Identification of a dual acting SARS-CoV-2 proteases inhibitor through in silico design and step-by-step biological characterization

COVID-19 pandemic, starting from the latest 2019, and caused by SARS-CoV-2 pathogen, led to the hardest health-socio-economic disaster in the last century. Despite the tremendous scientific efforts, mainly focused on the development of vaccines, identification of potent and efficient anti-SARS-CoV-2 therapeutics still represents an unmet need. Remdesivir, an anti-Ebola drug selected from a repurposing campaign, is the only drug approved, so far, for the treatment of the infection. Nevertheless, WHO in later 2020 has recommended against its use in COVID-19. In the present paper, we describe a step-bystep in silico design of a small library of compounds as main protease (MPpro) inhibitors. All the molecules were screened by an enzymatic assay on MPpro and, then, cellular activity was evaluated using Vero cells viral infection model. The cellular screening disclosed compounds 29 and 34 as in-vitro SARS-CoV-2 replication inhibitors at non-toxic concentrations (0.32 < EC50 < 5.98 mu M). To rationalize these results, additional in-vitro assays were performed, focusing on papain like protease (PLpro) and spike protein (SP) as potential targets for the synthesized molecules. This pharmacological workflow allowed the identification of compound 29, as a dual acting SARS-CoV-2 proteases inhibitor featuring micromolar inhibitory potency versus MPpro (IC50 = 1.72 mu M) and submicromolar potency versus PLpro (IC50 = 0.67 mu M), and of compound 34 as a selective SP inhibitor (IC50 = 3.26 mu M). (C) 2021 Elsevier Masson SAS. All rights reserved.

Automated summary

The COVID-19 pandemic caused by the SARS-CoV-2 pathogen has resulted in a major health-socio-economic disaster. Despite efforts in vaccine development, there is still a need for effective anti-SARS-CoV-2 therapeutics. This study describes the design of compounds as main protease inhibitors and identifies two compounds as inhibitors of SARS-CoV-2 replication, potentially offering new therapeutic options.