Identification of Inhibitors of SARS-CoV-2 3CL-Pro Enzymatic Activity Using a Small Molecule in Vitro Repurposing Screen

Maria Kuzikov;Elisa Costanzi;Jeanette Reinshagen;Francesca Esposito;Laura Vangeel;Markus Wolf;Bernhard Ellinger;Carsten Claussen;Gerd Geisslinger;Angela Corona;Daniela Iaconis;Carmine Talarico;Candida Manelfi;Rolando Cannalire;Giulia Rossetti;Jonas Gossen;Simone Albani;Francesco Musiani;Katja Herzog;Yang Ye;Barbara Giabbai;Nicola Demitri;Dirk Jochmans;Steven De Jonghe;Jasper Rymenants;Vincenzo Summa;Enzo Tramontano;Andrea R. Beccari;Pieter Leyssen;Paola Storici;Johan Neyts;Philip Gribbon;Andrea Zaliani

2021-01-01

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Abstract

Compound repurposing is an important strategy for the identification of effective treatment options against SARS-CoV-2 infection and COVID-19 disease. In this regard, SARS-CoV-2 main protease (3CL-Pro), also termed M-Pro, is an attractive drug target as it plays a central role in viral replication by processing the viral polyproteins pp1a and pp1ab at multiple distinct cleavage sites. We here report the results of a repurposing program involving 8.7 K compounds containing marketed drugs, clinical and preclinical candidates, and small molecules regarded as safe in humans. We confirmed previously reported inhibitors of 3CL-Pro and have identified 62 additional compounds with IC50 values below 1 μM and profiled their selectivity toward chymotrypsin and 3CL-Pro from the Middle East respiratory syndrome virus. A subset of eight inhibitors showed anticytopathic effect in a Vero-E6 cell line, and the compounds thioguanosine and MG-132 were analyzed for their predicted binding characteristics to SARS-CoV-2 3CL-Pro. The X-ray crystal structure of the complex of myricetin and SARS-Cov-2 3CL-Pro was solved at a resolution of 1.77 Å, showing that myricetin is covalently bound to the catalytic Cys145 and therefore inhibiting its enzymatic activity.