Small-Molecule Thioesters as SARS-CoV-2 Main Protease Inhibitors: Enzyme Inhibition, Structure-Activity Relationships, Antiviral Activity, and X-ray Structure Determination.

The main protease (M ^pro , 3CL ^pro ) of SARS-CoV-2 is an attractive target in coronaviruses because of its crucial involvement in viral replication and transcription. Here, we report on the design, synthesis, and structure-activity relationships of novel small-molecule thioesters as SARS-CoV-2 M ^pro inhibitors. Compounds 3w and 3x exhibited excellent SARS-CoV-2 M ^pro inhibition with k _inac / K _i of 58,700 M ^-1 s ^-1 ( K _i = 0.0141 μM) and 27,200 M ^-1 s ^-1 ( K _i = 0.0332 μM), respectively. In Calu-3 and Vero76 cells, compounds 3h , 3i, 3l , 3r , 3v , 3w , and 3x displayed antiviral activity in the nanomolar range without host cell toxicity. Co-crystallization of 3w and 3af with SARS-CoV-2 M ^pro was accomplished, and the X-ray structures showed covalent binding with the catalytic Cys145 residue of the protease. The potent SARS-CoV-2 Mpro inhibitors also inhibited the M ^pro of other beta-coronaviruses, including SARS-CoV-1 and MERS-CoV, indicating that they might be useful to treat a broader range of coronaviral infections.