C6-Alkoxy substituted penicillins are potent non-covalently binding inhibitors of the SARS-CoV-2 main protease

Abstract

Inhibition of the SARS-CoV-2 main protease (M^pro) by small-molecules is a validated strategy for COVID-19 treatment. There is a need for improved M^pro inhibitors, including because M^pro mutations can confer resistance to clinically used M^pro inhibitors. Previous work has revealed the potential of penicillin derivatives as covalently reacting M^pro inhibitors. Here we report studies on M^pro inhibition by C6-alkoxy substituted penicillin derivatives. The combined mass spectrometric and computational evidence imply most of the tested penicillin C6-alkoxy derivatives bind via non-covalent interactions at the M^pro active site, resulting in potent substrate-competitive inhibition. Some penicillin C6-alkoxy derivatives ((R)-, but not (S)-sulfoxides) manifest covalent reaction to different extents. Penicillins and related drugs are widely used antibiotics, acting via covalent reaction of their β-lactam with a nucleophilic serine in their transpeptidase targets to give an acyl–enzyme complex. The results imply penicillin derivatives can be developed to inhibit enzymes via mechanisms other than formation of stable acyl–enzyme complexes.