Molecular Basis for Antiviral Action of EDP‐235: A Potent and Selective SARS‐CoV‐2 3CLpro Inhibitor for the Treatment of Covid 19

Abstract

To date, there are no approved oral antiviral therapies that can be administered early in the course of COVID‐19 to suppress progression of the disease or for prophylaxis. EDP‐235 is a potent and selective inhibitor of SARS‐CoV‐2 3C‐like protease (3CLpro). EDP‐235 inhibits SARS‐CoV‐2 3CLpro protease activity with an IC50 of 5.8 ± 3.7 nM and retains its activity against variant 3CLpro proteins from multiple SARS‐CoV‐2 lineages (IC50range of 2.8­–5.8 nM). 3CLpro protease activity progress curves showed significant curvature in a time‐ and EDP‐235‐concentration‐dependent manner indicative of slow‐onset inhibition. Slow reversal of inhibition of SARS‐CoV‐2 3CLpro enzyme activity was observed in a jump dilution experiment. Michaelis‐Menten kinetic studies with a FRET peptide substrate in the presence of EDP‐235 indicated that EDP‐235 is a substrate‐competitive inhibitor of SARS‐CoV‐2 3CLpro with an overall dissociation constant Kiof 3.0 ± 1.6 nM. SARS‐CoV‐2 3CLpro was crystallized bound to a close analog of EDP‐235 and structure elucidation revealed that the ligand bound at the active site and interacted with side chains of conserved residues Cys‐145, His‐163, and Glu‐166. EDP‐235 also potently inhibits 3CLpro enzymes from other α‐coronaviruses (IC50range of 2–4 nM) and β‐coronaviruses (SARS‐CoV IC50 of 5.4 nM, MERS‐CoV IC50 of 70 nM) which cause disease in humans to date. EDP‐235 resistance mutations in HCoV‐229E map to the active site of 3CLpro close to the predicted binding site and offer additional support to the mechanism of inhibition. EDP‐235 also showed a favorable selectivity profile (>300 selectivity index) when tested against a panel of 30 mammalian proteases. In summary, EDP‐235 acts as a slow‐onset, slow‐reversible, substrate‐competitive inhibitor of SARS‐CoV‐2 3CLpro. The outstanding preclinical profile of EDP‐235 supports its further evaluation as an oral therapeutic for the management of COVID‐19.

On the origins of SARS-CoV-2 main protease inhibitors

“…Also of interest, the SARS-CoV-2 M pro inhibitor EDP-235 ( 85 ) from Enanta Pharmaceuticals 319 recently met the primary end point of a phase 2 clinical trial without the coadministration of ritonavir (NCT05616728). In this case, 13 patents from this company claim a very large variety of peptides sometimes featuring conformational locks related to the one depicted above.…”

Use of Antiviral Agents and other Therapies for COVID-19

“…83 Researchers at the German Center for Infection Research (DZIF) at the University of Lübeck have identified RHCDS-13b (structure not disclosed), a "second-generation" derivative derived from an ɑ-keto-amide through studies employing the main protease's crystal structure. 84 EDP-235 from Enanta 85 (structure not disclosed, IC 50 range of 2.8 to 5.8 nM against variant 3CLpro proteins from multiple SARS-CoV-2 lineage), 86 entered a phase I single and multiple ascending dose study to evaluate the safety, tolerability, and pharmacokinetics of EDP-235 in healthy volunteers in February 2022. In March 2022, the U.S. Food and Drug Administration granted Fast Track designation for EDP-235.…”

A Sustainable Synthesis of the SARS-CoV-2 Mpro Inhibitor Nirmatrelvir, the Active Ingredient in Paxlovid

“…2 Both aims have been achieved in record time, the latter initially being driven by investigations into repurposing existing drugs, which has produced relatively few effective treatment options. 3 The search for novel SARS-CoV-2 antivirals, on the other hand, has led to the development of molnupiravir by Merck, 4 EDP-235 by Enanta, 5 S-217622 by Shionogi, 6 and several others. 7 The first such antiviral to receive approval by the FDA, Paxlovid, is an orally-active combination of the SARS-CoV-2 main protease inhibitor nirmatrelvir (1; PF-07321332) and the HIV antiviral ritonavir, disclosed by Pfizer in late 2021.…”