Structure-based lead optimization of peptide-based vinyl methyl ketones as SARS-CoV-2 main protease inhibitors

Despite several major achievements in the development of vaccines and antivirals, the fight against SARS-CoV-2 and the health problems accompanying COVID-19 are still ongoing. SARS-CoV-2 main protease (Mpro), an essential viral cysteine protease, is a crucial target for the development of antiviral agents. A virtual screening analysis of in-house cysteine protease inhibitors against SARS-CoV-2 Mpro allowed us to identify two hits (i.e., 1 and 2) bearing a methyl vinyl ketone warhead. Starting from these compounds, we herein report the development of Michael acceptors targeting SARS-CoV-2 Mpro, which differ from each other for the warhead and for the amino acids at the P2 site. The most promising vinyl methyl ketone-containing analogs showed sub-micromolar activity against the viral protease. SPR38, SPR39, and SPR41 were fully characterized, and additional inhibitory properties towards hCatL, which plays a key role in the virus entry into host cells, were observed. SPR39 and SPR41 exhibited single-digit micromolar EC50 values in a SARS-CoV-2 infection model in cell culture. [Display omitted] •Peptide-based Michael acceptors as potent SARS-CoV-2 Mpro inhibitors.•From virtual screening to rational design.•Bi-phasic mechanism of action towards SARS-CoV-2 Mpro and human cathepsin L.•Multitarget ligands in the fight against COVID-19.•Antiviral activity against infected Huh-7-ACE2 cells.