Improved SARS-CoV-2 Mpro inhibitors based on feline antiviral drug GC376: Structural enhancements, increased solubility, and micellar studies

Replication of SARS-CoV-2, the coronavirus causing COVID-19, requires a main protease (M-pro) to cleave viral proteins. Consequently, Mpro is a target for antiviral agents. We and others previously demonstrated that GC376, a bisulfite prodrug with efficacy as an anti-coronaviral agent in animals, is an effective inhibitor of Mpro in SARS-CoV-2. Here, we report structure-activity studies of improved GC376 derivatives with nanomolar affinities and therapeutic indices >200. Crystallographic structures of inhibitor-M-pro complexes reveal that an alternative binding pocket in M-pro, S4, accommodates the P3 position. Alternative binding is induced by polar P3 groups or a nearby methyl. NMR and solubility studies with GC376 show that it exists as a mixture of stereoisomers and forms colloids in aqueous media at higher concentrations, a property not previously reported. Replacement of its Na+ counter ion with choline greatly increases solubility. The physical, biochemical, crystallographic, and cellular data reveal new avenues for Mpro inhibitor design. (C) 2021 Elsevier Masson SAS. All rights reserved.

Automated summary

The study demonstrates that GC376 and its derivatives are effective inhibitors of Mpro in SARS-CoV-2, with improved affinity and therapeutic index. Crystallographic analysis reveals an alternative binding pocket in M-pro and the mechanism of alternative binding induced by polar groups or a nearby methyl. Additionally, new findings on solubility and colloidal formation in aqueous media are reported, along with the use of choline to increase solubility.