover is limited by the duplex dissociation kinetics beyond probes longer than a 6-mer and proceeded fastest for a 5-mer PNA probe. Using a beacon architecture that masks the catalytic template, we show that this methodology can be used for nucleic acid sensing extending the analyte recognition beyond a 5-mer. The system proceeds with a catalytic efficiency of 105 M^−1 s^−1 and achieves turnover frequency of >100 h^−
Julie B. Trads; Thomas Tørring; Kurt V. Gothelf Acc. Chem. Res., 2017, 50, 6, 1367-1374 https://doi.org/10.1021/acs.accounts.6b00618AbstractConjugation of DNA to proteins is increasingly used in academia and industry to provide proteins with tags for identification or handles for hybridization to other DNA strands. Assay technologies such as immuno-PCR and proximity ligation and the imaging technology DNA-PAINT require DNA–protein conjugates. In DNA nanotechnology, the DNA handle is exploited to precisely position proteins by self-assembly. For these applications, site-selective conjugation is almost always desired because fully functional proteins are required to maintain the specificity of antibodies and the activity of enzymes. The introduction of a bioorthogonal handle at a specific position of a protein by recombinant techniques provides an excellent approach to site-specific conjugation, but for many laboratories and for applications where several proteins are to be labeled, the expression of recombinant proteins may be cumbersome. In recent years, a number of chemical methods that target conjugation to specific sites at native proteins have become available, and an overview of these methods is provided in this Account. Our laboratory has investigated DNA-templated protein conjugation (DTPC), which offers an alternative approach to site-selective conjugation of DNA to proteins. The method is inspired by the concept of DNA-templated synthesis where functional groups conjugated to DNA strands are preorganized by DNA hybridization to dramatically increase the reaction rate. In DPTC, we target metal binding sites in proteins to template selective covalent conjugation reactions. By chelation of a DNA–metal complex with a metal binding site of the protein, an electrophile on a second DNA strand is aligned for reaction with a lysine side chain on the protein in the proximity of the metal binding site. The method is quite general because approximately one-third of all wild-type proteins contain metal-binding sites, including many IgG antibodies, and it is also applicable to His-tagged proteins. This emerging field provides direct access to site-selective conjugates of DNA to commercially available proteins. In this Account, we introduce these methods to the reader and describe current developments and future aspects.
Robert A. Goodnow Jr; Christoph E. Dumelin; Anthony D. Keefe Nat. Rev. Drug Discov., 2017, 16(2), 131-147 https://doi.org/10.1038/nrd.2016.213AbstractDNA-encoded chemical library technologies are increasingly being adopted in drug discovery for hit and lead generation. DNA-encoded chemistry enables the exploration of chemical spaces four to five orders of magnitude more deeply than is achievable by traditional high-throughput screening methods. Operation of this technology requires developing a range of capabilities including aqueous synthetic chemistry, building block acquisition, oligonucleotide conjugation, large-scale molecular biological transformations, selection methodologies, PCR, sequencing, sequence data analysis and the analysis of large chemistry spaces.This Review provides an overview of the development and applications of DNA-encoded chemistry, highlighting the challenges and future directions for the use of this technology.
Philip A. Harris; Scott B. Berger; Jae U Jeong; Rakesh Nagilla; Deepak Bandyopadhyay; Nino Campobasso; Carol A. Capriotti; Julie A Cox; Lauren Dare; Xiaoyang Dong; Patrick M Eidam; Joshua N. Finger; Sandra J Hoffman; James Kang; Viera Kasparcova; Bryan W. King; Ruth Lehr; Yunfeng Lan; Lara K Leister; John D Lich; Thomas T. MacDonald; Nathan A Miller; Michael T. Ouellette; Christina S. Pao; Attiq Rahman; Michael A. Reilly; Alan R. Rendina; Elizabeth J. Rivera; Michelle C Schaeffer; Clark A. Sehon; Robert R. Singhaus; Helen H. Sun; Barbara A. Swift; Rachel D Totoritis; Anna Vossenkamper; Paris Ward; David D Wisnoski; Daohua Zhang; Robert W Marquis; Peter J. Gough; John Bertin J. Med. Chem., 2017, 60, 4, 1247-1261 https://doi.org/10.1021/acs.jmedchem.6b01751AbstractRIP1 regulates necroptosis and inflammation and may play an important role in contributing to a variety of human pathologies, including immune-mediated inflammatory diseases. Small-molecule inhibitors of RIP1 kinase that are suitable for advancement into the clinic have yet to be described. Herein, we report our lead optimization of a benzoxazepinone hit from a DNA-encoded library and the discovery and profile of clinical candidate GSK2982772 (compound 5), currently in phase 2a clinical studies for psoriasis, rheumatoid arthritis, and ulcerative colitis. Compound 5 potently binds to RIP1 with exquisite kinase specificity and has excellent activity in blocking many TNF-dependent cellular responses. Highlighting its potential as a novel anti-inflammatory agent, the inhibitor was also able to reduce spontaneous production of cytokines from human ulcerative colitis explants. The highly favorable physicochemical and ADMET properties of 5, combined with high potency, led to a predicted low oral dose in humans.
Seungkirl Ahn; Alem W. Kahsai; Biswaranjan Pani; Qin-Ting Wang; Shuai Zhao; Alissa L. Wall; Ryan T. Strachan; Dean P. Staus; Laura M. Wingler; Lillian D. Sun; Justine Sinnaeve; Minjung Choi; Ted Cho; Thomas T. Xu; Gwenn M. Hansen; Michael B. Burnett; Jane E. Lamerdin; Daniel L. Bassoni; Bryant J. Gavino; Gitte Husemoen; Eva K. Olsen; Thomas Franch; Stefano Costanzi; Xin Chen; Robert J. Lefkowitz Proc. Natl. Acad. Sci. USA, 2017, 114(7), 1708-1713 https://doi.org/10.1073/pnas.1620645114AbstractThe β2-adrenergic receptor (β2AR) has been a model system for understanding regulatory mechanisms of G-protein-coupled receptor (GPCR) actions and plays a significant role in cardiovascular and pulmonary diseases. Because all known β-adrenergic receptor drugs target the orthosteric binding site of the receptor, we set out to isolate allosteric ligands for this receptor by panning DNA-encoded small-molecule libraries comprising 190 million distinct compounds against purified human β2AR. Here, we report the discovery of a small-molecule negative allosteric modulator (antagonist), compound 15 [([4-((2S)-3-(((S)-3-(3-bromophenyl)-1-(methylamino)-1-oxopropan-2-yl)amino)-2-(2-cyclohexyl-2-phenylacetamido)-3-oxopropyl)benzamide], exhibiting a unique chemotype and low micromolar affinity for the β2AR. Binding of 15 to the receptor cooperatively enhances orthosteric inverse agonist binding while negatively modulating binding of orthosteric agonists. Studies with a specific antibody that binds to an intracellular region of the β2AR suggest that 15 binds in proximity to the G-protein binding site on the cytosolic surface of the β2AR. In cell-signaling studies, 15 inhibits cAMP production through the β2AR, but not that mediated by other Gs-coupled receptors. Compound 15 also similarly inhibits β-arrestin recruitment to the activated β2AR. This study presents an allosteric small-molecule ligand for the β2AR and introduces a broadly applicable method for screening DNA-encoded small-molecule libraries against purified GPCR targets. Importantly, such an approach could facilitate the discovery of GPCR drugs with tailored allosteric effects.
Bingbing Shi; Yu Zhou; Yiran Huang; Jianfu Zhang; Xiaoyu Li Bioorg. Med. Chem. Lett., 2017, 361-369 https://doi.org/10.1016/j.bmcl.2016.12.025AbstractDNA-encoded chemical library (DEL) has emerged as a powerful and versatile tool for ligand discovery in chemical biology research and in drug discovery. Encoding and selection methods are two of the most important technological aspects of DEL that can dictate the performance and utilities of DELs. In this digest, we have summarized recent advances on the encoding and selection strategies of DEL and also discussed the latest developments on DNA-encoded dynamic library, a new frontier in DEL research.
Yizhou Li; Gunther Zimmermann; Jörg Scheuermann; Dario Neri ChemBioChem, 2017, 18(9), 848-852 https://doi.org/10.1002/cbic.201600626AbstractPerformance perfection: We used a quantitative PCR methodology to rapidly characterize DNA‐encoded chemical library selections. This approach facilitates determination of optimal experimental conditions for the recovery of binding molecules from large combinatorial libraries. Information derived from this method complements that derived from high‐throughput sequencing of libraries, before and after selection. Abstract Phage‐display libraries and DNA‐encoded chemical libraries (DECLs) represent useful tools for the isolation of specific binding molecules from large combinatorial sets of compounds. With both methods, specific binders are recovered at the end of affinity capture procedures by using target proteins of interest immobilized on a solid support. However, although the efficiency of phage‐display selections is routinely quantified by counting the phage titer before and after the affinity capture step, no similar quantification procedures have been reported for the characterization of DECL selections. In this article, we describe the potential and limitations of quantitative PCR (qPCR) methods for the evaluation of selection efficiency by using a combinatorial chemical library with more than 35 million compounds. In the experimental conditions chosen for the selections, a quantification of DNA input/recovery over five orders of magnitude could be performed, revealing a successful enrichment of abundant binders, which could be confirmed by DNA sequencing. qPCR provided rapid information about the performance of selections, thus facilitating the optimization of experimental conditions.
Jeffrey W. Johannes; Stephanie Bates; Carl Beigie; Matthew A. Belmonte; John Breen; Shenggen Cao; Paolo A. Centrella; Matthew A. Clark; John W. Cuozzo; Christoph E. Dumelin; Andrew D. Ferguson; Sevan Habeshian; David Hargreaves; Camil Joubran; Steven Kazmirski; Anthony D. Keefe; Michelle L. Lamb; Haiye Lan; Yunxia Li; Hao Ma; Scott Mlynarski; Martin J. Packer; Philip B. Rawlins; Daniel W. Robbins; Haidong Shen; Eric A. Sigel; Holly H. Soutter; Nancy Su; Dawn M. Troast; Haiyun Wang; Kate F. Wickson; Chengyan Wu; Ying Zhang; Qiuying Zhao; Xiaolan Zheng; Alexander W. Hird ACS Med. Chem. Lett., 2017, 8, 239−244 https://doi.org/10.1021/acsmedchemlett.6b00464AbstractMcl-1 is a pro-apoptotic BH3 protein family member similar to Bcl-2 and Bcl-xL. Overexpression of Mcl-1 is often seen in various tumors and allows cancer cells to evade apoptosis. Here we report the discovery and optimization of a series of non-natural peptide Mcl-1 inhibitors. Screening of DNA-encoded libraries resulted in hit compound 1, a 1.5 μM Mcl-1 inhibitor. A subsequent crystal structure demonstrated that compound 1 bound to Mcl-1 in a β-turn conformation, such that the two ends of the peptide were close together. This proximity allowed for the linking of the two ends of the peptide to form a macrocycle. Macrocyclization resulted in an approximately 10-fold improvement in binding potency. Further exploration of a key hydrophobic interaction with Mcl-1 protein and also with the moiety that engages Arg256 led to additional potency improvements. The use of protein−ligand crystal structures and binding kinetics contributed to the design and understanding of the potency gains. Optimized compound 26 is a <3 nM Mcl-1 inhibitor, while inhibiting Bcl-2 at only 5 μM and Bcl-xL at >99 μM, and induces cleaved caspase-3 in MV4−11 cells with an IC50 of 3 μM after 6 h.
John W. Cuozzo; Paolo A. Centrella; Diana Gikunju; Sevan Habeshian; Christopher D. Hupp; Anthony D. Keefe; Eric A. Sigel; Holly H. Soutter; Heather A. Thomson; Ying Zhang; Matthew A. Clark ChemBioChem, 2017, 18(9), 864-871 https://doi.org/10.1002/cbic.201600573AbstractSub‐nanomolar BTK inhibitor: Both mechanism of action and potency were predicted by using multiple selection conditions to identify and characterize novel Bruton's tyrosine kinase (BTK) inhibitors from a DNA‐encoded library of 110 million compounds. The co‐crystal structure of the most potent compound demonstrated a novel binding mode. Abstract We have identified and characterized novel potent inhibitors of Bruton's tyrosine kinase (BTK) from a single DNA-encoded library of over 110 million compounds by using multiple parallel selection conditions, including variation in target concentration and addition of known binders to provide competition information. Distinct binding profiles were observed by comparing enrichments of library building block combinations under these conditions; one enriched only at high concentrations of BTK and was competitive with ATP, and another enriched at both high and low concentrations of BTK and was not competitive with ATP. A compound representing the latter profile showed low nanomolar potency in biochemical and cellular BTK assays. Results from kinetic mechanism of action studies were consistent with the selection profiles. Analysis of the co-crystal structure of the most potent compound demonstrated a novel binding mode that revealed a new pocket in BTK. Our results demonstrate that profile-based selection strategies using DNA-encoded libraries form the basis of a new methodology to rapidly identify small molecule inhibitors with novel binding modes to clinically relevant targets.
Lik Hang Yuen; Raphael M. Franzini ChemBioChem, 2017, 18(9), 829-836 https://doi.org/10.1002/cbic.201600567AbstractDiscovery with DECLs: Encoding with DNA sequence barcodes enables rapid and inexpensive synthesis and interrogation of compound libraries of unprecedented size. Screening DNA‐encoded chemical libraries (DECLs) is increasingly used in drug discovery, with applications in chemical biology under development. This minireview summarizes key achievements in the field and analyzes its challenges and opportunities. Abstract DNA‐encoded chemical libraries (DECLs) are pools of DNA‐tagged small molecules that enable facile screening and identification of bio‐macromolecule binders. The successful development of DECLs has led to their increasingly important role in drug development, and screening hits have entered clinical trials. In this review, we summarize the development and currently active research areas of DECLs with a focus on contributions from groups at academic institutes. We further look at opportunities and future directions of DECL research in medicinal chemistry and chemical biology based on the symbiotic relationship between academia and industry. Challenges associated with the application of DECLs in academic drug discovery are further discussed.
Lijun Fan; Christopher P. Davie ChemBioChem, 2017, 18(9), 843-847 https://doi.org/10.1002/cbic.201600563AbstractOn‐DNA epoxide opening: We report zirconium tetrakis(dodecyl sulfate)‐catalyzed ring opening of on‐DNA epoxides in water with amines, including anilines. Cyclization of the resulting on‐DNA β‐amino alcohols leads to a variety of non‐aromatic heterocycles. A DNA encoded library of β‐amino alcohols and their cyclization products was assembled and screened against biological targets of interest. Abstract DNA-encoded library technology (ELT) has spurred wide interest in the pharmaceutical industry as a powerful tool for hit and lead generation. In recent years a number of “DNA-compatible” chemical modifications have been published and used to synthesize vastly diverse screening libraries. Herein we report a newly developed, zirconium tetrakis(dodecyl sulfate)[Zr(DS)4] catalyzed ring-opening of on-DNA epoxides in water with amines, including anilines. Subsequent cyclization of the resulting on-DNA b-amino alcohols leads to a variety of biologically interesting, nonaromatic heterocycles. Under these conditions, a library of 137 million on-DNA b-amino alcohols and their cyclization products was assembled
Olivier Defert; Sandro Boland Expert Opin. Ther. Patents, 2017, 27, 4, 507-515 https://doi.org/10.1080/13543776.2017.1272579AbstractIntroduction: The Rho-kinases (ROCK), ROCK1 and ROCK2, are potent, widespread biochemical modulators which have been extensively studied. Due to the involvement of ROCKs in multiple biological processes, ROCK inhibitors have pleiotropic actions and may be of relevance for a number of therapeutic applications. The drawback is however that their use might be limited by occurrence of side effects. Areas covered: Since the publication of the latest review in 2014, there have been significant advances in the field of ROCK inhibitors. In this paper we reviewed the patents published between September 2013 and September 2016. Recent novel molecules will be described. and progress from the compounds series described in the previous review as well as any new expected therapeutic uses for ROCK inhibitors that popped up in the last three years will be examined. Expert opinion: While a number of potential applications in human for ROCK inhibitors have been reported, very few molecules are currently available to patients. In addition to fasudil, ripasudil (K-115, Kowa) was only recently approved in Japan for the treatment of glaucoma (2014). Notwithstanding some failures and subsequent discontinuation, the Pipeline of preclinical and clinical ROCK inhibitors remains significant.
Kimberly R. Mendes; Marie Lynne Malone; John Maina Ndungu; Irena Suponitsky-Kroyter; Valerie J. Cavett; Patrick J. McEnaney; Andrew B. MacConnell; Todd. M. Doran; Katharina Ronacher; Kim Stanley; Ofelia Utset; Gerhard Walzl; Brian M. Paegel; Thomas Kodadek ACS Chem. Biol., 2017, 12, 1, 234-243 https://doi.org/10.1021/acschembio.6b00855AbstractThe circulating antibody repertoire encodes a patient’s health status and pathogen exposure history, but identifying antibodies with diagnostic potential usually requires knowledge of the antigen(s). We previously circumvented this problem by screening libraries of bead-displayed small molecules against case and control serum samples to discover “epitope surrogates” (ligands of IgGs enriched in the case sample). Here, we describe an improved version of this technology that employs DNA-encoded libraries and highthroughput FACS-based screening to discover epitope surrogates that differentiate noninfectious/latent (LTB) patients from infectious/active TB (ATB) patients, which is imperative for proper treatment selection and antibiotic stewardship. Normal control/LTB (10 patients each, NCL) and ATB (10 patients) serum pools were screened against a library (5 × 10^6 beads, 448 000 unique compounds) using fluorescent antihuman IgG to label hit compound beads for FACS.Deep sequencing decoded all hit structures and each hit’s occurrence frequencies. ATB hits were pruned of NCL hits and prioritized for resynthesis based on occurrence and homology. Several structurally homologous families were identified and 16/21 resynthesized representative hits validated as selective ligands of ATB serum IgGs (p < 0.005). The native secreted TB protein Ag85B (though not the E. coli recombinant form) competed with one of the validated ligands for binding to antibodies,suggesting that it mimics a native Ag85B epitope. The use of DNA-encoded libraries and FACS-based screening in epitope surrogate discovery reveals thousands of potential hit structures. Distilling this list down to several consensus chemical structures yielded a diagnostic panel for ATB composed of thermally stable and economically produced small molecule ligands in place of protein antigens.
Amalie Kai Bentzen; Sine Reker Hadrup Cancer Immunol. Immunother., 2017, 66, 657–666 https://doi.org/10.1007/s00262-017-1971-5AbstractT cell-mediated recognition of peptide-major histocompatibility complex (pMHC) class I and II molecules is crucial for the control of intracellular pathogens and cancer, as well as for stimulation and maintenance of efficient cytotoxic responses. Such interactions may also play a role in the development of autoimmune diseases. Novel insights into this mechanism are crucial to understanding disease development and establishing new treatment strategies. MHC multimers have been used for detection of antigen-responsive T cells since the first report by Altman et al. showed that tetramerization of pMHC class I molecules provided sufficient stability to T cell receptor (TCR)-pMHC interactions, allowing detection of MHC multimer-binding T cells using flow cytometry. Since this breakthrough the scientific community has aimed for expanding the capacity of MHC multimer-based detection technologies to facilitate large-scale epitope discovery and immune monitoring in limited biological material. Screening of T cell specificity using large libraries of pMHC molecules is suitable for analyses of T cell recognition potentially at genome-wide levels rather than analyses restricted to a selection of model antigens. Such strategies provide novel insights into the immune specificities involved in disease development and response to immunotherapy, and extend fundamental knowledge related to T cell recognition patterns and cross-recognition by TCRs. MHC multimer-based technologies have now evolved from detection of 1–2 different T cell specificities per cell sample, to include more than 1000 evaluable pMHC molecules using novel technologies. Here, we provide an overview of MHC multimer-based detection technologies developed over two decades, focusing primarily on MHC class I interactions.
Kaja Rožman; Izidor Sosič; Raquel Fernandez; Robert J. Young; Alfonso Mendoza; Stanislav Gobec; Lourdes Encinas Drug Discov. Today, 2017, 22, 3, 492-502 https://doi.org/10.1016/j.drudis.2016.09.009AbstractThe increasing prevalence of multidrug-resistant strains of Mycobacterium tuberculosis is the main contributing factor in unfavorable outcomes in the treatment of tuberculosis. Studies suggest that direct inhibitors of InhA, an enoyl-ACP-reductase, might yield promising clinical candidates that can be developed into new antitubercular drugs. In this review, we describe the application of different hit-identification strategies to InhA, which clearly illustrate the druggability of its active site through distinct binding mechanisms. We further characterize four classes of InhA inhibitors that show novel binding modes, and provide evidence of their successful target engagement as well as their in vivo activity.
Hideto Maruyama; Ryota Oikawa; Mayu Hayakawa; Shono Takamori; Yasuaki Kimura; Naoko Abe; Genichiro Tsuji; Akira Matsuda; Satoshi Shuto; Yoshihiro Ito; Hiroshi Abe Nucleic Acids Research, 2017, 45, 12, 7042-7048 https://doi.org/10.1093/nar/gkx459AbstractWe developed a new approach for chemical ligation of oligonucleotides using the electrophilic phosphorothioester (EPT) group. A nucleophilic phosphorothioate group on oligonucleotides was converted into the EPT group by treatment with Sanger's reagent (1-fluoro-2,4-dinitrobenzene). EPT oligonucleotides can be isolated, stored frozen, and used for the ligation reaction. The reaction of the EPT oligonucleotide and an amino-modified oligonucleotide took place without any extra reagents at pH 7.0–8.0 at room temperature, and resulted in a ligation product with a phosphoramidate bond with a 39–85% yield. This method has potential uses in biotechnology and chemical biology.
Carl A. Machutta; Christopher S. Kollmann; Kenneth E. Lind; Xiaopeng Bai; Pan F. Chan; Jianzhong Huang; Lluis Ballell; Svetlana L. Belyanskaya; Gurdyal S. Besra; David Barros-Aguirre; Robert H. Bates; Paolo A. Centrella; Sandy S. Chang; Jing Chai; Anthony E. Choudhry; Aaron Coffin; Christopher P. Davie; Hongfeng Deng; Jianghe Deng; Yun Ding; Jason W. Dodson; David T. Fosbenner; Enoch N. Gao; Taylor L. Graham; Todd L. Graybill; Karen Ingraham; Walter P. Johnson; Bryan W. King; Christopher R Kwiatkowski; Joël Lelièvre; Yue Li; Xiaorong Liu; Quinn Lu; Ruth Lehr; Alfonso Mendoza-Losana; John Martin; Lynn McCloskey; Patti McCormick; Heather P. O’Keefe; Thomas O’Keeffe; Christina Pao; Christopher B. Phelps; Hongwei Qi; Keith Rafferty; Genaro S. Scavello; Matt S. Steiginga; Flora S. Sundersingh; Sharon M. Sweitzer; Lawrence M. Szewczuk; Amy Taylor; May Fern Toh; Juan Wang; Minghui Wang; Devan J. Wilkins; Bing Xia; Gang Yao; Jean Zhang; Jingye Zhou; Christine P. Donahue; Jeffrey A. Messer; David Holmes; Christopher C. Arico-Muendel; Andrew J. Pope; Jeffrey W. Gross; Ghotas Evindar Nat. Commun., 2017, 8, 16081 https://doi.org/10.1038/ncomms16081AbstractThe identification and prioritization of chemically tractable therapeutic targets is a significant challenge in the discovery of new medicines. We have developed a novel method that rapidly screens multiple proteins in parallel using DNA-encoded library technology (ELT). Initial efforts were focused on the efficient discovery of antibacterial leads against 119 targets from Acinetobacter baumannii and Staphylococcus aureus. The success of this effort led to the hypothesis that the relative number of ELT binders alone could be used to assess the ligandability of large sets of proteins. This concept was further explored by screening 42 targets from Mycobacterium tuberculosis. Active chemical series for six targets from our initial effort as well as three chemotypes for DHFR from M. tuberculosis are reported. The findings demonstrate that parallel ELT selections can be used to assess ligandability and highlight opportunities for successful lead and tool discovery.
Robert K. Y. Cheng; Cédric Fiez-Vandal; Oliver Schlenker; Karl Edman; Birte Aggeler; Dean G. Brown; Giles A. Brown; Robert M. Cooke; Christoph E. Dumelin; Andrew S. Doré; Stefan Geschwindner; Christoph Grebner; Nils-Olov Hermansson; Ali Jazayeri; Patrik Johansson; Louis Leong; Rudi Prihandoko; Mathieu Rappas; Holly Soutter; Arjan Snijder; Linda Sundström; Benjamin Tehan; Peter Thornton; Dawn Troast; Giselle Wiggin; Andrei Zhukov; Fiona H. Marshall; Niek Dekker Nature, 2017, 545, 112-115 https://doi.org/10.1038/nature22309AbstractProtease-activated receptors (PARs) are a family of G-protein-coupled receptors (GPCRs) that are irreversibly activated by proteolytic cleavage of the N terminus, which unmasks a tethered peptide ligand that binds and activates the transmembrane receptor domain, eliciting a cellular cascade in response to inflammatory signals and other stimuli. PARs are implicated in a wide range of diseases, such as cancer and inflammation1, 2, 3. PARs have been the subject of major pharmaceutical research efforts3 but the discovery of small-molecule antagonists that effectively bind them has proved challenging
Václav Navrátil; Jiří Schimer; Jan Tykvart; Tomáš Knedlík; Viktor Vik; Pavel Majer; Jan Konvalinka; Pavel Šácha Nucleic Acids Reaserch, 2017, 45, 2, e10 https://doi.org/10.1093/nar/gkw853AbstractHuman diseases are often diagnosed by determining levels of relevant enzymes and treated by enzyme inhibitors. We describe an assay suitable for both ultrasensitive enzyme quantification and quantitative inhibitor screening with unpurified enzymes. In the DNA-linked Inhibitor ANtibody Assay (DIANA), the target enzyme is captured by an immobilized antibody, probed with a small-molecule inhibitor attached to a reporter DNA and detected by quantitative PCR. We validate the approach using the putative cancer markers prostate-specific membrane antigen and carbonic anhydrase IX. We show that DIANA has a linear range of up to six logs and it selectively detects zeptomoles of targets in complex biological samples. DIANA's wide dynamic range permits determination of target enzyme inhibition constants using a single inhibitor concentration. DIANA also enables quantitative screening of small-molecule enzyme inhibitors using microliters of human blood serum containing picograms of target enzyme. DIANA's performance characteristics make it a superior tool for disease detection and drug discovery.
John W. Cuozzo; Paolo A. Centrella; Diana Gikunju; Sevan Habeshian; Christopher D. Hupp; Anthony D. Keefe; Eric A. Sigel; Holly H. Soutter; Heather A. Thomson; Ying Zhang; Matthew A. Clark PDBJ - 5U9Dhttps://pdbj.org/mine/summary/5u9d Abstract Tyrosine-protein kinase BTK, (R)-N-methyl-2-(3-((quinoxalin-6-ylamino)methyl)furan-2-carbonyl)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carboxamide, 1,2-ETHANEDIOL
Yun Ding; Jennifer L. DeLorey; Matthew A. Clark Bioconjugate Chem., 2016, 27, 11, 2597-2600 https://doi.org/10.1021/acs.bioconjchem.6b00541AbstractA novel Pd catalyst system, [(t-Bu)2P-(OH)]2PdCl2 (POPd) with the ligand sodium 2′-(dicyclohexylphosphino)-2,6-dimethoxy-[1,1′-biphenyl]-3-sulfonate, is reported. It effectively catalyzes the Suzuki-Miyaura coupling of challenging phenyl chlorides and pyrimidinyl chlorides that are covalently linked to a double-stranded DNA-template with various boronic acids/esters.
Gunther Zimmermann; Dario Neri Drug Discov. Today, 2016, 21(11), 1828-1834 https://doi.org/10.1016/j.drudis.2016.07.013AbstractDNA-encoded chemical libraries have emerged as a powerful tool for hit identification in the pharmaceutical industry and in academia. Similar to biological display techniques (such as phage display technology), DNA-encoded chemical libraries contain a link between the displayed chemical building block and an amplifiable genetic barcode on DNA. Using routine procedures, libraries containing millions to billions of compounds can be easily produced within a few weeks. The resulting compound libraries are screened in a single test tube against proteins of pharmaceutical interest and hits can be identified by PCR amplification of DNA barcodes and subsequent high-throughput sequencing.
Holly H. Souttera; Paolo A. Centrella; Matthew A. Clark; John W. Cuozzo; Christoph E. Dumelin; Marie-Aude Guie; Sevan Habeshian; Anthony D. Keefe; Kaitlyn M. Kennedy; Eric A. Sigel; Dawn M. Troast; Ying Zhang; Andrew D. Ferguson; Gareth Davies; Eleanor R. Stead; Jason Breed; Prashanti Madhavapeddi; Jon A. Read Proc. Natl. Acad. Sci. USA, 2016, 113(49), E7880-E7889 https://doi.org/10.1073/pnas.1610978113AbstractMillions of individuals are infected with and die from tuberculosis (TB) each year, and multidrug-resistant (MDR) strains of TB are increasingly prevalent. As such, there is an urgent need to identify novel drugs to treat TB infections. Current frontline therapies include the drug isoniazid, which inhibits the essential NADH-dependent enoyl-acyl-carrier protein (ACP) reductase, InhA. To inhibit InhA, isoniazid must be activated by the catalase-peroxidase KatG. Isoniazid resistance is linked primarily to mutations in the katG gene. Discovery of InhA inhibitors that do not require KatG activation is crucial to combat MDR TB. Multiple discovery efforts have been made against InhA in recent years. Until recently, despite achieving high potency against the enzyme, these efforts have been thwarted by lack of cellular activity. We describe here the use of DNA-encoded X-Chem (DEX) screening, combined with selection of appropriate physical properties, to identify multiple classes of InhA inhibitors with cell-based activity. The utilization of DEX screening allowed the interrogation of very large compound libraries (10^11 unique small molecules) against multiple forms of the InhA enzyme in a multiplexed format. Comparison of the enriched library members across various screening conditions allowed the identification of cofactor-specific inhibitors of InhA that do not require activation by KatG, many of which had bactericidal activity in cell-based assays.
Christopher C. Arico-Muendel Med. Chem. Commun., 2016, 7, 1898-1909 https://doi.org/10.1039/C6MD00341AAbstractDNA encoded library technology (ELT) provides access to broad chemical diversity through affinity selection. As an early adopter, GlaxoSmithKline enabled the development of ELT from proof of concept to full fledged contributor to the small molecule therapeutic pipeline. Unique benefits of the GSK incubation include uptake of large numbers of building blocks and privileged scaffolds, guidance on library design, and opportunities to add value beyond lead discovery. Soluble epoxide hydrolase and RIP1 kinase represent two case studies of ELT hits that have advanced into clinical studies. ELT also shows promise as a strategy to forecast target tractability. As an emerging technology, it benefits from incubation at the interface of industry and academia.
Tuan Trinh; Pongphak Chidchob; Hassan S. Bazzi; Hanadi F. Sleiman Chem. Commun., 2016, 52, 10914-10917 https://doi.org/10.1039/c6cc04970bAbstractWe report a micelle-templated method to enhance the reactivity of DNA with highly hydrophobic molecules. Lipids, chromophores and polymers can be conjugated to DNA in high yield and under mild conditions. This method expands the range of DNA-templated reactions for DNA-encoded libraries, oligonucleotide and drug delivery, nanopore mimetics and DNA nanotechnology.
Arun Shivalingam; Tom Brown Biochem. Soc. Trans., 2016, 44, 709-15 https://doi.org/10.1042/BST20160051AbstractNaturally occurring DNA is encoded by the four nucleobases adenine, cytosine, guanine and thymine. Yet minor chemical modifications to these bases, such as methylation, can significantly alter DNA function, and more drastic changes, such as replacement with unnatural base pairs, could expand its function. In order to realize the full potential of DNA in therapeutic and synthetic biology applications, our ability to ‘write’ long modified DNA in a controlled manner must be improved. This review highlights methods currently used for the synthesis of moderately long chemically modified nucleic acids (up to 1000 bp), their limitations and areas for future expansion.
Jean-Luc Ravanat; Thierry Douki Radiat. Phys. Chem., 2016, 128, 92-102 https://doi.org/10.1016/j.radphyschem.2016.07.007AbstractBoth UV and ionizing radiations damage DNA. Two main mechanisms, so-called direct and indirect pathways, are involved in the degradation of DNA induced by ionizing radiations. The direct effect of radiation corresponds to direct ionization of DNA (one electron ejection) whereas indirect effects are produced by reactive oxygen species generated through water radiolysis, including the highly reactive hydroxyl radicals, which damage DNA. UV (and visible) light damages DNA by again two distinct mechanisms. UVC and to a lesser extend UVB photons are directly absorbed by DNA bases, generating their excited states that are at the origin of the formation of pyrimidine dimers. UVA (and visible) light by interaction with endogenous or exogenous photosensitizers induce the formation of DNA damage through photosensitization reactions. The excited photosensitizer is able to induce either a one-electron oxidation of DNA (type I) or to produce singlet oxygen (type II) that reacts with DNA. In addition, through an energy transfer from the excited photosensitizer to DNA bases (sometime called type III mechanism) formation of pyrimidine dimers could be produced. Interestingly it has been shown recently that pyrimidine dimers are also produced by direct absorption of UVA light by DNA, even if absorption of DNA bases at these wavelengths is very low. It should be stressed that some excited photosensitizers (such as psoralens) could add directly to DNA bases to generate adducts. The review will described the differences and similarities in terms of damage formation (structure and mechanisms) between these two physical genotoxic agents.
Yun Ding; G. Joseph Franklin; Jennifer L. DeLorey; Paolo A. Centrella; Sibongile Mataruse; Matthew A. Clark; Steven R. Skinner; Svetlana L. Belyanskaya ACS Comb. Sci., 2016, 18, 625−629 https://doi.org/10.1021/acscombsci.6b00078AbstractDNA-encoded library technology (ELT) is a powerful tool for the discovery of new small-molecule ligands to various protein targets. Here we report the design and synthesis of biaryl DNA-encoded libraries based on the scaffold of 5-formyl 3-iodobenzoic acid. Three reactions on DNA template,acylation, Suzuki−Miyaura coupling and reductive amination, were applied in the library synthesis. The three cycle library of 3.5 million diversity has delivered potent hits for phosphoinositide 3-kinase α (PI3Kα).
Mirzo Kanoatov; Sergey N. Krylov Anal. Chem., 2016, 88, 14, 7421-7428 https://doi.org/10.1021/acs.analchem.6b02117AbstractKinetic capillary electrophoresis (KCE) methods are useful in the study of kinetics and equilibrium properties of interactions between DNA and its binding partners (ligands). KCE experiments are typically performed in a narrow set of “conventional” low-conductivity run buffers while DNA–ligand interactions in biological systems occur in physiological fluids, characterized by high ionic strengths. The nature and ionic strength of the buffer, in which DNA–ligand interaction occurs, can significantly influence the binding. Therefore, KCE experiments meant to study such interactions would greatly benefit if they could be performed in physiological buffers, such as phosphate buffered saline (PBS). No previous KCE studies of DNA used PBS as the run buffer. Here, we test the feasibility of using PBS as a KCE run buffer for analysis of DNA and show that its usage under standard KCE conditions renders DNA undetectable. We uncover the causes of this previously unreported detrimental effect and come up with a modification of KCE which allows one to overcome it. We apply the modified KCE method to an experimental model of a platelet-derived growth factor (PDGF) protein and its DNA aptamer, which was selected in PBS, and show that the results obtained in PBS run buffer are much closer to previously reported values than those which were obtained with a conventional low-conductivity capillary electrophoresis (CE) buffer.
Kyle E. Denton; Casey J. Krusemark Med. Chem. Commun., 2016, 7, 2020-2027 https://doi.org/10.1039/C6MD00288AAbstractAchieving sufficient enrichment of ligands from DNA-encoded libraries for detection can be difficult, particularly for low affinity ligands within highly complex libraries. To address this challenge, we present an approach for crosslinking DNA-linked ligands to target proteins using electrophilic or photoreactive groups. The approach involves the tethering of a ssDNA oligonucleotide to a DNA-encoded molecule to enable attachment of a reactive group post-synthetically via DNA hybridization. Crosslinking traps ligand–protein complexes while in solution and allows for stringent washing conditions to be applied in the subsequent purification. Five reactive groups (tosyl, NHS ester, sulfonyl fluoride, phenyl azide, and diazirine) were tested for crosslinking efficiency and specificity with three DNA-linked ligands to their target proteins. In a model selection, crosslinking resulted in improved enrichment of both high and a low affinity ligands in comparison to a selection with a solid-phase immobilized protein
Nestor Concha; Jianzhong Huang; Xiaopeng Bai; Andrew Benowitz; Pat Brady; LaShadric C. Grady; Luz Helena Kryn; David Holmes; Karen Ingraham; Qi Jin; Laura Pothier Kaushansky; Lynn McCloskey; Jeffrey A. Messer; Heather O’Keefe; Amish Patel; Alexander L. Satz; Robert H. Sinnamon; Jessica Schneck; Steve R. Skinner; Jennifer Summerfield; Amy Taylor; J. David Taylor; Ghotas Evindar; Robert A. Stavenger J. Med. Chem., 2016, 59, 15, 7299-7304 https://doi.org/10.1021/acs.jmedchem.6b00746AbstractUndecaprenyl pyrophosphate synthase (UppS) is an essential enzyme in bacterial cell wall synthesis. Here we report the discovery of Staphylococcus aureus UppS inhibitors from an Encoded Library Technology screen and demonstrate binding to the hydrophobic substrate site through cocrystallography studies. The use of bacterial strains with regulated uppS expression and inhibitor resistant mutant studies confirmed that the whole cell activity was the result of UppS inhibition, validating UppS as a druggable antibacterial target.
Nathan Fuller; Loredana Spadola; Scott Cowen; Joe Patel; Heike Schönherr; Qing Cao; Andrew McKenzie; Fredrik Edfeldt; Al Rabow; Robert Goodnow Drug Discov. Today, 2016, 21(8), 1272-83 https://doi.org/10.1016/j.drudis.2016.04.023AbstractModest success rates in fragment-based lead generation (FBLG) projects at AstraZeneca (AZ) prompted operational changes to improve performance. In this review, we summarize these changes, emphasizing the construction and composition of the AZ fragment library, screening practices and working model. We describe the profiles of the screening method for specific fragment subsets and statistically assess our ability to follow up on fragment hits through near-neighbor selection. Performance analysis of our second-generation fragment library (FL2) in screening campaigns illustrates the complementary nature of flat and 3D fragments in exploring protein-binding pockets and highlights our ability to deliver fragment hits using multiple screening techniques for various target classes. The new model has had profound impact on the successful delivery of lead series to drug discovery projects.
Asher Mullard Nat. Biotechnol., 2016, 34(5), 450-1 https://doi.org/10.1038/nbt0516-450bAbstractDiCE Molecules, a company with a new approach to DNA-encoded chemical libraries, signed a deal in March worth $2.3 billion with French pharma Sanofi. The Paris-based drugmaker will pay DiCE an initial $50 million to access DiCE’s ‘directed-evolution’ technology and up to $184 million in milestones per target. In January, Sanofi also inked a drug discovery deal with X-Chem, a biotech with a DNA-encoded drug discovery platform, based in Waltham, Massachusetts.
M. Klika Škopić; O. Bugain; K. Jung; S. Onstein; S. Brandherm; T. Kalliokoski; A.Brunschweiger Med. Chem. Commun., 2016, 7, 1957-1965 https://doi.org/10.1039/C6MD00243AAbstractSelection-based screening of large DNA-encoded libraries of drug-like small molecules is a validated method to identify bioactive compounds. Among the chemical space of bioactive compounds certain scaffold structures are well represented. These are commonly called “privileged scaffolds”. We have synthesized DNA-encoded libraries based on two representatives of these scaffolds, a benzodiazepine and a pyrazolopyrimidine, and additionally a third library based on propargyl glycine. All three core structures possess a carboxylic acid to couple them to aminolinker-modified DNA. For subsequent library synthesis they contained an amino function to which a set of carboxylic acid building blocks were coupled, and a terminal alkyne that was reacted with a set of azides to furnish triazoles. The two sets of building blocks, 114 carboxylic acids and 104 azides, were selected with the help of chemoinformatic methods in order to control the physicochemical properties of the final libraries, remove unwanted substructures, and maximize diversity. The set of building blocks contained desthiobiotin allowing for validation of library synthesis. The DNA-encoded libraries were synthesized by split-and-pool combinatorial chemistry yielding three libraries that contain 28,254 compounds together. For DNA barcoding, 5′-phosphorylated double-stranded coding DNA sequences with four base overhangs were ligated with T4 ligase. The resulting DNA-encoded libraries were compared to bioactivity databases and, though being based on core structures well-established in medicinal chemistry, showed novelty with respect to the known bioactive chemical space.
C. Zambaldo; J.-P. Daguer; J. Saarbach; S. Barluenga; N. Winssinger Med. Chem. Commun., 2016, 7, 1340-1351 https://doi.org/10.1039/C6MD00242KAbstractDNA-encoded chemical libraries are increasingly used to identify leads for drug discovery or chemical biology. Despite the resurging interest in covalent inhibitors, libraries are typically designed with synthon filtered out for reactive functionalities that can engage a target through covalent interactions. Herein, we report the synthesis of two libraries containing Michael acceptors to identify cysteine reactive ligands. We developed a simple procedure to discriminate between covalent and high affinity non-covalent inhibitors using DNA display of the library in a microarray format. The methodology was validated with known covalent and high affinity non-covalent kinase inhibitors. Screening of the library revealed novel covalent inhibitors for MEK2 and ERBB2.
L. K. Petersen; P. Blakskjær; A. Chaikuad; A. B. Christensen; J. Dietvorst; J. Holmkvist; S. Knapp; M. Kořínek; L. K. Larsen; A. E. Pedersen; S. Röhm; F. A. Sløk; N. J. V. Hansen Med. Chem. Commun., 2016, 7, 1332-1339 https://doi.org/10.1039/c6md00241bAbstractA highly specific and potent (7 nM cellular IC50) inhibitor of p38α kinase was identified directly from a 12.6 million membered DNA-encoded small molecule library. This was achieved using the high fidelity yoctoReactor technology (yR) for preparing the DNA-encoded library, and a homogeneous screening technique – the binder trap enrichment technology (BTE). Although structurally atypical to other kinase blockers,this inhibitor was found by X-ray crystallography to interact with the ATP binding site and provide strong distortion of the P-loop. Remarkably, it assumed an alternative binding mode as it lacks key features of known kinase inhibitors such as typical hinge binding motifs. Interestingly, the inhibitor bound assuming a canonical type-II (‘DFG-out’) binding mode by forming hinge hydrogen bonds with the backbone, showed excellent shape complementarity, and formed a number of specific polar interactions. Moreover, the crystal structure showed, that although buried in the p38α active site, the original DNA attachment point of the compound was accessible through a channel created by the distorted P-loop conformation. This study demonstrates the usability of DNA-encoded library technologies for identifying novel chemical matter with alternative binding modes to provide a good starting point for drug development
Robert A. Goodnow; Christopher P. Davie Med. Chem. Commun., 2016, 7, 1268-1270 https://doi.org/10.1039/C6MD90023BAbstractIntroduction about the First Boston Symposium of Encoded Library Platforms
Oliv Eidam; Alexander L. Satz Med. Chem. Commun., 2016, 7, 1323-1331 https://doi.org/10.1039/C6MD00221HAbstractDNA encoded library screens have gained recent interest as they allow for screening of millions of small molecules in a simple manner, with the goal of providing novel chemical starting points in target-based hit identification. Despite this interest, no publication describes the physical properties, novelty, or structural diversity of molecules derived from such screens, nor a comparison of productivity of different DNA encoded libraries. Here we address this gap by analysis of two DNA encoded library screens run against two protein targets employing mixtures of up to 16 different libraries. Fifty-seven exemplar small molecule compounds from 34 structurally distinct clusters were prioritized from the screening results, synthesized and tested for biochemical activity. Thirty-five of the 57 compounds possess significant biochemical activity (IC50 ≤ 10 μM). Seventeen of the 35 biochemically active compounds possess a molecular weight (MW) < 500 Dalton (Da) and clogP < 5, and 6 possess a MW < 400 Da and clogP < 4. None of the 57 DEL-derived compounds exist in the Roche corporate high throughput screening collection and public compound collections. Productivity per library was observed to be independent of library size. The most productive of the 16 investigated libraries was synthesized employing only simple chemistry. Physical properties of DEL-derived compounds correlate with average library properties when truncated sub-libraries are accounted for. Our analysis may help guide the design of future DNA encoded libraries
Yizhou Li; Elena Gabriele; Florent Samain; Nicholas Favalli; Filippo Sladojevich; Jörg Scheuermann; Dario Neri ACS Comb. Sci., 2016, 18, 8, 438-443 https://doi.org/10.1021/acscombsci.6b00058AbstractDNA-encoded combinatorial libraries are increasingly being used as tools for the discovery of small organic binding molecules to proteins of biological or pharmaceutical interest. In the majority of cases, synthetic procedures for the formation of DNA-encoded combinatorial libraries incorporate at least one step of amide bond formation between amino-modified DNA and a carboxylic acid. We investigated reaction conditions and established a methodology by using 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide, 1-hydroxy-7-azabenzotriazole and N,N′-diisopropylethylamine (EDC/HOAt/DIPEA) in combination, which provided conversions greater than 75% for 423/543 (78%) of the carboxylic acids tested. These reaction conditions were efficient with a variety of primary and secondary amines, as well as with various types of amino-modified oligonucleotides. The reaction conditions, which also worked efficiently over a broad range of DNA concentrations and reaction scales, should facilitate the synthesis of novel DNA-encoded combinatorial libraries.
Rachael R. Jetson; Casey J. Krusemark Angew. Chem. Int. Ed. Engl., 2016, 55(33), 9562-6 https://doi.org/10.1002/anie.201603387AbstractA sensing approach is applied to encode quantitative enzymatic activity information into DNA sequence populations. The method utilizes DNA‐linked peptide substrates as activity probes. Signal detection involves chemical manipulation of a probe population downstream of sample exposure and application of purifying, selective pressure for enzyme products. Selection‐induced changes in DNA abundance indicate sample activity. The detection of protein kinase, protease, and farnesyltransferase activities is demonstrated. The assays were employed to measure enzyme inhibition by small molecules and activity in cell lysates using parallel DNA sequencing or quantitative PCR. This strategy will allow the extensive infrastructure for genetic analysis to be applied to proteomic assays, which has a number of advantages in throughput, sensitivity, and sample multiplexing.
Jon A. Read; Jason Breed PDB Entry - 5G0T https://doi.org/10.2210/pdb5G0T/pdbAbstractClassification: OXIDOREDUCTASE Organism(s): Mycobacterium tuberculosis H37Rv Expression System: Escherichia coli BL21(DE3)
Jon A. Read; Jason Breed PDBe - 5g0u https://www.ebi.ac.uk/pdbe/entry/pdb/5g0uAbstractChains: A, B, C, D Length: 269 amino acids Theoretical weight: 28.55 KDa Source organism: Mycobacterium tuberculosis H37Rv Expression system: Escherichia coli BL21(DE3) UniProt: Canonical: Protein Data Bank in Europe - Knowledge Base P9WGR1 (Residues: 1-269; Coverage: 100%) Gene names: MTCY277.05, Rv1484, inhA Sequence domains: Enoyl-(Acyl carrier protein) reductase Structure domains: NAD(P)-binding Rossmann-like Domain
Jon A. Read; Jason Breed PDB Entry - 5G0S https://doi.org/10.2210/pdb5G0S/pdbAbstractClassification: OXIDOREDUCTASE Organism(s): Mycobacterium tuberculosis H37Rv Expression System: Escherichia coli BL21(DE3)
Jon A. Read; Jason Breed PDBe - 5g0w https://www.ebi.ac.uk/pdbe/entry/pdb/5g0wAbstractChains: A, B, C, D Length: 269 amino acids Theoretical weight: 28.55 KDa Source organism: Mycobacterium tuberculosis H37Rv Expression system: Escherichia coli BL21(DE3) UniProt: Canonical: Protein Data Bank in Europe - Knowledge Base P9WGR1 (Residues: 1-269; Coverage: 100%) Gene names: MTCY277.05, Rv1484, inhA Sequence domains: Enoyl-(Acyl carrier protein) reductase Structure domains: NAD(P)-binding Rossmann-like Domain
Jon A. Read; Jason Breed PDB Entry - 5G0V https://doi.org/10.2210/pdb5G0V/pdbAbstractClassification: OXIDOREDUCTASE Organism(s): Mycobacterium tuberculosis H37Rv Expression System: Escherichia coli BL21(DE3)
Takuya Machida; Som Dutt; Nicolas Winssinger Angew. Chem. Int. Ed. Engl., 2016, 55, 8595-8598 https://doi.org/10.1002/anie.201602751AbstractThe importance of spatial organization in short peptide catalysts is well recognized. We synthesized and screened a library of peptides flanked by peptide nucleic acids (PNAs) such that the peptide would be constrained in a hairpin loop upon hybridization. A screen for phosphatase activity led to the discovery of a catalyst with >25‐fold rate acceleration over the linear peptide. We demonstrated that the hybridization‐enforced folding of the peptide is necessary for activity, and designed a catalyst that is allosterically controlled using a complementary PNA sequence.
Willy Decurtins; Moreno Wichert; Raphael M Franzini; Fabian Buller; Michael A Stravs; Yixin Zhang; Dario Neri; Jörg Scheuermann Nat. Protoc., 2016, 11(4), 764-80 https://doi.org/10.1038/nprot.2016.039AbstractDNA-encoded chemical libraries (DECLs) are collections of organic compounds that are individually linked to different oligonucleotides, serving as amplifiable identification barcodes. As all compounds in the library can be identified by their DNA tags, they can be mixed and used in affinity-capture experiments on target proteins of interest. In this protocol, we describe the screening process that allows the identification of the few binding molecules within the multiplicity of library members. First, the automated affinity selection process physically isolates binding library members. Second, the DNA codes of the isolated binders are PCR-amplified and subjected to high-throughput DNA sequencing. Third, the obtained sequencing data are evaluated using a C++ program and the results are displayed using MATLAB software. The resulting selection fingerprints facilitate the discrimination of binding from nonbinding library members. The described procedures allow the identification of small organic ligands to biological targets from a DECL within 10 d.
Jiayin Bao; Svetlana M. Krylova; Leonid T. Cherney; Robert L. Hale; Svetlana L. Belyanskaya; Cynthia H. Chiu; Alex Shaginian; Christopher C. Arico-Muendel; Sergey N. Krylov Anal. Chem., 2016, 88, 10, 5498-5506 https://doi.org/10.1021/acs.analchem.6b00980AbstractSelection of target-binding ligands from DNA-encoded libraries of small molecules (DELSMs) is a rapidly developing approach in drug-lead discovery. Methods of kinetic capillary electrophoresis (KCE) may facilitate highly efficient homogeneous selection of ligands from DELSMs. However, KCE methods require accurate prediction of electrophoretic mobilities of protein–ligand complexes. Such prediction, in turn, requires a theory that would be applicable to DNA tags of different structures used in different DELSMs. Here we present such a theory. It utilizes a model of a globular protein connected, through a single point (small molecule), to a linear DNA tag containing a combination of alternating double-stranded and single-stranded DNA (dsDNA and ssDNA) regions of varying lengths. The theory links the unknown electrophoretic mobility of protein–DNA complex with experimentally determined electrophoretic mobilities of the protein and DNA. Mobility prediction was initially tested by using a protein interacting with 18 ligands of various combinations of dsDNA and ssDNA regions, which mimicked different DELSMs. For all studied ligands, deviation of the predicted mobility from the experimentally determined value was within 11%. Finally, the prediction was tested for two proteins and two ligands with a DNA tag identical to those of DELSM manufactured by GlaxoSmithKline. Deviation between the predicted and experimentally determined mobilities did not exceed 5%. These results confirm the accuracy and robustness of our model, which makes KCE methods one step closer to their practical use in selection of drug leads, and diagnostic probes from DELSMs.
Amalie Kai Bentzen; Andrea Marion Marquard; Rikke Lyngaa; Sunil Kumar Saini; Sofie Ramskov; Marco Donia; Lina Such; Andrew J. S. Furness; Nicholas McGranahan; Rachel Rosenthal; Per thor Straten; Zoltan Szallasi; Inge Marie Svane; Charles Swanton; Sergio A. Quezada; Søren Nyboe Jakobsen; Aron Charles Eklund; Sine Reker Hadrup Nat. Biotech., 2016, 34, 1037-1045 https://doi.org/10.1038/nbt.3662AbstractIdentification of the peptides recognized by individual T cells is important for understanding and treating immune-related diseases. Current cytometry-based approaches are limited to the simultaneous screening of 10–100 distinct T-cell specificities in one sample. Here we use peptide–major histocompatibility complex (MHC) multimers labeled with individual DNA barcodes to screen >1,000 peptide specificities in a single sample, and detect low-frequency CD8 T cells specific for virus- or cancer-restricted antigens. When analyzing T-cell recognition of shared melanoma antigens before and after adoptive cell therapy in melanoma patients, we observe a greater number of melanoma-specific T-cell populations compared with cytometry-based approaches. Furthermore, we detect neoepitope-specific T cells in tumor-infiltrating lymphocytes and peripheral blood from patients with non-small cell lung cancer. Barcode-labeled pMHC multimers enable the combination of functional T-cell analysis with large-scale epitope recognition profiling for the characterization of T-cell recognition in various diseases, including in small clinical samples.
Asher Mullard Nature, 2016, 530, 367–369 https://doi.org/10.1038/530367aAbstractDrug discovery is a daunting process that requires chemists to sift through millions of chemicals to find a single hit. DNA technology can dramatically speed up the search
Xia Tian; Gregory S. Basarab; Nidhal Selmi; Thierry Kogej; Ying Zhang; Matthew A. Clark; Robert A. Goodnow Med. Chem. Commun., 2016, 7, 1316-1322 https://doi.org/10.1039/C6MD00088FAbstractThe generation of novel chemical leads for clinical development is a constant challenge in the pharmaceutical industry. The synthesis of DNA-encoded libraries has emerged as a powerful method for hit and lead generation. We report the development of the tertiary amino effect reaction on DNA-tethered substrates. A variety of ortho-dialkylaminoaryl aldehydes undergo a cascade reaction involving a Knoevenagel condensation, a [1,5]-hydride shift, and a Mannich cyclization to give diversely substituted spirocycles. NMR analysis of substrates bearing an enriched double-13C label confirmed product formation. The net formation of two carbon–carbon bonds adds to the few examples of carbon–carbon forming reactions performed in presence of DNA-encoding systems.
Pamela A. Haile; Bartholomew J. Votta; Robert W. Marquis; Michael J. Bury; John F. Mehlmann; Robert Singhaus; Adam K. Charnley; Ami S. Lakdawala; Máire A. Convery; David B. Lipshutz; Biva M. Desai; Barbara Swift; Carol A. Capriotti; Scott B. Berger; Mukesh K. Mahajan; Michael A. Reilly; Elizabeth J. Rivera; Helen H. Sun; Rakesh Nagilla; Allison M. Beal; Joshua N. Finger; Michael N. Cook; Bryan W. King; Michael T. Ouellette; Rachel D. Totoritis; Maria Pierdomenico; Anna Negroni; Laura Stronati; Salvatore Cucchiara; Bartłomiej Ziółkowski; Anna Vossenkämper; Thomas T. MacDonald; Peter J. Gough; John Bertin; Linda N. Casillas J. Med. Chem., 2016, 59, 10, 4867-4880 https://doi.org/10.1021/acs.jmedchem.6b00211AbstractRIP2 kinase is a central component of the innate immune system and enables downstream signaling following activation of the pattern recognition receptors NOD1 and NOD2, leading to the production of inflammatory cytokines. Recently, several inhibitors of RIP2 kinase have been disclosed that have contributed to the fundamental understanding of the role of RIP2 in this pathway. However, because they lack either broad kinase selectivity or strong affinity for RIP2, these tools have only limited utility to assess the role of RIP2 in complex environments. We present, herein, the discovery and pharmacological characterization of GSK583, a next-generation RIP2 inhibitor possessing exquisite selectivity and potency. Having demonstrated the pharmacological precision of this tool compound, we report its use in elucidating the role of RIP2 kinase in a variety of in vitro, in vivo, and ex vivo experiments, further clarifying our understanding of the role of RIP2 in NOD1 and NOD2 mediated disease pathogenesis.
Thomas Kodadek; Patrick J. McEnaney Chem. Commun., 2016, 52, 6038-6059 https://doi.org/10.1039/C6CC00617EAbstractThere is great interest in the development of probe molecules and drug leads that would bind tightly and selectively to protein surfaces that are difficult to target with traditional molecules, such as those involved in protein–protein interactions. The currently available evidence suggests that this will require molecules that are larger and have quite different chemical properties than typical Lipinski-compliant molecules that target enzyme active sites. We describe here efforts to develop vast libraries of conformationally constrained oligomers as a potentially rich source of these molecules.
Alexander L. Satz ACS Comb. Sci., 2016, 18, 7, 415-424 https://doi.org/10.1021/acscombsci.6b00001AbstractSimulated screening of DNA encoded libraries indicates that the presence of truncated byproducts complicates the relationship between library member enrichment and equilibrium association constant (these truncates result from incomplete chemical reactions during library synthesis). Further, simulations indicate that some patterns observed in reported experimental data may result from the presence of truncated byproducts in the library mixture and not structure–activity relationships. Potential experimental methods of minimizing the presence of truncates are assessed via simulation; the relationship between enrichment and equilibrium association constant for libraries of differing purities is investigated. Data aggregation techniques are demonstrated that allow for more accurate analysis of screening results, in particular when the screened library contains significant quantities of truncates.
Marie L. Malone; Brian M. Paegel ACS Comb. Sci., 2016, 18, 182−187 https://doi.org/10.1021/acscombsci.5b00198AbstractDNA-encoded synthesis can generate vastly diverse screening libraries of arbitrarily complex molecules as long as chemical reaction conditions do not compromise DNA’s informational integrity, a fundamental constraint that “DNA-compatible” reaction development does not presently address. We devised DNA-encoded reaction rehearsal, an integrated analysis of reaction yield and impact on DNA, to acquire these key missing data. Magnetic DNA-functionalized sensor beads quantitatively report the % DNA template molecules remaining viable for PCR amplification after exposure to test reaction conditions. Analysis of solid-phase bond forming (e.g., Suzuki−Miyaura cross-coupling, reductive amination) and Deprotection reactions (e.g., allyl esters, silyl ethers) guided the definition and optimization of DNA-compatible reaction conditions (>90% yield, >30% viable DNA molecules), most notably in cases that involved known (H+, Pd) and more obscure (Δ, DMF) hazards to DNA integrity. The data provide an empirical yet mechanistically consistent and predictive framework for designing successful DNA-encoded reaction sequences for combinatorial library synthesis.
Gary R. Abel; Zachary A. Calabrese; Jeffrey Ayco; Jason E. Hein; Tao Ye Bioconjugate Chem., 2016, 27, 698−704 https://doi.org/10.1021/acs.bioconjchem.5b00665AbstractWe have used the quantitative polymerase chain reaction (qPCR) to measure the extent of oxidative DNA damage under varying reaction conditions used for copper(I)-catalyzed click chemistry. We systematically studied how the damage depends on a number of key reaction parameters, including the amounts of copper, ascorbate, and ligand used, and found that the damage is significant under nearly all conditions tested, including those commonly used for bioconjugation. Furthermore, we discovered that the addition of dimethyl sulfoxide, a known radical scavenger, into the aqueous mixture dramatically suppresses DNA damage during the reaction. We also measured the efficiency of cross-linking two short synthetic oligonucleotides via click chemistry, and found that the reaction could proceed reasonably efficiently even with DMSO present. This approach for screening both DNA damage and reactivity under a range of reaction conditions will be valuable for improving the biocompatibility of click chemistry, and should help to extend this powerful synthetic tool for both in vitro and in vivo applications.
Philip A. Harris; Bryan W. King; Deepak Bandyopadhyay; Scott B. Berger; Nino Campobasso; Carol A. Capriotti; Julie A. Cox; Lauren Dare; Xiaoyang Dong; Joshua N. Finger; LaShadric C. Grady; Sandra J. Hoffman; Jae U. Jeong; James Kang; Viera Kasparcova; Ami S. Lakdawala; Ruth Lehr; Dean E. McNulty; Rakesh Nagilla; Michael T. Ouellette; Christina S. Pao; Alan R. Rendina; Michelle C. Schaeffer; Jennifer D. Summerfield; Barbara A. Swift; Rachel D. Totoritis; Paris Ward; Aming Zhang; Daohua Zhang; Robert W. Marquis; John Bertin; Peter J. Gough J. Med. Chem., 2016, 59, 5, 2163-2178 https://doi.org/10.1021/acs.jmedchem.5b01898AbstractThe recent discovery of the role of receptor interacting protein 1 (RIP1) kinase in tumor necrosis factor (TNF)-mediated inflammation has led to its emergence as a highly promising target for the treatment of multiple inflammatory diseases. We screened RIP1 against GSK’s DNA-encoded small-molecule libraries and identified a novel highly potent benzoxazepinone inhibitor series. We demonstrate that this template possesses complete monokinase selectivity for RIP1 plus unique species selectivity for primate versus nonprimate RIP1. We elucidate the conformation of RIP1 bound to this benzoxazepinone inhibitor driving its high kinase selectivity and design specific mutations in murine RIP1 to restore potency to levels similar to primate RIP1. This series differentiates itself from known RIP1 inhibitors in combining high potency and kinase selectivity with good pharmacokinetic profiles in rodents. The favorable developability profile of this benzoxazepinone template, as exemplified by compound 14 (GSK’481), makes it an excellent starting point for further optimization into a RIP1 clinical candidate.
Raphael M. Franzini; Cassie Randolph J. Med. Chem., 2016, 59, 14, 6629-6644 https://doi.org/10.1021/acs.jmedchem.5b01874AbstractIn recent years, DNA-encoded chemical libraries (DECLs) have attracted considerable attention as a potential discovery tool in drug development. Screening encoded libraries may offer advantages over conventional hit discovery approaches and has the potential to complement such methods in pharmaceutical research. As a result of the increased application of encoded libraries in drug discovery, a growing number of hit compounds are emerging in scientific literature. In this review we evaluate reported encoded library-derived structures and identify general trends of these compounds in relation to library design parameters. We in particular emphasize the combinatorial nature of these libraries. Generally, the reported molecules demonstrate the ability of this technology to afford hits suitable for further lead development, and on the basis of them, we derive guidelines for DECL design.
Hazem Salamon; Mateja Klika Škopić; Kathrin Jung; Olivia Bugain; Andreas Brunschweiger ACS Chem. Biol., 2016, 11, 2, 296-307 https://doi.org/10.1021/acschembio.5b00981AbstractThe identification of bioactive compounds is a crucial step toward development of probes for chemical biology studies. Screening of DNA-encoded small molecule libraries (DELs) has emerged as a validated technology to interrogate vast chemical space. DELs consist of chimeric molecules composed of a low-molecular weight compound that is conjugated to a DNA identifier tag. They are screened as pooled libraries using selection to identify "hits." Screening of DELs has identified numerous bioactive compounds. Some of these molecules were instrumental in gaining a deeper understanding of biological systems. One of the main challenges in the field is the development of synthesis methodology for DELs.
Sofia Barluenga; Claudio Zambaldo; Heraklidia A. Ioannidou; Mihai Ciobanu; Pierre Morieux; Jean-Pierre Daguer; Nicolas Winssinger Bioorg. Med. Chem. Lett., 2016, 26(3), 1080-5 https://doi.org/10.1016/j.bmcl.2015.11.102AbstractDNA display of PNA-encoded libraries was used to pair fragments containing different phosphotyrosine surrogates with diverse triazoles. Microarray-based screening of the combinatorially paired fragment sets (62,500 combinations) against a prototypical phosphatase, PTP1B, was used to identify the fittest fragments. A focused library (10,000 members) covalently pairing identified fragments with linkers of different length and geometry was synthesized. Screening of the focused library against PTP1B and closely related TCPTP revealed orthogonal inhibitors. The selectivity of the identified inhibitors for PTP1B versus TCPT was confirmed by enzymatic inhibition assay.
Casey J. Krusemark; Nicolas P. Tilmans; Patrick O. Brown; Pehr B. Harbury PLoS ONE, 2016, 11(8), e0154765 https://doi.org/10.1371/journal.pone.0154765AbstractThe first demonstration that macromolecules could be evolved in a test tube was reported twenty-five years ago. That breakthrough meant that billions of years of chance discovery and refinement could be compressed into a few weeks, and provided a powerful tool that now dominates all aspects of protein engineering. A challenge has been to extend this scientific advance into synthetic chemical space: to enable the directed evolution of abiotic molecules. The problem has been tackled in many ways. These include expanding the natural genetic code to include unnatural amino acids, engineering polyketide and polypeptide synthases to produce novel products, and tagging combinatorial chemistry libraries with DNA. Importantly, there is still no small-molecule analog of directed protein evolution, i.e. a substantiated approach for optimizing complex (≥ 10^9 diversity) populations of synthetic small molecules over successive generations. We present a key advance towards this goal: a tool for genetically-programmed synthesis of small-molecule libraries from large chemical alphabets. The approach accommodates alphabets that are one to two orders of magnitude larger than any in Nature, and facilitates evolution within the chemical spaces they create. This is critical for small molecules, which are built up from numerous and highly varied chemical fragments. We report a proof-of-concept chemical evolution experiment utilizing an outsized genetic code, and demonstrate that fitness traits can be passed from an initial small-molecule population through to the great-grandchildren of that population. The results establish the practical feasibility of engineering synthetic small molecules through accelerated evolution.
Miguel Angel Aleman Garcia; Yuwei Hu; Itamar Willner Chem. Commun., 2016, 52, 2153-2156 https://doi.org/10.1039/C5CC08873AAbstractSwitchable β-cyclodextrin (β-CD)-induced hydrolysis of m-tert-butylphenyl acetate is demonstrated in the presence of supramolecular β-CD/adamantane oligonucleotide scaffolds. In one system, a duplex between a β-CD-functionalized nucleic acid and an adamantane-nucleic acid leads to a switchable catalytic system. In a second system, a β-CD/adamantane duplex is cooperatively generated by K+-stabilized G-quadruplex units. The binding of hemin to the second system yields a bifunctional DNA scaffold with alternate catalytic functions.
Aili L. Lazaar; Lucy Yang; Rebecca L. Boardley; Navin S. Goyal; Jonathan Robertson; Sandra J. Baldwin; David E. Newby; Ian B. Wilkinson; Ruth Tal‐Singer; Ruth J. Mayer; Joseph Cheriyan Br. J. Clin. Pharmacol., 2016, 81, 5, 971-979 https://doi.org/10.1111/bcp.12855AbstractAims: Endothelial‐derived epoxyeicosatrienoic acids may regulate vascular tone and are metabolized by soluble epoxide hydrolase enzymes (sEH). GSK2256294 is a potent and selective sEH inhibitor that was tested in two phase I studies. Methods: Single escalating doses of GSK2256294 2–20 mg or placebo were administered in a randomized crossover design to healthy male subjects or obese smokers. Once daily doses of 6 or 18 mg or placebo were administered for 14 days to obese smokers. Data were collected on safety, pharmacokinetics, sEH enzyme inhibition and blood biomarkers. Single doses of GSK2256294 10 mg were also administered to healthy younger males or healthy elderly males and females with and without food. Data on safety, pharmacokinetics and biliary metabolites were collected. Results: GSK2256294 was well‐tolerated with no serious adverse events (AEs) attributable to the drug. The most frequent AEs were headache and contact dermatitis. Plasma concentrations of GSK2256294 increased with single doses, with a half‐life averaging 25–43 h. There was no significant effect of age, food or gender on pharmacokinetic parameters. Inhibition of sEH enzyme activity was dose‐dependent, from an average of 41.9% on 2 mg (95% confidence interval [CI] –51.8, 77.7) to 99.8% on 20 mg (95% CI 99.3, 100.0) and sustained for up to 24 h. There were no significant changes in serum VEGF or plasma fibrinogen. Conclusions: GSK2256294 was well‐tolerated and demonstrated sustained inhibition of sEH enzyme activity. These data support further investigation in patients with endothelial dysfunction or abnormal tissue repair, such as diabetes, wound healing or COPD.
Hongfeng Deng; Jingye Zhou; Flora Sundersingh; Jeffrey A. Messer; Donald O. Somers; Myriam Ajakane; Christopher C. Arico-Muendel; Arthur Beljean; Svetlana L. Belyanskaya; Ryan Bingham; Emily Blazensky; Anne-Benedicte Boullay; Eric Boursier; Jing Chai; Paul Carter; Chun-Wa Chung; Alain Daugan; Yun Ding; Kenny Herry; Clare Hobbs; Eric Humphries; Christopher Kollmann; Van Loc Nguyen; Edwige Nicodeme; Sarah E. Smith; Nerina Dodic; Nicolas Ancellin ACS Med. Chem. Lett., 2016, 7, 4, 379-384 https://doi.org/10.1021/acsmedchemlett.5b00389AbstractTo identify BCATm inhibitors suitable for in vivo study, Encoded Library Technology (ELT) was used to affinity screen a 117 million member benzimidazole based DNA encoded library, which identified an inhibitor series with both biochemical and cellular activities. Subsequent SAR studies led to the discovery of a highly potent and selective compound, 1-(3-(5-bromothiophene-2-carboxamido)cyclohexyl)-N-methyl-2-(pyridin-2-yl)-1H-benzo[d]imidazole-5-carboxamide (8b) with much improved PK properties. X-ray structure revealed that 8b binds to the active site of BACTm in a unique mode via multiple H-bond and van der Waals interactions. After oral administration, 8b raised mouse blood levels of all three branched chain amino acids as a consequence of BCATm inhibition.
Dean G. Brown; Jonas Boström J. Med. Chem., 2016, 59, 10, 4443-4458 https://doi.org/10.1021/acs.jmedchem.5b01409AbstractAn analysis of chemical reactions used in current medicinal chemistry (2014), three decades ago (1984), and in natural product total synthesis has been conducted. The analysis revealed that of the current most frequently used synthetic reactions, none were discovered within the past 20 years and only two in the 1980s and 1990s (Suzuki-Miyaura and Buchwald-Hartwig). This suggests an inherent high bar of impact for new synthetic reactions in drug discovery. The most frequently used reactions were amide bond formation, Suzuki-Miyaura coupling, and SNAr reactions, most likely due to commercial availability of reagents, high chemoselectivity, and a pressure on delivery. We show that these practices result in overpopulation of certain types of molecular shapes to the exclusion of others using simple PMI plots. We hope that these results will help catalyze improvements in integration of new synthetic methodologies as well as new library design
Steven McGinn; David Bauer; Thomas Brefort; Liqin Dong; Afaf El-Sagheer; Abdou Elsharawy; Geraint Evans; Elin Falk-Sörqvist; Michael Forster; Simon Fredriksson; Peter Freeman; Camilla Freitag; Joachim Fritzsche; Spencer Gibson; Mats Gullberg; Marta Gut; Simon Heath; Isabelle Heath-Brun; Andrew J. Heron; Johannes Hohlbein; Rongqin Ke; Owen Lancaster; Ludovic Le Reste; Giovanni Maglia; Rodolphe Marie; Florence Mauger; Florian Mertes; Marco Mignardi; Lotte Moens; Jelle Oostmeijer; Ruud Out; Jonas Nyvold Pedersen; Fredrik Persson; Vincent Picaud; Dvir Rotem; Nadine Schracke; Jennifer Sengenes; Peer F. Stähler; Björn Stade; David Stoddart; Xia Teng; Colin D. Veal; Nathalie Zahra; Hagan Bayley; Markus Beier; Tom Brown; Cees Dekker; Björn Ekström; Henrik Flyvbjerg; Andre Franke; Simone Guenther; Achillefs N. Kapanidis; Jane Kaye; Anders Kristensen; Hans Lehrach; Jonathan Mangion; Sascha Sauer; Emile Schyns; Jörg Tost; Joop M. L. M. van Helvoort; Pieter J. van der Zaag; Jonas O. Tegenfeldt; Anthony J. Brookes; Kalim Mir; Mats Nilsson; James P. Willcocks; Ivo G. Gut N. Biotechnol., 2016, 33(3), 311-30 https://doi.org/10.1016/j.nbt.2015.10.003AbstractThe REvolutionary Approaches and Devices for Nucleic Acid analysis (READNA) project received funding from the European Commission for 41/2 years. The objectives of the project revolved around technological developments in nucleic acid analysis. The project partners have discovered, created and developed a huge body of insights into nucleic acid analysis, ranging from improvements and implementation of current technologies to the most promising sequencing technologies that constitute a 3(rd) and 4(th) generation of sequencing methods with nanopores and in situ sequencing, respectively
L. K. Petersen; P. Blakskjær; A. Chaikuad; A. B. Christensen; J. Dietvorst; J. Holmkvist; S. Knapp; M. Kořínek; L. K. Larsen; A. E. Pedersen; S. Röhm; F. A. Sløk; N. J. V. Hansen PDBJ - 5LARhttps://pdbj.org/mine/summary/5lar Abstract Mitogen-activated protein kinase 14, 5-azanyl-~{N}-[[4-[[(2~{S})-1-azanyl-4-cyclohexyl-1-oxidanylidene-butan-2-yl]carbamoyl]phenyl]methyl]-1-phenyl-pyrazole-4-carboxamide
Dixita Limbachiya; Bansari Rao; Manish K. Gupta arXiv:1607.00266https://arxiv.org/abs/1607.00266 Abstract The idea of computing with DNA was given by Tom Head in 1987, however in 1994 in a seminal paper, the actual successful experiment for DNA computing was performed by Adleman. The heart of the DNA computing is the DNA hybridization, however, it is also the source of errors. Thus the success of the DNA computing depends on the error control techniques. The classical coding theory techniques have provided foundation for the current information and communication technology (ICT). Thus it is natural to expect that coding theory will be the foundational subject for the DNA computing paradigm. For the successful experiments with DNA computing usually we design DNA strings which are sufficiently dissimilar. This leads to the construction of a large set of DNA strings which satisfy certain combinatorial and thermodynamic constraints. Over the last 16 years, many approaches such as combinatorial, algebraic, computational have been used to construct such DNA strings. In this work, we survey this interesting area of DNA coding theory by providing key ideas of the area and current known results.
Philip A. Harris; Bryan W. King; Deepak Bandyopadhyay; Scott B. Berger; Nino Campobasso; Carol A. Capriotti; Julie A. Cox; Lauren Dare; Xiaoyang Dong; Joshua N. Finger; LaShadric C. Grady; Sandra J. Hoffman; Jae U. Jeong; James Kang; Viera Kasparcova; Ami S. Lakdawala; Ruth Lehr; Dean E. McNulty; Rakesh Nagilla; Michael T. Ouellette; Christina S. Pao; Alan R. Rendina; Michelle C. Schaeffer; Jennifer D. Summerfield; Barbara A. Swift; Rachel D. Totoritis; Paris Ward; Aming Zhang; Daohua Zhang; Robert W. Marquis; John Bertin; Peter J. Gough PDBJ - 5HX6https://pdbj.org/mine/summary/5hx6 Abstract Receptor-interacting serine/threonine-protein kinase 1, 5-benzyl-N-[(3S)-5-methyl-4-oxo-2,3,4,5-tetrahydro-1,5-benzoxazepin-3-yl]-1,2-oxazole-3-carboxamide
Trisha Gura Science, 2015, 350, 1139-1140 https://doi.org/10.1126/science.350.6265.1139AbstractIn the painstaking work of synthesizing vast numbers of compounds and identifying those that are the best candidate drugs, researchers have cultivated a capable new lab assistant: DNA. At a meeting last month just outside Boston, chemists and biologists discussed the promise of DNA-encoded chemical libraries (DELs), which rely on the unique talents of DNA to track, select, and even synthesize compounds that bind to enzymes, receptors, and other biological targets. The technology is allowing basic scientists and small companies to generate impressive libraries of molecules, on a scale once reserved for big pharma, and select from them the most useful compounds. Faster, cheaper, and more versatile than traditional screening methods, DELs are a potential game-changer for academics who want to probe the workings of biological molecules, and they have already yielded drug candidates entering clinical trials.
Zining Wu; Todd L. Graybill; Xin Zeng; Michael Platchek; Jean Zhang; Vera Q. Bodmer; David D. Wisnoski; Jianghe Deng; Frank T. Coppo; Gang Yao; Alex Tamburino; Genaro Scavello; G. Joseph Franklin; Sibongile Mataruse; Katie L. Bedard; Yun Ding; Jing Chai; Jennifer Summerfield; Paolo A. Centrella; Jeffrey A. Messer; Andrew J. Pope; David I. Israel ACS Comb. Sci., 2015, 17, 12, 722-731 https://doi.org/10.1021/acscombsci.5b00124AbstractDNA-encoded small-molecule library technology has recently emerged as a new paradigm for identifying ligands against drug targets. To date, this technology has been used with soluble protein targets that are produced and used in a purified state. Here, we describe a cell-based method for identifying small-molecule ligands from DNA-encoded libraries against integral membrane protein targets. We use this method to identify novel, potent, and specific inhibitors of NK3, a member of the tachykinin family of G-protein coupled receptors (GPCRs). The method is simple and broadly applicable to other GPCRs and integral membrane proteins. We have extended the application of DNA-encoded library technology to membrane-associated targets and demonstrate the feasibility of selecting DNA-tagged, small-molecule ligands from complex combinatorial libraries against targets in a heterogeneous milieu, such as the surface of a cell.
Afaf H. El-Sagheer; Tom Brown Q. Rev. Biophys., 2015, 48(4), 429-36 https://doi.org/10.1017/S0033583515000141AbstractWe describe the development of a chemical process based on the CuAAC reaction (click chemistry) to ligate DNA strands and produce an unnatural triazole backbone linkage. The chemical reaction is templated by a complementary DNA splint which accelerates the reaction and provides the required specificity. The resultant 1,4-triazole linkage is read through by DNA and RNA polymerases and is biocompatible in bacterial and human cells. This work has implications for the synthesis of chemically modified genes and other large modified DNA and RNA constructs
Gang Li; Wenlu Zheng; Zitian Chen; Yu Zhou; Yu Liu; Junrui Yang; Yanyi Huang; Xiaoyu Li Chem. Sci., 2015, 6, 7097-7104 https://doi.org/10.1039/C5SC02467FAbstractWe report a method for the preparation and selection of DNA-encoded dynamic libraries (DEDLs). The library is composed of two sets of DNA-linked small molecules that are under dynamic exchange through DNA hybridization. Addition of the protein target shifted the equilibrium, favouring the assembly of high affinity bivalent binders. Notably, we introduced a novel locking mechanism to stop the dynamic exchange and "freeze" the equilibrium, thereby enabling downstream hit isolation and decoding by PCR amplification and DNA sequencing. Our DEDL approach has circumvented the limitation of library size and realized the analysis and selection of large dynamic libraries. In addition, this method also eliminates the requirement for modified and immobilized target proteins.
Adam K. Charnley; Máire A. Convery; Ami Lakdawala Shah; Emma Jones; Philip Hardwicke; Angela Bridges; Michael Ouellette; Rachel Totoritis; Benjamin Schwartz; Bryan W. King; David D. Wisnoski; James Kang; Patrick M. Eidam; Bartholomew J. Votta; Peter J. Gough; Robert W. Marquis; John Bertin; Linda Casillas Bioorg. Med. Chem., 2015, 23(21), 7000-6 https://doi.org/10.1016/j.bmc.2015.09.038AbstractReceptor interacting protein 2 (RIP2) is an intracellular kinase and key signaling partner for the pattern recognition receptors NOD1 and NOD2 (nucleotide-binding oligomerization domain-containing proteins 1 and 2). As such, RIP2 represents an attractive target to probe the role of these pathways in disease. In an effort to design potent and selective inhibitors of RIP2 we established a crystallographic system and determined the structure of the RIP2 kinase domain in an apo form and also in complex with multiple inhibitors including AMP-PCP (b,c-Methyleneadenosine 50-triphosphate, a non-hydrolysable adenosine triphosphate mimic) and structurally diverse ATP competitive chemotypes identified via a high-throughput screening campaign. These structures represent the first set of diverse RIP2-inhibitor co-crystal structures and demonstrate that the protein possesses the ability to adopt multiple DFG-in as well as DFG-out and C-helix out conformations. These structures reveal key protein–inhibitor structural insights and serve as the foundation for establishing a robust structure-based drug design effort to identify bothpotent and highly selective inhibitors of RIP2 kinase.
Andrew B. MacConnell; Patrick J. McEnaney; Valerie J. Cavett; Brian M. Paegel ACS Comb. Sci., 2015, 17(9), 518-34 https://doi.org/10.1021/acscombsci.5b00106AbstractThe promise of exploiting combinatorial synthesis for small molecule discovery remains unfulfilled due primarily to the "structure elucidation problem": the back-end mass spectrometric analysis that significantly restricts one-bead-one-compound (OBOC) library complexity. The very molecular features that confer binding potency and specificity, such as stereochemistry, regiochemistry, and scaffold rigidity, are conspicuously absent from most libraries because isomerism introduces mass redundancy and diverse scaffolds yield uninterpretable MS fragmentation. Here we present DNA-encoded solid-phase synthesis (DESPS), comprising parallel compound synthesis in organic solvent and aqueous enzymatic ligation of unprotected encoding dsDNA oligonucleotides. Computational encoding language design yielded 148 thermodynamically optimized sequences with Hamming string distance ≥ 3 and total read length <100 bases for facile sequencing. Ligation is efficient (70percent yield), specific, and directional over 6 encoding positions. A series of isomers served as a testbed for DESPS's utility in split-and-pool diversification. Single-bead quantitative PCR detected 9 × 10^4 molecules/bead and sequencing allowed for elucidation of each compound's synthetic history. We applied DESPS to the combinatorial synthesis of a 75 645-member OBOC library containing scaffold, stereochemical and regiochemical diversity using mixed-scale resin (160-μm quality control beads and 10-μm screening beads). Tandem DNA sequencing/MALDI-TOF MS analysis of 19 quality control beads showed excellent agreement (<1 ppt) between DNA sequence-predicted mass and the observed mass. DESPS synergistically unites the advantages of solid-phase synthesis and DNA encoding, enabling single-bead structural elucidation of complex compounds and synthesis using reactions normally considered incompatible with unprotected DNA. The widespread availability of inexpensive oligonucleotide synthesis, enzymes, DNA sequencing, and PCR make implementation of DESPS straightforward, and may prompt the chemistry community to revisit the synthesis of more complex and diverse libraries.
Hongfang Yang; Patricia F. Medeiros; Kaushik Raha; Patricia Elkins; Kenneth E. Lind; Ruth Lehr; Nicholas D. Adams; Joelle L. Burgess; Stanley J. Schmidt; Steven D. Knight; Kurt R. Auger; Michael D. Schaber; G. Joseph Franklin; Yun Ding; Jennifer L. DeLorey; Paolo A. Centrella; Sibongile Mataruse; Steven R. Skinner; Matthew A. Clark; John W. Cuozzo; Ghotas Evindar PDBJ - 4YKN https://pdbj.org/mine/summary/4yknAbstractPhosphatidylinositol 3-kinase regulatory subunit alpha,Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform fusion protein, 3-(6-methoxypyridin-3-yl)-5-[({4-[(5-methyl-1,3,4-thiadiazol-2-yl)sulfamoyl]phenyl}amino)methyl]benzoic acid
Wenlu Zheng; Gang Li; Xiaoyu Li Arch. Pharm. Res., 2015, 38, 1661-1685 https://doi.org/10.1007/s12272-015-0635-2AbstractWe have summarized these approaches in two categories: noise reduction and comparative distinction. This review also highlights the importance of choosing an integrated approach combining multiple methods to achieving success in target identification.
Raphael M. Franzini; Stefan Biendl; Gediminas Mikutis; Florent Samain; Jörg Scheuermann; Dario Neri ACS Comb. Sci., 2015, 17, 7, 393-398 https://doi.org/10.1021/acscombsci.5b00072AbstractThe potential of DNA-encoded combinatorial libraries (DECLs) as tools for hit discovery crucially relies on the availability of methods for their synthesis at acceptable purity and quality. Incomplete reactions in the presence of DNA can noticeably affect the purity of DECLs and methods to selectively remove unreacted oligonucleotide-based starting products would likely enhance the quality of DECL screening results. We describe an approach to selectively remove unreacted oligonucleotide starting products from reaction mixtures and demonstrate its applicability in the context of acylation of amino-modified DNA. Following an amide bond forming reaction, we treat unreacted amino-modified DNAs with biotinylating reagents and isolate the corresponding biotinylated oligonucleotides from the reaction mixture by affinity capture on streptavidin-coated sepharose. This approach, which yields the desired DNA-conjugate at enhanced purity, can be applied both to reactions performed in solution and to procedures in which DNA is immobilized on an anion exchange solid support.
Hongfeng Deng; Jingye Zhou; Flora S. Sundersingh; Jennifer Summerfield; Don Somers; Jeffrey A. Messer; Alexander L. Satz; Nicolas Ancellin; Christopher C. Arico-Muendel; Katie L. (Sargent) Bedard; Arthur Beljean; Svetlana L. Belyanskaya; Ryan Bingham; Sarah E. Smith; Eric Boursier; Paul Carter; Paolo A. Centrella; Matthew A. Clark; Chun-wa Chung; Christopher P. Davie; Jennifer L. Delorey; Yun Ding; G. Joseph Franklin; LaShadric C. Grady; Kenny Herry; Clare Hobbs; Christopher S. Kollmann; Barry A. Morgan; Laura J. (Pothier) Kaushansky; Quan Zhou ACS Med. Chem. Lett., 2015, 6, 8, 919-924 https://doi.org/10.1021/acsmedchemlett.5b00179AbstractAs a potential target for obesity, human BCATm was screened against more than 14 billion DNA encoded compounds of distinct scaffolds followed by off-DNA synthesis and activity confirmation. As a consequence, several series of BCATm inhibitors were discovered. One representative compound (R)-3-((1-(5-bromothiophene-2-carbonyl)pyrrolidin-3-yl)oxy)-N-methyl-2'-(methylsulfonamido)-[1,1'-biphenyl]-4-carboxamide (15e) from a novel compound library synthesized via on-DNA Suzuki-Miyaura cross-coupling showed BCATm inhibitory activity with IC50 = 2.0 μM. A protein crystal structure of 15e revealed that it binds to BCATm within the catalytic site adjacent to the PLP cofactor. The identification of this novel inhibitor series plus the establishment of a BCATm protein structure provided a good starting point for future structure-based discovery of BCATm inhibitors.
Alexander L. Satz; Jianping Cai; Yi Chen; Robert A. Goodnow; Felix Gruber; Agnieszka Kowalczyk; Ann Petersen; Goli Naderi-Oboodi; Lucja Orzechowski; Quentin Strebel Bioconjugate Chem., 2015, 26, 8, 1623-1632 https://doi.org/10.1021/acs.bioconjchem.5b00239AbstractComplex mixtures of DNA encoded small molecules may be readily interrogated via high-throughput sequencing. These DNA encoded libraries (DELs) are commonly used to discover molecules that interact with pharmaceutically relevant proteins. The chemical diversity displayed by the library is key to successful discovery of potent, novel, and drug-like chemical matter. The small molecule moieties of DELs are generally synthesized though a multistep process, and each chemical step is accomplished while it is simultaneously attached to an encoding DNA oligomer. Hence, library chemical diversity is often limited to DNA compatible synthetic reactions. Herein, protocols for 24 reactions are provided that have been optimized for high-throughput production of DELs. These protocols detail the multistep synthesis of benzimidazoles, imidazolidinones, quinazolinones, isoindolinones, thiazoles, and imidazopyridines. Additionally, protocols are provided for a diverse range of useful chemical reactions including BOC deprotection (under pH neutral conditions), carbamylation, and Sonogashira coupling. Last, step-by-step protocols for synthesizing functionalized DELs from trichloronitropyrimidine and trichloropyrimidine scaffolds are detailed.
Tuomo Kalliokoski ACS Comb. Sci., 2015, 17, 10, 600-607 https://doi.org/10.1021/acscombsci.5b00063AbstractCombinatorial libraries are synthesized by combining smaller reagents (building blocks), the price of which is an important component of the total costs associated with the synthetic exercise. A significant portion of commercially available reagents are too expensive for large scale work. In this study, 13 commonly used reagent classes in combinatorial library synthesis (primary and secondary amines, carboxylic acids, alcohols, ketones, aldehydes, boronic acids, acyl halides, sulfonyl chlorides, isocyanates, isothiocyanates, azides and chloroformates) were analyzed with respect to the cost, physicochemical properties (molecular weight and calculated logP), chemical diversity, and 3D-likeness using a large data set. The results define the chemical space accessible under a constraint of limited financial resources.
Milka Kostic Chem. Biol., 2015, 22, 427-428 https://doi.org/10.1016/j.chembiol.2015.04.007AbstractEach month, Chemistry & Biology Select highlights a selection of research reports from the recent literature. These highlights are a snapshot of interesting research done across the field of chemical biology. This installment was scheduled to be published in the March 2015 issue; however, unforeseen circumstances caused a delay in publication. Nevertheless, we think that you will enjoy reading about the research despite the delay. Our March 2015 selection includes reports on the potential new role for bile acids, dual-pharmacophore encoded self-assembling chemical libraries, how to control the Ebola virus, and how bacteria use uracil to control the host.
Yun Ding; Heather O’Keefe; Jennifer L. DeLorey; David I. Israel; Jeffrey A. Messer; Cynthia H. Chiu; Steven R. Skinner; Rosalie E. Matico; Monique F. Murray-Thompson; Fan Li; Matthew A. Clark; John W. Cuozzo; Christopher C. Arico-Muendel; Barry A. Morgan ACS Med. Chem. Lett., 2015, 6, 8, 888-893 https://doi.org/10.1021/acsmedchemlett.5b00138AbstractThe aggrecan degrading metalloprotease ADAMTS-4 has been identified as a novel therapeutic target for osteoarthritis. Here, we use DNA-encoded Library Technology (ELT) to identify novel ADAMTS-4 inhibitors from a DNA-encoded triazine library by affinity selection. Structure-activity relationship studies based on the selection information led to the identification of potent and highly selective inhibitors. For example, 4-(((4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)-6-(((4-methylpiperazin-1-yl)methyl)amino)-1,3,5-triazin-2-yl)amino)methyl)-N-ethyl-N-(m-tolyl)benzamide has IC50 of 10 nM against ADAMTS-4, with >1000-fold selectivity over ADAMT-5, MMP-13, TACE, and ADAMTS-13. These inhibitors have no obvious zinc ligand functionality.
Edgar R. Wood; Randy Bledsoe; Jing Chai; Philias Daka; Hongfeng Deng; Yun Ding; Sarah Harris-Gurley; Luz Helena Kryn; Eldridge Nartey; James Nichols; Robert T. Nolte; Ninad Prabhu; Cecil Rise; Timothy Sheahan; J. Brad Shotwell; Danielle Smith; Vince Tai; J. David Taylor; Ginger Tomberlin; Liping Wang; Bruce Wisely; Shihyun You; Bing Xia; Hamilton Dickson J. Biol. Chem., 2015, 290(32), 19681-96 https://doi.org/10.1074/jbc.M115.653113Abstract2′,5′-Oligoadenylate synthetase (OAS) enzymes and RNase-L constitute a major effector arm of interferon (IFN)-mediated antiviral defense. OAS produces a unique oligonucleotide second messenger, 2′,5′-oligoadenylate (2-5A), that binds and activates RNase-L. This pathway is down-regulated by virus-and host-encoded enzymes that degrade 2-5A. Phosphodiesterase 12 (PDE12) was the first cellular 2-5A-degrading enzyme to be purified and described at a molecular level. Inhibition of PDE12 may up-regulate the OAS/RNase-L pathway in response to viral infection resulting in increased resistance to a variety of viral pathogens. We generated a PDE12-null cell line, HeLaΔPDE12, using transcription activator-like effector nuclease-mediated gene inactivation. This cell line has increased 2-5A levels in response to IFN and poly(I-C), a double-stranded RNA mimic compared with the parental cell line. Moreover, HeLaΔPDE12 cells were resistant to viral pathogens, including encephalomyocarditis virus, human rhinovirus, and respiratory syncytial virus. Based on these results, we used DNA-encoded chemical library screening to identify starting points for inhibitor lead optimization. Compounds derived from this effort raise 2-5A levels and exhibit antiviral activity comparable with the effects observed with PDE12 gene inactivation. The crystal structure of PDE12 complexed with an inhibitor was solved providing insights into the structure-activity relationships of inhibitor potency and selectivity.
Florent Samain; Torun Ekblad; Gediminas Mikutis; Nan Zhong; Mauro Zimmermann; Angela Nauer; Davor Bajic; Willy Decurtins; Jörg Scheuermann; Peter J. Brown; Jonathan Hall; Susanne Gräslund; Herwig Schüler; Dario Neri; Raphael M. Franzini J. Med. Chem., 2015, 58, 12, 5143-5149 https://doi.org/10.1021/acs.jmedchem.5b00432AbstractWe describe the synthesis and screening of a DNA-encoded chemical library containing 76230 compounds. In this library, sets of amines and carboxylic acids are directly linked producing encoded compounds with compact structures and drug-like properties. Affinity screening of this library yielded inhibitors of the potential pharmaceutical target tankyrase 1, a poly(ADP-ribose) polymerase. These compounds have drug-like characteristics, and the most potent hit compound (X066/Y469) inhibited tankyrase 1 with an IC50 value of 250 nM.
Robert Chen; Harneet S. Rishi; Vladimir Potapov; Masaki R. Yamada; Vincent J. Yeh; Thomas Chow; Celia L. Cheung; Austin T. Jones; Terry D. Johnson; Amy E. Keating; William C. DeLoache; John E. Dueber ACS Synth. Biol., 2015, 4, 11, 1205-1216 https://doi.org/10.1021/acssynbio.5b00060AbstractDramatic progress has been made in the design and build phases of the design–build–test cycle for engineering cells. However, the test phase usually limits throughput, as many outputs of interest are not amenable to rapid analytical measurements. For example, phenotypes such as motility, morphology, and subcellular localization can be readily measured by microscopy, but analysis of these phenotypes is notoriously slow. To increase throughput, we developed microscopy-readable barcodes (MiCodes) composed of fluorescent proteins targeted to discernible organelles. In this system, a unique barcode can be genetically linked to each library member, making possible the parallel analysis of phenotypes of interest via microscopy. As a first demonstration, we MiCoded a set of synthetic coiled-coil leucine zipper proteins to allow an 8 × 8 matrix to be tested for specific interactions in micrographs consisting of mixed populations of cells. A novel microscopy-readable two-hybrid fluorescence localization assay for probing candidate interactions in the cytosol was also developed using a bait protein targeted to the peroxisome and a prey protein tagged with a fluorescent protein. This work introduces a generalizable, scalable platform for making microscopy amenable to higher-throughput library screening experiments, thereby coupling the power of imaging with the utility of combinatorial search paradigms.
Jörg Scheuermann; Dario Neri Curr. Opin. Chem. Biol., 2015, 26, 99-103 https://doi.org/10.1016/j.cbpa.2015.02.021AbstractIn contrast to single-pharmacophore DNA-encoded libraries, where only one chemical moiety is linked to DNA, dual-pharmacophore DNA-encoded chemical libraries feature the display of two independent small-molecules in close proximity. This, in principle, allows to explore adjacent epitopes on a pharmaceutical target of choice and hence the discovery of simultaneously binding pairs of fragments, by virtue of the chelate effect.
Christian Heinis; Greg Winter Curr. Opin. Chem. Biol., 2015, 26, 89-98 https://doi.org/10.1016/j.cbpa.2015.02.008AbstractThe use of powerful technologies for generating and screening DNA-encoded protein libraries has helped drive the development of proteins as pharmaceutical ligands. However the development of peptides as pharmaceutical ligands has been more limited. Although encoded peptide libraries are typically several orders of magnitude larger than classical chemical libraries, can be more readily screened, and can give rise to higher affinity ligands, their use as pharmaceutical ligands is limited by their intrinsic properties. Two of the intrinsic limitations include the rotational flexibility of the peptide backbone and the limited number (20) of natural amino acids. However these limitations can be overcome by use of chemical modification. For example, the libraries can be modified to introduce topological constraints such as cyclization linkers, or to introduce new chemical entities such as small molecule ligands, fluorophores and photo-switchable compounds. This article reviews the chemistry involved, the properties of the peptide ligands, and the new opportunities offered by chemical modification of DNA-encoded peptide libraries.
Anthony D. Keefe; Matthew A. Clark; Christopher D. Hupp; Alexander Litovchick; Ying Zhang Curr. Opin. Chem. Biol., 2015, 26, 80-88 https://doi.org/10.1016/j.cbpa.2015.02.015AbstractThe generation of DNA-encoded chemical libraries requires the unimolecular association of multiple encoding oligonucleotides with encoded chemical entities during combinatorial synthesis processes. This has traditionally been achieved using enzymatic ligation. We discuss a range of chemical ligation methods that provide alternatives to enzymatic ligation. These chemical ligation methods include the generation of modified internucleotide linkages that support polymerase translocation and other modified linkages that while not supporting the translocation of polymerases can also be used to generate individual cDNA molecules containing encoded chemical information specifying individual library members. We also describe which of these approaches have been successfully utilized for the preparation of DNA-encoded chemical libraries and those that were subsequently used for the discovery of inhibitors.
Sofia Barluenga; Nicolas Winssinger Acc Chem. Res., 2015, 48, 5, 1319-1331 https://doi.org/10.1021/acs.accounts.5b00109AbstractThe programmability of oligonucleotide hybridization offers an attractive platform for the design of assemblies with emergent properties or functions. Developments in DNA nanotechnologies have transformed our thinking about the applications of nucleic acids. Progress from designed assemblies to functional outputs will continue to benefit from functionalities added to the nucleic acids that can participate in reactions or interactions beyond hybridization. In that respect, peptide nucleic acids (PNAs) are interesting because they combine the hybridization properties of DNA with the modularity of peptides. In fact, PNAs form more stable duplexes with DNA or RNA than the corresponding natural homoduplexes. The high stability achieved with shorter oligomers (an 8-mer is sufficient for a stable duplex at room temperature) typically results in very high sequence fidelity in the hybridization with negligible impact of the ionic strength of the buffer due to the lack of electrostatic repulsion between the duplex strands. The simple peptidic backbone of PNA has been shown to be tolerant of modifications with substitutions that further enhance the duplex stability while providing opportunities for functionalization. Moreover, the metabolic stability of PNAs facilitates their integration into systems that interface with biology. Over the past decade, there has been a growing interest in using PNAs as biosupramolecular tags to program assemblies and reactions. A series of robust templated reactions have been developed with functionalized PNA. These reactions can be used to translate DNA templates into functional polymers of unprecedented complexity, fluorescent outputs, or bioactive small molecules. Furthermore, cellular nucleic acids (mRNA or miRNA) have been harnessed to promote assemblies and reactions in live cells. The tolerance of PNA synthesis also lends itself to the encoding of small molecules that can be further assembled on the basis of their nucleic acid sequences. It is now well-established that hybridization-based assemblies displaying two or more ligands can interact synergistically with a target biomolecule. These assemblies have now been shown to be functional in vivo. Similarly, PNA-tagged macromolecules have been used to prepare bioactive assemblies and three-dimensional nanostructures. Several technologies based on DNA-templated synthesis of sequence-defined polymers or DNA-templated display of ligands have been shown to be compatible with reiterative cycles of selection/amplification starting with large libraries of DNA templates, bringing the power of in vitro evolution to synthetic molecules and offering the possibility of exploring uncharted molecular diversity space with unprecedented scope and speed.
Alexander L. Satz ACS Chem. Biol., 2015, 10, 10, 2237-2245 https://doi.org/10.1021/acschembio.5b00378AbstractDNA encoded library (DEL) technology allows for rapid generation of extremely large numbers of small molecules and is often used to find novel chemical starting points for pharmaceutically relevant proteins. DEL selection output consists of a list of small-molecule structures and enrichment levels. It is widely presumed that molecules with greater enrichment will have larger equilibrium association constants, and follow-up efforts are triaged accordingly. Herein we describe a simple mathematical model used to simulate DEL selections. Simulations predict that enrichment levels will correlate poorly with equilibrium association constants when selections use high concentrations of protein or lower quality DELs (high variance in final product synthetic yields). A potentially superior technique is demonstrated to qualitatively assess equilibrium association constants directly from sequencing data. This technique requires conducting selections over a range of protein concentrations, so that the influence of synthetic yield can be accounted for.
Florent Samain; Giulio Casi Curr. Opin. Chem. Biol., 2015, 26, 72-79 https://doi.org/10.1016/j.cbpa.2015.02.009AbstractConventional chemotherapeutic drugs do not selectively localize to tumors, causing undesired toxicities to healthy organs, and precluding the escalation to therapeutically active regimens. The selective delivery at sites of disease of potent effector molecules represents a promising strategy for the treatment of cancer and other diseases. High affinity antibodies towards disease-associated antigens are currently the vehicles of choice for the targeted delivery of payloads. Low molecular weight ligands have the potential to overcome some of the intrinsic limitations associated with antibodies, and have recently been proposed for the development of a novel class of targeted therapeutics. However, the identification of binding molecules, which display high affinity properties and exquisite specificity against protein of therapeutic interest, remains a great challenge. DNA-encoded chemical library technology relies on small molecule libraries of unprecedented size to identify high affinity ligands towards specific target proteins, and could help in the development of next generation targeted cytotoxics.
Peter Blakskjaer; Tara Heitner; Nils Jakob Vest Hansen Curr. Opin. Chem. Biol., 2015, 26, 62-71 https://doi.org/10.1016/j.cbpa.2015.02.003AbstractDNA-encoded small-molecule library (DEL) technology allows vast drug-like small molecule libraries to be efficiently synthesized in a combinatorial fashion and screened in a single tube method for binding, with an assay readout empowered by advances in next generation sequencing technology. This approach has increasingly been applied as a viable technology for the identification of small-molecule modulators to protein targets and as precursors to drugs in the past decade. Several strategies for producing and for screening DELs have been devised by both academic and industrial institutions. This review highlights some of the most significant and recent strategies along with important results. A special focus on the production of high fidelity DEL technologies with the ability to eliminate screening noise and false positives is included: using a DNA junction called the Yoctoreactor, building blocks (BBs) are spatially confined at the center of the junction facilitating both the chemical reaction between BBs and encoding of the synthetic route. A screening method, known as binder trap enrichment, permits DELs to be screened robustly in a homogeneous manner delivering clean data sets and potent hits for even the most challenging targets.
Yong Zhang; Benjamin A. Seigal; Nicholas K. Terrett; Randy L. Talbott; Joseph Fargnoli; Joseph G. Naglich; Charu Chaudhry; Shana L. Posy; Ragini Vuppugalla; Georgia Cornelius; Ming Lei; Chunlei Wang; Yingru Zhang; Robert J. Schmidt; Donna D. Wei; Michael M. Miller; Martin P. Allen; Ling Li; Percy H. Carter; Gregory D. Vite; Robert M. Borzilleri ACS Med. Chem. Lett., 2015, 6, 7, 770-775 https://doi.org/10.1021/acsmedchemlett.5b00091AbstractA series of dimeric macrocyclic compounds were prepared and evaluated as antagonists for inhibitor of apoptosis proteins. The most potent analogue 11, which binds to XIAP and c-IAP proteins with high affinity and induces caspase-3 activation and ultimately cell apoptosis, inhibits growth of human melanoma and colorectal cell lines at low nanomolar concentrations. Furthermore, compound 11 demonstrated significant antitumor activity in the A875 human melanoma xenograft model at doses as low as 2 mg/kg on a q3d schedule.
Francesco V. Reddavide; Weilin Lin; Sarah Lehnert; Yixin Zhang Angew. Chem. Int. Ed. Engl., 2015, 54(27), 7924-8 https://doi.org/10.1002/ange.201501775AbstractDynamic combinatorial chemistry (DCC) explores the thermodynamic equilibrium of reversible reactions. Its application in the discovery of protein binders is largely limited by difficulties in the analysis of complex reaction mixtures. DNA-encoded chemical library (DECL) technology allows the selection of binders from a mixture of up to billions of different compounds; however, experimental results often show low a signal-to-noise ratio and poor correlation between enrichment factor and binding affinity. Herein we describe the design and application of DNA-encoded dynamic combinatorial chemical libraries (EDCCLs). Our experiments have shown that the EDCCL approach can be used not only to convert monovalent binders into high-affinity bivalent binders, but also to cause remarkably enhanced enrichment of potent bivalent binders by driving their insitu synthesis. We also demonstrate the application of EDCCLs in DNA-templated chemical reactions. Selection goes dynamic: Dynamic combinatorial chemistry explores the thermodynamic equilibrium of reversible reactions. DNA-encoded chemical libraries enable the selection of binders from compound mixtures. Now the advantages of DNA-encoded dynamic combinatorial chemical libraries for the selection of protein binders from a myriad of compounds (see picture) and for DNA-templated reactions are demonstrated.
Alexandre Novoa; Nicolas Winssinger Beilstein J. Org. Chem., 2015, 11, 707-719 https://doi.org/10.3762/bjoc.11.81AbstractGlycans (carbohydrate portion of glycoproteins and glycolipids) frequently exert their function through oligomeric interactions involving multiple carbohydrate units. In efforts to recapitulate the diverse spatial arrangements of the carbohydrate units, assemblies based on hybridization of nucleic acid conjugates have been used to display simplified ligands with tailored interligand distances and valences. The programmability of the assemblies lends itself to a combinatorial display of multiple ligands. Recent efforts in the synthesis and applications of such conjugates are discussed
Alix I. Chan; Lynn M. McGregor; David R. Liu Curr. Opin. Chem. Biol., 2015, 26, 55-61 https://doi.org/10.1016/j.cbpa.2015.02.010AbstractDriven by the need for new compounds to serve as biological probes and leads for therapeutic development and the growing accessibility of DNA technologies including high-throughput sequencing, many academic and industrial groups have begun to use DNA-encoded chemical libraries as a source of bioactive small molecules. In this review, we describe the technologies that have enabled the selection of compounds with desired activities from these libraries. These methods exploit the sensitivity of in vitro selection coupled with DNA amplification to overcome some of the limitations and costs associated with conventional screening methods. In addition, we highlight newer techniques with the potential to be applied to the highthroughput evaluation of DNA-encoded chemical libraries.
William H. Connors; Stephen P. Hale; Nicholas K. Terrett Curr. Opin. Chem. Biol., 2015, 26, 42-47 https://doi.org/10.1016/j.cbpa.2015.02.004AbstractConformationally constrained macrocyclic molecules can present functionally diverse chemical groups distributed over a relatively large molecular surface. This class of molecule is well suited to bind to the extended interface surfaces typical of protein–protein interactions that make up key therapeutically relevant pathways. Large numbers of macrocycles can be generated using DNA-encoded technologies to yield chemically diverse libraries where individual macrocycles are identifiable by a unique covalently attached DNA sequence. Recent developments in this field have revealed library-generated macrocycles possessing potent affinity against tough targets such as XIAP, IL17 and IDE. This review highlights recent progression toward developing drug-like macrocycles and as illustration, advances against these targets.
Raphael M. Franzini; Angela Nauer; Jörg Scheuermann; Dario Neri Chem. Commun., 2015, 51, 8014-8016 https://doi.org/10.1039/C5CC01230AAbstractParallel affinity screening of a DNA-encoded chemical library against rat, bovine and human serum albumin allowed the identification of small-molecule ligands with distinctive binding specificities to the individual proteins.
Mickel J. Hansen; Willem A. Velema; Michael M. Lerch; Wiktor Szymanski; Ben L. Feringa Chem. Soc. Rev., 2015,44, 3358-3377 https://doi.org/10.1039/C5CS00118HAbstractPhotocleavable protecting groups (PPGs) are extensively used in chemical and biological sciences. In their application, advantage is taken of using light as an external, non-invasive stimulus, which can be delivered with very high spatiotemporal precision. More recently, orthogonally addressing multiple PPGs, in a single system and with different wavelengths of light, has been explored. This approach allows one to independently control multiple functionalities in an external, non-invasive fashion. In this tutorial review, we discuss the design principles for dynamic systems involving wavelength-selective deprotection, focusing on the choice and optimization of PPGs, synthetic methods for their introduction and strategies for combining multiple PPGs into one system. Finally, we illustrate the design principles with representative examples, aiming at providing the reader with an instructive overview on how the wavelength-selective cleavage of photoprotecting groups can be applied in materials science, organic synthesis and biological systems.
Gang Li; Wenlu Zheng; Ying Liu; Xiaoyu Li Curr. Opin. Chem. Biol., 2015, 26, 25-33 https://doi.org/10.1016/j.cbpa.2015.01.004AbstractAmong various types of DNA-encoded chemical libraries, DNA-templated library takes advantage of the sequence-specificity of DNA hybridization, enabling not only highly effective DNA-templated chemical reactions, but also high fidelity in library encoding. This brief review summarizes recent advances that have been made on the encoding strategies for DNA-templated libraries, and it also highlights their respective advantages and limitations for the preparation of DNA-encoded libraries.
Claudio Zambaldo; Sofia Barluenga; Nicolas Winssinger Curr. Opin. Chem. Biol., 2015, 26, 8-15 https://doi.org/10.1016/j.cbpa.2015.01.005AbstractPeptide nucleic acid (PNA)-encoded chemical libraries along with DNA-encoded libraries have provided a powerful new paradigm for library synthesis and ligand discovery. PNA-encoding stands out for its compatibility with standard solid phase synthesis and the technology has been used to prepare libraries of peptides, heterocycles and glycoconjugates. Different screening formats have now been reported including selection-based and microarray-based methods that have yielded specific ligands against diverse target classes including membrane receptors, lectins and challenging targets such as Hsp70.
Hongfang Yang; Patricia F. Medeiros; Kaushik Raha; Patricia Elkins; Kenneth E. Lind; Ruth Lehr; Nicholas D. Adams; Joelle L. Burgess; Stanley J. Schmidt; Steven D. Knight; Kurt R. Auger; Michael D. Schaber; G. Joseph Franklin; Yun Ding; Jennifer L. DeLorey; Paolo A. Centrella; Sibongile Mataruse; Steven R. Skinner; Matthew A. Clark; John W. Cuozzo; Ghotas Evindar ACS Med. Chem. Lett., 2015, 6, 5, 531-536 https://doi.org/10.1021/acsmedchemlett.5b00025AbstractIn the search of PI3K p110α wild type and H1047R mutant selective small molecule leads, an encoded library technology (ELT) campaign against the desired target proteins was performed which led to the discovery of a selective chemotype for PI3K isoforms from a three-cycle DNA encoded library. An X-ray crystal structure of a representative inhibitor from this chemotype demonstrated a unique binding mode in the p110α protein.
Alexander Litovchick; Christoph E. Dumelin; Sevan Habeshian; Diana Gikunju; Marie-Aude Guié; Paolo A. Centrella; Ying Zhang; Eric A. Sigel; John W. Cuozzo; Anthony D. Keefe; Matthew A. Clark Sci. Rep., 2015, 5, 10916 https://doi.org/10.1038/srep10916AbstractA chemical ligation method for construction of DNA-encoded small-molecule libraries has been developed. Taking advantage of the ability of the Klenow fragment of DNA polymerase to accept templates with triazole linkages in place of phosphodiesters, we have designed a strategy for chemically ligating oligonucleotide tags using cycloaddition chemistry. We have utilized this strategy in the construction and selection of a small molecule library, and successfully identified inhibitors of the enzyme soluble epoxide hydrolase.
Christopher C. Arico-Muendel; Zhengrong Zhu; Hamilton Dickson; Derek Parks; Jesse Keicher; Jianghe Deng; Leah Aquilani; Frank Coppo; Todd Graybill; Kenneth Lind; Andrew Peat; Michael Thomson Antimicrob Agents Chemother., 2015, 9(6), 3450-9 https://doi.org/10.1128/AAC.00070-15AbstractTo identify novel antivirals to the hepatitis C virus (HCV) NS4B protein, we utilized encoded library technology (ELT), which enables purified proteins not amenable to standard biochemical screening methods to be tested against large combinatorial libraries in a short period of time. We tested NS4B against several DNA-encoded combinatorial libraries (DEL) and identified a single DEL feature that was subsequently progressed to off-DNA synthesis. The most active of the initial synthesized compounds had 50% inhibitory concentrations (IC50s) of 50 to 130 nM in a NS4B radioligand binding assay and 300 to 500 nM in an HCV replicon assay. Chemical optimization yielded compounds with potencies as low as 20 nM in an HCV genotype 1b replicon assay, 500 nM against genotype 1a, and 5 μM against genotype 2a. Through testing against other genotypes and genotype 2a-1b chimeric replicons and from resistance passage using the genotype 1b replicon, we confirmed that these compounds were acting on the proposed first transmembrane region of NS4B. A single sequence change (F98L) was identified as responsible for resistance, and it was thought to largely explain the relative lack of potency of this series against genotype 2a. Unlike other published series that appear to interact with this region, we did not observe sensitivity to amino acid substitutions at positions 94 and 105. The discovery of this novel compound series highlights ELT as a valuable approach for identifying direct-acting antivirals to nonenzymatic targets.
Jean-Pierre Daguer; Claudio Zambaldo; Daniel Abegg; Sofia Barluenga; Cynthia Tallant; Susanne Müller; Alexander Adibekian; Nicolas Winssinger Angew. Chem. Int. Ed. Engl., 2015, 54, 6057-6061 https://doi.org/10.1002/anie.201412276AbstractThe regulation of transcriptional programs by epigenetic readers (bromodomains) has been linked to the development of several pathologies. Notably, it has been implicated in the regulation of cellular growth and evasion of apoptosis, in cancer as well as in inflammation. The discovery of small-molecule probes to dissect the role of bromodomains is thus important. We demonstrate that specific cysteine residues conserved across the bromodomains can be harnessed for covalent trapping. We report the discovery of two small molecules that form a covalent bond with cysteine residues conserved across the bromodomain family, analyze the subset of bromodomains that can be addressed through covalent binding, and show proteomic analyses enabled by the enrichment of bromodomains from native lysates.
Benjamin A. Seigal; William H. Connors; Andrew Fraley; Robert M. Borzilleri; Percy H. Carter; Stuart L. Emanuel; Joseph Fargnoli; Kyoung Kim; Ming Lei; Joseph G. Naglich; Matthew E. Pokross; Shana L. Posy; Henry Shen; Neha Surti; Randy Talbott; Yong Zhang; Nicholas K. Terrett J. Med. Chem., 2015, 58, 6, 2855-2861 https://doi.org/10.1021/jm501892gAbstractAffinity selection screening of macrocycle libraries derived from DNA-programmed chemistry identified XIAP BIR2 and BIR3 domain inhibitors that displace bound pro-apoptotic caspases. X-ray cocrystal structures of key compounds with XIAP BIR2 suggested potency-enhancing structural modifications. Optimization of dimeric macrocycles with similar affinity for both domains were potent pro-apoptotic agents in cancer cell lines and efficacious in shrinking tumors in a mouse xenograft model.
Jiayin Bao; Svetlana M. Krylova; Leonid T. Cherney; Robert L. Hale; Svetlana L. Belyanskaya; Cynthia H. Chiu; Christopher C. Arico-Muendel; Sergey N. Krylov Anal. Chem., 2015, 87, 4, 2474-2479 https://doi.org/10.1021/ac504504cAbstractSelection of protein binders from highly diverse combinatorial libraries of DNA-encoded small molecules is a highly promising approach for discovery of small-molecule drug leads. Methods of kinetic capillary electrophoresis provide the high efficiency of partitioning required for such selection but require the knowledge of electrophoretic mobility of the protein–ligand complex. Here we present a theoretical approach for an accurate estimate of the electrophoretic mobility of such complexes. The model is based on a theory of the thin double layer and corresponding expressions used for the mobilities of a rod-like short oligonucleotide and a sphere-like globular protein. The model uses empirical values of mobilities of free protein, free ligand, and electroosmotic flow. The model was tested with a streptavidin–dsDNA complex linked through biotin (small molecule). The deviation of the prediction from the experimental mobility did not exceed 4%, thus confirming that not only is the model adequate but it is also accurate. This model will facilitate reliable use of KCE methods for selection of drug leads from libraries of DNA-encoded small molecules
Paolo Mellini; Sergio Valente; Antonello Mai Expert Opin. Ther. Patents, 2015, 25, 1, 5-15 https://doi.org/10.1517/13543776.2014.982532AbstractIntroduction: Since 2000 sirtuins (SIRT1–7) have gained growing attention for their connections with many biological processes such as cellular metabolism regulation, neuroprotection, apoptosis, inflammation, and cancer progression. In particular, SIRT1 has been the most studied isoform, not only for its role during caloric restriction but also as target in prevention of aging-related diseases. SIRT inhibition can be useful for treating cancer, HIV infection or muscular diseases, SIRT activation can exert positive effects in aging-related disorders such as metabolism, cardiovascular, and neurodegenerative diseases. Areas covered: This review includes the patents about sirtuin modulation released during the 2012 – 2014 period, and covers the potential therapeutic uses of known sirtuin modulators as well as new related small molecules in various disease contexts. Expert opinion: The effective role of sirtuins in cancer is still controversial, because some of them seem to have tumor-promoter as well as tumor-suppressor properties. Thus, few patents describing SIRT inhibitors have been found in 2012 – 2014 period. Despite the still active debate on their role as direct or indirect activators of SIRT1, sirtuin-activating compounds are actually subjected to intense research for the ability to treat neurodegenerative diseases, metabolic disorders, inflammation, vascular system injuries, wound healing and endothelial dysfunctions. A great number of clinical trials are reported with either SIRT inhibitors or activators, thus it is possible that in the foreseeable future one or more of them will enter in the clinical arena.
László Csaba Lengyel; Gellért Sipos; Tamás Sipőcz; Teréz Vágó; György Dormán; János Gerencsér; Gergely Makara; Ferenc Darvas Org. Process Res. Dev., 2015, 19, 3, 399-409 https://doi.org/10.1021/op500354zAbstractIn the present paper we report the synthesis of condensed pyrimidone heterocycles (including novel ones) prepared by the Gould–Jacobs reaction using an in-house-built vacuum-to-high pressure multipurpose “three-mode” pyrolysis reactor. Four of the ring systems have not been described in the literature to date. The pyrolysis reactor has (i) a flash vacuum pyrolysis (FVP) module that applies high vacuum (10–3 mbar), letting the starting material through the reactor chamber heated up to 1000 °C; (ii) a pneumatic spray pyrolysis (PSP) module that can inject nonvolatile reactants to the heated reactor zone; and (iii) a high-pressure pyrolysis (HPP) continuous-flow module that operates from atmospheric to 400 bar pressure and between room temperature and 600 °C. The capabilities of the pyrolysis reactor were demonstrated by comparison experiments on two different condensed pyrimidone bicyclic ring systems, and the established reaction conditions were then successfully applied to the synthesis of another six condensed pyrimidone bicyclic systems.
Raphael M. Franzini; Torun Ekblad; Nan Zhong; Moreno Wichert; Willy Decurtins; Angela Nauer; Mauro Zimmermann; Florent Samain; Jörg Scheuermann; Peter J. Brown; Jonathan Hall; Susanne Gräslund; Herwig Schüler; Dario Neri Angew. Chem. Int. Ed. Engl., 2015, 54(13), 3927-31 https://doi.org/10.1002/anie.201410736AbstractMethods for the rapid and inexpensive discovery of hit compounds are essential for pharmaceutical research and DNA-encoded chemical libraries represent promising tools for this purpose. We here report on the design and synthesis of DAL-100K, a DNA-encoded chemical library containing 103 200 structurally compact compounds. Affinity screening experiments and DNA-sequencing analysis provided ligands with nanomolar affinities to several proteins, including prostate-specific membrane antigen and tankyrase 1. Correlations of sequence counts with binding affinities and potencies of enzyme inhibition were observed and enabled the identification of structural features critical for activity. These results indicate that libraries of this type represent a useful source of small-molecule binders for target proteins of pharmaceutical interest and information on structural features important for binding.
Weilin Lin; Francesco V. Reddavide; Veselina Uzunova; Fatih Nadi Gür; Yixin Zhang Anal. Chem., 2015, 87, 2, 864-868 https://doi.org/10.1021/ac503960zAbstractDNA-encoded chemical library (DECL) technology has emerged as a new avenue in the field of drug discovery. Combined with high-throughput sequencing, DECL selection experiments can provide not only many lead compounds but also insights into the structure-affinity relationship. However, the counts of individual DNA codes reflect, but cannot be used to precisely rank, the binding affinities of the corresponding compounds to protein targets. Herein, we describe a chip-based approach to realize an automated high-throughput assay for the kinetic characterization of the interaction between DNA-conjugated small organic compounds and protein targets. Importantly, this method can be applied to both single-pharmacophore DECLs and self-assembled dual-pharmacophore DECLs.
Frederick W. Goldberg; Jason G. Kettle; Thierry Kogej; Matthew W. D. Perry; Nick P. Tomkinson Drug Discov. Today, 2015, 20, 11-17 https://doi.org/10.1016/j.drudis.2014.09.023AbstractOne pragmatic way to improve compound quality, while enhancing and accelerating drug discovery projects, is the ability to access a high quality, novel, diverse building block collection. Here, we outline general principles that should be applied to ensure that a building block collection has the greatest impact on drug discovery projects, by discussing design principles for novel reagents and what types of reagents are popular with medicinal chemists in general. We initiated a program in 2009 to address this, which has already delivered three candidate drugs, and the success of that program provides evidence that focussing on building block design is a useful strategy for drug discovery.
Andrew D. Foster; James D. Ingram; Eilidh K. Leitch; Katherine R. Lennard; Eliot L. Osher; Ali Tavassoli J. Biomol. Screen., 2015, 20(5), 563-76 https://doi.org/10.1177/1087057114566803AbstractThe identification of initial hits is a crucial stage in the drug discovery process. Although many projects adopt high-throughput screening of small-molecule libraries at this stage, there is significant potential for screening libraries of macromolecules created using chemical biology approaches. Not only can the production of the library be directly interfaced with a cell-based assay, but these libraries also require significantly fewer resources to generate and maintain. In this context, cyclic peptides are increasingly viewed as ideal scaffolds and have proven capability against challenging targets such as protein-protein interactions. Here we discuss a range of methods used for the creation of cyclic peptide libraries and detail examples of their successful implementation
William M. Hewitt; Siegfried S. F. Leung; Cameron R. Pye; Alexandra R. Ponkey; Maria Bednarek; Matthew P. Jacobson; R. Scott Lokey J. Am. Chem. Soc., 2015, 137, 2, 715-721 https://doi.org/10.1021/ja508766bAbstractDrug design efforts are turning to a new generation of therapeutic targets, such as protein−protein interactions (PPIs), that had previously been considered “undruggable” by typical small molecules. There is an emerging view that accessing these targets will require molecules that are larger and more complex than typical small molecule drugs. Here, we present a methodology for the discovery of geometrically diverse, membrane permeable cyclic peptide scaffolds based on the synthesis and permeability screening of a combinatorial library, followed by deconvolution of membrane-permeable scaffolds to identify cyclic peptides with good to excellent passive cell permeabilities.We use a combination of experimental and computational approaches to investigate structure-permeability relationships in one of these scaffolds, and uncover structural and conformational factors that govern passive membrane diffusion in a related set of cyclic peptide diastereomers. Further, we investigate the dependency of permeability on side-chain identity of one of these scaffolds through single-point diversifications to show the adaptability of these scaffolds toward development of permeabilitybiased libraries suitable for bioactivity screens. Overall, our results demonstrate that many novel, cell permeable scaffolds exist beyond those found in extant natural products, and that such scaffolds can be rapidly identified using a combination of synthesis and deconvolution which can, in principle, be applied to any type of macrocyclic template
Moreno Wichert; Nikolaus Krall; Willy Decurtins; Raphael M. Franzini; Francesca Pretto; Petra Schneider; Dario Neri; Jörg Scheuermann Nat. Chem., 2015, 7(3), 241-9 https://doi.org/10.1038/nchem.2158AbstractIn contrast to standard fragment-based drug discovery approaches, dual-display DNA-encoded chemical libraries have the potential to identify fragment pairs that bind simultaneously and benefit from the chelate effect. However, the technology has been limited by the difficulty in unambiguously decoding the ligand pairs from large combinatorial libraries. Here we report a strategy that overcomes this limitation and enables the efficient identification of ligand pairs that bind to a target protein. Small organic molecules were conjugated to the 5′ and 3′ ends of complementary DNA strands that contain a unique identifying code. DNA hybridization followed by an inter-strand code-transfer created a stable dual-display DNA-encoded chemical library of 111,100 members. Using this approach we report the discovery of a low micromolar binder to alpha-1-acid glycoprotein and the affinity maturation of a ligand to carbonic anhydrase IX, an established marker of renal cell carcinoma. The newly discovered subnanomolar carbonic anhydrase IX binder dramatically improved tumour targeting performance in vivo.
Yun Ding; Matthew A. Clark ACS Comb. Sci., 2015, 17(1), 1-4 https://doi.org/10.1021/co5001037AbstractThe Suzuki-Miyaura cross-coupling is one of the most widely employed reactions in medicinal chemistry. To apply this reaction to DNA-encoded library technology (ELT), an alternative approach in the discovery of small molecule hits and leads, we explored the Suzuki-Miyaura cross-coupling on DNA-linked aryl halides. Pd(PPh3)4 was demonstrated to be an effective catalyst for cross-coupling with on-DNA halide substrates under aqueous conditions. It efficiently catalyzes the coupling of phenyl halides (iodide or bromide) and pyridinyl bromides with various boronic acids/esters, including challenging heterocyclic boronic acids/esters.
Ying Liu; Wenlu Zheng; Wan Zhang; Nan Chen; Yang Liu; Li Chen; Xiaozhou Zhou; Xingshuo Chen; Haifeng Zheng; Xiaoyu Li Chem. Sci., 2015, 6, 745 https://doi.org/10.1039/C4SC01953AAbstractCharacterization of transcription factor-DNA interaction is of high importance in elucidating the molecular mechanisms of gene transcriptions. DNA-based affinity probes were developed to capture and identify transcription factors by covalent crosslinking; however, the requirement of a crosslinker on the affinity probe remains a disadvantage, as the crosslinker itself often interferes with the protein–DNA interactions. We report a dual-probe method able to capture DNA-binding transcription factors with unmodified protein-binding sites in scenarios where conventional probes have failed. We have also shown the method's converse application in selecting specific transcription factor-binding DNA sequences from a probe library and its extension to studying proteins recognizing epigenetic marks. This study may provide a new tool for exploring DNA-binding proteins in biology
J.-P. Daguer; C. Zambaldo; M. Ciobanu; P. Morieux; Sofia Barluenga; Nicolas Winssinger Chem. Sci., 2015, 6, 739 https://doi.org/10.1039/C4SC01654HAbstractFragment-based lead discovery has proven to be a powerful method in the drug discovery process. The combinatorial output that is accessible by combining fragments is very attractive; however, identifying fragment pairs that bind synergistically and linking them productively can be challenging. Several technologies have now been established to prepare and screen nucleic acid-encoded libraries (ssDNA, dsDNA, PNA), and it has been shown that pairs of molecules combined by hybridization can bind synergistically to a target. Herein we apply this concept to combinatorially pair two libraries of small molecule fragments, use the fittest fragments supplemented with closely related analogs to build a focused library covalently linking the fragments with different spacers, and apply this strategy to the discovery of a potent ligand for Hsp70.
Huw D. Lewis; John Liddle; Jim E. Coote; Stephen J. Atkinson; Michael D. Barker; Benjamin D. Bax; Kevin L. Bicker; Ryan P. Bingham; Matthew Campbell; Yu Hua Chen; Chun-wa Chung; Peter D. Craggs; Rob P. Davis; Dirk Eberhard; Gerard Joberty; Kenneth E. Lind; Kelly Locke; Claire Maller; Kimberly Martinod; Chris Patten; Oxana Polyakova; Cecil E. Rise; Martin Rüdiger; Robert J. Sheppard; Daniel J. Slade; Pamela Thomas; Jim Thorpe; Gang Yao; Gerard Drewes; Denisa D. Wagner; Paul R. Thompson; Rab K. Prinjha; David M. Wilson Nat. Chem. Biol., 2015, 11(3), 189-91 https://doi.org/10.1038/nchembio.1735AbstractPAD4 has been strongly implicated in the pathogenesis of autoimmune, cardiovascular and oncological diseases through clinical genetics and gene disruption in mice. New selective PAD4 inhibitors binding a calcium-deficient form of the PAD4 enzyme have validated the critical enzymatic role of human and mouse PAD4 in both histone citrullination and neutrophil extracellular trap formation for, to our knowledge, the first time. The therapeutic potential of PAD4 inhibitors can now be explored.
Hongfeng Deng; Jingye Zhou; Flora S. Sundersingh; Jennifer Summerfield; Don Somers; Jeffrey A. Messer; Alexander L. Satz; Nicolas Ancellin; Christopher C. Arico-Muendel; Katie L. (Sargent) Bedard; Arthur Beljean; Svetlana L. Belyanskaya; Ryan Bingham; Sarah E. Smith; Eric Boursier; Paul Carter; Paolo A. Centrella; Matthew A. Clark; Chun-wa Chung; Christopher P. Davie; Jennifer L. Delorey; Yun Ding; G. Joseph Franklin; LaShadric C. Grady; Kenny Herry; Clare Hobbs; Christopher S. Kollmann; Barry A. Morgan; Laura J. (Pothier) Kaushansky; Quan Zhou PDBJ - 5CR5https://pdbj.org/mine/summary/5cr5 Abstract Branched-chain-amino-acid aminotransferase, mitochondrial, PYRIDOXAL-5'-PHOSPHATE, 3-({(3R)-1-[(5-bromothiophen-2-yl)carbonyl]pyrrolidin-3-yl}oxy)-N-methyl-2'-[(methylsulfonyl)amino]biphenyl-4-carboxamide
Haoxing Wu; Brandon T. Cisneros; Christian M. Cole; Neal K. Devaraj J. Am. Chem. Soc., 2014, 136, 17942-17945 https://doi.org/dx.doi.org/10.1021/ja510839rAbstractTetrazine ligations have proven to be a powerful bioorthogonal technique for the detection of many labeled biomolecules, but the ligating nature of these reactions can limit reaction turnover in templated chemistry. We have developed a transfer reaction between 7-azabenzonorbornadiene derivatives and fluorogenic tetrazines that facilitates turnover amplification of the fluorogenic response in nucleic acid-templated reactions. Fluorogenic tetrazine-mediated transfer (TMT) reaction probes can be used to detect DNA and microRNA (miRNA) templates to 0.5 and 5 pM concentrations, respectively. The endogenous oncogenic miRNA target mir-21 could be detected in crude cell lysates and detected by imaging in live cells. Remarkably, the technique is also able to differentiate between miRNA templates bearing a single mismatch with high signal to background. We imagine that TMT reactions could find wide application for amplified fluorescent detection of clinically relevant nucleic acid templates.
Yi-Chao Huang; Cheng Cao; Xiang-Long Tan; Xiaoyu Li; Lei Liu Org. Chem. Front., 2014, 1, 1050-1054 https://doi.org/10.1039/C4QO00217BAbstractPNA–peptide conjugates are useful molecular tools in chemical biology and biotechnology. Although several approaches have been developed to synthesize PNA–peptide conjugates, more efficient methods are still needed. In this report a new pNZ (p-nitrobenzyloxycarbonyl)/bis-Boc strategy was developed as an alternative backbone/nucleobase protecting group method. The mild deprotection conditions of pNZ group and pNZ’s full orthogonality with Fmoc solid-phase synthesis enable a new dimension of synthetic flexibility and practicality to generate versatile PNA–peptide conjugates
Christian Heinis Nat. Chem. Biol., 2014, 10(9), 696-8 https://doi.org/10.1038/nchembio.1605AbstractThe application of macrocycles to previously undruggable targets has aroused a great deal of interest in this structural class. Recent studies advance our understanding of the way macrocycles bind protein targets and provide new strategies and tools to generate peptide-based macrocycles
Alexandre Novoa; Takuya Machida; Sofia Barluenga; Anne Imberty; Nicolas Winssinger ChemBioChem, 2014, 15(14), 2058-65 https://doi.org/10.1002/cbic.201402280AbstractCarbohydrate arrays: We report a highly miniaturized synthetic format to access a nucleic‐acid‐encoded hetero‐glycoconjugate libraries with an unprecedented diversity in the combinations of carbohydrates, linkers, and capping groups. Novel information about plant and bacterial lectin specificity was obtained after conversion of this library to a microarray format. Abstract Identification of selective and synthetically tractable ligands to glycan‐binding proteins is important in glycoscience. Carbohydrate arrays have had a tremendous impact on profiling glycan‐binding proteins and as analytical tools. We report a highly miniaturized synthetic format to access nucleic‐acid‐encoded hetero‐glycoconjugate libraries with an unprecedented diversity in the combinations of glycans, linkers, and capping groups. Novel information about plant and bacterial lectin specificity was obtained by microarray profiling, and we show that a ligand identified on the array can be converted to a high‐affinity soluble ligand by straightforward chemistry.
Pratyusha Mandal; Scott B. Berger; Sirika Pillay; Kenta Moriwaki; Chunzi Huang; Hongyan Guo; John D. Lich; Joshua Finger; Viera Kasparcova; Bart Votta; Michael Ouellette; Bryan W. King; David Wisnoski; Ami S. Lakdawala; Michael P. DeMartino; Linda N. Casillas; Pamela A. Haile; Clark A. Sehon; Robert W. Marquis; Jason Upton; Lisa P. Daley-Bauer; Linda Roback; Nancy Ramia; Cole M. Dovey; Jan E. Carette; Francis Ka-Ming Chan; John Bertin; Peter J. Gough; Edward S. Mocarski; William J. Kaiser Molecular Cell, 2014, 56, 481–495 https://doi.org/10.1016/j.molcel.2014.10.021AbstractReceptor-interacting protein kinase 3 (RIP3 or RIPK3) has emerged as a central player in necroptosis and a potential target to control inflammatory disease. Here, three selective small-molecule compounds are shown to inhibit RIP3 kinase-dependent necroptosis, although their therapeutic value is undermined by a surprising, concentration-dependent induction of apoptosis. These compounds interact with RIP3 to activate caspase 8 (Casp8) via RHIM-driven recruitment of RIP1 (RIPK1) to assemble a Casp8-FADD-cFLIP complex completely independent of pronecrotic kinase activities and MLKL. RIP3 kinase-dead D161N mutant induces spontaneous apoptosis independent of compound, whereas D161G, D143N, and K51A mutants, like wild-type, only trigger apoptosis when compound is present. Accordingly, RIP3-K51A mutant mice (Rip3K51A/K51A) are viable and fertile, in stark contrast to the perinatal lethality of Rip3D161N/D161N mice. RIP3 therefore holds both necroptosis and apoptosis in balance through a Ripoptosome-like platform. This work highlights a common mechanism unveiling RHIM-driven apoptosis by therapeutic or genetic perturbation of RIP3.
Raphael M. Franzini; Florent Samain; Maaly Abd Elrahman; Gediminas Mikutis; Angela Nauer; Mauro Zimmermann; Jörg Scheuermann; Jonathan Hall; Dario Neri Bioconjugate Chem., 2014, 25(8), 1453-61 https://doi.org/10.1021/bc500212nAbstractDNA-encoded chemical libraries are collections of small molecules, attached to DNA fragments serving as identification barcodes, which can be screened against multiple protein targets, thus facilitating the drug discovery process. The preparation of large DNA-encoded chemical libraries crucially depends on the availability of robust synthetic methods, which enable the efficient conjugation to oligonucleotides of structurally diverse building blocks, sharing a common reactive group. Reactions of DNA derivatives with amines and/or carboxylic acids are particularly attractive for the synthesis of encoded libraries, in view of the very large number of building blocks that are commercially available. However, systematic studies on these reactions in the presence of DNA have not been reported so far. We first investigated conditions for the coupling of primary amines to oligonucleotides, using either a nucleophilic attack on chloroacetamide derivatives or a reductive amination on aldehyde-modified DNA. While both methods could be used for the production of secondary amines, the reductive amination approach was generally associated with higher yields and better purity. In a second endeavor, we optimized conditions for the coupling of a diverse set of 501 carboxylic acids to DNA derivatives, carrying primary and secondary amine functions. The coupling efficiency was generally higher for primary amines, compared to secondary amine substituents, but varied considerably depending on the structure of the acids and on the synthetic methods used. Optimal reaction conditions could be found for certain sets of compounds (with conversions >80percent), but multiple reaction schemes are needed when assembling large libraries with highly diverse building blocks. The reactions and experimental conditions presented in this article should facilitate the synthesis of future DNA-encoded chemical libraries, while outlining the synthetic challenges that remain to be overcome.
Cheng Cao; Peng Zhao; Ze Li; Zitian Chen; Yanyi Huang; Yu Bai; Xiaoyu Li Chem. Commun., 2014, 50, 10997-10999 https://doi.org/10.1039/C4CC03380AAbstractWe report a novel method for the synthesis of DNA-encoded libraries without the need for discrete DNA template. Reactant DNAs self-assemble to enable chemical reactions and photo-cleavage transfers the product to the DNA terminus, making it suitable for the subsequent affinity-based selection and hit deconvolution.
Peng Zhao; Zitian Chen; Yizhou Li; Dawei Sun; Yuan Gao; Yanyi Huang; Xiaoyu Li Angew. Chem. Int. Ed. Engl., 2014, 53(38), 10056-9 https://doi.org/10.1002/anie.201404830AbstractThe selection of DNA-encoded libraries against biological targets has become an important discovery method in chemical biology and drug discovery, but the requirement of modified and immobilized targets remains a significant disadvantage. With a terminal protection strategy and ligand-induced photo-crosslinking, we show that iterated selections of DNA-encoded libraries can be realized with unmodified and non-immobilized protein targets.
Gang Li; Yu Liu; Xuerong Yu; Xiaoyu Li Bioconjugate Chem., 2014, 25, 6, 1172-1180 https://doi.org/10.1021/bc500195wAbstractCharacterization of small molecule (SM)–protein interaction is of high importance in biomedical research such as target identification and proteomic profiling. Photo-cross-linking is a powerful and straightforward strategy to covalently capture SM’s binding proteins. The DNA-based photoaffinity labeling method is able to capture SM’s protein targets with high specificity but suffers low cross-linking efficiency, which limits its utility for low abundance and low affinity proteins. After screening a variety of cross-linkers, by utilizing the multivalency effect, the cross-linking efficiency was improved by nearly 7-fold without compromising probe specificity. The generality and performance of multivalent photoaffinity probes have been validated with a variety of SM-protein pairs in the complexity of cell lysates.
Leonard O. Hargiss; G. Greg Zipp; Theodore C. Jessop; Xuejun Sun; Philip Keyes; David B. Rawlins; Zhi Liang; Kum Joo Park; Huizhong Gu J. Chromatogr. B, 2014, 971, 120-5 https://doi.org/10.1016/j.jchromb.2014.09.001AbstractAn ultra high-pressure liquid chromatography/mass spectrometry (UHPLC/MS) separation and analysis method has been devised for open access analysis of synthetic reactions used in the production of DNA-encoded chemical libraries. The aqueous mobile phase is 100mM hexafluoroisopropanol and 8.6mM triethylamine; the organic mobile phase is methanol. The UHPLC separation uses a C18 OST column (50mm×2.1mm×1.7μm) at 60°C, with a flow rate of 0.6mL/min. Gradient concentration is from 10 to 40percent B in 1.0min, increasing to 95percent B at 1.2min. Cycle time was about 5min. This method provides a detection limit of a 20-mer oligonucleotide by mass spectrometry of better than 1pmol on-column. Linear UV response for 20-mer extends from 2 to 200pmol/μL in concentration, same-day relative average deviations are less than 5percent and bias (observed minus expected) is less than 10percent. Deconvoluted mass spectra are generated for components in the predicted mass range using a maximum entropy algorithm. Mass accuracy of deconvoluted spectra is typically 20ppm or better for isotopomers of oligonucleotides up to 7000Da.
Dhruvin R. Shah; Rahul P. Modh; Kishor H. Chikhalia Future Med. Chem., 2014, 6(4), 463–477 https://doi.org/10.4155/fmc.13.212AbstractThis review summarizes recent reports on s-triazine and its respective analogs from the medicinal chemistry angle. Due to its high reactivity and binding characteristic towards various enzymes, s-triazine has attracted attention. This is combined with facile synthesis and interesting pharmacology. The triazine class demonstrates wide biological applications – including antimicrobial, antituberculosis, anticancer, antiviral and antimalarial. In this article the library of s-triazine-based molecular designs has been collated with respective bioactivity. Compounds are further compared with other heterocyclic/nontriazine moieties to correlate the efficiency of privileged s-triazine. We hope this article may assist chemists in their drug design and discovery efforts.
Julia Michaelis; Alexander Roloff; Oliver Seitz Org. Biomol. Chem., 2014, 12, 2821-2833 https://doi.org/10.1039/C4OB00096JAbstractNucleic acid-templated reactions enable the design of conditional reaction systems, in which bond formation occurs only when a particular DNA or RNA molecule is present. Such reaction systems are currently being explored for applications in DNA/RNA diagnosis, drug screening and as a means to design gene expression specific therapy. However, biological nucleic acid templates usually have low abundance. Therefore, either the targeted nucleic acid template has to be multiplied by means of an amplification step or the template itself has to act as a catalyst which amplifies product formation. This critical review highlights the recent advancements in nucleic acid-templated reactions that proceed with turnover in template and thereby provide a means of amplification. Improvements in reaction engineering and the development of new chemistries have pushed the limits from 101 to 102–103 turnovers. This includes reaction systems that lead to the ligation of oligonucleotides or to the interconversion of appended functional groups beyond ligation as well as templated chemistries that enable the activation of catalysts for subsequent triggering of reactions between non-nucleotidic substrates. The present limitations and future opportunities are discussed.
Alexander Litovchick; Matthew A. Clark; Anthony D. Keefe Artif. DNA PNA XNA, 2014, 5(1), e27896 https://doi.org/10.4161/adna.27896AbstractThe affinity-mediated selection of large libraries of DNA-encoded small molecules is increasingly being used to initiate drug discovery programs. We present universal methods for the encoding of such libraries using the chemical ligation of oligonucleotides. These methods may be used to record the chemical history of individual library members during combinatorial synthesis processes. We demonstrate three different chemical ligation methods as examples of information recording processes (writing) for such libraries and two different cDNA-generation methods as examples of information retrieval processes (reading) from such libraries. The example writing methods include uncatalyzed and Cu(I)-catalyzed alkyne-azide cycloadditions and a novel photochemical thymidine-psoralen cycloaddition. The first reading method "relay primer-dependent bypass" utilizes a relay primer that hybridizes across a chemical ligation junction embedded in a fixed-sequence and is extended at its 3'-terminus prior to ligation to adjacent oligonucleotides. The second reading method "repeat-dependent bypass" utilizes chemical ligation junctions that are flanked by repeated sequences. The upstream repeat is copied prior to a rearrangement event during which the 3'-terminus of the cDNA hybridizes to the downstream repeat and polymerization continues. In principle these reading methods may be used with any ligation chemistry and offer universal strategies for the encoding (writing) and interpretation (reading) of DNA-encoded chemical libraries.
Lynn M. McGregor; Tara Jain; David R. Liu J. Am. Chem. Soc., 2014, 136, 3264-3270 https://doi.org/10.1021/ja412934tAbstractWe describe the development and validation of interaction determination using unpurified proteins (IDUP), a method that selectively amplifies DNA sequences identifying ligand+target pairs from a mixture of DNA-linked small molecules and unpurified protein targets in cell lysates. By operating in cell lysates, IDUP preserves native post-translational modifications and interactions with endogenous binding partners, thereby enabling the study of difficult-to-purify targets and increasing the potential biological relevance of detected interactions compared with methods that require purified proteins. In IDUP, target proteins are associated with DNA oligonucleotide tags either non-covalently using a DNA-linked antibody or covalently using a SNAP-tag. Ligand–target binding promotes hybridization of a self-priming hairpin that is extended by a DNA polymerase to create a DNA strand that contains sequences identifying both the target and its ligand. These sequences encoding ligand+target pairs are selectively amplified by PCR and revealed by high-throughput DNA sequencing. IDUP can respond to the effect of affinity-modulating adaptor proteins in cell lysates that would be absent in ligand screening or selection methods using a purified protein target. This capability was exemplified by the 100-fold amplification of DNA sequences encoding FRB+rapamycin or FKBP+rapamycin in samples overexpressing both FRB and FKBP (FRB·rapamycin+FKBP, Kd ≈ 100 fM; FKBP·rapamycin+FRB, Kd = 12 nM). In contrast, these sequences were amplified 10-fold less efficiently in samples overexpressing either FRB or FKBP alone (rapamycin+FKBP, Kd ≈ 0.2 nM; rapamcyin+FRB, Kd = 26 μM). Finally, IDUP was used to process a model library of DNA-linked small molecules and a model library of cell lysates expressing SNAP-target fusions combined in a single sample. In this library×library experiment, IDUP resulted in enrichment of sequences corresponding to five known ligand+target pairs ranging in binding affinity from Kd = 0.2 nM to 3.2 μM out of 67,858 possible combinations, with no false positive signals enriched to the same extent as that of any of the bona fide ligand+target pairs.
Raphael M. Franzini; Dario Neri; Jörg Scheuermann Acc Chem. Res., 2014, 47(4), 1247-55 https://doi.org/10.1021/ar400284tAbstractDNA-encoded chemical libraries (DECLs) represent a promising tool in drug discovery. DECL technology allows the synthesis and screening of chemical libraries of unprecedented size at moderate costs. In analogy to phage-display technology, where large antibody libraries are displayed on the surface of filamentous phage and are genetically encoded in the phage genome, DECLs feature the display of individual small organic chemical moieties on DNA fragments serving as amplifiable identification barcodes. The DNA-tag facilitates the synthesis and allows the simultaneous screening of very large sets of compounds (up to billions of molecules), because the hit compounds can easily be identified and quantified by PCR-amplification of the DNA-barcode followed by high-throughput DNA sequencing. Several approaches have been used to generate DECLs, differing both in the methods used for library encoding and for the combinatorial assembly of chemical moieties. For example, DECLs can be used for fragment-based drug discovery, displaying a single molecule on DNA or two chemical moieties at the extremities of complementary DNA strands. DECLs can vary substantially in the chemical structures and the library size. While ultralarge libraries containing billions of compounds have been reported containing four or more sets of building blocks, also smaller libraries have been shown to be efficient for ligand discovery. In general, it has been found that the overall library size is a poor predictor for library performance and that the number and diversity of the building blocks are rather important indicators. Smaller libraries consisting of two to three sets of building blocks better fulfill the criteria of drug-likeness and often have higher quality. In this Account, we present advances in the DECL field from proof-of-principle studies to practical applications for drug discovery, both in industry and in academia. DECL technology can yield specific binders to a variety of target proteins and is likely to become a standard tool for pharmaceutical hit discovery, lead expansion, and Chemical Biology research. The introduction of new methodologies for library encoding and for compound synthesis in the presence of DNA is an exciting research field and will crucially contribute to the performance and the propagation of the technology.
Christopher S. Kollmann; Xiaopeng Bai; Ching-Hsuan Tsai; Hongfang Yang; Kenneth E. Lind; Steven R. Skinner; Zhengrong Zhu; David I. Israel; John W. Cuozzo; Barry A. Morgan; Koichi Yuki; Can Xie; Timothy A. Springer; Motomu Shimaoka; Ghotas Evindar Bioorg. Med. Chem., 2014, 22, 7, 2353-2365 https://doi.org/10.1016/j.bmc.2014.01.050AbstractAbstract The inhibition of protein–protein interactions remains a challenge for traditional small molecule drug discovery. Here we describe the use of DNA-encoded library technology for the discovery of small molecules that are potent inhibitors of the interaction between lymphocyte function-associated antigen 1 and its ligand intercellular adhesion molecule 1. A DNA-encoded library with a potential complexity of 4.1 billion compounds was exposed to the I-domain of the target protein and the bound ligands were affinity selected, yielding an enriched small-molecule hit family. Compounds representing this family were synthesized without their DNA encoding moiety and found to inhibit the lymphocyte function-associated antigen 1/intercellular adhesion molecule-1 interaction with submicromolar potency in both ELISA and cell adhesion assays. Re-synthesized compounds conjugated to DNA or a fluorophore were demonstrated to bind to cells expressing the target protein. Graphical abstract Encoded library technology (ELT) was utilized to identify a class of compounds that disrupt the interaction between lymphocyte function-associated antigen-1 (LFA-1) and its ligand intercellular adhesion molecule-1 (ICAM-1) at submicromolar potency in both ELISA and cell adhesion assays.
John W. Cuozzo; Holly H. Soutter J. Biomol. Screen., 2014, 1-14 https://doi.org/10.1177/1087057114520975AbstractProduction of novel soluble and membrane-localized protein targets for functional and affinity-based screening has often been limited by the inability of traditional protein-expression systems to generate recombinant proteins that have properties similar to those of their endogenous counterparts. Such targets have often been labeled as challenging. Although biological validation of these challenging targets for specific disease areas may be strong, discovery of small-molecule modulators can be greatly delayed or completely halted due to target-expression issues. In this article, the limitations of traditional protein-expression systems will be discussed along with new systems designed to overcome these challenges. Recent work in this field has focused on two major areas for both soluble and membrane targets: construct-design strategies to improve expression levels and new hosts that can carry out the posttranslational modifications necessary for proper target folding and function. Another area of active research has been on the reconstitution of solubilized membrane targets for both structural analysis and screening. Finally, the potential impact of these new systems on the output of small-molecule screening campaigns will be discussed.
Qinpeng Shen; Lifen Zhou; Yijia Yuan; Yan Huang; Binbin Xiang; Chunyan Chen; Zhou Nie; Shouzhuo Yao Biosens. Bioelectron., 2014, 55, 187-194 https://doi.org/10.1016/j.bios.2013.12.019AbstractA novel homogenous fluorescent sensor for signal-on detection of Cu2+ has been developed based on intra-molecular G-quadruplex formed by DNA-templated click reaction and crystal violet (CV) as label-free signal reporter. The clickable DNA probe consists of two G-rich strands (A and B) bearing azide and alkyne group, respectively, and a template strand (C) locating two proximate reactants by pairing with A and B. The sequences of A and B are derived from asymmetric split of the G-quadruplex sequence (TTAGGG)4. In the presence of Cu2+, the whole G-quadruplex sequence A–B is generated by chemical ligation of A and B via copper ion-catalyzed alkyne-azide cycloaddition, then released from template by toehold strand displacement, and consequently forming a stable intra-molecular G-quadruplex, which binds with CV to generate a strong fluorescent signal. Oppositely, weak fluorescence was obtained without Cu2+ because of unstable intermolecular G-quadruplex formed by A and B and lack of lateral loop connection. Therefore, the Cu2+ can be sensitively and specifically detected by the fluorescence of the CV-stained G-quadruplex with a low detection limit of 65 nM and a linear range of 0.1–3 µM. This method rationally integrated the DNA-templated synthesis and G-quadruplex structure-switch, presenting a simple and promising approach for biosensor development.
Lourdes Encinas; Heather O’Keefe; Margarete Neu; Modesto J. Remuiñán; Amish M. Patel; Ana Guardia; Christopher P. Davie; Natalia Pérez-Macías; Hongfang Yang; Maire A. Convery; Jeff A. Messer; Esther Pérez-Herrán; Paolo A. Centrella; Daniel Álvarez-Gómez; Matthew A. Clark; Sophie Huss; Gary K. O’Donovan; Fátima Ortega-Muro; William McDowell; Pablo Castañeda; Christopher C. Arico-Muendel; Stane Pajk; Joaquín Rullás; Iñigo Angulo-Barturen; Emilio Álvarez-Ruíz; Alfonso Mendoza-Losana; Lluís Ballell Pages; Julia Castro-Pichel; Ghotas Evindar J. Med. Chem., 2014, 57, 4, 1276-1288 https://doi.org/10.1021/jm401326jAbstractTuberculosis (TB) is one of the world's oldest and deadliest diseases, killing a person every 20 s. InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis, is the target of the frontline antitubercular drug isoniazid (INH). Compounds that directly target InhA and do not require activation by mycobacterial catalase peroxidase KatG are promising candidates for treating infections caused by INH resistant strains. The application of the encoded library technology (ELT) to the discovery of direct InhA inhibitors yielded compound 7 endowed with good enzymatic potency but with low antitubercular potency. This work reports the hit identification, the selected strategy for potency optimization, the structure-activity relationships of a hundred analogues synthesized, and the results of the in vivo efficacy studies performed with the lead compound 65.
Juan Pablo Maianti; Amanda McFedries; Zachariah H. Foda; Ralph E. Kleiner; Xiu Quan Du; Malcolm A. Leissring; Wei-Jen Tang; Maureen J. Charron; Markus A. Seeliger; Alan Saghatelian; David R. Liu Nature, 2014, 511, 94-98 https://doi.org/10.1038/nature13297AbstractDespite decades of speculation that inhibiting endogenous insulin degradation might treat type-2 diabetes, and the identification of IDE (insulin-degrading enzyme) as a diabetes susceptibility gene, the relationship between the activity of the zinc metalloprotein IDE and glucose homeostasis remains unclear. Although Ide(-/-) mice have elevated insulin levels, they exhibit impaired, rather than improved, glucose tolerance that may arise from compensatory insulin signalling dysfunction. IDE inhibitors that are active in vivo are therefore needed to elucidate IDE's physiological roles and to determine its potential to serve as a target for the treatment of diabetes. Here we report the discovery of a physiologically active IDE inhibitor identified from a DNA-templated macrocycle library. An X-ray structure of the macrocycle bound to IDE reveals that it engages a binding pocket away from the catalytic site, which explains its remarkable selectivity. Treatment of lean and obese mice with this inhibitor shows that IDE regulates the abundance and signalling of glucagon and amylin, in addition to that of insulin. Under physiological conditions that augment insulin and amylin levels, such as oral glucose administration, acute IDE inhibition leads to substantially improved glucose tolerance and slower gastric emptying. These findings demonstrate the feasibility of modulating IDE activity as a new therapeutic strategy to treat type-2 diabetes and expand our understanding of the roles of IDE in glucose and hormone regulation.
Aidan G. Gilmartin; Thomas H. Faitg; Mark Richter; Arthur Groy; Mark A. Seefeld; Michael G. Darcy; Xin Peng; Kelly Federowicz; Jingsong Yang; Shu-Yun Zhang; Elisabeth Minthorn; Jon-Paul Jaworski; Michael Schaber; Stan Martens; Dean E. McNulty; Robert H. Sinnamon; Hong Zhang; Robert B. Kirkpatrick; Neysa Nevins; Guanglei Cui; Beth Pietrak; Elsie Diaz; Amber Jones; Martin Brandt; Benjamin Schwartz; Dirk A. Heerding; Rakesh Kumar Nat. Chem. Biol., 2014, 10(3), 181-7 https://doi.org/10.1038/nchembio.1427AbstractAlthough therapeutic interventions of signal-transduction cascades with targeted kinase inhibitors are a well-established strategy, drug-discovery efforts to identify targeted phosphatase inhibitors have proven challenging. Herein we report a series of allosteric, small-molecule inhibitors of wild-type p53-induced phosphatase (Wip1), an oncogenic phosphatase common to multiple cancers. Compound binding to Wip1 is dependent on a 'flap' subdomain located near the Wip1 catalytic site that renders Wip1 structurally divergent from other members of the protein phosphatase 2C (PP2C) family and that thereby confers selectivity for Wip1 over other phosphatases. Treatment of tumor cells with the inhibitor GSK2830371 increases phosphorylation of Wip1 substrates and causes growth inhibition in both hematopoietic tumor cell lines and Wip1-amplified breast tumor cells harboring wild-type TP53. Oral administration of Wip1 inhibitors in mice results in expected pharmacodynamic effects and causes inhibition of lymphoma xenograft growth. To our knowledge, GSK2830371 is the first orally active, allosteric inhibitor of Wip1 phosphatase.
Yizhou Li; Peng Zhao; Mingda Zhang; Xianyuan Zhao; Xiaoyu Li J. Am. Chem. Soc., 2013, 135, 17727-17730 https://doi.org/10.1021/ja409936rAbstractWe report a DNA-templated synthesis method that allows construction of the entire DNA-encoded library with a single DNA template. Taking advantage of deoxyinosine's indiscriminate base-pairing property, we designed a "universal template" that is capable of directing chemical reactions with multiple reactant DNAs with different sequences. In combination with other design features including photocleavable linkers and direct encoding by the reactant DNA, we demonstrated the capabilities of the universal template in library synthesis, target selection, and hit decoding. Our method can be generally and straightforwardly applied to prepare a variety of chemically diverse DNA-encoded libraries.
Lukas Lercher; Joanna F. McGouran; Benedikt M. Kessler; Christopher J. Schofield; Benjamin G. Davis Angew. Chem. Int. Ed. Engl., 2013, 52(40), 10553–10558 https://doi.org/10.1002/anie.201304038AbstractA method for Pd‐catalyzed Suzuki–Miyaura cross‐coupling to iododeoxyuridine (IdU) in DNA is described. Key to the reactivity is the choice of the ligand and the buffer. A covalent [Pd]–DNA intermediate was isolated and characterized. Photocrosslinking probes were generated to trap proteins that bind to epigenetic DNA modifications.
Kalyan K. Sadhu; Nicolas Winssinger Chimia, 2013, 67(12-13), 905-9 https://doi.org/10.2533/chimia.2013.905AbstractRecent progress in the synthesis, screening, and self-assembly of nucleic acid-tagged peptidic libraries is reviewed.
Lifen Zhou; Qinpeng Shen; Peng Zhao; Bingbing Xiang; Zhou Nie; Yan Huang; Shouzhuo Yao Methods, 2013, 64, 3, 299-304 https://doi.org/10.1016/j.ymeth.2013.09.001AbstractA novel DNA-templated click chemistry strategy for homogenous fluorescent detection of Cu2+ has been developed based on click ligation-dependent DNA structure switch and the selective quenching ability of graphene oxide (GO) nanosheet. The clickable duplex probe consists of two DNA strands with alkyne and azide group, respectively, and Cu+-catalyzed alkyne–azide cycloaddition (CuAAC) reaction can chemically ligate these two strands. Toehold sequence displacement was consequently exploited to achieve DNA structure transformation bearing fluorescent tag FAM. Cu2+-induced chemical ligation caused the probe transfer to hybrid structure with single stranded DNA (ssDNA) tail, while only duplex structure was obtained without Cu2+. This structural difference can be probed by GO-based fluorescence detection due to the preferential binding of GO to ssDNA. Under the optimum conditions, this sensor can sensitively and specifically detect Cu2+ with a low detection limit of 58 nM and a linear range of 0.1–10 μM. This new strategy is highly sensitive and selective for Cu2+ detection because of the great specificity of click chemistry and super-quenching ability of GO. Moreover, with the aid of high efficient DNA templated synthesis, the detection process requires only about half an hour which is much quicker than previous click-chemistry-based Cu2+ sensors.
Robert E. Martell; David G. Brooks; Yan Wang; Keith Wilcoxen Clin. Pharmacol., 2013, 35, 9, 1271-1281 https://doi.org/10.1016/j.clinthera.2013.08.005AbstractBackground: Once a promising drug target is identified, the steps to actually discover and optimize a drug are diverse and challenging. Objective: The goal of this study was to provide a road map to navigate drug discovery. Methods: Review general steps for drug discovery and provide illustrating references. Results: A number of approaches are available to enhance and accelerate target identification and validation. Consideration of a variety of potential mechanisms of action of potential drugs can guide discovery efforts. The hit to lead stage may involve techniques such as high-throughput screening, fragment-based screening, and structure-based design, with informatics playing an ever-increasing role. Biologically relevant screening models are discussed, including cell lines, 3-dimensional culture, and in vivo screening. The process of enabling human studies for an investigational drug is also discussed. Conclusions: Drug discovery is a complex process that has significantly evolved in recent years.
Alexander Roloff; Oliver Seitz ChemBioChem, 2013, 14, 2322-2328 https://doi.org/10.1002/cbic.201300516AbstractProgrammable interactions allow nucleic acid molecules to template chemical reactions by increasing the effective molarities of appended reactive groups. DNA/RNA‐triggered reactions can proceed, in principle, with turnover in the template. The amplification provided by the formation of many product molecules per template is a valuable asset when the availability of the DNA or RNA target is limited. However, turnover is usually impeded by reaction products that block access to the template. Product inhibition is most severe in ligation reactions, where products after ligation have dramatically increased template affinities. We introduce a potentially generic approach to reduce product inhibition in nucleic acid‐programmed ligation reactions. A DNA‐triggered ligation–cyclization sequence (“cycligation”) of bifunctional peptide nucleic acid (PNA) conjugates affords cyclic ligation products. Melting experiments revealed that product cyclization is accompanied by a pronounced decrease in template affinity compared to linear ligation products. The reaction system relies upon haloacetylated PNA‐thioesters and isocysteinyl‐PNA‐cysteine conjugates, which were ligated on a DNA template according to a native chemical ligation mechanism. Dissociation of the resulting linear product‐template duplex (induced by, for example, thermal cycling) enabled product cyclization through sulfur‐halide substitution. Both ligation and cyclization are fast reactions (ligation: 86 % yield after 20 min, cyclization: quantitative after 5 min). Under thermocycling conditions, the DNA template was able to trigger the formation of new product molecules when fresh reactants were added. Furthermore, cycligation produced 2–3 times more product than a conventional ligation reaction with substoichiometric template loads (0.25–0.01 equiv). We believe that cyclization of products from DNA‐templated reactions could ultimately afford systems that completely overcome product inhibition.
Sofia Barluenga; Nicolas Winssinger ChemBioChem, 2013, 14(15), 1927-8 https://doi.org/10.1002/cbic.201300416AbstractJust so far apart and no further: Small molecules target ID by DNA tagging. Nucleic acid hybridization has been used to pair a photo-crosslinking group with a small molecule of interest. Following photoactivation, the protein(s) interacting with the small molecule are covalently linked to a nucleic acid tag.
Amelie Heuer-Jungemann; Robert Kirkwood; Afaf H. El-Sagheer; Tom Brown; Antonios G. Kanaras Nanoscale, 2013, 5, 7209-7212 https://doi.org/10.1039/C3NR02362AAbstractWe demonstrate a new method to program the ligation of single stranded DNA-modified gold nanoparticles using copper-free click chemistry. Gold nanoparticles functionalized with a discrete number of 3′-azide or 5′-alkyne modified oligonucleotides, can be brought together via a splint strand and covalently ‘clicked’, in a simple one-pot reaction. This new approach to the assembly of gold nanoparticles is inherently advantageous in comparison to the traditional enzymatic ligation. The chemical ligation is specific and takes place at room temperature by simply mixing the particles without the need for special enzymatic conditions. The yield of ‘clicked’ nanoparticles can be as high as 92%. The ease of the copper-free, ‘click-ligation’ method allows for its universal applicability and opens up new avenues in programmed nanoparticle organization.
Nicolas Winssinger Chimia, 2013, 67, 340-348 https://doi.org/10.2533/chimia.2013.340AbstractThe predictability of nucleic acid hybridization offers an attractive platform to program the assembly of tagged ligands or reactants. Hybridization can be used to display multiple ligands in order to gain affinity and/or selectivity through the cooperative interaction of each ligand. Additionally, hybridization of tagged reagents increases their effective concentration and accelerates reactions. In both cases, an oligonucleotide directs an assembly to yield a functional output in the form of enhanced binding, inhibition, or reaction; for example, a reaction can be used to unmask a fluorophore or a bioactive molecule. This review provides an account of our research in this area as well as future directions.
Hongkai Zhang; Kyungmoo Yea; Jia Xie; Diana Ruiz; Ian A. Wilson; Richard A. Lerner Chem. Biol., 2013, 20(5), 734-41 https://doi.org/10.1016/j.chembiol.2013.04.012AbstractWe describe a system for direct selection of antibodies that are receptor agonists. Combinatorial antibody libraries in lentiviruses are used to infect eukaryotic cells that contain a fluorescent reporter system coupled to the receptor for which receptor agonist antibodies are sought. In this embodiment of the method, very large numbers of candidate antibodies expressing lentivirus and eukaryotic reporter cells are packaged together in a format where each is capable of replication, thereby forging a direct link between genotype and phenotype. Following infection, cells that fluoresce are sorted and the integrated genes encoding the agonist antibodies recovered. We validated the system by illustrating its ability to generate rapidly potent antibody agonists that are complete thrombopoietin phenocopies. The system should be generalizable to any pathway where its activation can be linked to production of a selectable phenotype
Jieqiong Qiu; Afaf H. El-Sagheer; Tom Brown Chem. Commun., 2013, 49, 6959-6961 https://doi.org/10.1039/C3CC42451KAbstractOligonucleotides have been ligated efficiently on solid-phase using CuAAC and SPAAC chemistry to produce up to 186-mer triazole linked DNA products. Multiple sequential ligation reactions can be carried out by using a masked azide approach. This work suggests a novel modular approach to the synthesis of large complex oligonucleotide analogues.
Jia Xie; Hongkai Zhang; Kyungmoo Yea; Richard A. Lerner Proc. Natl. Acad. Sci. USA, 2013, 110(20), 8099-104 https://doi.org/10.1073/pnas.1306263110AbstractWe report here the generation of antibody agonists from intracellular combinatorial libraries that transdifferentiate human stem cells. Antibodies that are agonists for the granulocyte colony stimulating factor receptor were selected from intracellular libraries on the basis of their ability to activate signaling pathways in reporter cells. We used a specialized "near neighbor" approach in which the entire antibody library and its target receptor are cointegrated into the plasma membranes of a population of reporter cells. This format favors unusual interactions between receptors and their protein ligands and ensures that the antibody acts in an autocrine manner on the cells that produce it. Unlike the natural granulocyte-colony stimulating factor that activates cells to differentiate along a predetermined pathway, the isolated agonist antibodies transdifferentiated human myeloid lineage CD34+ bone marrow cells into neural progenitors. This transdifferentiation by agonist antibodies is different from more commonly used methods because initiation is agenetic. Antibodies that act at the plasma membrane may have therapeutic potential as agents that transdifferentiate autologous cells
Gang Li; Ying Liu; Yu Liu; Li Chen; Siyu Wu; Yang Liu; Xiaoyu Li Angew. Chem. Int. Ed. Engl., 2013, 52(36), 9544-9 https://doi.org/10.1002/anie.201302161AbstractDNA-templated affinity labeling: Accurate characterization of small-molecule (SM)–protein interactions is one of the most important but also very challenging tasks in chemical biology and drug discovery. A novel method named DNA-programmed photoaffinity labeling is reported. The introduction of DNA encoding and template effect enable multiplexed protein labeling by multiple probes.
Reema K. Thalji; Jeff J. McAtee; Svetlana L. Belyanskaya; Martin Brandt; Gregory D. Brown; Melissa H. Costell; Yun Ding; Jason W. Dodson; Steve H. Eisennagel; Rusty E. Fries; Jeffrey W. Gross; Mark R. Harpel; Dennis A. Holt; David I. Israel; Larry J. Jolivette; Daniel Krosky; Hu Li; Quinn Lu; Tracy Mandichak; Theresa Roethke; Christine G. Schnackenberg; Benjamin Schwartz; Lisa M. Shewchuk; Wensheng Xie; David J. Behm; Stephen A. Douglas; Ami L. Shaw; Joseph P. Marino Jr. Bioorg. Med. Chem. Lett., 2013, 23, 3584-3588 https://doi.org/10.1016/j.bmcl.2013.04.019Abstract1-(1,3,5-Triazin-yl)piperidine-4-carboxamide inhibitors of soluble epoxide hydrolase were identified from high through-put screening using encoded library technology. The triazine heterocycle proved to be a critical functional group, essential for high potency and P450 selectivity. Phenyl group substitution was important for reducing clearance, and establishing good oral exposure. Based on this lead optimization work, 1-[4-methyl-6-(methylamino)-1,3,5-triazin-2-yl]-N-{[[4-bromo-2-(trifluoromethoxy)]-phenyl]methyl}-4-piperidinecarboxamide (27) was identified as a useful tool compound for in vivo investigation. Robust effects on a serum biomarker, 9, 10-epoxyoctadec-12(Z)-enoic acid (the epoxide derived from linoleic acid) were observed, which provided evidence of robust in vivo target engagement and the suitability of 27 as a tool compound for study in various disease models.
Jeremy S. Disch; Ghotas Evindar; Cynthia H. Chiu; Charles A. Blum; Han Dai; Lei Jin; Eli Schuman; Kenneth E. Lind; Svetlana L. Belyanskaya; Jianghe Deng; Frank Coppo; Leah Aquilani; Todd L. Graybill; John W. Cuozzo; Siva Lavu; Cheney Mao; George P. Vlasuk; Robert B. Perni J. Med. Chem., 2013, 56, 3666−3679 https://doi.org/10.1021/jm400204kAbstractThe sirtuins SIRT1, SIRT2, and SIRT3 are NAD(+) dependent deacetylases that are considered potential targets for metabolic, inflammatory, oncologic, and neurodegenerative disorders. Encoded library technology (ELT) was used to affinity screen a 1.2 million heterocycle enriched library of DNA encoded small molecules, which identified pan-inhibitors of SIRT1/2/3 with nanomolar potency (e.g., 11c: IC50 = 3.6, 2.7, and 4.0 nM for SIRT1, SIRT2, and SIRT3, respectively). Subsequent SAR studies to improve physiochemical properties identified the potent drug like analogues 28 and 31. Crystallographic studies of 11c, 28, and 31 bound in the SIRT3 active site revealed that the common carboxamide binds in the nicotinamide C-pocket and the aliphatic portions of the inhibitors extend through the substrate channel, explaining the observable SAR. These pan SIRT1/2/3 inhibitors, representing a novel chemotype, are significantly more potent than currently available inhibitors, which makes them valuable tools for sirtuin research.
Mohosin Sarkar; Bruce D. Pascal; Caitlin Steckler; Claudio Aquino; Glenn C. Micalizio; Thomas Kodadek; Michael J. Chalmers J. Am. Soc. Mass Spectrom., 2013, 24, 1026-1036 https://doi.org/10.1007/s13361-013-0633-xAbstractScreening of bead-based split and pool combinatorial chemistry libraries is a powerful approach to aid the discovery of new chemical compounds able to interact with, andmodulate the activities of, protein targets of interest. Split and pool synthesis provides for large and well diversified chemical libraries, in this case comprised of oligomers generated from a well-defined starting set. At the end of the synthesis,each bead in the library displays many copies of a unique oligomer sequence.Because the sequence of the oligomer is not known at the time of screening,methods for decoding of the sequence of each screening “hit” are essential. Here we describe an electron-transfer dissociation (ETD) based tandem mass spectrometry approach for the decoding of mass-encoded split and pool libraries. We demonstrate that the newly described “chiral oligomers of pentenoic amides (COPAs)” yield non-sequence-specific product ions upon collisional activated dissociation; however, complete sequence information can be obtained with ETD. To aid in the decoding of libraries from MS andMS/MS data, we have incorporated 79Br/81Br isotope “tags” to differentiate N- and C-terminal product ions. In addition, we have created “Hit-Find,” a software program that allows users to generate libraries in silico. The user can then search all possiblemembers of the chemical library for those that fall within a user-defined mass error.
Kalyan K. Sadhu; Manuel Röthlingshöfer; Nicolas Winssinger Isr. J. Chem., 2013, 53, 75-86 https://doi.org/10.1002/ijch.201200100AbstractThe programmability of oligonucleotide recognition offers an attractive platform for directing the assembly of ligands that can interact cooperatively with a target or the assembly of reactive partner that can engage in chemical reactions. In both cases, an oligonucleotide directs the assembly, which yields a functional output. In terms of the controlled display of ligands, several examples have shown a significant increase in binding upon ligand assembly using small molecule fragments, peptides, glycans, and protein fragments. Combinatorial approaches to identifying the optimal pairing have also been reported. In terms of nucleic acidtriggered reactions, several robust chemistries have been reported, leading to the unmasking of different fluorophores or bioactive molecules as well as the synthesis of a bioactive compound in response to DNA or even cellular RNA. Herein we present the historical context of this work and summarize recent developments in this area.
Christian Hejesen; Lars K. Petersen; Nils Jakob V. Hansenb; Kurt V. Gothelf Org. Biomol. Chem., 2013, 11(15), 2493-7 https://doi.org/10.1039/C3OB27504CAbstractDNA-directed synthesis of encoded combinatorial libraries of small organic compounds most often involves transfer of organic building blocks from one DNA strand to another. This requires cleavable linkers to enable cleavage of the link to the original DNA strand from which the building block is transferred. Relatively few cleavable linkers are available for DNA-directed synthesis and most often they leave an amino group at the organic molecule. Here we have extended the application of aryltriazenes as traceless linkers for DNA-directed synthesis. After reaction of one building block with a building block at another DNA strand the triazene linker is cleaved and reduced with hypophosphorous acid in high yield to leave the aryl group with a hydrogen in place of the triazene i.e. without a functional group trace. It was also demonstrated that alternatively the triazene could be converted to an azide, which was used in a cycloaddition reaction. The linker is generally stable at pH > 7 and could be stored for several months in a freezer without significant degradation.
Ryan Hili; Jia Niu; David R. Liu J. Am. Chem. Soc., 2013, 135, 98-101 https://doi.org/10.1021/ja311331mAbstractWe developed a method to translate DNA sequences into densely functionalized nucleic acids by using T4 DNA ligase to mediate the DNA-templated polymerization of 5'-phosphorylated trinucleotides containing a wide variety of appended functional groups. This polymerization proceeds sequence specifically along a DNA template and can generate polymers of at least 50 building blocks (150 nucleotides) in length with remarkable efficiency. The resulting single-stranded highly modified nucleic acid is a suitable template for primer extension using deep vent (exo-) DNA polymerase, thereby enabling the regeneration of template DNA. We integrated these capabilities to perform iterated cycles of in vitro translation, selection, and template regeneration on libraries of modified nucleic acid polymers.
Christopher C. Arico-Muendel; Heather Blanchette; Dennis R. Benjamin; Teresa M. Caiazzo; Paolo A. Centrella; Jennifer DeLorey; Elisabeth G. Doyle; Steven R. Johnson; Matthew T. Labenski; Barry A. Morgan; Gary O’Donovan; Amy A. Sarjeant; Steven Skinner; Charles D. Thompson; Sarah T. Griffin; William Westlin; Kerry F. White ACS Med. Chem. Lett., 2013, 4, 4, 381-386 https://doi.org/10.1021/ml3003633AbstractSemisynthetic analogues of fumagillin, 1, inhibit methionine aminopeptidase-2 (MetAP2) and have entered the clinic for the treatment of cancer. An optimized fumagillin analogue, 3 (PPI-2458), was found to be orally active, despite containing a spiroepoxide function that formed a covalent linkage to the target protein. In aqueous acid, 3 underwent ring-opening addition of water and HCl, leading to four products, 4–7, which were characterized in detail. The chlorohydrin, but not the diol, products inhibited MetAP2 under weakly basic conditions, suggesting reversion to epoxide as a step in the mechanism. In agreement, chlorohydrin 6 was shown to revert rapidly to 3 in rat plasma. In an ex vivo assay, rats treated with purified acid degradants demonstrated inhibition of MetAP2 that correlated with the biochemical activity of the compounds. Taken together, the results indicate that degradation of the parent compound was compensated by the formation of active equivalents leading to a pharmacologically useful level of MetAP2 inhibition.
Katarzyna Gorska; Nicolas Winssinger Angew. Chem. Int. Ed. Engl., 2013, 52, 6820-6843 https://doi.org/10.1002/anie.201208460AbstractThe programmability of oligonucleotide recognition offers an attractive platform to direct the assembly of reactive partners that can engage in chemical reactions. Recently, significant progress has been made in both the breadth of chemical transformations and in the functional output of the reaction. Herein we summarize these recent progresses and illustrate their applications to translate oligonucleotide instructions into functional materials and novel architectures (conductive polymers, nanopatterns, novel oligonucleotide junctions); into fluorescent or bioactive molecule using cellular RNA; to interrogate secondary structures or oligonucelic acids; or a synthetic oligomer.
Ghotas Evindar; Hongfeng Deng; Sylvie G. Bernier; Elisabeth Doyle; Jeanine Lorusso; Barry A. Morgan; William F. Westlin Bioorg. Med. Chem. Lett., 2013, 23, 2, 472-475 https://doi.org/10.1016/j.bmcl.2012.11.053AbstractIn the quest to discover a potent and selective class of direct agonists to the sphingosine-1-phosphate receptor, we explored the carboxylate functional group as a replacement to previously reported lead phosphates. This has led to the discovery of potent and selective direct agonists with moderate to substantial in vivo lymphopenia. The previously reported selectivity enhancing moiety (SEM) and selectivity enhancing orientation (SEO) in the phenylamide and phenylimidazole scaffolds were crucial to obtaining selectivity for S1P receptor subtype 1 over 3.
Julia Michaelis; Atsushi Maruyama; Oliver Seitz Chem. Commun., 2013, 49, 618-620 https://doi.org/10.1039/C2CC36162KAbstractMost DNA-templated reactions suffer from product inhibition. We explored a DNA-triggered fluorophor transfer reaction and demonstrated that comb-type polylysine–polydextran copolymers increase the turnover in template by promoting strand exchange.
Alexander Roloff; Oliver Seitz Chem. Sci., 2013, 4, 432-436 https://doi.org/10.1039/C2SC20961FAbstractDNA directed chemistry is commonly performed by using nanomolar amounts of DNA templates. Herein we introduce a method that allows the use of attomolar template loads. A DNA templated native chemical ligation yields to the covalent fixation of two fluorophores while the template is being produced during polymerase chain reaction.
Claudia Percivalle; Jean-François Bartolo; Sylvain Ladame Org. Biomol. Chem., 2013, 11, 16-26 https://doi.org/10.1039/C2OB26163DAbstractWidespread in nature, oligonucleotide-templated reactions of phosphodiester bond formation have inspired chemists who are now applying this elegant strategy to the catalysis of a broad range of otherwise inefficient reactions. This review highlights the increasing diversity of chemical reactions that can be efficiently catalysed by an oligonucleotide template, using Watson–Crick base-pairing to bring both reagents in close enough proximity to react, thus increasing significantly their effective molarity. The applications of this elegant concept for nucleic acid sensing and controlled organic synthesis will also be discussed.
Patricia L. Podolin; Brian J. Bolognese; Joseph F. Foley; Edward Long III; Brian Peck; Sandra Umbrecht; Xiaojun Zhang; Penny Zhu; Benjamin Schwartz; Wensheng Xie; Chad Quinn; Hongwei Qi; Sharon Sweitzer; Stephanie Chen; Marc Galop; Yun Ding; Svetlana L. Belyanskaya; David I. Israel; Barry A. Morgan; David J. Behm; Joseph P. Marino Jr.; Edit Kurali; Mary S. Barnette; Ruth J. Mayer; Catherine L. Booth-Genthe; James F. Callahan Prostaglandins Other Lipid Mediat., 2013, 104-105, 25-31 https://doi.org/10.1016/j.prostaglandins.2013.02.001AbstractSoluble epoxide hydrolase (sEH, EPHX2) metabolizes eicosanoid epoxides, including epoxyeicosatrienoic acids (EETs) to the corresponding dihydroxyeicosatrienoic acids (DHETs), and leukotoxin (LTX) to leukotoxin diol (LTX diol). EETs, endothelium-derived hyperpolarizing factors, exhibit potentially beneficial properties, including anti-inflammatory effects and vasodilation. A novel, potent, selective inhibitor of recombinant human, rat and mouse sEH, GSK2256294A, exhibited potent cell-based activity, a concentration-dependent inhibition of the conversion of 14,15-EET to 14,15-DHET in human, rat and mouse whole blood in vitro, and a dose-dependent increase in the LTX/LTX diol ratio in rat plasma following oral administration. Mice receiving 10 days of cigarette smoke exposure concomitant with oral administration of GSK2256294A exhibited significant, dose-dependent reductions in pulmonary leukocytes and keratinocyte chemoattractant (KC, CXCL1) levels. Mice receiving oral administration of GSK2256294A following 10 days of cigarette smoke exposure exhibited significant reductions in pulmonary leukocytes compared to vehicle-treated mice. These data indicate that GSK2256294A attenuates cigarette smoke-induced inflammation by both inhibiting its initiation and/or maintenance and promoting its resolution. Collectively, these data indicate that GSK2256294A would be an appropriate agent to evaluate the role of sEH in clinical studies, for example in diseases where cigarette smoke is a risk factor, such as chronic obstructive pulmonary disease (COPD) and cardiovascular disease
Nikolaus Krall; Jörg Scheuermann; Dario Neri Angew. Chem. Int. Ed. Engl., 2013, 52(5), 1384-402 https://doi.org/10.1002/anie.201204631AbstractThe targeted delivery of potent cytotoxic agents has emerged as a promising strategy for the treatment of cancer and other serious conditions. Traditionally, antibodies against markers of disease have been used as drug-delivery vehicles. More recently, lower molecular weight ligands have been proposed for the generation of a novel class of targeted cytotoxics with improved properties. Advances in this field crucially rely on efficient methods for the identification and optimization of organic molecules capable of high-affinity binding and selective recognition of target proteins. The advent of DNA-encoded chemical libraries allows the construction and screening of compound collections of unprecedented size. In this Review, we survey developments in the field of small ligand-based targeted cytotoxics and show how innovative library technologies will help develop the drugs of the future. On target: Antibodies have emerged as promising vehicles for the targeted delivery of potent cytotoxic agents to sites of disease. This Review surveys how the use of smaller organic molecules can yield targeted constructs with improved properties and how DNA-encoded library technologies will facilitate the discovery of the necessary ligands (see scheme).
Tian Xie; Wei Peng; Yexing Liu; Chuangye Yan; Yigong Shi PDB Entry - 4ITJhttps://doi.org/10.2210/pdb4ITJ/pdb Abstract Classification: TRANSFERASE/TRANSFERASE INHIBITOR Organism(s): Homo sapiens Expression System: Spodoptera frugiperda Mutation(s): Yes
Sivaraman Dandapani; Gerard Rosse; Noel Southall; Joseph M. Salvino; Craig J. Thomas Curr. Protoc. Chem. Biol., 2012, 4, 177-191 https://doi.org/10.1002/9780470559277.ch110252AbstractThe selection, acquisition and use of high quality small molecule libraries for screening is an essential aspect of drug discovery and chemical biology programs. Screening libraries continue to evolve as researchers gain a greater appreciation of the suitability of small molecules for specific biological targets, processes and environments. The decisions surrounding the make-up of any given small molecule library is informed by a multitude of variables and opinions vary on best-practices. The fitness of any collection relies upon upfront filtering to avoiding problematic compounds, assess appropriate physicochemical properties, install the ideal level of structural uniqueness and determine the desired extent of molecular complexity. These criteria are under constant evaluation and revision as academic and industrial organizations seek out collections that yield ever improving results from their screening portfolios. Practical questions including cost, compound management, screening sophistication and assay objective also play a significant role in the choice of library composition. This overview attempts to offer advice to all organizations engaged in small molecule screening based upon current best practices and theoretical considerations in library selection and acquisition.
Hongkai Zhang; Ian A. Wilson; Richard A. Lerner Proc. Natl. Acad. Sci. USA, 2012, 109(39), 15728-15733 https://doi.org/10.1073/pnas.1214275109AbstractA method is presented that uses combinatorial antibody libraries to endow cells with new binding energy landscapes for the purpose of regulating their phenotypes. Antibodies that are expressed in cells infected with a lentiviral combinatorial antibody library are selected directly for function rather than only for binding. The potential diversity space can be very large because more than one lentivirus can infect a single cell. Thus, the initial combinatorial diversity of ~1.0 × 10(11) members generated by the random association of antibody heavy and light chains is greatly increased by the reassortment of the antibody Fv domains themselves inside cells. The power of the system is illustrated by its ability to select unusual antibodies. Here, the selected antibodies are potent erythropoietin agonists whose ontogeny depends on recombination at the protein level of pairs of antibodies expressed in the same cell to generate heterodimeric bispecific antibodies. The obligate synergy between the different binding specificities of the antibody's monomeric subunits appears to replicate the asymmetric binding mechanism of authentic erythropoietin
Yizhou Li; Mingda Zhang; Chi Zhang; Xiaoyu Li Chem. Commun., 2012, 48, 9513-9515 https://doi.org/10.1039/C2CC35230CAbstractA system capable of performing both DNA-templated chemical reactions and detection of bond formations is reported. Photocleavable DNA templates direct reactions. Products from bond-forming events re-ligate original templates, amplifiable by PCR, therefore distinguishing bond formation from background. This system provides a novel approach for discovering potential new chemical reactions.
Nicolas Winssinger Artif. DNA PNA XNA, 2012, 3, 105-108 https://doi.org/10.4161/adna.21108AbstractOver the past decade, several technologies have emerged to access nucleic acid-tagged libraries and select the fittest compound within such libraries. This perspective focuses on recent development with PNA-tagged small molecules displayed on DNA templates for screening purposes and to probe the optimal geometry in multivalent interactions.
Shiyu Chen; Julia Morales‐Sanfrutos; Alessandro Angelini; Brian Cutting; Christian Heinis ChemBioChem, 2012, 13(7), 1032-1038 https://doi.org/10.1002/cbic.201200049AbstractCyclizing peptides: Small chemical linkers in bicyclic peptides were found to have a prominent structural role. Exchange of a linker in a potent bicyclic peptide antagonist significantly altered the structure and activity of the macrocycle. Combinatorial libraries of structurally diverse peptide macrocycles offer a rich source for the development of high‐affinity ligands to targets of interest. In this work we have developed linkers for the generation of genetically encoded bicyclic peptides and tested whether the peptides cyclised by them have significant variations in their backbone conformations. Two new cyclisation reagents, each containing three thiol‐reactive groups, efficiently and selectively cyclised linear peptides containing three cysteine moieties. When the mesitylene linker of the bicyclic peptide PK15, a potent inhibitor of plasma kallikrein (K i=2 nM ), was replaced by the new linkers, its inhibitory activity dropped by a factor of more than 1000, suggesting that the linkers impose different conformations on the peptide. Indeed, structural analysis by solution‐state NMR revealed different NOE constraints in the three bicyclic peptides, indicating that these relatively small linkers at the centres of bicyclic peptide structures significantly influence the conformations of the peptides. These results demonstrate the prominent structural role of linkers in peptide macrocycles and suggest that application of different cyclisation linkers in a combinatorial fashion could be an attractive means to generate topologically diverse macrocycle libraries.
Dalila Chouikhi; Mihai Ciobanu; Claudio Zambaldo; Vincent Duplan; Sofia Barluenga; Nicolas Winssinger Chem. Eur. J., 2012, 18(40), 12698-704 https://doi.org/10.1002/chem.201201337AbstractNucleic acid-encoded libraries are emerging as an attractive and highly miniaturized format for the rapid identification of protein ligands. An important criterion in the synthesis of nucleic acid encoded libraries is the scope of reactions that can be used to introduce molecular diversity and devise divergent pathways for diversity-oriented synthesis (DOS). To date, the protecting group strategies that have been used in peptide nucleic acid (PNA) encoded synthesis (PES) have limited the choice of reactions used in the library synthesis to just a few prototypes. Herein, we describe the preparation of PNA monomers with a protecting group combination (Mtt/Boc) that is orthogonal to Fmoc-based synthesis and compatible with a large palette of reactions that have been productively used in DOS (palladium cross-couplings, metathesis, reductive amination, amidation, heterocycle formation, nucleophilic addition, conjugate additions, Pictet-Spengler cyclization). We incorporate γ-modifications in the PNA backbone that are known to enhance hybridization and solubility. We demonstrate the robustness of this strategy with a library synthesis that is characterized by MALDI MS analysis at every step.
Hongfeng Deng; Heather O’Keefe; Christopher P. Davie; Kenneth E. Lind; Raksha A. Acharya; G. Joseph Franklin; Jonathan Larkin; Rosalie Matico; Michael Neeb; Monique M. Thompson; Thomas Lohr; Jeffrey W. Gross; Paolo A. Centrella; Gary K. O’Donovan; Katie L. (Sargent) Bedard; Kurt van Vloten; Sibongile Mataruse; Steven R. Skinner; Svetlana L. Belyanskaya; Tiffany Y. Carpenter; Todd W. Shearer; Matthew A. Clark; John W. Cuozzo; Christopher C. Arico-Muendel; Barry A. Morgan J. Med. Chem., 2012, 55, 16, 7061-7079 https://doi.org/10.1021/jm300449xAbstractThe metalloprotease ADAMTS-5 is considered a potential target for the treatment of osteoarthritis. To identify selective inhibitors of ADAMTS-5, we employed encoded library technology (ELT), which enables affinity selection of small molecule binders from complex mixtures by DNA tagging. Selection of ADAMTS-5 against a four-billion member ELT library led to a novel inhibitor scaffold not containing a classical zinc-binding functionality. One exemplar, (R)-N-((1-(4-(but-3-en-1-ylamino)-6-(((2-(thiophen-2-yl)thiazol-4-yl)methyl)amino)-1,3,5-triazin-2-yl)pyrrolidin-2-yl)methyl)-4-propylbenzenesulfonamide (8), inhibited ADAMTS-5 with IC50 = 30 nM, showing >50-fold selectivity against ADAMTS-4 and >1000-fold selectivity against ADAMTS-1, ADAMTS-13, MMP-13, and TACE. Extensive SAR studies showed that potency and physicochemical properties of the scaffold could be further improved. Furthermore, in a human osteoarthritis cartilage explant study, compounds 8 and 15f inhibited aggrecanase-mediated 374ARGS neoepitope release from aggrecan and glycosaminoglycan in response to IL-1β/OSM stimulation. This study provides the first small molecule evidence for the critical role of ADAMTS-5 in human cartilage degradation.
Mikko J. Vainio; Thierry Kogej; Florian Raubacher J. Chem. Inf. Model., 2012, 52, 7, 1777-1786 https://doi.org/10.1021/ci300157mAbstractAn early stage drug discovery project needs to identify a number of chemically diverse and attractive compounds. These hit compounds are typically found through high-throughput screening campaigns. The diversity of the chemical libraries used in screening is therefore important. In this study, we describe a virtual high-throughput screening system called Virtual Library. The system automatically “recycles” validated synthetic protocols and available starting materials to generate a large number of virtual compound libraries, and allows for fast searches in the generated libraries using a 2D fingerprint based screening method. Virtual Library links the returned virtual hit compounds back to experimental protocols to quickly assess the synthetic accessibility of the hits. The system can be used as an idea generator for library design to enrich the screening collection and to explore the structure–activity landscape around a specific active compound.
Markus Leimbacher; Yixin Zhang; Luca Mannocci; Michael Stravs; Tim Geppert; Jörg Scheuermann; Gisbert Schneider; Dario Neri Chem. Eur. J., 2012, 18, 7729-7737 https://doi.org/10.1002/chem.201200952AbstractLibraries of chemical compounds individually coupled to encoding DNA tags (DNA-encoded chemical libraries) hold promise to facilitate exceptionally efficient ligand discovery. We constructed a high-quality DNAencoded chemical library comprising 30 000 drug-like compounds; this was screened in 170 different affinity capture experiments. High-throughput sequencing allowed the evaluation of 120 million DNA codes for a systematic analysis of selection strategies and statistically robust identification of binding molecules. Selections performed against the tumor-associated antigen carbonic anhydrase IX (CA IX) and the pro-inflammatory cytokine interleukin-2 (IL-2) yielded potent inhibitors with exquisite target specificity. The binding mode of the revealed pharmacophore against IL-2 was confirmed by molecular docking. Our findings suggest that DNA-encoded chemical libraries allow the facile identification of drug-like ligands principally to any protein of choice, including molecules capable of disrupting high-affinity protein-protein interactions.
Phillip J. Milnes; Mireya L. McKee; Jonathan Bath; Lijiang Song; Eugen Stulz; Andrew J. Turberfield; Rachel K. O'Reilly Chem. Commun., 2012, 48, 5614-5616 https://doi.org/10.1039/C2CC31975FAbstractUsing a strand exchange mechanism we have prepared, by DNA templated chemistry, two 10-mers with defined and tunable monomer sequences. An optimized reaction protocol achieves 85% coupling yield per step, demonstrating that DNA-templated chemistry is a powerful tool for the synthesis of macromolecules with full sequence control.
Aya Shibata; Hiroshi Abe; Yoshihiro Ito Molecules, 2012, 17(3), 2446-2463 https://doi.org/10.3390/molecules17032446AbstractOligonucleotide-templated reactions are useful for applying nucleic acid sensing. Various chemistries for oligonucleotide-templated reaction have been reported so far. Major scientific interests are focused on the development of signal amplification systems and signal generation systems. We introduce the recent advances of oligonucleotide-templated reaction in consideration of the above two points.
Chi Zhang; Yizhou Li; Mingda Zhang; Xiaoyu Li Tetrahedron, 2012, 68, 5152-5156 https://doi.org/10.1016/j.tet.2012.04.032AbstractA model study of DNA-directed peptide ligation has been developed by transferring fluorescent reporting group from small molecule thioester to a DNA strand (template DNA) in the presence of a thiolfunctionalized DNA strand (auxiliary DNA), mimicking the Native Chemical Ligation (NCL) reaction. This DNA-directed transfer shows dependence on the sequence complementarity of the two DNA strands, with in situ generated 4-thiolphenylmethyl functionalized oligonucleotide as the auxiliary DNA strand, under mild basic condition (pH¼8.5), and with tris(2-carboxyethyl) phosphine hydrochloride (TCEP) as a reducing agent. Reactions with different amino acid a-thioesters resulted in varied transfer efficiencies from glycine to a-substituted amino acids. This study has provided the basic foundation to use DNA-programmed chemistry toward the chemical synthesis or unnatural modification of protein molecules.
Melissa M. Kemp; Michel Weïwer; Angela N. Koehler Bioorg. Med. Chem., 2012, 20(6), 1979-89 https://doi.org/10.1016/j.bmc.2011.11.071Abstract2011 marks the 10-year anniversary of milestone manuscripts describing drafts of the human genome sequence. Over the past decade, a number of new proteins have been linked to disease-many of which fall into classes that have been historically considered challenging from the perspective of drug discovery. Several of these newly associated proteins lack structural information or strong annotation with regard to function, making development of conventional in vitro functional assays difficult. A recent resurgence in the popularity of simple small molecule binding assays has led to new approaches that do not require knowledge of protein structure or function in advance. Here we briefly review selected methods for executing binding assays that have been used successfully to discover small-molecule probes or drug candidates
Afaf H. El-Sagheer; Tom Brown Acc Chem. Res., 2012, 45(8), 1258-67 https://doi.org/10.1021/ar200321nAbstractBiochemical strategies that use a combination of synthetic oligonucleotides, thermostable DNA polymerases, and DNA ligases can produce large DNA constructs up to 1 megabase in length. Although these ambitious targets are feasible biochemically, comparable technologies for the chemical synthesis of long DNA strands lag far behind. The best available chemical approach is the solid-phase phosphoramidite method, which can be used to assemble DNA strands up to 150 bases in length. Beyond this point, deficiencies in the chemistry make it impossible to produce pure DNA. A possible alternative approach to the chemical synthesis of large DNA strands is to join together carefully purified synthetic oligonucleotides by chemical methods. Click ligation by the copper-catalyzed azide-alkyne (CuAAC) reaction could facilitate this process. In this Account, we describe the synthesis, characterization, and applications of oligonucleotides prepared by click ligation. The alkyne and azide oligonucleotide strands can be prepared by standard protocols, and the ligation reaction is compatible with a wide range of chemical modifications to DNA and RNA. We have employed click ligation to synthesize DNA constructs up to 300 bases in length and much longer sequences are feasible. When the resulting triazole linkage is placed in a PCR template, various DNA polymerases correctly copy the entire base sequence. We have also successfully demonstrated both in vitro transcription and rolling circle amplification through the modified linkage. This linkage has shown in vivo biocompatibility: an antibiotic resistance gene containing triazole linkages functions in E. coli . Using click ligation, we have synthesized hairpin ribozymes up to 100 nucleotides in length and a hammerhead ribozyme with the triazole linkage located at the substrate cleavage site. At the opposite end of the length scale, click-ligated, cyclic mini-DNA duplexes have been used as models to study base pairing. Cyclic duplexes have potential therapeutic applications. They have extremely high thermodynamic stability, have increased resistance to enzymatic degradation, and have been investigated as decoys for regulatory proteins. For potential nanotechnology applications, we have synthesized double stranded DNA catenanes by click ligation. Other researchers have studied covalently fixed multistranded DNA constructs including triplexes and quadruplexes.
Gabriella Gentile; Giancarlo Merlo; Alfonso Pozzan; Giovanni Bernasconi; Benjamin Bax; Paul Bamborough; Angela Bridges; Paul Carter; Margarete Neu; Gang Yao; Caroline Brough; Geoffrey Cutler; Aaron Coffinc; Svetlana L. Belyanskaya Bioorg. Med. Chem. Lett., 2012, 22, 1989–1994 https://doi.org/10.1016/j.bmcl.2012.01.034Abstract5-Aryl-4-carboxamide-1,3-oxazoles are a novel, potent and selective series of GSK-3 inhibitors. The optimization of the series to yield compounds with cell activity and brain permeability is described.
Manuel Röthlingshöfer; Katarzyna Gorska; Nicolas Winssinger Org. Lett., 2012, 14, 2, 482-485 https://doi.org/10.1021/ol203029tAbstractHybridization-based reactions have attracted significant attention. The nucleic acid templated photocatalyzed azide reduction using catalytic amounts of a [Ru(bpy)(2)phen](2+) conjugate is reported. The reaction could be performed with as little as 2% of the Ru nucleic acid probe and was shown to productively unquench 7-azido-coumarin as well as uncage a small molecule.
Quinn Lu; Amy M. Quinn; Mehul P. Patel; Simon F. Semus; Alan P. Graves; Deepak Bandyopadhyay; Andrew J. Pope; Sara H. Thrall J. Biomol. Screen., 2012, 17, 5, 555-571 https://doi.org/10.1177/1087057112437763AbstractEpigenetic gene regulation is a critical process controlling differentiation and development, the malfunction of which may underpin a variety of diseases. In this article, we review the current landscape of small-molecule epigenetic modulators including drugs on the market, key compounds in clinical trials, and chemical probes being used in epigenetic mechanistic studies. Hit identification strategies for the discovery of small-molecule epigenetic modulators are summarized with respect to writers, erasers, and readers of histone marks. Perspectives are provided on opportunities for new hit discovery approaches, some of which may define the next generation of therapeutic intervention strategies for epigenetic processes.
Jean-Pierre Daguer; Mihai Ciobanu; Sofia Barluenga; Nicolas Winssinger Org. Biomol. Chem., 2012, 10, 1502-5 https://doi.org/10.1039/C2OB06880JAbstractDehydrocholic acid was identified as a selective streptavidin binder from a PNA-tagged library. Isothermal calorimetry titration measurements showed this interaction to be entropically driven. Peptides tagged with dehydrocholic acid can be captured on a streptavidin resin and released under thermal conditions.
Rebecca M. Weisinger; Robert J. Marinelli; S. Jarrett Wrenn; Pehr B. Harbury PLoS ONE, 2012, 7, e32299 https://doi.org/10.1371/journal.pone.0032299AbstractHybrid combinatorial chemistry strategies that use DNA as an information-carrying medium are proving to be powerful tools for molecular discovery. In order to extend these efforts, we present a highly parallel format for DNA-programmed chemical library synthesis. The new format uses a standard microwell plate footprint and is compatible with commercially available automation technology. It can accommodate a wide variety of combinatorial synthetic schemes with up to 384 different building blocks per chemical step. We demonstrate that fluidic routing of DNA populations in the highly parallel format occurs with excellent specificity, and that chemistry on DNA arrayed into 384 well plates proceeds robustly, two requirements for the high-fidelity translation and efficient in vitro evolution of small molecules.
Rebecca M. Weisinger; S. Jarrett Wrenn; Pehr B. Harbury PLoS ONE, 2012, 7(3), e28056 https://doi.org/10.1371/journal.pone.0028056AbstractA large body of in vitro evolution work establishes the utility of biopolymer libraries comprising 10^10 to 10^15 distinct molecules for the discovery of nanomolar-affinity ligands to proteins.[1,2,3,4,5] Small-molecule libraries of comparable complexity will likely provide nanomolar-affinity small-molecule ligands.[6,7] Unlike biopolymers, small molecules can offer the advantages of cell permeability, low immunogenicity, metabolic stability, rapid diffusion and inexpensive mass production. It is thought that such desirable in vivo behavior is correlated with the physical properties of small molecules, specifically a limited number of hydrogen bond donors and acceptors, a defined range of hydrophobicity, and most importantly, molecular weights less than 500 Daltons.[8] Creating a collection of 10^10 to 10^15 small molecules that meet these criteria requires the use of hundreds to thousands of diversity elements per step in a combinatorial synthesis of three to five steps. With this goal in mind, we have reported a set of mesofluidic devices that enable DNA-programmed combinatorial chemistry in a highly parallel 384-well plate format. Here, we demonstrate that these devices can translate DNA genes encoding 384 diversity elements per coding position into corresponding small-molecule gene products. This robust and efficient procedure yields small molecule-DNA conjugates suitable for in vitro evolution experiments.
Qinpeng Shen; Shiyun Tang; Wenhua Li; Zhou Nie; Zhuoliang Liu; Yan Huang; Shouzhuo Yao Chem. Commun., 2012, 48, 281-283 https://doi.org/10.1039/C1CC16049DAbstractA novel fluorescent strategy has been developed for sensitive turn-on detection of Cu2+ based on high efficiency of DNA-templated organic synthesis, great specificity of alkyne–azide click reaction to the catalysis of copper ions and the sequential strand displacement for signal transduction.
George Georghiou; Ralph E. Kleiner; Michael Pulkoski-Gross; David R. Liu; Markus A. Seeliger Nat. Chem. Biol., 2012, 8(4), 366-74 https://doi.org/10.1038/nchembio.792AbstractProtein kinases are attractive therapeutic targets, but their high sequence and structural conservation complicates the development of specific inhibitors. We recently identified, in a DNA-templated macrocycle library, inhibitors with unusually high selectivity among Src-family kinases. Starting from these compounds, we developed and characterized in molecular detail potent macrocyclic inhibitors of Src kinase and its cancer-associated 'gatekeeper' mutant. We solved two cocrystal structures of macrocycles bound to Src kinase. These structures reveal the molecular basis of the combined ATP- and substrate peptide-competitive inhibitory mechanism and the remarkable kinase specificity of the compounds. The most potent compounds inhibit Src activity in cultured mammalian cells. Our work establishes that macrocycles can inhibit protein kinases through a bisubstrate-competitive mechanism with high potency and exceptional specificity, reveals the precise molecular basis for their desirable properties and provides new insights into the development of Src-specific inhibitors with potential therapeutic relevance.
Christian Bergsdorf; Johannes Ottl Expert Opin. Drug Discov., 2011, 5, 1095-1107 https://doi.org/10.1517/17460441.2010.524641AbstractImportance of the field: The generation of new chemical leads as a starting point for drug development is a critical step in pharmaceutical drug discovery. High-throughput screening and the attached processes have rapidly evolved over the past few years to become one of the main sources for new leads by testing large compound libraries for activity against a target of interest in biochemical in vitro tests using the recombinant protein or cell-based assays. Very recently, the traditional functional assay read-out technologies are being complemented by biophysical methods which directly measure the physical interaction (affinity) between a low molecular weight compound and a target protein. These technologies are receiving increasing attention and application for affinity screening and increasingly complement and augment the more classical activity screens. Today, such biophysical techniques are applied in hit identification as well as later stages such as hit validation, optimization and lead optimization phase. Areas covered in this review: This review focuses on the principle and application of selected affinity-based screening technologies, especially those which increasingly have been used in different phases of the lead finding process over the past few years. Furthermore, we highlight how throughput, robustness and information content of the discussed methods guide and determine their impact in lead finding and how to make the best use of them. What the reader will gain: The reader will gain an insight into the very broad spectrum of biophysical affinity screening methods and its high potential to support the generation of new leads. As a consequence, the reader will be able to judge which affinity method is of advantage at a certain lead discovery phase. Take home message: Biophysical methods are very powerful tools to identify new hits and/or validate/optimize a hit to a lead. Those technologies often offer novel ways of screening complementing available classical screening technologies. An integrated, holistic approach using the combination of functional read-out technologies with different biophysical methods enables a project team to efficiently promote and progress the most promising chemotypes.
Luca Mannocci; Markus Leimbacher; Moreno Wichert; Jörg Scheuermann; Dario Neri Chem. Commun., 2011, 47, 12747-12753 https://doi.org/10.1039/C1CC15634AAbstractThe identification of specific binding molecules is a central problem in chemistry, biology and medicine. Therefore, technologies, which facilitate ligand discovery, may substantially contribute to a better understanding of biological processes and to drug discovery. DNA-encoded chemical libraries represent a new inexpensive tool for the fast and efficient identification of ligands to target proteins of choice. Such libraries consist of collections of organic molecules, covalently linked to a unique DNA tag serving as an amplifiable identification bar code. DNA-encoding enables the in vitro selection of ligands by affinity capture at sub-picomolar concentrations on virtually any target protein of interest, in analogy to established selection methodologies like antibody phage display. Multiple strategies have been investigated by several academic and industrial laboratories for the construction of DNA-encoded chemical libraries comprising up to millions of DNA-encoded compounds. The implementation of next generation high-throughput sequencing enabled the rapid identification of binding molecules from DNA-encoded libraries of unprecedented size. This article reviews the development of DNA-encoded library technology and its evolution into a novel drug discovery tool, commenting on challenges, perspectives and opportunities for the different experimental approaches.
Warren R.J.D. Galloway; David R. Spring Chem. Biol., 2011, 18(10), 1209-10 https://doi.org/10.1016/j.chembiol.2011.10.002AbstractThe development of a method for the amplification of PNA tags (Svensen et al., in this issue of Chemistry & Biology) should expand the range of biological targets amenable to screening using PNA-encoded combinatorial libraries and thus facilitate the discovery of new biologically useful agents.
Nina Svensen; Juan José Díaz-Mochón; Mark Bradley Chem. Biol., 2011, 18(10), 1284-9 https://doi.org/10.1016/j.chembiol.2011.07.017AbstractThe ability to screen and identify new ligands for cell surface receptors has been a long-standing goal as it might allow targeting of pharmaceutically relevant receptors, such as integrins or G protein coupled receptors. Here, we present a method to amplify hits from a library of PNA-tagged peptides. To this end, human cells, overexpressing either integrins or the CCR6 receptor, were treated with a 10,000 member PNA-encoded peptide library. Extraction of the PNA tags from the surface of the cells was followed by a PNA-tag to DNA translation and amplification enabling decoding of the tags via microarray hybridization. This approach to ligand discovery facilitates screening for differences in surface-receptor ligands and/or receptor expression between different cell types, and opens up a practical approach to PNA-tag amplification.
Christian Heinis Chimia, 2011, 65(9), 677-9 https://doi.org/10.2533/chimia.2011.677AbstractLigands based on bicyclic peptides can combine favourable properties of antibodies (good binding affinity and target specificity) and small molecule ligands (stability, access to chemical synthesis, diffusion properties) and might be suitable molecular structures for the development of therapeutics. By using a combinatorial methodology based on phage display and a chemical cyclisation reaction, we are generating bicyclic peptide antagonists of protein targets with therapeutic applications in mind.
Mihai Ciobanu; Kuo-Ting Huang; Jean-Pierre Daguer; Sofia Barluenga; Olivier Chaloin; Evelyne Schaeffer; Christopher G. Mueller; Daniel A. Mitchell; Nicolas Winssinger Chem. Commun., 2011, 47, 9321-9323 https://doi.org/10.1039/C1CC13213JAbstractWe report the synthesis of a nucleic acid-encoded carbohydrate library, its combinatorial self-assembly into 37,485 pairs and a screen against DC-SIGN leading to the identification of consensus ligand motifs. A prototypical example from the selected pairs was shown to have enhanced binding. A dendrimer incorporating the selected motifs inhibited gp120's binding to dendritic cells with higher efficiency than mannan.
Thomas Kodadek Chem. Commun., 2011, 47, 9757-9763 https://doi.org/10.1039/C1CC12102BAbstractCombinatorial chemistry provides a powerful tool for the rapid creation of large numbers of synthetic compounds. Ideally, these libraries should be a rich source of bioactive molecules, but there is the general feeling that the initial promise of combinatorial chemistry has not yet been realized. In particular, enthusiasm for conducting unbiased (non-structure-guided) screens of large libraries for protein or RNA ligands has waned. A central challenge in this area is to devise methods for the synthesis of chemically diverse, high-quality libraries of molecules with many of the desirable features of natural products. These include diverse functionality, a significant representation of chiral sp3 centers that provide conformational bias to the molecule, significant skeletal diversity, and good pharmacokinetic properties. However, these libraries must be easy to make from cheap, readily available building blocks, ideally those that would support convenient hit optimization/structure reactivity relationship studies. Meeting these challenges will not be easy. Here I review some recent advances in this area and provide some thoughts on likely important developments in the next few years.
Yu He; David R. Liu J. Am. Chem. Soc., 2011, 133, 26, 9972-9975 https://doi.org/10.1021/ja201361tAbstractWe developed a sequential strand-displacement strategy for multistep DNA-templated synthesis (DTS) and used it to mediate an efficient six-step DTS that proceeded in 35% overall yield (83% average yield per step). The efficiency of this approach and the fact that the final product remains linked to a DNA sequence that fully encodes its reaction history suggests its utility for the translation of DNA sequences into high-complexity synthetic libraries suitable for in vitro selection.
Nina Svensen; Juan José Díaz-Mochón; Mark Bradley Chem. Commun., 2011, 47, 7638-7640 https://doi.org/10.1039/C1CC11668AAbstractCells over-expressing integrins or CCR6 were incubated on a DNA microarray, pre-hybridized with a 10 000 member PNA-encoded peptide library allowing novel cell specific ligands for integrins and CCR6 to be identified
Eelco Ruijter; Rachel Scheffelaar; Romano V. A. Orru Angew. Chem. Int. Ed., 2011, 50, 6234-6246 https://doi.org/10.1002/anie.201006515AbstractMulticomponent reactions have become increasingly popular as tools for the rapid generation of small-molecule libraries. However, to ensure sufficient molecular diversity and complexity, there is a continuous need for novel reactions. Although serendipity has always played an important role in the discovery of novel (multicomponent) reactions, rational design strategies have become much more important over the past decade. In this Review, we present an overview of general strategies that allow the design of novel multicomponent reactions. The challenges and opportunities for the future will be discussed.
Nina Svensen; Juan José Díaz‐Mochón; Kevin Dhaliwal; Songsak Planonth; Michael Dewar; J. Douglas Armstrong; Mark Bradley Angew. Chem. Int. Ed. Engl., 2011, 50, 6133-6136 https://doi.org/10.1002/anie.201101804AbstractTo the point: The identification of peptides to optimize both the delivery and tumor penetration of existing cancer drugs in a cell‐selective manner would be highly desirable. The screening of a peptide nucleic acid (PNA)‐encoded peptide library (see picture) now allows the identification of versatile cell homing peptides for any cell type of interest.
Ralph E. Kleiner; Christoph E. Dumelin; David R. Liu Chem. Soc. Rev., 2011, 40, 5707-5717 https://doi.org/10.1039/C1CS15076FAbstractResearchers seeking to improve the efficiency and cost effectiveness of the bioactive small-molecule discovery process have recently embraced selection-based approaches, which in principle offer much higher throughput and simpler infrastructure requirements compared with traditional small-molecule screening methods. Since selection methods benefit greatly from an information-encoding molecule that can be readily amplified and decoded, several academic and industrial groups have turned to DNA as the basis for library encoding and, in some cases, library synthesis. The resulting DNA-encoded synthetic small-molecule libraries, integrated with the high sensitivity of PCR and the recent development of ultra high-throughput DNA sequencing technology, can be evaluated very rapidly for binding or bond formation with a target of interest while consuming minimal quantities of material and requiring only modest investments of time and equipment. In this tutorial review we describe the development of two classes of approaches for encoding chemical structures and reactivity with DNA: DNA-recorded library synthesis, in which encoding and library synthesis take place separately, and DNA-directed library synthesis, in which DNA both encodes and templates library synthesis. We also describe in vitro selection methods used to evaluate DNA-encoded libraries and summarize successful applications of these approaches to the discovery of bioactive small molecules and novel chemical reactivity
Alessandro Angelini; Christian Heinis Curr. Opin. Chem. Biol., 2011, 15(3), 355-61 https://doi.org/10.1016/j.cbpa.2011.03.009AbstractThe genetic encoding of polypeptides with biological display systems enables the facile generation and screening of very large combinatorial libraries of molecules. By post-translationally modifying the encoded polypeptides, chemically and structurally more diverse molecules beyond linear amino acid polymers can be generated. The first post-translational modification applied to encoded polypeptides, the oxidation of cysteine residues to form disulfide bridges, is a natural one and was used to cyclise short peptides soon after the invention of phage display. Recently a range of non-natural chemical strategies for the post-translational modification of encoded polypeptide repertoires were applied to generate optical biosensors, semisynthetic polypeptides, peptide-drug conjugates, redox-insensitive monocyclic peptides or multicyclic peptides, and these strategies are reviewed in this article
Franziska Diezmann; Oliver Seitz Chem. Soc. Rev., 2011, 40, 5789-5801 https://doi.org/10.1039/C1CS15054EAbstractThe self-assembly of nanosized DNA templates—based on formation of duplex, triplex, quadruplex or even pentaplex structures—provides unique opportunities for the controlled presentation of appended functional units. Recently, researchers have recognized the potential of such DNA scaffolds to address questions in the life sciences. In this critical review the focus is on the exploration of proteins. It is shown how different scaffolds can be used to control localization, structure and bioactivity of proteins and protein ligands. Further examples demonstrate that DNA-based recognition can even be used to trigger the formation of protein targeted molecules. Potential and existing applications in protein detection, drug discovery, structural characterization of protein targets as well as in the design of nucleic acid responsive pharmacophores are discussed (107 references).
Katarzyna Gorska; Alex Manicardi; Sofia Barluenga; Nicolas Winssinger Chem. Commun., 2011, 47, 4364-4366 https://doi.org/10.1039/C1CC10222BAbstractNucleic acid templated reactions have attracted significant attention for nucleic acid sensing. Herein we report a general design which extends the potential of nucleic acid templated reactions to unleash the function of a broad diversity of small molecules such as a transcription factor agonist, a cytotoxic or a fluorophore.
Steffen Renner; Maxim Popov; Ansgar Schuffenhauer; Hans-Joerg Roth; Werner Breitenstein; Andreas Marzinzik; Ian Lewis; Philipp Kraste; Florian Nigsch; Jeremy Jenkins; Edgar Jacoby Future Med. Chem., 2011, 3(6), 751-766 https://doi.org/10.4155/fmc.11.15AbstractThe design of a high-quality screening collection is of utmost importance for the early drug-discovery process and provides, in combination with high-quality assay systems, the foundation of future discoveries. Herein, we review recent trends and observations to successfully expand the access to bioactive chemical space, including the feedback from hit assessment interviews of high-throughput screening campaigns; recent successes with chemogenomics target family approaches, the identification of new relevant target/domain families, diversity-oriented synthesis and new emerging compound classes, and non-classical approaches, such as fragment-based screening and DNA-encoded chemical libraries. The role of in silico library design approaches are emphasized.
Jungkyu K. Lee; Young Hwan Jung; Jeffery B.-H. Tok; Zhenan Bao ACS Nano, 2011, 5, 3, 2067-2074 https://doi.org/10.1021/nn1032455AbstractInvestigation of robust and efficient pathways to build DNA−organic molecule hybrid structures is fundamentally important to realize controlled placement of single molecules for potential applications, such as single-molecule electronic devices. Herein, we report a systematic investigation of synthetic processes for preparing organic molecule−DNA building blocks and their subsequent elongation to generate precise micrometer-sized structures. Specifically, optimal cross-coupling routes were identified to enable chemical linkages between three different organic molecules, namely, polyethylene glycol (PEG), poly(p-phenylene ethynylene) (PPE), and benzenetricarboxylate, with single-stranded (ss) DNA. The resulting DNA−organic molecule hybrid building blocks were purified and characterized by both denaturing gel electrophoresis and electrospray ionization mass spectrometry (ESI-MS). The building blocks were subsequently elongated through both the DNA hybridization and ligation processes to prepare micrometer-sized double-stranded (ds) DNA−organic molecule hybrid structures. The described synthetic procedures should facilitate future syntheses of various hybrid DNA-based organic molecular structures.
Jean Pierre Daguer; Mihai Ciobanu; Susana Alvarez; Sofia Barluenga; Nicolas Winssinger Chem. Sci., 2011, 2, 625-632 https://doi.org/10.1039/C0SC00574FAbstractThe discovery of small molecule probes which selectively modulate biological pathways is a cornerstone in the development of new therapeutics. Progress in our ability to access libraries of biologically relevant small molecules in conjunction with streamlined screening technologies have also enabled a more systematic approach to chemical biology. Nevertheless, the current state of the art still requires a large infrastructure and only a small fraction of the proteome has been addressed thus far. The emergence of technologies based on nucleic acid encoding of small molecules presents a new screening paradigm. We describe a method based on DNA-templated combinatorial display of PNA-encoded drug fragments affording 62500 combinations which can be amplified following a selection. This concept was demonstrated with a screen against a representative target, carbonic anhydrase, by iterative cycles of affinity selection, amplification of DNA template and “translation” back into selected library members. The results show a clear convergence towards combinations which, upon resynthesis as covalent adducts, proved to bind cooperatively to carbonic anhydrase.
Fabian Buller; Martina Steiner; Katharina Frey; Dennis Mircsof; Jörg Scheuermann; Markus Kalisch; Peter Bühlmann; Claudiu T. Supuran; Dario Neri ACS Chem. Biol., 2011, 6(4), 336-44 https://doi.org/10.1021/cb1003477AbstractDNA-encoded chemical libraries, i.e., collections of compounds individually coupled to distinctive DNA fragments serving as amplifiable identification barcodes, represent a new tool for the de novo discovery of small molecule ligands to target proteins of pharmaceutical interest. Here, we describe the design and synthesis of a novel DNA-encoded chemical library containing one million small molecules. The library was synthesized by combinatorial assembly of three sets of chemical building blocks using Diels-Alder cycloadditions and by the stepwise build-up of the DNA barcodes. Model selections were performed to test library performance and to develop a statistical method for the analysis of high-throughput sequencing data. A library selection against carbonic anhydrase IX revealed a new class of submicromolar bis(sulfonamide) inhibitors. One of these inhibitors was synthesized in the absence of the DNA-tag and showed accumulation in hypoxic tumor tissue sections in vitro and tumor targeting in vivo.
Raphael M. Franzini; Eric T. Kool Chem. Eur. J., 2011, 17(7), 2168-2175 https://doi.org/10.1002/chem.201002426AbstractWe report a new strategy for template-mediated fluorogenic chemistry that results in enhanced performance for the fluorescence detection of nucleic acids. In this approach, two successive templated reactions are required to induce a fluorescence signal, rather than only one. These novel fluorescein-labeled oligonucleotide probes, termed 2-STAR probes, contain two quencher groups tethered by separate reductively cleavable linkers. When a 2-STAR quenched probe binds adjacent to either two successive mono triphenyl-phosphine (TPP)-DNAs or a dual TPP-DNA, the two quenchers are released, resulting in a fluorescence signal. Because of the requirement for two consecutive reactions, 2-STAR probes display an unprecedented level of sequence-specificity for template-mediated probe designs. At the same time, background emission generated by off-template reactions or incomplete quenching is among the lowest of any fluorogenic reactive probes for the detection of DNA or RNA.
Yiyun Chen; Adam S. Kamlet; Jonathan B. Steinman; David R. Liu Nat. Chem., 2011, 3(2), 146-53 https://doi.org/10.1038/nchem.932AbstractUsing a system that accelerates the serendipitous discovery of new reactions by evaluating hundreds of DNA-encoded substrate combinations in a single experiment, we explored a broad range of reaction conditions for new bond-forming reactions. We discovered reactivity that led to a biomolecule-compatible, Ru(II)-catalysed azide-reduction reaction induced by visible light. In contrast to current azide-reduction methods, this reaction is highly chemoselective and is compatible with alcohols, phenols, acids, alkenes, alkynes, aldehydes, alkyl halides, alkyl mesylates and disulfides. The remarkable functional group compatibility and mild conditions of the reaction enabled the azide reduction of nucleic acid and oligosaccharide substrates, with no detectable occurrence of side reactions. The reaction was also performed in the presence of a protein enzyme without the loss of enzymatic activity, in contrast to two commonly used azide-reduction methods. The visible-light dependence of this reaction provides a means of photouncaging functional groups, such as amines and carboxylates, on biological macromolecules without using ultraviolet irradiation.
Kuo‐Ting Huang; Katarzyna Gorska; Susana Alvarez; Sofia Barluenga; Nicolas Winssinger ChemBioChem, 2011, 12(1), 56-60 https://doi.org/10.1002/cbic.201000567AbstractComplex glycan arrays: A general method for preparing thioglycosides from native oligosaccharides and coupling them to peptide nucleic acids (PNAs) is reported. An array of 625 glycans was assembled by hybridization of two PNA‐encoded libraries of 25 glycan fragments. The cooperativity of the fragment for carbohydrate binding proteins was validated with lectins of known specificity.
Paul Edwards Drug Discov. Today, 2010, 15, 15–16, 690-691 https://doi.org/10.1016/j.drudis.2010.06.013AbstractMolecules with biological activity emerge in living systems through cycles of translation, selection, and amplification with mutation. Scientists have adopted features of biological evolution to create DNA, RNA, and protein molecules with tailor-made binding or catalytic properties. The benefits to the discovery of functional molecules by applying translation and selection-based methods has provided the impetus to develop new approaches to addressing translating DNA sequences into structures not necessarily compatible with polymerase enzymes or ribosomal machinery. For example, a ‘DNA display’ method in which resin-bound DNA hybridization directs split-andpool combinatorial synthesis, as well as the use of DNA display to generate libraries of linear peptides and peptoids has been developed
Paul Edwards Drug Discov. Today, 2010, 15, 15–16, 690-691 https://doi.org/10.1016/j.drudis.2010.06.013AbstractMedicinal chemists often search for small molecules that can bind to specific targets. A solution chosen when the target protein displays enzymatic activity, or the researchers have labeled ligands that can be used in displacement assays, is the use of large libraries of chemical compounds that can be screened individually in order to identify novel ligands. These highthroughput screening approaches can be expensive, both in terms of target protein requirements and costs in synthesizing the library, such as in chemical synthesis and the use of robotics. In this regard, the use of DNA fragments as amplifiable "bar-codes" for the identification of chemical compounds in a library represents an attractive avenue for the synthesis and screening of large combinatorial libraries
Jennifer Berger; Michael Oberhuber Chem. Biodivers., 2010, 7, 2581-2615 https://doi.org/10.1002/cbdv.201000108AbstractIn this review, we will focus on chemistry that is controlled by a NA templating effect without requiring a macromolecular catalyst. The possibilities that NAs offer to promote and control the formation (and rarely cleavage) of chemical bonds will be presented and discussed. Particular attention will be devoted to the added value of NAs for chemistry, since several excellent reviews on NA templating have been published in recent years [9 – 12]. In the tradition of chemical biology [13], we will discuss how the biological principles that are inherent to NAs can be used to advance chemistry, which, in turn, fuels advances in biology. Most examples will concern DNA templates, but RNA and some NA analogs will also be treated.
Jörg Scheuermann; Dario Neri ChemBioChem, 2010, 11(7), 931-7 https://doi.org/10.1002/cbic.201000066AbstractDNA decoder: The Collection of organic molecules, individually coupled to distinctive oligonucleatides, is generally referred to as "DNA-encoded chemical library". In full analogy to phage disply technology, these libraries can be panned on immobilized target protains and analyzed (before and after selection) by suitable "decoding" methods (for example, DNA-sequencing).
Samu Melkko; Luca Mannocci; Christoph E. Dumelin; Alessandra Villa; Roberto Sommavilla; Yixin Zhang; Markus G. Grütter; Nadine Keller; Lutz Jermutus; Ronald H. Jackson; Jörg Scheuermann; Dario Neri ChemMedChem, 2010, 5, 4, 584-590 https://doi.org/10.1002/cmdc.200900520AbstractBcl-xL is an antiapoptotic member of the Bcl-2 protein family and an attractive target for the development of anticancer agents. Here we describe the isolation of binders to Bcl-xL from a DNA-encoded chemical library using affinity-capture selections and massively parallel high-throughput sequencing of >30 000 sequence tags of library members. The most potent binder identified, compound 19/93 [(R)-3-(amido indomethacin)-4-(naphthalen-1-yl)butanoic acid], bound to Bcl-xL with a dissociation constant (Kd) of 930 nm and was able to compete with a Bak-derived BH3 peptide, an antagonist of Bcl-xL function.
Kui Lu; Qun-Peng Duan; Li Ma; Dong-Xin Zhao Bioconjugate Chem., 2010, 21, 2, 187-202 https://doi.org/10.1021/bc900158sAbstractThe use of synthetic oligonucleotides and their mimics to inhibit gene expression by hybridizing with their target sequences has been hindered by their poor cellular uptake and inability to reach the nucleus. Covalent postsynthesis or solid-phase conjugation of peptides to oligonucleotides offers a possible solution to these problems. As feasible chemistry is a prerequisite for biological studies, development of efficient and reproducible approaches for convenient preparation of peptide−oligonucleotide conjugates has become a subject of considerable importance. The present review gives an account of the main synthetic methods available to prepare covalent conjugation of peptides to oligonucleotides.
Lynn M. McGregor; David J. Gorin; Christoph E. Dumelin; David R. Liu J. Am. Chem. Soc., 2010, 132, 44, 15522-15524 https://doi.org/10.1021/ja107677qAbstractInteraction-dependent PCR (IDPCR) is a solution-phase method to identify binding partners from combined libraries of small-molecule ligands and targets in a single experiment. Binding between DNA-linked targets and DNA-linked ligands induces formation of an extendable duplex. Extension links codes that identify the ligand and target into one selectively amplifiable DNA molecule. In a model selection, IDPCR resulted in the enrichment of DNA encoding all five known protein−ligand pairs out of 67 599 possible sequences.
Yu He; David R. Liu Nat. Nanotech., 2010, 5, 778-782 https://doi.org/10.1038/nnano.2010.190AbstractMultistep synthesis in the laboratory typically requires numerous reaction vessels, each containing a different set of reactants. In contrast, cells are capable of performing highly efficient and selective multistep biosynthesis under mild conditions with all reactants simultaneously present in solution. If the latter approach could be applied in the laboratory, it could improve the ease, speed and efficiency of multistep reaction sequences. Here, we show that a DNA mechanical device—a DNA walker moving along a DNA track—can be used to perform a series of amine acylation reactions in a single solution without any external intervention. The products of these reactions are programmed by the sequence of the DNA track, but they are not related to the structure of DNA. Moreover, they are formed with speeds and overall yields that are significantly greater than those previously achieved by multistep DNA-templated small-molecule synthesis.
Balint Kintses; Liisa D van Vliet; Sean RA Devenish; Florian Hollfelder Curr. Opin. Chem. Biol., 2010, 14, 5, 548-555 https://doi.org/10.1016/j.cbpa.2010.08.013AbstractMiniaturization of the classical test tube to picoliter dimensions is possible in monodisperse water-in-oil droplets that are generated in microfluidic devices. The establishment of standard unit operations for droplet handling and the ability to carry out experiments with DNA, proteins, cells and organisms provides the basis for the design of more complex workflows to address biological challenges. The emerging experimental format makes possible a quantitative readout for large numbers of experiments with a precision comparable to the macroscopic scale. Directed evolution, diagnostics and compound screening are areas in which the first steps are being taken toward the long-term goal of transforming the way we design and carry out experiments.
Natalie R. Gassman; J. Patrick Nelli; Samrat Dutta; Adam Kuhn; Keith Bonin; Zbigniew Pianowski; Nicolas Winssinger; Martin Guthold; Jed C. Macosko J. Mol. Recognit., 2010, 23(5), 414-422 https://doi.org/10.1002/jmr.1007AbstractThe lack of efficient identification and isolation methods for specific molecular binders has fundamentally limited drug discovery. Here, we have developed a method to select peptide nucleic acid (PNA) encoded molecules with specific functional properties from combinatorially generated libraries. This method consists of three essential stages: (1) creation of a Lab-on-BeadTM library, a one-bead, one-sequence library that, in turn, displays a library of candidate molecules, (2) fluorescence microscopy-aided identification of single target-bound beads and the extraction - wet or dry - of these beads and their attached candidate molecules by a micropipette manipulator, and (3) identification of the target-binding candidate molecules via amplification and sequencing. This novel integration of techniques harnesses the sensitivity of DNA detection methods and the multiplexed and miniaturized nature of molecule screening to efficiently select and identify target-binding molecules from large nucleic acid encoded chemical libraries. Beyond its potential to accelerate assays currently used for the discovery of new drug candidates, its simple bead-based design allows for easy screening over a variety of prepared surfaces that can extend this technique's application to the discovery of diagnostic reagents and disease markers.
Mireya L. McKee; Phillip J. Milnes; Jonathan Bath; Eugen Stulz; Andrew J. Turberfield; Rachel K. O'Reilly Angew. Chem. Int. Engl., 2010, 49(43), 7948-7951 https://doi.org/10.1002/anie.201002721AbstractBiomimetic : A strand displacement mechanism was designed to permit DNA‐templated synthesis of functional oligomers of arbitrary length (see scheme). Key features of the mechanism are that successive coupling reactions take place in near‐identical environments and that purification is only necessary in the last synthesis step.
Luca Mannocci; Samu Melkko; Fabian Buller; Ilona Molnàr; Jean-Paul Gapian Bianké; Christoph E. Dumelin; Jörg Scheuermann; Dario Neri Bioconjugate Chem., 2010, 21, 10, 1836-1841 https://doi.org/10.1021/bc100198xAbstractCollections of chemical compounds, individually attached to unique DNA fragments serving as amplifiable identification bar codes, are generally referred to as "DNA-encoded chemical libraries". Such libraries can be used for the de novo isolation of binding molecules against target proteins of interest. Here, we describe the synthesis and use of a DNA-encoded library based on benzamidine analogues, which allowed the isolation of a trypsin inhibitor with an IC50 value of 3.0 nM, thus representing a >10 000-fold potency improvement compared to the parental compound. The novel trypsin inhibitor displayed an excellent selectivity toward other serine proteases. This study indicates that DNA-encoded libraries can be used for the facile "affinity maturation" of suboptimal binding compounds, thus facilitating drug development.
Ralph E. Kleiner; Christoph E. Dumelin; Gerald C. Tiu; Kaori Sakurai; David R. Liu J. Am. Chem. Soc., 2010, 132, 33, 11779-11791 https://doi.org/10.1021/ja104903xAbstractDNA-templated organic synthesis enables the translation of DNA sequences into synthetic small-molecule libraries suitable for in vitro selection. Previously, we described the DNA-templated multistep synthesis of a 13,824-membered small-molecule macrocycle library. Here, we report the discovery of small molecules that modulate the activity of kinase enzymes through the in vitro selection of this DNA-templated small-molecule macrocycle library against 36 biomedically relevant protein targets. DNA encoding selection survivors was amplified by PCR and identified by ultra-high-throughput DNA sequencing. Macrocycles corresponding to DNA sequences enriched upon selection against several protein kinases were synthesized on a multimilligram scale. In vitro assays revealed that these macrocycles inhibit (or activate) the kinases against which they were selected with IC(50) values as low as 680 nM. We characterized in depth a family of macrocycles enriched upon selection against Src kinase, and showed that inhibition was highly dependent on the identity of macrocycle building blocks as well as on backbone conformation. Two macrocycles in this family exhibited unusually strong Src inhibition selectivity even among kinases closely related to Src. One macrocycle was found to activate, rather than inhibit, its target kinase, VEGFR2. Taken together, these results establish the use of DNA-templated synthesis and in vitro selection to discover small molecules that modulate enzyme activities, and also reveal a new scaffold for selective ATP-competitive kinase inhibition.
Fabian Buller; Luca Mannocci; Jörg Scheuermann; Dario Neri Bioconjugate Chem., 2010, 21(9), 1571-80 https://doi.org/10.1021/bc1001483AbstractDNA-encoded chemical libraries represent a novel avenue for the facile discovery of small molecule ligands against target proteins of biological or pharmaceutical importance. Library members consist of small molecules covalently attached to unique DNA fragments that serve as amplifiable identification barcodes. This encoding allows the in vitro selection of ligands at subpicomolar concentrations from large library populations by affinity capture on a target protein of interest, in analogy to established technologies for the selection of binding polypeptides (e.g., antibodies). Different library formats have been explored by various groups, allowing the construction of chemical libraries comprising up to millions of DNA-encoded compounds. Libraries before and after selection have been characterized by PCR amplification of the DNA codes and subsequent relative quantification of library members using high-throughput sequencing. The most enriched compounds have then been further analyzed in biological assays, in the presence or in the absence of linked DNA. This article reviews experimental strategies used for the construction of DNA-encoded chemical libraries, revealing how selection, decoding, and hit validation technologies have been used for drug discovery programs.
Scott K. Silverman Angew. Chem. Int. Engl., 2010, 49, 7180-7201 https://doi.org/10.1002/anie.200906345AbstractDNA (deoxyribonucleic acid) is the genetic material common to all of Earth’s organisms. Our biological understanding of DNA is extensive and well‐exploited. In recent years, chemists have begun to develop DNA for nonbiological applications in catalysis, encoding, and stereochemical control. This Review summarizes key advances in these three exciting research areas, each of which takes advantage of a different subset of DNA’s useful chemical properties.
Nicholas K.Terrett Drug Discov. Today Technol., 2010, 7(2), e95-e146 https://doi.org/10.1016/j.ddtec.2010.06.002AbstractMacrocycles are found widely in nature where they fulfill numerous specific functions. However they have been generally underexploited as drug molecules, as they are larger than more conventional ‘Rule of 5’ compliant molecules and their synthesis and screening has been considered a challenge. Consequently most pharmaceutical companies have very few macrocycles in their screening files, and yet these compounds can have potent and selective pharmacological activity, and exhibit drug-like properties such as cell membrane permeability and oral bioavailability. To permit the further investigation of macrocyclic drugs, several groups have developed diverse methods for the rapid synthesis and screening of macrocyclic libraries
Matthew A. Clark Curr. Opin. Chem. Biol., 2010, 14(3), 396-403 https://doi.org/10.1016/j.cbpa.2010.02.017AbstractOver the past 10 years, a handful of academic and industrial research groups have developed strategies for the synthesis and interrogation of DNA-encoded small-molecule libraries. These strategies can be divided into those in which DNA directs small-molecule synthesis and those in which it records the synthesis. These libraries have started to yield novel modulators of biological targets, including: SH3-domain-binding peptoids, macrocyclic peptide-based Bcl-XL/BH3 interaction disruptors, ligands for TNF, albumin, streptavidin and others, and small-molecule kinase inhibitors. The DNA-encoded library field holds the potential to address the general problem of biological ligand discovery, including pharmaceutical lead generation.
Matthew E. Welsch; Scott A. Snyder; Brent R. Stockwell Curr. Opin. Chem. Biol., 2010, 14(3), 347-361 https://doi.org/10.1016/j.cbpa.2010.02.018AbstractThis review explores the concept of using privileged scaffolds to identify biologically active compounds through building chemical libraries. We hope to accomplish three main objectives: to provide one of the most comprehensive listings of privileged scaffolds; to reveal through four selected examples the present state of the art in privileged scaffold library synthesis (in hopes of inspiring new and even more creative approaches); and also to offer some thoughts on how new privileged scaffolds might be identified and exploited.
Fabian Buller; Martina Steiner; Jörg Scheuermann; Luca Mannocci; Ina Nissen; Manuel Kohler; Christian Beisel; Dario Neri Bioorg. Med. Chem. Lett., 2010, 20(14), 4188-92 https://doi.org/10.1016/j.bmcl.2010.05.053AbstractDNA-encoded chemical libraries are large collections of small organic molecules, individually coupled to DNA fragments that serve as amplifiable identification bar codes. The isolation of specific binders requires a quantitative analysis of the distribution of DNA fragments in the library before and after capture on an immobilized target protein of interest. Here, we show how Illumina sequencing can be applied to the analysis of DNA-encoded chemical libraries, yielding over 10 million DNA sequence tags per flow-lane. The technology can be used in a multiplex format, allowing the encoding and subsequent sequencing of multiple selections in the same experiment. The sequence distributions in DNA-encoded chemical library selections were found to be similar to the ones obtained using 454 technology, thus reinforcing the concept that DNA sequencing is an appropriate avenue for the decoding of library selections. The large number of sequences obtained with the Illumina method now enables the study of very large DNA-encoded chemical libraries (>500,000 compounds) and reduces decoding costs.
Yevgeny Brudno; Michael E. Birnbaum; Ralph E Kleiner; David R. Liu Nat. Chem. Biol., 2010, 6, 148-155 https://doi.org/10.1038/nchembio.280AbstractMethods to evolve synthetic, rather than biological, polymers could significantly expand the functional potential of polymers that emerge from in vitro evolution. Requirements for synthetic polymer evolution include (i) sequence-specific polymerization of synthetic building blocks on an amplifiable template, (ii) display of the newly translated polymer strand in a manner that allows it to adopt folded structures, (iii) selection of synthetic polymer libraries for desired binding or catalytic properties and (iv) amplification of template sequences that survive selection in a manner that allows subsequent translation. Here we report the development of such a system for peptide nucleic acids (PNAs) using a set of 12 PNA pentamer building blocks. We validated the system by performing six iterated cycles of translation, selection and amplification on a library of 4.3 × 10^8 PNA-encoding DNA templates and observed >1,000,000-fold overall enrichment of a template encoding a biotinylated (streptavidin-binding) PNA. These results collectively provide an experimental foundation for PNA evolution in the laboratory.
Dario Neri; André W. Brändli Nat. Chem. Biol., 2009, 5, 452–453 https://doi.org/10.1038/nchembio0709-452AbstractA new methodology combining small molecules and phage-displayed peptides enables the isolation of chemically modified bicyclic peptides capable of high-affinity recognition of target proteins.
Christopher Arico-Muendela; Paolo A. Centrella; Brooke D. Contonio; Barry A. Morgan; Gary O’Donovan; Christopher L. Paradise; Steven R. Skinner; Barbara Sluboski; Jennifer L. Svendsen; Kerry F. White; Anjan Debnath; Jiri Gut; Nathan Wilson; James H. McKerrow; Joseph L. DeRisi; Philip J. Rosenthal; Peter K. Chiang Bioorg. Med. Chem. Lett., 2009, 19, 19, 5128-5131 https://doi.org/10.1016/j.bmcl.2009.07.029AbstractFumagillin, an irreversible inhibitor of MetAP2, has been shown to potently inhibit growth of malaria parasites in vitro. Here, we demonstrate activity of fumagillin analogs with an improved pharmacokinetic profile against malaria parasites, trypanosomes, and amoebas. A subset of the compounds showed efficacy in a murine malaria model. The observed SAR forms a basis for further optimization of fumagillin based inhibitors against parasitic targets by inhibition of MetAP2.
David J. Gorin; Adam S. Kamlet; David R. Liu J. Am. Chem. Soc., 2009, 131, 26, 9189-9191 https://doi.org/10.1021/ja903084aAbstract(Figure Presented) In vitro selection is a key component of efforts to discover functional nucleic acids and small molecules from libraries of DNA, RNA, and DNA-encoded small molecules. Such selections have been widely used to evolve RNA and DNA catalysts and, more recently, to discover new reactions from DNA-encoded libraries of potential substrates. While effective, current strategies for selections of bond-forming and bond-cleaving reactivity are generally indirect, require the synthesis of biotin-linked substrates, and involve multiple solution-phase and solid-phase manipulations. In this work we report the successful development and validation of reactivity-dependent PCR (RDPCR), a new method that more directly links bond formation or bond cleavage with the amplification of desired sequences and that obviates the need for solid-phase capture, washing, and elution steps. We show that RDPCR can be used to select for bond formation in the context of reaction discovery and for bond cleavage in the context of protease activity profiling.
Fabian Buller; Yixin Zhang; Jörg Scheuermann; Juliane Schäfer; Peter Bühlmann; Dario Neri Chem. Biol., 2009, 16, 10, 1075-1086 https://doi.org/10.1016/j.chembiol.2009.09.011AbstractDNA-encoded chemical libraries are promising tools for the discovery of ligands toward protein targets of pharmaceutical relevance. DNA-encoded small molecules can be enriched in affinity-based selections and their unique DNA "barcode" allows the amplification and identification by high-throughput sequencing. We describe selection experiments using a DNA-encoded 4000-compound library generated by Diels-Alder cycloadditions. High-throughput sequencing enabled the identification and relative quantification of library members before and after selection. Sequence enrichment profiles corresponding to the "bar-coded" library members were validated by affinity measurements of single compounds. We were able to affinity mature trypsin inhibitors and identify a series of albumin binders for the conjugation of pharmaceuticals. Furthermore, we discovered a ligand for the antiapoptotic Bcl-xL protein and a class of tumor necrosis factor (TNF) binders that completely inhibited TNF-mediated killing of L-M fibroblasts in vitro.
Mohane Selvaraj Coumar; Chun Hei Antonio Cheung; Jang-Yang Chang; Hsing-Pang Hsieh Expert Opin. Ther. Patents, 2009, 19, 3, 321-356 https://doi.org/10.1517/13543770802646949AbstractThis review is one of a two part series on Aurora kinase inhibitors. The other part ‘Aurora Kinase Inhibitors in Clinical and Preclinical Testing’ will appear in Expert Opinion in Investigational Drugs. Background: Aurora-A, Aurora-B and Aurora-C, members of serine/threonine kinase family, play an important role in mitosis. They are essential for spindle assembly, centrosome maturation, chromosomal segregation and cytokinesis during mitosis. Abnormalities in the mitotic process as a result of overexpression/amplification of Aurora kinase have been linked to genomic instability leading to tumorigenesis. Hence, the use of Aurora kinase small-molecule inhibitors as a potential molecular-targeted therapeutic intervention for cancer is being pursued. Objective: A number of reviews focus on the biology of Aurora kinase; a few focus on the medicinal chemistry aspect of Aurora kinase inhibitor development. Here, we review the medicinal chemistry aspect of Aurora kinase inhibitors, with a particular emphasis on the patent literature. Method: The Scifinder® and Delphion® databases were used to search the literature for Aurora kinase inhibitors. Approximately 150 patents and 700 journal references are available, most of them published in the last 5 years. Conclusion/results: Analysis of the literature reveals three common strategies utilized by different groups in developing Aurora kinase inhibitors. These are discussed in detail and could be of use to medicinal chemists in laying out new strategies for developing novel Aurora kinase inhibitors.
Zhengrong Zhu; John W. Cuozzo J. Biomol. Screen., 2009, 14(10), 1157-64 https://doi.org/10.1177/1087057109350114AbstractHigh-throughput affinity-based technologies are rapidly growing in use as primary screening methods in drug discovery. In this review, their principles and applications are described and their impact on small-molecule drug discovery is evaluated.In general, these technologies can be divided into 2 groups: those that detect binding interactions by measuring changes to the protein target and those that detect bound compounds. Technologies detecting binding interactions by focusing on the protein have limited throughput but can reveal mechanistic information about the binding interaction; technologies detecting bound compounds have very high throughput, some even significantly higher than current high-throughput screening technologies,but offer limited information about the binding interaction. In addition, the appropriate use of affinity-based technologiesis discussed. Finally, nanotechnology is predicted to generate a significant impact on the future of affinity-based technologies.
Ghotas Evindar; Alexander L. Satz; Sylvie G. Bernier; Malcolm J. Kavarana; Elisabeth Doyle; Jeanine Lorusso; Nazbeh Taghizadeh; Keith Halley; Amy Hutchings; Michael S. Kelley; Albion D. Wright; Ashis K. Saha; Gerhard Hannig; Barry A. Morgan; William F. Westlin Bioorg. Med. Chem. Lett., 2009, 19, 8, 2315-2319 https://doi.org/10.1016/j.bmcl.2009.02.073AbstractIn pursuit of potent and selective sphingosine-1-phosphate receptor agonists, we have utilized previously reported phenylamide and phenylimidazole scaffolds to explore extensive side-chain modifications to generate new molecular entities. A number of designed molecules demonstrate good selectivity and excellent in vitro and in vivo potency in both mouse and rat models. Oral administration of the lead molecule 11c (PPI-4667) demonstrated potent and dose-responsive lymphopenia.
Matthew A. Clark; Raksha A. Acharya; Christopher C. Arico-Muendel; Svetlana L. Belyanskaya; Dennis R. Benjamin; Neil R. Carlson; Paolo A. Centrella; Cynthia H. Chiu; Steffen P. Creaser; John W. Cuozzo; Christopher P. Davie; Yun Ding; G. Joseph Franklin; Kurt D. Franzen; Malcolm L. Gefter; Steven P. Hale; Nils Jakob Vest Hansen; David I. Israel; Jinwei Jiang; Malcolm J. Kavarana; Michael S. Kelley; Christopher S. Kollmann; Fan Li; Kenneth Lind; Sibongile Mataruse; Patricia F. Medeiros; Jeffrey A. Messer; Paul Myers; Heather O'Keefe; Matthew C. Oliff; Cecil E. Rise; Alexander L. Satz; Steven R. Skinner; Jennifer L. Svendsen; Lujia Tang; Kurt van Vloten; Richard W. Wagner; Gang Yao; Baoguang Zhao; Barry A. Morgan Nat. Chem. Biol., 2009, 5, 9, 647-654 https://doi.org/10.1038/nchembio.211AbstractBiochemical combinatorial techniques such as phage display, RNA display and oligonucleotide aptamers have proven to be reliable methods for generation of ligands to protein targets. Adapting these techniques to small synthetic molecules has been a long-sought goal. We report the synthesis and interrogation of an 800-million-member DNA-encoded library in which small molecules are covalently attached to an encoding oligonucleotide. The library was assembled by a combination of chemical and enzymatic synthesis, and interrogated by affinity selection. We describe methods for the selection and deconvolution of the chemical display library, and the discovery of inhibitors for two enzymes: Aurora A kinase and p38 MAP kinase.
Margit Haahr Hansen; Peter Blakskjær; Lars Kolster Petersen; Tara Heitner Hansen; Jonas Westergaard Højfeldt; Kurt Vesterager Gothelf; Nils Jakob Vest Hansen J. Am. Chem. Soc., 2009, 131, 3, 1322-1327 https://doi.org/10.1021/ja808558aAbstractThe center of DNA three-way junctions, constituting a yoctoliter (10 -24 L) volume, is applied as an efficient reactor to create DNA-encoded libraries of chemical products. Amino acids and short peptides are linked to oligonucleotides via cleavable and noncleavable linkers. The oligonucleotide sequences contain two universal assembling domains at the center and a distal codon sequence specific for the attached building block. Stepwise self-assembly and chemical reactions of these conjugates in a combinatorial fashion create a library of pentapeptides in DNA three-way junctions in a single reaction vessel. We demonstrate the formation of an evenly distributed library of 100 peptides. Each library member contains a short synthetic peptide attached to a unique genetic code creating the necessary "genotype-phenotype" linkage essential to the process of in vitro molecular evolution. Selective enrichment of the [Leu]-enkephalin peptide from an original frequency of 1 in 10 million in a model library to a final frequency of 1.7percent in only two rounds of affinity selection is described and demonstrates successful molecular evolution for a non-natural system.
Andrei P. Drabovich; Maxim V. Berezovski; Michael U. Musheev; Sergey N. Krylov Anal. Chem., 2009, 81(1), 490-494 https://doi.org/10.1021/ac8023813AbstractThe development of drugs and diagnostics with desirable characteristics requires smart small-molecule ligands—ligands with predefined binding parameters of interaction with the target. Here, we propose a general approach for selection of such ligands from highly diverse combinatorial libraries of small molecules by methods of kinetic capillary electrophoresis (KCE). We deduct three fundamental requirements for the combinatorial library to suit the KCE-based selection of smart ligands and suggest a universal design of the library for selecting smart small-molecule ligands: every small molecule in the library is tagged with DNA that encodes the structure of the molecule. Finally, we use several DNA-tagged small molecules, which represent a hypothetical library, to prove experimentally selection of smart small-molecule ligands by the proposed approach.
Ghotas Evindar; Sylvie G. Bernier; Malcolm J. Kavarana; Elisabeth Doyle; Jeanine Lorusso; Michael S. Kelley; Keith Halley; Amy Hutchings; Albion D. Wright; Ashis K. Sahaa; Gerhard Hannig; Barry A. Morgan; William F. Westlin Bioorg. Med. Chem. Lett., 2009, 19, 2, 369-372 https://doi.org/10.1016/j.bmcl.2008.11.072AbstractIn the design of potent and selective sphingosine-1-phosphate receptor agonists, we were able to identify two series of molecules based on phenylamide and phenylimidazole analogs of FTY-720. Several designed molecules in these scaffolds have demonstrated selectivity for S1P receptor subtype 1 versus 3 and excellent in vivo activity in mouse. Two molecules PPI-4621 (4b) and PPI-4691 (10a), demonstrated dose responsive lymphopenia, when administered orally.
Christopher C. Arico-Muendel; Dennis R. Benjamin; Teresa M. Caiazzo; Paolo A. Centrella; Brooke D. Contonio; Charles M. Cook; Elisabeth G. Doyle; Gerhard Hannig; Matthew T. Labenski; Lily L. Searle; Kenneth Lind; Barry A. Morgan; Gary Olson; Christopher L. Paradise; Christopher Self; Steven R. Skinner; Barbara Sluboski; Jennifer L. Svendsen; Charles D. Thompson; William Westlin; Kerry F. White J. Med. Chem., 2009, 52, 24, 8047-8056 https://doi.org/10.1021/jm901260kAbstractInhibition of methionine aminopeptidase-2 (MetAP2) represents a novel approach to antiangiogenic therapy. We describe the synthesis and activity of fumagillin analogues that address the pharmacokinetic and safety liabilities of earlier candidates in this compound class. Two-step elaboration of fumagillol with amines yielded a diverse series of carbamates at C6 of the cyclohexane spiroepoxide. The most potent of these compounds exhibited subnanomolar inhibition of cell proliferation in HUVEC and BAEC assays. Although a range of functionalities were tolerated at this position, α-trisubstituted amines possessed markedly decreased inhibitory activity, and this could be rationalized by modeling based on the known fumagillin-MetAP2 crystal structure. The lead compound resulting from these studies, (3R,4S,5S,6R)-5-methoxy-4-((2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl)-1-oxaspiro[2.5]octan-6-yl (R)-1-amino-3-methyl-1-oxobutan-2-ylcarbamate, (PPI-2458), demonstrated an improved pharmacokinetic profile relative to the earlier clinical candidate TNP-470, and has advanced into phase I clinical studies in non-Hodgkin’s lymphoma and solid cancers.
Tara Renee Heitner; Nils Jakob Vest Hansen Expert Opin. Drug Discov., 2009, 4(11), 1201-13 https://doi.org/10.1517/17460440903206940AbstractBackground: The field of DNA-encoded technology offers a cutting edge approach to creating 10^9-10^12-size small molecule libraries for the rapid identification of drug-like hits. The YoctoReactor ® (yR) is the newest DNA-encoded technology and features an innovative and fundamentally different design. Objective: This technology evaluation presents the basic principles of the yR drug discovery technology platform and discusses its potential as an alternative to current hit discovery methods where high quality and selective drug-like hits can be delivered together with instant structure activity relationships (SAR). Conclusion: The yR controls and encodes billions of different synthesis combinatorially through self-assembly of DNA into three-dimensional DNA junctions. The center of these junctions forms a yoctoliter (10-24 L) volume reactor where single-molecule, DNA- encoded reactions can take place. yR technology circumvents the high throughput screening process (HTS). Instead, drug-like families of hits are rapidly identified by molecular evolution in a single tube approach making it highly parallelizable and scalable. DNA-encoded technologies such as the yR, therefore, represent a solution to many of the bottlenecks in current early phase discovery and in handling the vast emerging opportunities in functional genomics, systems biology and proteomics.
Katarzyna Gorska; Kuo-Ting Huang; Olivier Chaloin; Nicolas Winssinger Angew. Chem. Int. Ed. Engl., 2009, 48, 7695-7700 https://doi.org/10.1002/anie.200903328AbstractAll under control: The programmability of hybridization has been utilized to generate a combinatorial library of structures that emulate the topologies of complex carbohydrates interacting with an antibody that shows broad‐spectrum activity against HIV. This simple method involves attaching oligosaccharides tagged with peptide nucleic acids onto DNA templates in a controlled manner (see schematic picture).
Matthew A. Clark; Raksha A. Acharya; Christopher C. Arico-Muendel; Svetlana L. Belyanskaya; Dennis R. Benjamin; Neil R. Carlson; Paolo A. Centrella; Cynthia H. Chiu; Steffen P. Creaser; John W. Cuozzo; Christopher P. Davie; Yun Ding; G. Joseph Franklin; Kurt D. Franzen; Malcolm L. Gefter; Steven P. Hale; Nils Jakob Vest Hansen; David I. Israel; Jinwei Jiang; Malcolm J. Kavarana; Michael S. Kelley; Christopher S. Kollmann; Fan Li; Kenneth Lind; Sibongile Mataruse; Patricia F. Medeiros; Jeffrey A. Messer; Paul Myers; Heather O'Keefe; Matthew C. Oliff; Cecil E. Rise; Alexander L. Satz; Steven R. Skinner; Jennifer L. Svendsen; Lujia Tang; Kurt van Vloten; Richard W. Wagner; Gang Yao; Baoguang Zhao; Barry A. Morgan PDBJ - 3HA8https://pdbj.org/mine/summary/3ha8 Abstract Mitogen-activated protein kinase 14, N~2~-{4-[6-(3,4-dihydroquinolin-1(2H)-ylcarbonyl)-1H-benzimidazol-1-yl]-6-ethoxy-1,3,5-triazin-2-yl}-3-(2,2-dimethyl-4H-1,3-benzodioxin-6-yl)-N-methyl-L-alaninamide
Jennifer M. Heemstra; David R. Liu J. Am. Chem. Soc., 2009, 131(32), 11347-9 https://doi.org/10.1021/ja904712tAbstractThe templated synthesis of nucleic acids has previously been achieved through the backbone ligation of preformed nucleotide monomers or oligomers. In contrast, here we demonstrate templated nucleic acid synthesis using a base-filling approach in which individual bases are added to abasic sites of a peptide nucleic acid (PNA). Because nucleobase substrates in this approach are not self-reactive, a base-filling approach may reduce the formation of nontemplated reaction products. Using either reductive amination or amine acylation chemistries, we observed efficient and selective addition of each of the four nucleobases to an abasic site in the middle of the PNA strand. We also describe the addition of single nucleobases to the end of a PNA strand through base filling, as well as the tandem addition of two bases to the middle of the PNA strand. These findings represent an experimental foundation for nonenzymatic information transfer through base filling.
Luca Mannocci; Yixin Zhang; Jörg Scheuermann; Markus Leimbacher; Gianluca De Bellis; Ermanno Rizzi; Christoph Dumelin; Samu Melkko; Dario Neri Proc. Natl. Acad. Sci. USA, 2008, 105(46), 17670-5 https://doi.org/10.1073/pnas.0805130105AbstractDNA encoding facilitates the construction and screening of large chemical libraries. Here, we describe general strategies for the stepwise coupling of coding DNA fragments to nascent organic molecules throughout individual reaction steps as well as the first implementation of high-throughput sequencing for the identification and relative quantification of the library members. The methodology was exemplified in the construction of a DNA-encoded chemical library containing 4,000 compounds and in the discovery of binders to streptavidin, matrix metalloproteinase 3, and polyclonal human IgG.
Edward M. Driggers; Stephen P. Hale; Jinbo Lee; Nicholas K. Terrett Nat. Rev. Drug Discov., 2008, 7, 608-624 https://doi.org/10.1038/nrd2590AbstractMacrocyclic natural products have evolved to fulfil numerous biochemical functions, and their profound pharmacological properties have led to their development as drugs. A macrocycle provides diverse functionality and stereochemical complexity in a conformationally pre-organized ring structure. This can result in high affinity and selectivity for protein targets, while preserving sufficient bioavailability to reach intracellular locations. Despite these valuable characteristics, and the proven success of more than 100 marketed macrocycle drugs derived from natural products, this structural class has been poorly explored within drug discovery. This is in part due to concerns about synthetic intractability and non-drug-like properties. This Review describes the growing body of data in favour of macrocyclic therapeutics, and demonstrates that this class of compounds can be both fully drug-like in its properties and readily prepared owing to recent advances in synthetic medicinal chemistry.
Brian N. Tse; Thomas M. Snyder; Yinghua Shen; David R. Liu J. Am. Chem. Soc., 2008, 130(46), 15611-26 https://doi.org/10.1021/ja805649fAbstractDNA-templated organic synthesis enables the translation, selection, and amplification of DNA sequences encoding synthetic small-molecule libraries. Previously we described the DNA-templated multistep synthesis and model in vitro selection of a pilot library of 65 macrocycles. In this work, we report several key developments that enable the DNA-templated synthesis of much larger (>10,000-membered) small-molecule libraries. We developed and validated a capping-based approach to DNA-templated library synthesis that increases final product yields, simplifies the structure and preparation of reagents, and reduces the number of required manipulations. To expand the size and structural diversity of the macrocycle library, we augmented the number of building blocks in each DNA-templated step from 4 to 12, selected 8 different starting scaffolds which result in 4 macrocycle ring sizes and 2 building-block orientations, and confirmed the ability of the 36 building blocks and 8 scaffolds to generate DNA-templated macrocycle products. We computationally generated and experimentally validated an expanded set of codons sufficient to support 1728 combinations of step 1, step 2, and step 3 building blocks. Finally, we developed new high-resolution LC/MS analysis methods to assess the quality of large DNA-templated small-molecule libraries. Integrating these four developments, we executed the translation of 13,824 DNA templates into their corresponding small-molecule macrocycles. Analysis of the resulting libraries is consistent with excellent (>90%) representation of desired macrocycle products and a stringent test of sequence specificity suggests a high degree of sequence fidelity during translation. The quality and structural diversity of this expanded DNA-templated library provides a rich starting point for the discovery of functional synthetic small-molecule macrocycles.
Fabian Buller; Luca Mannocci; Yixin Zhang; Christoph E. Dumelin; Jörg Scheuermann; Dario Neri Bioorg. Med. Chem. Lett., 2008, 18, 5926-5931 https://doi.org/10.1016/j.bmcl.2008.07.038AbstractDNA-encoded chemical libraries are increasingly being employed for the identification of binding molecules to protein targets of pharmaceutical relevance. Here, we describe the synthesis and characterization of a DNA-encoded chemical library, consisting of 4000 compounds generated by Diels-Alder cycloaddition reactions. The compounds were encoded with unique DNA fragments which were generated through a stepwise assembly process and serve as amplifiable bar codes for the identification and relative quantification of library members.
Scott K. Silverman Chem. Commun., 2008, 3467-3485 https://doi.org/10.1039/B807292MAbstractThe discovery of naturally occurring catalytic RNA (RNA enzymes, or ribozymes) in the 1980s immediately revised the view of RNA as a passive messenger that solely carries information from DNA to proteins. Because DNA and RNA differ only by the absence or presence of a 2′-hydroxyl group on each ribose ring of the polymer, the question of ‘catalytic DNA?’ arises. Although no natural DNA catalysts have been reported, since 1994 many artificial DNA enzymes, or ‘deoxyribozymes’, have been described. Deoxyribozymes offer insight into the mechanisms of natural and artificial ribozymes. DNA enzymes are also used as tools for in vitro and in vivo biochemistry, and they are key components of analytical sensors. This review focuses primarily on catalytic DNA for synthetic applications. Broadly defined, deoxyribozymes may have the greatest potential for catalyzing reactions in which the high selectivities of ‘enzymes’ are advantageous relative to traditional small-molecule catalysts. Although the scope of DNA-catalyzed synthesis is currently limited in most cases to oligonucleotide substrates, recent efforts have began to expand this frontier in promising new directions.
Srinivasu Pothukanuri; Zbigniew Pianowski; Nicolas Winssinger Eur. J. Org. Chem., 2008, 3141-3148 https://doi.org/10.1002/ejoc.200800141AbstractWe present a thorough investigation of six types of protecting groups for the terminal nitrogen atom and five protecting groups on the nucleobases of peptide nucleic acids for fully orthogonal synthesis with Fmoc. Peptide nucleic acids (PNAs) hybridize to natural oligonucleotides according toWatson and Crick base-pairing rules. The robustness of PNA oligomers and ease of synthesis have made them an attractive platform to encode small or macromolecules for microarraying purposes and other applications based on programmable self assembly. A cornerstone of these endeavors is the orthogonality of PNA synthesis with other chemistries. Herein, we present a thorough investigation of six types of protecting groups for the terminal nitrogen atom (Alloc, Teoc, 4-N3Cbz, Fmoc, 4-OTBSCbz, and Azoc) and five protecting groups on the nucleobases (Cl-Bhoc, FBhoc,Teoc, 4-OMeCbz, and Boc).
Zbigniew L. Pianowski; Nicolas Winssinger Chem. Soc. Rev., 2008, 37, 1330-1336 https://doi.org/10.1039/B706610BAbstractThis tutorial review serves as an introduction to the use of oligonucleotides and in particular peptide nucleic acids (PNAs) to encode function beyond heredity. Applications in chemical biology are reviewed starting with the use of nucleic acid tags to program self-assembled microarrays of small and macromolecules, followed by the use of nucleic acid templated reactions for the purpose of DNA or RNA sensing and finally, the use of nucleic acid templates to display ligands.
Jörg Scheuermann; Christoph E. Dumelin; Samu Melkko; Yixin Zhang; Luca Mannocci; Madalina Jaggi; Jens Sobek; Dario Neri Bioconjugate Chem., 2008, 19, 3, 778-785 https://doi.org/10.1021/bc7004347AbstractEncoded self-assembling chemical (ESAC) libraries are characterized by the covalent display of chemical moieties at the extremity of self-assembling oligonucleotides carrying a unique DNA sequence for the identification of the corresponding chemical moiety. We have used ESAC library technology in a two-step selection procedure for the identification of novel inhibitors of stromelysin-1 (MMP-3), a matrix metalloproteinase involved in both physiological and pathological tissue remodeling processes, yielding novel inhibitors with micromolar potency.
Christoph E. Dumelin; Sabrina Trüssel; Fabian Buller; Eveline Trachsel; Frank Bootz; Yixin Zhang; Luca Mannocci; Susanne C. Beck; Mihaela Drumea‐Mirancea; Mathias W. Seeliger; Christof Baltes; Thomas Müggler; Felicitas Kranz; Markus Rudin; Samu Melkko; Jörg Scheuermann; Dario Neri Angew. Chem. Int. Ed. Engl., 2008, 47, 3196-3201 https://doi.org/10.1002/anie.200704936AbstractSeeing eye to eye: Plasma‐protein binding is effective in improving the pharmacokinetic properties of otherwise short‐lived molecules. One compound in a class of small portable albumin binders can be used to improve the in vivo circulatory half‐life of two widely used contrast agents. It improves the imaging performance of fluorescein in angiographic analysis of the retina of mice (see picture).
Thomas M. Snyder; Brian N. Tse; David R. Liu J. Am. Chem. Soc., 2008, 130(4), 1392-401 https://doi.org/10.1021/ja076780uAbstractDNA-templated organic synthesis enables the translation, selection, and amplification of DNA sequences encoding synthetic small-molecule libraries. As the size of DNA-templated libraries increases, the possibility of forming intramolecularly base-paired structures within templates that impede templated reactions increases as well. To achieve uniform reactivity across many template sequences and to computationally predict and remove any problematic sequences from DNA-templated libraries, we have systematically examined the effects of template sequence and secondary structure on DNA-templated reactivity. By testing a series of template sequences computationally designed to contain different degrees of internal secondary structure, we observed that high levels of predicted secondary structure involving the reagent binding site within a DNA template interfere with reagent hybridization and impair reactivity, as expected. Unexpectedly, we also discovered that templates containing virtually no predicted internal secondary structure also exhibit poor reaction efficiencies. Further studies revealed that a modest degree of internal secondary structure is required to maximize effective molarities between reactants, possibly by compacting intervening template nucleotides that separate the hybridized reactants. Therefore, ideal sequences for DNA-templated synthesis lie between two undesirable extremes of too much or too little internal secondary structure. The relationship between effective molarity and intervening nucleic acid secondary structure described in this work may also apply to nucleic acid sequences in living systems that separate interacting biological molecules.
Ralph E. Kleiner; Yevgeny Brudno; Michael E. Birnbaum; David R. Liu J. Am. Chem. Soc., 2008, 130(14), 4646-59 https://doi.org/10.1021/ja0753997AbstractThe DNA-templated polymerization of synthetic building blocks provides a potential route to the laboratory evolution of sequence-defined polymers with structures and properties not necessarily limited to those of natural biopolymers. We previously reported the efficient and sequence-specific DNA-templated polymerization of peptide nucleic acid (PNA) aldehydes. Here, we report the enzyme-free, DNA-templated polymerization of side-chain-functionalized PNA tetramer and pentamer aldehydes. We observed that polymerization of tetramer and pentamer PNA building blocks with a single lysine-based side chain at various positions in the building block could proceed efficiently and sequence specifically. In addition, DNA-templated polymerization also proceeded efficiently and in a sequence-specific manner with pentamer PNA aldehydes containing two or three lysine side chains in a single building block to generate more densely functionalized polymers. To further our understanding of side-chain compatibility and expand the capabilities of this system, we also examined the polymerization efficiencies of 20 pentamer building blocks each containing one of five different side-chain groups and four different side-chain regio- and stereochemistries. Polymerization reactions were efficient for all five different side-chain groups and for three of the four combinations of side-chain regio- and stereochemistries. Differences in the efficiency and initial rate of polymerization correlate with the apparent melting temperature of each building block, which is dependent on side-chain regio- and stereochemistry but relatively insensitive to side-chain structure among the substrates tested. Our findings represent a significant step toward the evolution of sequence-defined synthetic polymers and also demonstrate that enzyme-free nucleic acid-templated polymerization can occur efficiently using substrates with a wide range of side-chain structures, functionalization positions within each building block, and functionalization densities.
Yumei Huang; James M. Coull J. Am. Chem. Soc., 2008, 130, 11, 3238-3239 https://doi.org/10.1021/ja0753602AbstractA hemicyanine fluorescent dye was generated by a diamine-catalyzed, DNA-templated, aldol-type condensation from nonfluorescent precursors. Studies of reaction rate and yield as a function of catalyst structure indicated the diamine catalyst operates in a concerted mechanism on both reaction components. Our findings expand the scope of reactions that can be performed by DPC and demonstrate that de novo chemical synthesis of labels can be coupled to biological recognition events in a homogeneous format with essentially no background.
Yeow Meng Chee; San Ling IEEE Trans. Inf. Theory, 2008, 54, 1 https://doi.org/10.1109/TIT.2007.911167AbstractThe design of large libraries of oligonucleotides having constant-content and satisfying Hamming distance constraints between oligonucleotides and their Watson-Crick complements is important in reducing hybridization errors in DNA computing, DNA microarray technologies, and molecular bar coding. Various techniques have been studied for the construction of such oligonucleotide libraries, ranging from algorithmic constructions via stochastic local search to theoretical constructions via coding theory. A new stochastic local search method is introduced, which yields improvements for more than one third of the benchmark lower bounds of Gaborit and King (2005) for n-mer oligonucleotide libraries when n les 14. Several optimal libraries are also found by computing maximum cliques on certain graphs.
Delphine Pouchain; Juan J. Díaz-Mochón; Laurent Bialy; Mark Bradley ACS Chem. Biol., 2007, 2, 12, 810-818 https://doi.org/10.1021/cb700199kAbstractA 10,000 member peptide nucleic acid (PNA) encoded peptide library was prepared, treated with the Abelson tyrosine kinase (Abl), and decoded using a DNA microarray and a fluorescently labeled secondary antiphosphotyrosine antibody. A dual-color approach ensured internal referencing for each and every member of the library and the generation of robust data sets. Analysis identified 155 peptides (out of 10,000) that were strongly phosphorylated by Abl in full agreement with known Abl specificities. BLAST analysis identified known cellular Abl substrates such as c-Jun amino-terminal kinase as well as new potential target proteins such as the G-protein coupled receptor kinase 6 and diacylglycerol kinase gamma. To illustrate the generalization of this approach, two other tyrosine kinases, human epidermal growth factor 2 (Her2) and vascular endothelial growth factor receptor 2/kinase insert domain protein receptor (VEGFR2/KDR), were profiled allowing characterization of specific peptide sequences known to interact with these kinases; under these conditions Her2 was demonstrated to have a marked preference for d-proline perhaps offering a unique means of targeting and inhibiting this kinase.
Samu Melkko; Christoph E. Dumelin; Jörg Scheuermann; Dario Neri Drug Discov. Today, 2007, 12, 465-471 https://doi.org/10.1016/j.drudis.2007.04.007AbstractThe isolation of specific binding molecules is a central problem in the drug discovery process and might be useful for the elucidation of the biological function of proteins identified in genome and proteome research. Libraries of organic molecules, conjugated covalently to DNA tags that serve as identification bar codes, have been proposed recently as a way to identify ligands of target proteins of choice efficiently. Here, we analyze the different strategies for constructing DNA-encoded chemical libraries, and the potential and challenges of this promising technology.
S. Jarrett Wrenn; Pehr B. Harbury Annu. Rev. Biochem., 2007, 76, 331-349 https://doi.org/10.1146/annurev.biochem.76.062205.122741AbstractIn modern academic and industrial laboratories, evolutionary strategies are used routinely to identify biopolymers with novel activities. Large libraries of nucleic acids (∼10^15) or peptides and proteins (∼10^13) can be subjected to multiple rounds of selective pressure, amplification, and diversification, yielding individual sequences with desirable properties. Although the evolutionary approach is a powerful search tool, the chemical nature of biopolymers is not suited for all purposes. Application of evolutionary strategies to libraries of arbitrary chemical composition would overcome this problem, and radically change traditional small-molecule discovery. The chemical make-up of in vitro evolution libraries has necessarily been limited, because library synthesis relies on enzymes. A great deal of current research focuses on expanding the chemical repertoire of in vitro evolution by (a) broadening enzyme substrate specificities to include unnatural building blocks, or (b) developing methods to translate DNA sequences into multistep organic syntheses. We discuss the strengths and weaknesses of the approaches, review the successes, and consider the future of chemical evolution as a tool.
Mary M. Rozenman; Matthew W. Kanan; David R. Liu J. Am. Chem. Soc., 2007, 129(48), 14933-8 https://doi.org/10.1021/ja074155jAbstractWe have developed and applied an approach to reaction discovery that takes advantage of DNA encoding, DNA-programmed assembly of substrate pairs, in vitro selection, and PCR amplification, yet does not require reaction conditions that support DNA hybridization. This system allows the simultaneous evaluation of >200 potential bond-forming combinations of substrates in a single experiment and can be applied in a range of solvent and temperature conditions. In an initial application, we applied this system to explore Au(III)-mediated chemistry and uncovered a simple, mild method for the selective Markovnikov-type hydroarylation of vinyl arenes and trisubstituted olefins with indoles.
Mary M. Rozenman; Brian R. McNaughton; David R. Liu Curr. Opin. Chem. Biol., 2007, 11, 3, 259-268 https://doi.org/10.1016/j.cbpa.2007.05.016AbstractMolecular evolution has been widely applied in the laboratory to generate novel biological macromolecules. The principles underlying evolution have more recently been used to address problems in the chemical sciences, including the discovery of functional synthetic small molecules, catalysts, materials and new chemical reactions. The application of these principles in dynamic combinatorial chemistry and in efforts involving small molecule–nucleic acid conjugates has facilitated the evaluation of large numbers of candidate structures or reactions for desired characteristics. These early efforts suggest the promise of pairing evolutionary approaches with synthetic chemistry.
S. Jarrett Wrenn; Rebecca M. Weisinger; David R. Halpin; Pehr B. Harbury J. Am. Chem. Soc., 2007, 129, 13137-13143 https://doi.org/10.1021/ja073993aAbstractThe recognition and catalytic properties of biopolymers derive from an elegant evolutionary mechanism, whereby the genetic material encoding molecules with superior functional attributes survives a selective pressure and is propagated to subsequent generations. This process is routinely mimicked in vitro to generate nucleic-acid or peptide ligands and catalysts. Recent advances in DNA-programmed organic synthesis have raised the possibility that evolutionary strategies could also be used for small-molecule discovery, but the idea remains unproven. Here, using DNA-programmed combinatorial chemistry, a collection of 100 million distinct compounds is synthesized and subjected to selection for binding to the N-terminal SH3 domain of the proto-oncogene Crk. Over six generations, the molecular population converges to a small number of novel SH3 domain ligands. Remarkably, the hits bind with affinities similar to those of peptide SH3 ligands isolated from phage libraries of comparable complexity. The evolutionary approach has the potential to drastically simplify and accelerate small-molecule discovery.
Samu Melkko; Yixin Zhang; Christoph E. Dumelin; Jörg Scheuermann; Dario Neri Angew. Chem. Int. Ed. Engl., 2007, 46(25), 4671-4 https://doi.org/10.1002/anie.200700654AbstractMaturing is easy to do: Annealing benzamidine-oligonucleotide conjugates with a library of DNA-encoded compounds allows the affinity capture of pharmacophores that are capable of binding to exosites adjacent to the primary substrate-binding pocket of the serine protease trypsin. Selected conjugates show an improvement in IC50 values of several orders of magnitude compared with the starting benzamidine.
François Debaene; Julien A.Da Silva; Zbigniew Pianowski; Fernando J.Duran; Nicolas Winssinger Tetrahedron, 2007, 63, 6577-6586 https://doi.org/10.1016/j.tet.2007.03.033AbstractSeven PNA-encoded combinatorial libraries targeting proteases and phosphatases with covalent reversible and irreversible mechanism-based inhibitors were prepared. The libraries were synthesized using modified PNA monomers, which dramatically increase the water solubility of the libraries in biologically relevant buffers. The libraries were shown to selectively inhibit targeted enzymes.
Ravindra Kumar; Afaf El-Sagheer; John Tumpane; Per Lincoln; L. Marcus Wilhelmsson; Tom Brown J. Am. Chem. Soc., 2007, 129, 21, 6859-6864 https://doi.org/10.1021/ja070273vAbstractThe copper-catalyzed azide−alkyne cycloaddition reaction has been used for the template-mediated chemical ligation of two oligonucleotide strands, one with a 5‘-alkyne and the other with a 3‘-azide, to produce a DNA strand with an unnatural backbone at the ligation point. A template-free click-ligation reaction has been used for the intramolecular circularization of a single stranded oligonucleotide which was used as a template for the synthesis of a covalently closed DNA catenane.
Juan J. Díaz-Mochón; Laurent Bialy; Mark Bradley Chem. Commun., 2006, 3984-3986 https://doi.org/10.1039/B609029JAbstractA 10,000 member PNA-encoded library of FRET based peptides was synthesised for global analysis of protease cleavage specificity; analysis was achieved using a DNA microarray and consumed minimal quantities of enzyme (60 pmole) and library (3.5 nmole).
Hugo D. Urbina; François Debaene; Bernard Jost; Christine Bole-Feysot; Daniel E. Mason; Petr Kuzmic; Jennifer L. Harris; Nicolas Winssinger ChemBioChem, 2006, 7, 1790-1797 https://doi.org/10.1002/cbic.200600242AbstractSmall-molecule microarrays are attractive for chemical biology as they permit the analysis of hundreds to thousands of interactions in a highly miniaturized format. Methods to prepare small-molecule microarrays from combinatorial libraries by a self-assembly process based on the sequence-specific hybridization of peptide nucleic acid (PNA) encoded libraries to oligonucleotide arrays are presented. A systematic study of the dynamic range for multiple detection agents, including direct fluorescence of attached fluorescein and cyanine-3 dyes, antibody-mediated fluorescence amplification, and biotin-gold nanoparticle detection, demonstrated that individual PNA-encoded probes can be detected to concentrations of 10 pM on the oligonucleotide microarrays. Furthermore, a new method for parallel processing of biological samples by using gel-based separation of probes is presented. The methods presented in this report are exemplified through profiling two closely related cysteine proteases, cathepsin K and cathepsin F, across a 625-member PNA-encoded tetrapeptide acrylate library. A series of the specific cathepsin K and F inhibitors identified from the library were kinetically characterized and shown to correlate with the observed microarray profile, thus validating the described methods. Importantly, it was shown that this method could be used to obtain orthogonal inhibitors that displayed greater than tenfold selectivity for these closely related cathepsins
Adam P. Silverman; Eric T. Kool Chem. Rev., 2006, 106, 9, 3775-3789 https://doi.org/10.1021/cr050057+AbstractThe aim of this review is to summarize recent advances in the development of chemical reactions that are triggered by hybridization of an oligonucleotidesa synthetic short strand of DNAsto a target RNA or DNA strand and outline how these reactions are being used, or could be used, in detecting and identifying such targets in a biological sample or cell. First, we will briefly discuss the basic principles of DNA and RNA molecular recognition that underlie this research. We will then outline the historical development of such templated reactions and then proceed to delineate the current state of the art. Finally, we will predict some future areas of special promise and mention some exciting new technologies that are under development in this field.
Nicolas Winssinger; Jennifer L. Harris Expert Rev. Proteomics, 2006, 2, 6, 937-947 https://doi.org/10.1586/14789450.2.6.937AbstractThe availability of complete genome sequences from numerous organisms has provided investigators with the challenge of assigning physiologic functions to the encoded gene products. To facilitate this process, multiple technologies have been developed to profile the transcriptome and the proteome, including methods to monitor the function of enzymes in complex biologic systems. These methods typically target specific classes of enzymes and attempt to correlate the enzymatic activity with the specific phenotype of interest. Here, technologies to measure enzymatic activity on a subproteomic scale are reviewed, including the authors’ own efforts, which are based on self-assembled microarrays utilizing peptide nucleic acid-encoded small-molecule libraries.
Naoki Kanoh; Hiroyuki Osada J. Synth. Org. Chem., 2006, 64, 6, 639-650 https://doi.org/10.5059/yukigoseikyokaishi.64.639AbstractThe identification of small molecule ligands for cellular proteins of biologically importance has not only provided new insights in biology, but has also provided organic chemists with the opportunity to create new synthetic molecules which efficiently regulate specific protein-protein interactions and biological processes. Small-molecule microarrays are recently developed as platforms to screen such small molecule ligands for proteins of interest. The small-molecule microarray technology is highly dependent on chemistry to immobilize small molecules on a variety of solid surfaces, and synthetic organic chemistry will continue to play a significant role in this field. This review covers the recent advances in the field with particular emphasis on the immobilization chemistry, as well as technical advances in the platform optimization and the successful applications of this platform.
Mary M. Rozenman1; David R. Liu ChemBioChem, 2006, 7(2), 253-6 https://doi.org/10.1002/cbic.200500413AbstractWhen DNA duplexes are preformed in water, then DNA-templated synthesis (DTS) can take place efficiently and sequence-specifically in organic solvents with low or minimal water content. This simple method enables reactions that are inaccessible in water, such as pyrrolidine-catalyzed aldol condensation, to be performed. The conditions needed to perform DTS in organic solvents also support DNA-templated reactions that are known to take place in aqueous solution.
Christoph E. Dumelin; Jörg Scheuermann; Samu Melkko; Dario Neri Bioconjugate Chem., 2006, 17, 2, 366-370 https://doi.org/10.1021/bc050282yAbstractDNA-encoded libraries of small organic molecules facilitate the construction of large, encoded self-assembling chemical libraries for the identification of high-affinity binders to protein targets. We have constructed a library of 477 chemical compounds, coupled to 48 mer-oligonucleotides, each containing a unique six-base sequence serving as "bar-code" for the identification of the chemical moiety. The functionality of the library was confirmed by selection and amplification of both high- and low-affinity binding molecules specific to streptavidin.
Samu Melkko; Christoph E. Dumelin; Jörg Scheuermann; Dario Neri Chem. Biol., 2006, 13, 2, 225-231 https://doi.org/10.1016/j.chembiol.2005.12.006AbstractThe simultaneous interaction of the binding moieties of a bidentate ligand on adjacent epitopes of a target protein represents an attractive avenue for the discovery of specific, high-affinity binders. We used short DNA fragments in heteroduplex format to scaffold pairs of binding molecules with defined spatial arrangements. Iminobiotin derivates were coupled either via bifunctional linkers or by using various oligonucleotides, thus allowing monovalent or bivalent binding to streptavidin. We determined the binding affinities of the synthesized constructs in solution. We also investigated the efficiency of recovery of superior bidentate ligands in affinity capture experiments, by using both radioactive counts and DNA microarrays as readouts. This analysis confirmed the suitability of the DNA heteroduplex as a scaffold for the identification of synergistic pairs of binding moieties, capable of a high-affinity interaction with protein targets by virtue of the chelate effect.
Paula A. Sacca; Angelo Fontana; Javier M. Montserrat; Adolfo M. Iribarren Chem. Biodivers., 2004, 1, 595-600 https://doi.org/10.1002/cbdv.200490049AbstractThe basis for further development of combinatorial libraries of modified oligonucleotides tagged by a codifying sequence is discussed. The chemistry involved in the orthogonal synthesis of both strands and some representative examples of building blocks are presented.
Christopher T. Calderone; David R. Liu Angew. Chem. Int. Ed. Engl., 2005, 44(45),7383-6 https://doi.org/10.1002/anie.200502899AbstractTwo unique features of DNA-templated synthesis were integrated to prepare complex small molecules by using iterated branching reaction pathways: the direction of multiple, otherwise “incompatible”, reaction types in one reaction vessel and the direction of sequence-programmed subsets of reaction intermediates to appropriate reagents.
Thomas M. Snyder; David R. Liu Angew. Chem. Int. Ed. Engl., 2005, 44(45), 7379-82 https://doi.org/10.1002/anie.200502879AbstractA triolefin and a tripeptide were prepared in a single solution (see scheme) by ordered multistep synthesis. Temperature-sensitive variations in DNA secondary structure were used to orchestrate a series of effective molarity changes among four reactants of comparable intrinsic reactivity, resulting predominantly in one ordered product out of many possibilities. This approach mimics the selective control of effective molarity at precise moments during biosynthesis.
Jennifer L. Harris; Nicolas Winssinger Chem. Eur. J., 2005, 11, 6792-6801 https://doi.org/10.1002/chem.200500305AbstractMicroarray-based technologies have attracted attention in chemical biology by virtue of their miniaturized format, which is well suited to probe ligand-protein interactions or investigate enzymatic activity in complex biological mixtures. A number of research groups have reported the preparation of surfaces on microarrays with specific functional groups to chemoselectively attach small molecules from libraries. We have developed an alternative method whereby libraries are encoded with peptide nucleic acid (PNA), such that libraries which exist as mixtures in solution self-assemble into an organized microarray through hybridization to produce readily available DNA arrays. This allows libraries synthesized by split and mix methods to be decoded in a single step. An asset of this method compared to direct spotting is that libraries can be used in solution for bioassays prior to self-assembly into the microarray format.
K. Ingrid Sprinzb; Debarati M. Tagore; Andrew D. Hamilton Bioorg. Med. Chem. Lett., 2005, 15, 17, 3908-3911 https://doi.org/10.1016/j.bmcl.2005.05.094AbstractA library of bidentate fragments linked through an oligonucleotide duplex was tested for binding to streptavidin. When one fragment was biotin, only biotin-containing duplexes were selected by streptavidin but when heated above the melting temperature, only bidentate biotin ligands were obtained. Thermal denaturation experiments showed that the melting temperature, thus stability, of the monodentate versus bidentate binding ligand increased from 59 to 71 °C in the presence of streptavidin. Substituting biotin with 2-iminobiotin led to the exclusion of all other duplexes by the bidentate iminobiotin duplex in binding streptavidin.
Marina Lovrinovic; Christof M. Niemeyer Angew. Chem. Int. Ed. Engl., 2005, 44, 3179–3183 https://doi.org/10.1002/anie.200500645AbstractDecoding technology: Besides mainstream applications in transcription profiling, DNA microarrays are currently being explored as tools for the deconvolution of synthetic libraries. Current examples include the discovery of a new type of organic reaction and the screening of peptide nucleic acid encoded inhibitor libraries to investigate new proteolytic activities from dust mite extracts (see picture).
Juan José Díaz-Mochón; Laurent Bialy; Lise Keinicke; Mark Bradley Chem. Commun., 2005, 1384-1386 https://doi.org/10.1039/B415847DAbstractEnzymatic modifications of split and mix libraries were followed by “pulling down” onto a 2-dimensional DNA microarray, via PNA tagging; this allowed complete library interrogation of all members of the split and mix library.
Philippe Gaborit; Oliver D. King Theor. Comput. Sci., 2005, 334, 99-113 https://doi.org/10.1016/j.tcs.2004.11.004AbstractIn this paper we translate in terms of coding theory constraints that are used in designing DNA codes for use in DNA computing or as bar-codes in chemical libraries. We propose new constructions for DNA codes satisfying either a reverse-complement constraint, a GC-content constraint, or both, that are derived from additive and linear codes over four-letter alphabets. We focus in particular on codes over , and we construct new DNA codes that are in many cases better (sometimes far better) than previously known codes. We provide updated tables up to length 20 that include these codes as well as new codes constructed using a combination of lexicographic techniques and stochastic search.
Kaori Sakurai; Thomas M. Snyder; David R. Liu J. Am. Chem. Soc., 2005, 127(6), 1660-1661 https://doi.org/10.1021/ja0432315AbstractDNA-templated organic synthesis (DTS) has previously been used primarily to direct coupling reactions between two DNA-linked reactants. In some cases, reactants are difficult or impossible to tether to DNA oligonucleotides. The development of strategies that enable non-DNA linked small-molecule reagents to participate in sequence-programmed synthesis therefore would significantly expand the capabilities of DTS. We developed efficient DNA-templated functional group transformations of template-linked azides into corresponding amines, carboxylic acids, and thiols. The application of these reactions to a single-solution mixture of four template-linked organic azides enabled each azide to be transformed sequence specifically into a sulfonamide, carbamate, urea, or thiourea using small-molecule sulfonyl chloride, chloroformate, isocyanate, or isothiocyanate reagents not tethered to DNA. Only the four desired products were observed, without formation of any of the 12 possible undesired cross-products. Our results represent a new approach to small molecule diversification in a DNA-programmed manner.
Nicolas Winssinger; Robert Damoiseaux; David C. Tully; Bernhard H. Geierstanger; Keith Burdick; Jennifer L. Harris Chem. Biol., 2004, 11(10),1351-60 https://doi.org/10.1016/j.chembiol.2004.07.015AbstractOur current understanding of the role and regulation of protease activity in normal and pathogenic processes is limited by our ability to measure and deconvolute their enzymatic activity. To address this limitation, an approach was developed that utilizes rhodamine-based fluorogenic substrates encoded with PNA tags. The PNA tags address each of the substrates to a predefined location on an oligonucleotide microarray through hybridization, thus allowing the deconvolution of multiple signals from a solution. A library of 192 protease substrates was prepared by split and mix combinatorial synthesis. The methodology and validation of this approach for profiling proteolytic activity from single proteases and from those in crude cell lysates as well as clinical blood samples is described.
Jennifer Harris; Daniel E. Mason; Jun Li; Keith W. Burdick; Bradley J. Backes; Teresa Chen; Aaron Shipway; Gino Van Heeke; Lucy Gough; Amir Ghaemmaghami; Farouk Shakib; François Debaene; Nicolas Winssinger Chem. Biol., 2004, 11(10), 1361-72 https://doi.org/10.1016/j.chembiol.2004.08.008AbstractEnzymatic activity in the fecal droppings from the house dust mite has been postulated to contribute to the elicited allergic response. Screening dust mite extracts through 137,180 tetrapeptide fluorogenic substrates allowed for the characterization of proteolytic substrate specificity from the potential cysteine and serine proteases in the extract. The extract was further screened against a 4000 member peptide nucleic acid (PNA) encoded inhibitor library designed to target cysteine proteases using microarray detection. Affinity chromatography coupled with mass spectrometry identified Der p 1 as one of the proteases targeted by the PNA inhibitors in the dust mite lysate. A phenotypic readout of Der p 1 function in allergy progression was demonstrated by the inhibition of CD25 cleavage from T cells by dust mite extract that had been treated with the Der p 1 inhibitor identified from the PNA-encoded inhibitor library
François Debaene; Lorenzo Mejias; Jennifer L. Harris; Nicolas Winssinger Tetrahedron, 2004, 60, 8677-8690 https://doi.org/10.1016/j.tet.2004.05.107AbstractPeptide nucleic acids (PNAs) have been used to encode a combinatorial library whereby each compound is labeled with a PNA tag which reflects its synthetic history and localizes the compound upon hybridization to an oligonucleotide array. We report herein the full synthetic details for a 4000 member PNA-encoded library targeted towards cysteine protease
Zev J. Gartner; Brian N. Tse; Rozalina Grubina; Jeffrey B. Doyon; Thomas M. Snyder; David R. Liu Science, 2004, 305(5690), 1601-5 https://doi.org/10.1126/science.1102629AbstractThe translation of nucleic acid libraries into corresponding synthetic compounds would enable selection and amplification principles to be applied to man-made molecules. We used multistep DNA-templated organic synthesis to translate libraries of DNA sequences, each containing three "codons," into libraries of sequence-programmed synthetic small-molecule macrocycles. The resulting DNA-macrocycle conjugates were subjected to in vitro selections for protein affinity. The identity of a single macrocycle possessing known target protein affinity was inferred through the sequence of the amplified DNA template surviving the selection. This work represents the translation, selection, and amplification of libraries of nucleic acids encoding synthetic small molecules rather than biological macromolecules.
Xiaoyu Li; David R. Liu Angew. Chem. Int. Ed. Engl., 2004, 43(37), 4848-70 https://doi.org/10.1002/anie.200400656AbstractIn contrast to the approach commonly taken by chemists, nature controls chemical reactivity by modulating the effective molarity of highly dilute reactants through macromolecule-templated synthesis. Nature's approach enables complex mixtures in a single solution to react with efficiencies and selectivities that cannot be achieved in conventional laboratory synthesis. DNA-templated organic synthesis (DTS) is emerging as a surprisingly general way to control the reactivity of synthetic molecules by using nature's effective-molarity-based approach. Recent developments have expanded the scope and capabilities of DTS from its origins as a model of prebiotic nucleic acid replication to its current ability to translate DNA sequences into complex small-molecule and polymer products of multistep organic synthesis. An understanding of fundamental principles underlying DTS has played an important role in these developments. Early applications of DTS include nucleic acid sensing, small-molecule discovery, and reaction discovery with the help of translation, selection, and amplification methods previously available only to biological molecules.
Matthew W. Kanan; Mary M. Rozenman; Kaori Sakurai; Thomas M. Snyder; David R. Liu Nature, 2004, 431(7008), 545-9 https://doi.org/10.1038/nature02920AbstractCurrent approaches to reaction discovery focus on one particular transformation. Typically, researchers choose substrates based on their predicted ability to serve as precursors for the target structure, then evaluate reaction conditions for their ability to effect product formation. This approach is ideal for addressing specific reactivity problems, but its focused nature might leave many areas of chemical reactivity unexplored. Here we report a reaction discovery approach that uses DNA-templated organic synthesis and in vitro selection to simultaneously evaluate many combinations of different substrates for bond-forming reactions in a single solution. Watson-Crick base pairing controls the effective molarities of substrates tethered to DNA strands; bond-forming substrate combinations are then revealed using in vitro selection for bond formation, PCR amplification and DNA microarray analysis. Using this approach, we discovered an efficient and mild carbon-carbon bond-forming reaction that generates an enone from an alkyne and alkene using an inorganic palladium catalyst. Although this approach is restricted to conditions and catalysts that are at least partially compatible with DNA, we expect that its versatility and efficiency will enable the discovery of additional reactions between a wide range of substrates.
David R. Liu PLoS Biol., 2004, 2(7), E223 https://doi.org/10.1371/journal.pbio.0020223AbstractWhile significant remaining challenges face efforts to develop and apply chemical translation, the promise of marrying evolution and organic synthesis is an irresistible combination for some researchers. The work of Harbury and co-workers described in this issue represents the latest approach to the very ancient problem of translating replicable information into functional structures.
David R. Halpin; Pehr B. Harbury PLoS Biol., 2004, 2, E173 https://doi.org/10.1371/journal.pbio.0020173AbstractRecently reported technologies for DNA-directed organic synthesis and for DNA computing rely on routing DNA populations through complex networks. The reduction of these ideas to practice has been limited by a lack of practical experimental tools. Here we describe a modular design for DNA routing genes, and routing machinery made from oligonucleotides and commercially available chromatography resins. The routing machinery partitions nanomole quantities of DNA into physically distinct subpools based on sequence. Partitioning steps can be iterated indefinitely, with worst-case yields of 85% per step. These techniques facilitate DNA-programmed chemical synthesis, and thus enable a materials biology that could revolutionize drug discovery
David R. Halpin; Juanghae A Lee; S. Jarrett Wrenn; Pehr B. Harbury PLoS Biol., 2004, 2, E175 https://doi.org/10.1371/journal.pbio.0020175AbstractDNA-directed synthesis represents a powerful new tool for molecular discovery. Its ultimate utility, however, hinges upon the diversity of chemical reactions that can be executed in the presence of unprotected DNA. We present a solid-phase reaction format that makes possible the use of standard organic reaction conditions and common reagents to facilitate chemical transformations on unprotected DNA supports. We demonstrate the feasibility of this strategy by comprehensively adapting solid-phase 9-fluorenylmethyoxycarbonyl–based peptide synthesis to be DNA-compatible, and we describe a set of tools for the adaptation of other chemistries. Efficient peptide coupling to DNA was observed for all 33 amino acids tested, and polypeptides as long as 12 amino acids were synthesized on DNA supports. Beyond the direct implications for synthesis of peptide–DNA conjugates, the methods described offer a general strategy for organic synthesis on unprotected DNA. Their employment can facilitate the generation of chemically diverse DNA-encoded molecular populations amenable to in vitro evolution and genetic manipulation.
David R. Halpin; Pehr B. Harbury PLoS Biol., 2004, 2, E174 https://doi.org/10.1371/journal.pbio.0020174AbstractBiological in vitro selection techniques, such as RNA aptamer methods and mRNA display, have proven to be powerful approaches for engineering molecules with novel functions. These techniques are based on iterative amplification of biopolymer libraries, interposed by selection for a desired functional property. Rare, promising compounds are enriched over multiple generations of a constantly replicating molecular population, and subsequently identified. The restriction of such methods to DNA, RNA, and polypeptides precludes their use for small-molecule discovery. To overcome this limitation, we have directed the synthesis of combinatorial chemistry libraries with DNA "genes," making possible iterative amplification of a nonbiological molecular species. By differential hybridization during the course of a traditional split-and-pool combinatorial synthesis, the DNA sequence of each gene is read out and translated into a unique small-molecule structure. This "chemical translation" provides practical access to synthetic compound populations 1 million-fold more complex than state-of-the-art combinatorial libraries. We carried out an in vitro selection experiment (iterated chemical translation, selection, and amplification) on a library of 10(6) nonnatural peptides. The library converged over three generations to a high-affinity protein ligand. The ability to genetically encode diverse classes of synthetic transformations enables the in vitro selection and potential evolution of an essentially limitless collection of compound families, opening new avenues to drug discovery, catalyst design, and the development of a materials science "biology."
Samu Melkko; Jörg Scheuermann; Christoph E Dumelin; Dario Neri Nat. Biotechnol., 2004, 22, 568-574 https://doi.org/10.1038/nbt961AbstractThe isolation of molecules capable of high-affinity and specific binding to biological targets is a central problem in chemistry, biology and pharmaceutical sciences. Here we describe the use of encoded self-assembling chemical (ESAC) libraries for the facile identification of molecules that bind macromolecular targets. ESAC technology uses libraries of organic molecules linked to individual oligonucleotides that mediate the self-assembly of the library and provide a code associated with each organic molecule. After panning ESAC libraries on the biomolecular target of interest, the 'binding code' of the selected compounds can be 'decoded' by a number of experimental techniques (e.g., hybridization on oligonucleotide microarrays). The potential of this technology was demonstrated by the affinity maturation (>40-fold) of binding molecules to human serum albumin and bovine carbonic anhydrase, leading to binders with dissociation constants in the nanomolar range.
Xiaoyu Li; Zev J. Gartner; Brian N. Tse; David R. Liu J. Am. Chem. Soc., 2004, 126(16), 5090-5092 https://doi.org/10.1021/ja049666%2BAbstractThe translation of DNA into synthetic molecules enables their manipulation by powerful evolution-based methods previously available only to proteins and nucleic acids. The development of increasingly sophisticated DNA-templated small-molecule syntheses is crucial to broadening the scope of this approach. Here, we report the translation of DNA templates into monocyclic and bicyclic N-acyloxazolidines using multistep DNA-templated organic synthesis. Second-generation template architectures, used for the first time in a multistep DNA-templated synthesis, together with reactions and linker cleavage strategies not previously described in a DNA-templated format, were crucial to the successful translation. The products generated in this work represent the most complex small molecules to date synthesized in a DNA sequence-programmed manner and provide the basis for DNA-templated synthetic heterocycle libraries.
Daniel M. Rosenbaum; David R. Liu J. Am. Chem. Soc., 2003, 125(46),13924-13925 https://doi.org/10.1021/ja038058bAbstractOn the basis of the distance-dependence of DNA-templated reductive amination reactions and of recent findings of D. Lynn and co-workers, we developed DNA-templated polymerizations of synthetic peptide nucleic acid (PNA) aldehydes. The coupling reactions proceed in a highly efficient and sequence-specific manner, even in the presence of mixtures of PNA aldehydes of different sequence. Synthetic peptide nucleic acid polymers containing as many as 40 PNA units (representing 10 consecutive coupling reactions) were formed efficiently. The ease of preparing PNAs containing tailor-made functional groups together with these findings raises the possibility of evolving synthetic sequence-defined polymers by iterated cycles of translation, selection, PCR amplification, and diversification previously available only to biological macromolecules.
Jeffrey B. Doyon; Thomas M. Snyder; David R. Liu J. Am. Chem. Soc., 2003, 125(41), 12372-12373 https://doi.org/10.1021/ja036065uAbstractWe have developed in vitro selections for DNA-linked synthetic small molecules with protein binding affinity and specificity. These selections require only generally accessible equipment, offer high degrees of enrichment of active molecules from mixtures of predominantly inactive species, can be applied to a variety of unrelated proteins, and require approximately 10^8-fold less material than existing synthetic molecule screening methods. Iterating these selections multiplies the net enrichment of active molecules, enabling enormous overall enrichment factors exceeding 10^6 to be achieved. Further, the selections can be adapted to select for binding specificity in addition to binding affinity. The application of methods described in this work may play a key role in the discovery of desired molecules from DNA-templated synthetic libraries.
Xiaoyu Li; David R. Liu J. Am. Chem. Soc., 2003, 125(34),10188-10189 https://doi.org/10.1021/ja035379eAbstractDNA-templated synthesis is a surprisingly general strategy for controlling chemical reactivity that enables synthetic products to be manipulated in ways previously available only to biological macromolecules. The chiral nature of the DNA template raises the possibility that DNA-templated synthesis can proceed stereoselectively. Here, we show that DNA-templated substitution reactions can exhibit stereoselectivity without the assistance of chiral groups other than those present in DNA. By characterizing changes in stereoselectivity as a result of systematic changes in the structure of the template−reagent complexes, we begin to reveal the origins of the observed stereoselectivity. We propose that the conformations of nucleotides adjacent to the reactants are largely responsible for stereoselectivity. Indeed, template and reagent sequences that can adopt either a left-handed Z-form DNA helix or a normal right-handed B-form DNA helix generate opposite stereoselectivities in the Z-form and B-form even though they share the same covalent structure and the same absolute stereochemistry. Our findings establish ways in which the chirality of an information carrier can be transmitted to the stereochemistry of encoded products through templated synthesis.
Zev J. Gartner; Rozalina Grubina; Christopher T. Calderone; David R. Liu Angew. Chem. Int. Ed. Engl., 2003, 42(12), 1370-5 https://doi.org/10.1002/anie.200390351AbstractReactivity can be rationally manipulated in DNA-templated synthesis by altering the design of template architectures: a) The omega (Ω) architecture enables DNA-templated reactions to be encoded by bases distal from the reactive end of the template, regardless of the distance dependence of the reaction. b) The T architecture enables two DNA-templated reactions to take place on a single template in one solution, as illustrated.
M. Gallo; J. M. Montserrat; A. M. Iribarren Braz. J. Med. Biol. Res., 2003, 36(2), 143-151 http://dx.doi.org/10.1590/S0100-879X2003000200001AbstractOligonucleotides have a wide range of applications in fields such as biotechnology, molecular biology, diagnosis and therapy. However, the spectrum of uses can be broadened by introducing chemical modifications into their structures. The most prolific field in the search for new oligonucleotide analogs is the antisense strategy, where chemical modifications confer appropriate characteristics such as hybridization, resistance to nucleases, cellular uptake, selectivity and, basically, good pharmacokinetic and pharmacodynamic properties. Combinatorial technology is another research area where oligonucleotides and their analogs are extensively employed. Aptamers, new catalytic ribozymes and deoxyribozymes are RNA or DNA molecules individualized from a randomly synthesized library on the basis of a particular property. They are identified by repeated cycles of selection and amplification, using PCR technologies. Modified nucleotides can be introduced either during the amplification procedure or after selection.
Dan C. Tulpan; Holger H. Hoos; Anne E. Condon DNA Computing., 2002, 229-241 https://doi.org/10.1007/3-540-36440-4_20AbstractWe present results on the performance of a stochastic local search algorithm for the design of DNA codes, namely sets of equallength words over the nucleotides alphabet A,C,G, T that satisfy certain combinatorial constraints. Using empirical analysis of the algorithm, we gain insight on goodd esign principles. We report several cases in which our algorithm finds word sets that match or exceed the best previously known constructions.
Christopher T. Calderone; James W. Puckett; Zev J. Gartner; David R. Liu Angew. Chem. Int. Ed. Engl., 2002, 41(21), 4104-8 https://www.ncbi.nlm.nih.gov/pubmed/12412096AbstractDiversification without spatial separation can be achieved using DNA-templated synthesis. Twelve reactants with functional groups of similar reactivity linked to different DNA sequences (R, R′) were subjected to several different reaction types within the same solution and only six sequence-programmed products out of the more than 28 possibilities were formed (see scheme).
Zev J. Gartner; Matthew W. Kanan; David R. Liu J. Am. Chem. Soc., 2002, 124(35), 10304-6 https://doi.org/10.1021/ja027307dAbstractThe translation of DNA sequences into synthetic products is a key requirement of our approach to evolving synthetic molecules through iterated cycles of translation, selection, and amplification. Here we report general linker and purification strategies for sequence-specific DNA-templated synthesis that collectively enable the product of a DNA-templated reaction to be isolated and to undergo subsequent DNA-templated reactions. Using these strategies, we have achieved the first multistep nucleic acid-templated small-molecule syntheses to generate two different molecules. In addition to representing a method for translating DNA templates sequence-specifically into corresponding multistep synthetic products, our findings also provide experimental support for previously proposed models invoking multistep nucleic acid-templated synthesis as mediating the prebiotic translation of replicable information into the earliest functional molecules.
Arwen Brenneman; Anne Condon Theor. Comput. Sci., 2002, 287, 1, 39-58 https://doi.org/10.1016/S0304-3975(02)00135-4AbstractThe design of DNA or RNA strands for DNA computations poses many new questions in algorithms and coding theory. DNA strand design also arises in use of molecular bar codes to manipulate and identify individual molecules in complex chemical libraries, and to attach molecules to DNA chips. We survey several formulations of the DNA strand design problem, along with results and open questions in this area.
Nicolas Winssinger; Scott Ficarro; Peter G. Schultz; Jennifer L. Harris Proc. Natl. Acad. Sci. USA, 2002, 99(17), 11139-44 https://doi.org/10.1073/pnas.172286899AbstractThe regulation of protein function through posttranslational modification, local environment, and protein-protein interaction is critical to cellular function. The ability to analyze on a genome-wide scale protein functional activity rather than changes in protein abundance or structure would provide important new insights into complex biological processes. Herein, we report the application of a spatially addressable small molecule microarray to an activity-based profile of proteases in crude cell lysates. The potential of this small molecule-based profiling technology is demonstrated by the detection of caspase activation upon induction of apoptosis, characterization of the activated caspase, and inhibition of the caspase-executed apoptotic phenotype using the small molecule inhibitor identified in the microarray-based profile.
Zev J. Gartner; Matthew W. Kanan; David R. Liu Angew. Chem. Int. Ed. Engl., 2002, 41(10), 1796-800 https://www.ncbi.nlm.nih.gov/pubmed/19750721AbstractPowerful reactions such as Wittig olefinations, nitro-aldol additions, dipolar cycloadditions, and Heck coupling reactions can be mediated by DNA templates. The yields of several DNA-templated reaction products are independent of the number of bases (n=0 or 10) separating the annealed reactive groups (as an example, the denaturing polyacrylamide gel electrophoresis of a DNA-templated Wittig reaction is shown).
Ming Li; Hye Jin Lee; Anne E. Condon; Robert M. Corn Langmuir, 2002, 18, 3, 805-812 https://doi.org/10.1021/la0112209AbstractA template−map design strategy for generating sets of non-interacting DNA oligonucleotides for applications in DNA arrays and biosensors is demonstrated. This strategy is used to create a set of oligonucleotides of size s with length l that possess at least n base mismatches with the complements of all the other members in the set. These “DNA word” sets are denoted as nbm l-mers or l:n sets. To regularize the thermodynamic stability of the perfectly matched hybridized DNA duplexes, the l-mers chosen for all the sets are required to have an approximately 50% G/C content. To achieve good discrimination between each DNA word in each set generated using the template−map strategy, it is required that n should be approximately equal to l/2 or higher. The template−map strategy can be used in a straightforward manner to create DNA word sets for cases when l = 4k and n = 2k, where k is an integer. Specific examples of 4k:2k sets are designed: an 8:4 set (s = 224), a 12:6 set (s = 528), a 16:8 set (s = 960), and a 20:10 set (s = 1520). These sets are further optimized to achieve the narrowest possible distribution of melting temperatures by selecting the best set after permutation of the templates and maps over all possible configurations. To demonstrate the viability of this methodology, a non-interacting set of four specific 6bm 12mers have been chosen, synthesized, and used in an SPR imaging measurement of the hybridization adsorption onto a DNA array. The template−map strategy is also applied to generate DNA word sets for cases where l ≠ 4k. In these cases, the creation of the maps and templates is more complicated, but possible. The templates and maps for three additional types of sets are created: (4k − 1):(2k − 1), (4k + 1):2k, and (4k − 2):(2k − 1). Specific examples are given for l = 7, 9, and 10: DNA word sets of 7:3 (s = 224), 9:4 (s = 360), and 10:5 (s = 132).
Joelle Pelletier; Sachdev Sidhu Curr. Opin. Biotechnol., 2001, 12, 340-347 https://doi.org/10.1016/S0958-1669(00)00225-1AbstractExtremely diverse, DNA-encoded libraries of peptides and proteins have been constructed that include a linkage between each polypeptide and the encoding DNA. Library members can be selected by virtue of a particular binding specificity, and their protein sequence can be deduced from the sequence of the cognate DNA. Such combinatorial biology methods have proven invaluable in both identifying natural protein-protein interactions and also in mapping the specificities and energetics of these interactions in fine detail.
Nicolas Winssinger; Jennifer L. Harris; Bradley J. Backes; Peter G. Schultz Angew. Chem. Int. Ed. Engl., 2001, 40(17), 3152-3155 https://www.ncbi.nlm.nih.gov/pubmed/?term=From+Split%E2%80%90Pool+Libraries+to+Spatially+Addressable+Microarrays+and+Its+Application+to+Functional+Proteomic+ProfilingAbstractSmall molecules encoded with peptidonucleic acid (PNA) were used to probe protein function in a microarray format. The PNA tag served to encode the synthetic history of the small molecule and to positionally encode the identity of the small molecule by its location upon hybridization to an oligonucleotide microarray.
Amit Marathe; Anne E. Condon; Robert M. Corn J. Comput. Biol., 2004, 8, 3, 201-219 https://doi.org/10.1089/10665270152530818AbstractWe consider the problem of designing DNA codes, namely sets of equi-length words over the alphabet {A, C, G, T} that satisfy certain combinatorial constraints. This problem is motivated by the task of reliably storing and retrieving information in synthetic DNA strands for use in DNA computing or as molecular bar codes in chemical libraries. The primary constraints that we consider, defined with respect to a parameter d, are as follows: for every pair of words w, x in a code, there are at least d mismatches between w and x if w ≠ x and also between the reverse of w and the Watson-Crick complement of x. Extending classical results from coding theory, we present several upper and lower bounds on the maximum size of such DNA codes and give methods for constructing such codes. An additional constraint that is relevant to the design of DNA codes is that the free energies and enthalpies of the code words, and thus the melting temperatures, be similar. We describe dynamic programming algorithms that can (a) calculate the total number of words of length n whose free energy value, as approximated by a formula of Breslauer et al. (1986) falls in a given range, and (b) output a random such word. These algorithms are intended for use in heuristic algorithms for constructing DNA codes.
Zev J. Gartner; David R. Liu J. Am. Chem. Soc., 2001, 123, 6961-6963 https://doi.org/10.1021/ja015873nAbstractDespite their limited chemical functionality, proteins and nucleic acids dominate the solutions to many complex chemical problems because they can be evolved through iterated cycles of diversification, selection, and amplification. Researchers have demonstrated extensively that proteins and nucleic acids initially lacking desired activities can be mutated, amplified, and reselected to afford evolved molecules with greatly enhanced properties. We are interested in creating amplifiable and evolvable libraries of non-natural small molecules by developing methods to translate DNA into synthetic structures. Achieving this goal requires using DNA to direct chemical reactions sequence-specifically in a manner much more general than has been reported thus far. Researchers have previously demonstrated the ability of nucleic acid templates to promote the coupling of adjacently annealed oligonucleotides to form nucleic acids and nucleic acid analogues. We hypothesized that the proximity effect provided by DNA-templated synthesis can be used to generate libraries of synthetic small molecules unrelated in structure to the DNA backbone in one-pot, parallel reactions.
Fabrice Guillier; David Orain; Mark Bradley Chem. Rev., 2000, 100, 2091−2157 https://doi.org/10.1021/cr980040+AbstractThe massive increase in the number of papers describing the use of polymeric supports in organic synthesis over the past decade is a vivid demonstration of its impact in the chemical community. Few other changes in synthetic chemistry methodology have displayed such a growing passion or had such a profound influence on the way synthetic chemistry is carried out. The advantages gained by this methodology are striking, with four main factors contributing to the popularity of the technique. (i) The ease of chemistry. Reactions can be accomplished in only three steps: addition of reagents, filtering, and washing the resin, thus allowing many simple automated procedures to be developed. (ii) The elimination of purification steps en route. For each step of a multiple-step synthesis, the only purification needed is a resin-washing step. Only the final product of cleavage needs to be purified. (iii) In a solid-phase synthesis, high concentrations of reagents can be used to drive reactions to completion. (iv) The straightforward nature of parallel solid-phase synthesis.
Michael McPherson; Martin C. Wright; Peter A. Lohse Synlett, 1999, S1, 978-980 https://doi.org/10.1055/s-1999-3107AbstractA convergent strategy for the synthesis of an RNA-peptide conjugate is presented. Regioselective ligation between a 5′-modified RNA harboring a 5′-thioester group and a peptide carrying an N-terminal cysteine afforded an RNA-peptide conjugate under physiological conditions without the need for protecting groups.
Michal Lebl J. Comb. Chem., 1999, 1, 1, 3-24 https://doi.org/10.1021/cc9800327AbstractIt is appropriate to start this new journal with an article about the history of this particular fieldsespecially when the field is so new that even the basic terminology is not completely defined.† When I was asked to write an article about the history of combinatorial chemistry, I believed that it would not be too difficult since I was engaged in this exciting new technology from the start of the first combinatorial chemistry company, Selectide Corporation in Tucson. To make this historical retrospective more objective, I decided to consult with several scientists who published in this field at the early stages and whose articles may be considered as having formed the new way of thinking about the techniques used to develop new drugs and catalysts and an unlimited variety of materials. After reading the responses of my colleagues, it was clear that when we were building Selectide in 1991, there were scientists already heavily engaged in creating and using diversity in various forms. Some people were exploring this new field without even realizing it. I can document this fact from my own experience. In 1988, we published a paper with Giorgio Fassina and Irwin Chaiken1 in which we synthesized a mixture of analogues of oxytocin by coupling a mixture of amino acids in one step of the synthesis and applying this mixture to the column with immobilized neurophysin. The analogues with an affinity toward neurophysin were retained on the column, and after elution their structures were analyzed. None of us realized the potential of this technique for the development of new drugs; at that time we were “entrenched” in the approach of making one compound at a time, analyzing it, and evaluating it biologically. However, even in the 1990s the world was not ready to accept the idea of building libraries of organic molecules and screening them to find interesting compounds. It took another several years before an academic course of combinatorial chemistry was offered to students at The University of Louisville in 1996 by Professor Arno Spatola.
Mark F. Harper; Christopher G. Newton Expert Opin. Ther. Patents, 1999, 9, 5, 583-624 https://doi.org/10.1517/13543776.9.5.583AbstractPatent applications published June 1997 - November 1998 are reviewed to see how trends are evolving with time. Patents are divided into three subtypes: enabling chemistries, equipment and technologies, and therapeutic inventions using libraries. More than 170 patents reviewed testify to this explosive area. Nonetheless, many patents may require their claims amending before granting, and may also be very difficult to defend. Attention is drawn to articles from patent attorneys discussing the difficulties in prosecuting patents in this technology.
Jean-Luc Fauchère; Jean A. Boutin; Jean-Michel Henlin; Nathalie Kucharczyk; Jean-Claude Ortuno Chemometr. Intell. Lab. Syst., 1998, 43, 43-68 https://doi.org/10.1016/S0169-7439(98)00082-3AbstractThe principles of the combinatorial synthesis of large compound libraries are described. It is shown that these technologies can afford unprecedented numbers of chemical entities thus increasing the molecular diversity of the universe of organic compounds. Library design is considered in view of the generation of a maximized diversity for a given set of synthetic products. The advantages of solid phase organic chemistry are stressed and compared to the achievements of solution procedures for the production of large compound collections. The trend from the truly combinatorial synthesis of millions of biooligomers towards the parallel synthesis of individual small organics is stated and exemplified. Finally, an overview is given of the methods developed for the structural identification of the active component(s) in compound libraries and arrays, including iterative deconvolution via re-synthesis, positional scanning or structural analysis of the ligand on a single polymer bead. Strategies are suggested for both lead discovery and lead optimization.
Anthony W. Czarnik Curr. Opin. Chem. Biol., 1997, 1, 60-66 https://doi.org/10.1016/S1367-5931(97)80109-3AbstractAssuming that the chemical reactions used to synthesize a combinatorial library member are successful, then knowledge of the specific reaction sequence is equivalent to knowing the member's chemical identity. Because the determination of chemical identity is typically not automatable and requires a substantial amount of material, schemes that encode a member's reaction history onto the reaction platform are of value. The primary benefits of encoding are relational nomenclature (all methods) and automated handling (some methods). Encoding methods evaluated to date are spatial, graphical, chemical, spectrometric, electronic, and physical.
Derek Maclean; John R. Schullek; Martin M. Murphy; Zhi-Jie Ni; Eric M. Gordon; Mark A. Gallop Proc. Natl. Acad. Sci. USA, 1997, 94(7), 2805-10 https://doi.org/10.1073/pnas.94.7.2805AbstractThe application of a new encoding technology for drug discovery is described. A combinatorial library of mercaptoacyl pyrrolidines has been prepared on a beaded polymeric support. Each polymer bead carries one library constituent in association with an oligomeric "tag," the structure of which is a record of the specific reagents from which that library member was prepared. After the ligands were solubilized, an array of such beads was screened for angiotensin-converting enzyme inhibitory activity, and the structures of active pyrrolidines were deduced by analysis of the associated tags at sub-picomole levels. Several extremely potent enzyme inhibitors were identified, many from multiple beads. The most potent inhibitor was found to have a Ki of 160 pM, approximately 3-fold more active than captopril in the same assay. Direct comparison with iterative deconvolution shows that the encoded screening strategy is a much more efficient means for extracting information from such compound collections, producing more data on a larger number of active structures.
X. Williard; I. Pop; L. Bourel; D. Horvath; R. Baudelle; P. Melnyk; B. Déprez; A. Tartar Eur. J. Org. Chem., 1996, 31, 2, 87-98 https://doi.org/10.1016/0223-5234(96)80441-7AbstractAlthough knowledge-based de novo design of high affinity ligands for receptors or enzymes appears to be the most rational approach to the discovery of new pharmacologically active substances, high throughput random screening of compounds with no known structural similarity to the natural ligand has proved to be a highly efficient method. In order to increase the number of new molecules, methods of combinatorial synthesis have been considerably developed in recent years. In this review we will examine some of the main strategies for generating large arrays of diverse molecular entities, the screening of such libraries, the methods used to identify an active compound, and the chemical technologies used in the development of this very promising source of new drugs.
Magda Stanková; Michal Lebl Mol. Divers., 1996, 2, 75-80 https://doi.org/10.1007/BF01718703AbstractThe decreasing reactivity of tri-, di- and monochlorotriazine was utilized for the solid-phase construction of a combinatorial library with three randomized positions, using 20 amino acids and 50 amines as building blocks. The first chlorine atom was selectively substituted by coupling a large excess of trichlorotriazine to the support-bound amino acid, thus avoiding simultaneous substitution of the second chlorine. The second and third diversity positions were selectively introduced by coupling amines at different temperatures. Mixtures of model compounds were synthesized and analyzed, showing the correct representation of all expected components. A library composed of 12 000 compounds was generated using this method.
Richard K. Bruick; Philip E. Dawson; Stephen B. H. Kent; Nassim Usman; Gerald F. Joyce Chem. Biol., 1996, 3, 1, 49-56 https://doi.org/10.1016/S1074-5521(96)90084-8AbstractBackground: Oligonucleotide-peptide conjugates have several applications, including their potential use as therapeutic agents. We developed a strategy for the chemical ligation of unprotected peptides to oligonucleotides in aqueous solution. The two compounds are joined via a stable amide bond in a template-directed reaction. Results: Peptides, ending in a carboxy-terminal thioester, were converted to thioester-linked oligonucleotide-peptide intermediates. The oligonucleotide portion of the intermediate binds to a complementary oligonucleotide template, placing the peptide in close proximity to an adjacent template-bound oligonucleotide that terminates in a 3′ amine. The ensuing reaction results in the efficient formation of an amide-linked oligonucleotide-peptide conjugate. Conclusions: An oligonucleotide template can be used to direct the ligation of peptides to oligonucleotides via a highly stable amide linkage. The ligation reaction is sequence-specific, allowing the simultaneous ligation of multiple oligonucleotide-peptide pairs.
Gordon Lowe Chem. Soc. Rev., 1995, 24, 309-317 https://doi.org/10.1039/CS9952400309AbstractCombinatorial chemistry is a novel and innovative way of rapidly generating a large number of related compounds. The solid phase method of synthesis, first introduced by Merrifield for peptide synthesis some 30 years ago, is generally used for preparing combinatorial chemical libraries. The solid phase method has many advantages over solution phase chemistry, most notably that excess reagents may be used to drive reactions rapidly to completion and then by-products and excess reagents removed by a simple washing procedure. The whole process is capable of automation and instrumentation is commercially available for this purpose. Although the solid phase method of synthesis initially received much criticism, primarily because intermediates are not characterized, as reagents and protecting groups for peptide synthesis improved and the purification of peptides by HPLC became routine, the method was accepted as the preferred method of peptide synthesis. The solid phase method has also been successfully developed for the synthesis of oligonucleotides which are now routinely used in molecular biology.
Yasunori Kinoshita; Koichi Nishigaki Nucleic Acids Symp. Ser., 1995, 34, 201-202 https://www.ncbi.nlm.nih.gov/pubmed/8841622AbstractA synthesis method of oligonucleotides is proposed for generating the code regions for ECC (Encoded Combinatorial Chemistry). It exploits T4 RNA ligase with the use of ribonucleotide-containing-DNAs (RCDs). Preliminary success to ligate four fragments by this method is reported.
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