Highly contagious SARS-CoV-2 coronavirus has infected billions of people worldwide with flu-like symptoms since its emergence in 2019. It has caused deaths of several million people. The viral main protease (Mpro) is essential for SARS-CoV-2 replication and therefore a drug target. Several series of covalent inhibitors of Mpro were designed and synthesized. Structure–activity relationship studies show that (1) several chloroacetamide- and epoxide-based compounds targeting Cys145 are potent inhibitors with IC50 values as low as 0.49 μM and (2) Cys44 of Mpro is not nucleophilic for covalent inhibitor design. High-resolution X-ray studies revealed the protein–inhibitor interactions and mechanisms of inhibition. It is of interest that Cys145 preferably attacks the more hindered Cα atom of several epoxide inhibitors. Chloroacetamide inhibitor 13 and epoxide inhibitor 30 were found to inhibit cellular SARS-CoV-2 replication with an EC68 (half-log reduction of virus titer) of 3 and 5 μM. These compounds represent new pharmacological leads for anti-SARS-CoV-2 drug development.
