DNA-encoded chemistry technology yields expedient access to SARS-CoV-2 M ^pro inhibitors

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has killed more than 4 million humans globally, but there is no bona fide Food and Drug Administration–approved drug-like molecule to impede the COVID-19 pandemic. The sluggish pace of traditional therapeutic discovery is poorly suited to producing targeted treatments against rapidly evolving viruses. Here, we used an affinity-based screen of 4 billion DNA-encoded molecules en masse to identify a potent class of virus-specific inhibitors of the SARS-CoV-2 main protease (Mpro) without extensive and time-consuming medicinal chemistry. CDD-1714, the initial three-building-block screening hit (molecular weight [MW] = 542.5 g/mol), was a potent inhibitor (inhibition constant [Ki] = 20 nM). CDD-1713, a smaller two-building-block analog (MW = 353.3 g/mol) of CDD-1714, is a reversible covalent inhibitor of Mpro (Ki = 45 nM) that binds in the protease pocket, has specificity over human proteases, and shows in vitro efficacy in a SARS-CoV-2 infectivity model. Subsequently, key regions of CDD-1713 that were necessary for inhibitory activity were identified and a potent (Ki = 37 nM), smaller (MW = 323.4 g/mol), and metabolically more stable analog (CDD-1976) was generated. Thus, screening of DNA-encoded chemical libraries can accelerate the discovery of efficacious drug-like inhibitors of emerging viral disease targets.

Gain-of-Signal Assays for Probing Inhibition of SARS-CoV-2 M pro /3CL pro in Living Cells

“…It is therefore notable that two nonpeptide chemotypes also tested positive. CDD-1976, identified recently in a DNA-encoded library screen ( 39 ), shows strong inhibitory activity at lower concentrations and blunted activity at higher concentrations due to cytotoxicity ( Fig. 5a ).…”

“…While some reported SARS2 M pro inhibitors have gone through rigorous characterization and structure-activity relationships have been established, other candidates have been identified through high-throughput screens in drug repurposing efforts, and on-target efficacies remain controversial ( 9 , 38 – 46 ). We therefore used our luciferase-based gain-of-signal assay to directly compare candidate inhibitors and help shed light on this important and rapidly growing area.…”

“…Compounds were purchased as powders from the commercial vendors listed in Table S1 and resuspended in DMSO to stock concentrations of 10 mM. CDD-1976 was provided by the Young lab ( 39 ). The integrity (purity and molecular weight) of all compounds that tested negative in the system described here was confirmed by HPLC ( Table S1 ).…”

Newly Emerging Strategies in Antiviral Drug Discovery: Dedicated to Prof. Dr. Erik De Clercq on Occasion of His 80th Anniversary

“…Considering the high variability of virus and the complexity of pathogenic mechanisms, DNA-encoded chemistry technology [ 39 ], genome editing technology [ 40 ], nucleic acid aptamer technology [ 41 ] and ligand discovery based on protein self-assembly [ 42 ] are expected to provide references for the development of potent antiviral drugs. Moreover, natural products have complex structures and diverse physiological activities, providing unlimited resources to discover novel antiviral drugs [ 43 , 44 ].…”

Recent advances in DNA-encoded dynamic libraries

“… 20 Very recently, researchers selected a 4-billion DEL against the SARS-CoV-2 main protease (M pro ) and obtained potent inhibitors with high potential for clinical development. 56 …”