Haipeng Yu, Xiaoran Wang, Xinyu Wang, Yizhen Yin

Chemistry - A European Journal

DOI:10.1002/chem.71680

Abstract

Covalent drugs have attracted increasing attention in modern drug discovery because of their prolonged target engagement, enhanced pharmacological efficacy, and potential to address traditionally challenging protein targets. The rapid development of electrophilic warhead chemistry, chemical biology methods, and high-throughput screening technologies has shifted covalent drug discovery from serendipitous identification to target-oriented and rational design. In recent years, phage display, mRNA display, and DNA-encoded library technologies have enabled the efficient de novo discovery of covalent peptides and small molecules, expanding the accessible chemical space for targeting cysteine, lysine, tyrosine, serine, and other reactive residues. Meanwhile, artificial intelligence has further accelerated covalent drug research by supporting covalent database construction, binding-site prediction, virtual screening, molecular generation, and structure-based ligand optimization. This article reviews recent advances in high-throughput strategies for targeted covalent drug discovery, with a focus on phage display, mRNA display, DNA-encoded libraries, and artificial intelligence (AI)-assisted approaches. We further discuss current challenges and future opportunities in the field and provide our perspectives on emerging directions for next-generation covalent ligand discovery.

Danial Yazdan, A. Michael Downey, Ana Gimeno, Caishun Li, Edward N. Schmidt, Jeffrey Y. K. Wong, Caleb Loo, Jaesoo Jung, Ryan Qiu, Ewa Lis, Lily Lindmeier, June Ereño-Orbea, Jesús Jiménez-Barbero, Matthew S. Macauley, Ratmir Derda

Journal of the American Chemical Society

DOI: 10.1021/jacs.6c14068

Abstract

Glycan-binding proteins (GBPs) are among the most difficult drug targets, limiting clinical progress against therapeutically important GBPs. We employed bicyclic genetically encoded libraries (BiGELs), produced by the chemical modification of phage-displayed peptide libraries with twofold symmetric linchpins, to discover inhibitors of therapeutically relevant Siglec-7:GD3 interactions. Next-generation sequencing (NGS) analysis of BiGEL panning against Siglec-7 yielded 815 candidates, of which 23 hits yielded K_D = 1–100 μM, as determined by surface plasmon resonance (SPR). Competitive enzyme-linked immunosorbent assays (ELISA) identified leads that disrupted the Siglec-7:GD3 interaction with IC₅₀ = 3-300 μM. Machine learning models trained on NGS datasets identified additional inhibitors with equivalent potencies. Alanine scans of 8c (SWCRPATVNC, IC₅₀ = 3.8 μM) and 12c (SFCHYPTHVC, IC₅₀ = 11 μM) identified residues crucial for activity. Ring reshaping studies of 8c highlighted the critical role of bicyclic topology, yielding analog 46e (SAAAAAWCRPATVNC, IC₅₀ = 9.5 μM), which was further evolved into 67e (STVTQHWCRPATVNC). The multivalent display of lead bicycles alongside ∼100 glycans in a Liquid Glycan Array (LiGA) enabled the comparison of binding to Siglec-7-expressing cells. LiGA assays confirmed the binding of the bicycles to Siglec-7 but revealed considerable nonspecific interactions with receptor-negative cells. Saturation transfer difference nuclear magnetic resonance (STD-NMR) spectroscopy revealed that 46e binds to Siglec-7 at a site distinct from the V-Ig domain, suggesting inhibition through an allosteric site. Together, these results demonstrate that BiGEL enables the discovery of bicyclic peptides for undruggable Siglec targets, but highlights future challenges in molecular discoveries that aim to identify small, noncarbohydrate inhibitors of GBPs.

Tianxiong Mi, Lijun Fan, Amber Hackler, Diego B. Diaz, Chang Qi, Errol L. G. Samuel, Qi Gao, Tao Meng, Xingjian Xu, Erwin G. Abucayon, Edward N. DiNunzio, Marcelo J. Murai, Natalya Pissarnitski, Jordan De Jesus Silva, Christopher Sondey, Jack D. Scott

Journal of the American Chemical Society

DOI: 10.1021/jacs.6c07049

Abstract

Rapid, aqueous macrocyclization strategies that proceed in high conversion are valuable for ultralarge macrocyclic peptide (MP) library synthesis via display and DNA-encoded library (DEL) technology. Here we report an on-DNA macrocyclization based on sulfur(VI) fluoride exchange (SuFEx) that unites above features. This approach embeds a phenol and an aryl sulfonyl fluoride within a DNA-tagged peptide to accelerate SuFEx and trigger intramolecular cyclization immediately upon dissolution in basic aqueous buffer. MPs ranging from 15 to 52 membered rings bearing diverse amino acids were synthesized efficiently. The on-DNA conditions readily translate off DNA to furnish sulfonate and sulfonamide linked MPs. NMR analyses showed SuFEx-derived biaryl linkers act as conformational tuner: sequential changes in aryl substitution and linker length shift backbone conformations from extended strands to rigid turns. Leveraging this chemistry, we designed and synthesized ultralarge MP libraries via DEL technology. DEL screening followed by off-DNA hit validation identified a potent, de novo MP inhibitor of receptor-interacting serine/threonine kinase 1 (RIPK1). Collectively, these findings establish SuFEx cyclization as a robust, DEL-compatible strategy for programmable macrocycle design and drug discovery.

Ana L. Aguirre, Amanda W. Dombrowski, Noah P. Tu, Shubhendu S. Karandikar, Ying Wang

The Journal of Organic Chemistry 

DOI:  10.1021/acs.joc.6c00134

Abstract

In this work, we present the creation and implementation of a ChemBead-enabled, on-DNA high-throughput experimentation (HTE) platform to identify more general and mild reaction conditions for challenging C–N couplings. The platform was developed by adapting our small-molecule system to address the inherent challenges of on-DNA chemistry. The results generated by the platform were compared to the standard operating procedure for DNA-encoded library (DEL) production to ensure the translation of any promising conditions. The platform was used to discover a condition that has not been previously reported for on-DNA C–N coupling. This condition is best suited for secondary amines and select primary amines on short DNA substrates; primary aliphatic amines on elongated DNA-tagged aryl halide conjugates remain a challenge. This newly identified condition can be applied to DNA-encoded library constructions

James Wellnitz , Brandon Novy , Shu-Hang Lin , Travis Maxfield , Matthew Axtman , Tina M. Leisner , Eric M. Merten , Ivanna Zhilinskaya , Devan J. Shell , Kellyn M. Schroeder , Jacqueline L. Norris-Drouin , P. Brian Hardy , Kenneth H. Pearce , Konstantin I. Popov

Journal of Cheminformatics

DOI: 10.1186/s13321-026-01296-1

Abstract

DNA-encoded library (DEL) technology has become a powerful tool in modern drug discovery. However, fully harnessing its potential requires the use of extensive computational methods, which are often available only through proprietary software. This restricts accessibility for small teams lacking robust informatics support, hindering the growth of the technology. Objective We introduce DELi, an open-source DEL informatics platform developed for library design, next-generation sequencing (NGS) read processing, and binding data analysis. Implementation DELi offers a simple and flexible configuration setup that is readily customizable, while remaining easy to use and supported by a clear user interface. To showcase its capabilities, we used DELi to design a custom benzimidazole-based DEL (UNCDEL006), and performed proof-of-concept selection experiments against bromodomain-containing protein 4 (BRD4). Results The DELi decoding and analysis modules identified top-performing compounds, leading to the off-DNA synthesis of UNC11951. This compound was confirmed as a nanomolar BRD4 binder via isothermal titration calorimetry (ITC) and demonstrated thermal shift stabilization as observed by differential scanning fluorimetry (DSF). These results demonstrate DELi as an effective tool for DEL design and analysis. Availability DELi is written in Python and made available through PyPi and GitHub, alongside the open-sourced UNCDEL006 library. Scientific contribution DELi addresses the need for accessible computational tools for DEL by integrating robust analysis methods within a cohesive, open-source platform. It provides a fully documented and experimentally validated end-to-end workflow, improving reproducibility and lowering the barrier to entry for researchers adopting DEL technology.

Yeongjoo Suh, Kyung-Jin Cho, Hyun Jin Kim, Hongjun Jeon

The Journal of Organic Chemistry

DOI: 10.1021/acs.joc.6c00675

Abstract

DNA-encoded library (DEL) synthesis necessitates robust on-DNA chemical transformations that remain dependable throughout multistep library construction, especially for elongated DNA constructs. This study assessed the efficiency of commonly used on-DNA nitro reduction reactions under conditions relevant to practical DEL synthesis to evaluate the impact of DNA tag elongation. Although iron(II)-, diboron-, palladium(II)-, and dithionite-mediated reductions showed high efficiency for short DNA constructs, their performances varied significantly with DNA tag elongation. The generality of these reduction conditions was further explored across a diverse set of nitro-containing substrates, and their chemoselectivities toward additional functional groups were assessed to reflect realistic DEL design considerations. These findings underscore the importance of evaluating on-DNA chemistry beyond minimal tag systems and offer practical guidance for selecting suitable nitro reduction conditions for reliable DEL construction.