Discovery and Crystallographic Studies of Nonpeptidic Piperazine Derivatives as Covalent SARS-CoV-2 Main Protease Inhibitors.

The spread of SARS-CoV-2 keeps threatening human life and health, and small-molecule antivirals are in demand. The main protease (M pro ) is an effective and highly conserved target for anti-SARS-CoV-2 drug design. Herein, we report the discovery of potent covalent non-peptide-derived M pro inhibitors. A series of covalent compounds with a piperazine scaffold containing different warheads were designed and synthesized. Among them, GD-9 was identified as the most potent compound with a significant enzymatic inhibition of M pro (IC 50 = 0.18 μM) and good antiviral potency against SARS-CoV-2 (EC 50 = 2.64 μM), similar to that of remdesivir (EC 50 = 2.27 μM). Additionally, GD-9 presented favorable target selectivity for SARS-CoV-2 M pro versus human cysteine proteases. The X-ray co-crystal structure confirmed our original design concept showing that GD-9 covalently binds to the active site of M pro . Our nonpeptidic covalent inhibitors provide a basis for the future development of more efficient COVID-19 therapeutics.