Identification of a novel inhibitor of SARS-CoV-2 main protease: an in silico, biochemical, and cell-based approach.

The recurrent nature of coronavirus outbreaks, severity of the COVID-19 pandemic, rapid emergence of novel variants, and concerns over the effectiveness of existing vaccines against novel variants have highlighted the need to develop therapeutic interventions. Targeted efforts to identify inhibitors of crucial viral proteins are the preferred strategy. In this study, we screened FDA-approved and natural product libraries using in silico approach for potential hits against the SARS-CoV-2 main protease (Mpro) and experimentally validated their potency using in vitro biochemical and cell-based assays. Seven potential hits were identified through in silico screening and were subsequently evaluated in SARS-CoV-2-based cell-free assays, followed by testing in the HCoV-229E-based culture system. Of the tested compounds, 4-(3,4-dihydroxyphenyl)-6,7-dihydroxy-1-isopropyl-1H-benzofuro[3,2-b]pyrazolo[4,3-e]pyridin-3(2H)-one (PubChem CID:71755304, hereafter referred to as STL522228) exhibited significant antiviral activity. Subsequently, its potential as a novel COVID therapeutic molecule was validated in the SARS-CoV-2-culture system, where STL522228 demonstrated superior antiviral activity (EC  = 0.44 μm) compared to Remdesivir (EC  = 0.62 μm). Based on these findings, we report the strong anti-coronavirus activity of STL522228, and propose that it as a promising pan-coronavirus Mpro inhibitor for further experimental and preclinical validation.