Targeting SARS-CoV-2 Main Protease (MPro) with Kinase Inhibitors: A Promising Approach for Discovering Antiviral and Anti-inflammatory Molecules against SARS-CoV-2

Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus infected over 688 million people worldwide, causing public health concern and approximately 6.8 million deaths due to COVID-19. COVID-19, especially severe cases, is characterized by exacerbated lung inflammation with an increase of pro-inflammatory cytokines. In addition to antiviral drugs, there is a need for anti-inflammatory therapies to treat all phases of COVID-19. One of the most attractive drug targets for COVID-19 is the SARS-CoV-2 main protease (MPro), an enzyme responsible for cleaving polyproteins formed after the translation of viral RNA, which is essential for viral replication. MPro inhibitors, therefore, have the potential to stop viral replication and act as antiviral drugs. Considering that several kinase inhibitors are known for their action in inflammatory pathways, this could also be investigated toward a potential antiinflammatory treatment for COVID-19. Therefore, the use of kinase inhibitors against SARS-CoV-2 MPro may be a promising strategy to find molecules with dual activity�antiviral and anti-inflammatory. Considering this, the potential of six kinase inhibitors against SARS-CoV-2 MPro were evaluated in silico and in vitro, including Baricitinib, Tofacitinib, Ruxolitinib, BIRB-796, Skepinone-L, and Sorafenib. To assess the inhibitory potential of the kinase inhibitors, a continuous fluorescent-based enzyme activity assay was optimized with SARS-CoV-2 MPro and MCA-AVLQSGFR-K(Dnp)-K-NH2 (substrate). BIRB-796 and Baricitinib were identified as inhibitors of SARS-CoV-2 MPro, presenting IC 50 values of 7.99 and 25.31 μM, respectively. As they are also known for their anti-inflammatory action, both are prototype compounds with the potential to present antiviral and anti-inflammatory activity against SARS-CoV-2 infection.

Identification of SARS-CoV-2 Main Protease Inhibitors Using Chemical Similarity Analysis Combined with Machine Learning

“…First, from the literature, we identified anti-coronavirus compounds with diverse mechanisms of action ( Table S1 ) to be used as references for similarity searching from a compound screening library with ~17 million molecules of bioactive compounds, natural products, and peptides [ 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 ]. Of the 62 reference molecules, thirteen ( Table S2 ) have been shown to inhibit at least one of two SARS-CoV-2 proteases [ 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 ]. Using three different molecular fingerprints, we identified 632,149 unique molecules that had similarity values greater than the mean plus three standard deviations (SDs) for all three of the molecular fingerprints.…”

Kinetic comparison of all eleven viral polyprotein cleavage site processing events by SARS-CoV-2 main protease using a linked protein FRET platform

“…Efforts to develop an improved SARS-CoV-2 M pro assay that delivers improved consistency while maintaining high sensitivity are ongoing ( 41 , 50 ); however, the use of peptidomimetic substrates linked with small molecular fluorophores ( e.g. EDANS/Dabcyl ( 46 ), FAM/Dabcyl ( 50 ), MCA/Dnp ( 51 ), and so on.) remains problematic.…”