Psoralidin acts as a dual protease inhibitor against PL and M of SARS-CoV-2.

The emergence of new coronavirus variants and concerns about vaccine effectiveness against these novel variants emphasize the need for broad-spectrum therapeutics targeting conserved coronaviral non-structural proteins. Accordingly, a virtual library of 178 putative inhibitors targeting SARS-CoV-2 Papain-like protease (PL) was compiled through a systematic review of published literature and subsequently screened using molecular docking. Selected hits were analyzed for protease inhibitory activities, binding strength, and antiviral activities against HCoV229E-based surrogate system and subsequently against SARS-CoV-2 for validation. Differences in potential modes of action were investigated using an HCoV229E-based system, combined with in silico and biophysical methods against SARS-CoV-2 system. Of the 178 hits, 13 molecules showed superior docking scores against PL and met the inclusion criteria for further investigations. Of these, seven showed notable inhibitory activities against PL. Particularly, both Psoralidin and Corylifol-A exhibited superior and, importantly, dual activities against SARS-CoV-2 M. Both molecules were found to be biologically active against HCoV229E and SARS-CoV-2; however, Psoralidin exhibited more consistent effects and was relatively well-tolerated. Detailed in silico analyses of their interactions with the two proteases identified differences in their modes of action, primarily due to differences in their binding of PL. Based on these findings, we propose Psoralidin as a potential candidate for further development as a broad-spectrum antiviral and Corylifol-A as an ideal candidate for lead optimization.