Structure-Based Virtual Screening and Biochemical Validation to Discover a Potential Inhibitor of the SARS-CoV-2 Main Protease

Abstract

The recent pandemic caused by SARS-CoV-2 has led the world to a standstill, causing a medical and economic crisis worldwide. This crisis has triggered an urgent need to discover a possible treatment strategy against this novel virus using already-approved drugs. The main protease (Mpro) of this virus plays a critical role in cleaving the translated polypeptides that makes it a potential drug target against COVID-19. Taking advantage of the recently discovered three-dimensional structure of Mpro, we screened approved drugs from the Drug Bank to find a possible inhibitor against Mpro using computational methods and further validating them with biochemical studies. The docking and molecular dynamics study revealed that DB04983 (denufosol) showed the best glide docking score, −11.884 kcal/mol, and MM-PBSA binding free energy, −10.96 kcal/mol. Cobicistat, cangrelor (previous computational studies in our lab), and denufosol (current study) were tested for the in vitro inhibitory effects on Mpro. The IC 50 values of these drugs were ∼6.7 μM, 0.9 mM, and 1.3 mM, respectively, while the values of dissociation constants calculated using surface plasmon resonance were ∼2.1 μM, 0.7 mM, and 1.4 mM, respectively. We found that cobicistat is the most efficient inhibitor of Mpro both in silico and in vitro. In conclusion, cobicistat, which is already an FDA-approved drug being used against HIV, may serve as a good inhibitor against the main protease of SARS-CoV-2 that, in turn, can help in combating COVID-19, and these results can also form the basis for the rational structure-based drug design against COVID-19.

Validation and invalidation of SARS-CoV-2 main protease inhibitors using the Flip-GFP and Protease-Glo luciferase assays

“… Not a M pro inhibitor Nelfinavir >20 2 118 ± 18 30 –0.60 >10 >240 3.3 30 (Nluc)0.77 ± 0.32 24 3.1 ± 0.06 34 N.A. Not a M pro inhibitor Cobicistat >20 6.7 ± 0.6 39 –0.65 >20 >240 (Nluc)2.74 ± 0.20 24 N.A. Not a M pro inhibitor Calcium channel blocker Manidipine 64.2 ± 9.8 4.81 ± 1.87 40 0.45 >10 >240 N.A.…”

Recent Developments in Free Energy Calculations for Drug Discovery

“…The emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global health crisis with over 2 million deaths worldwide, compelling rapid drug development for potential therapeutics. Several major protein targets have been identified for inhibition of SARS-CoV-2 function and surveyed through molecular simulation for predicted binding affinity with repurposed and novel drugs, these include the RNA dependent RNA polymerase ( Procacci et al, 2020 ; Wakchaure et al, 2020 ) (RdRp) that replicates the RNA genome, the main protease ( Macchiagodena et al, 2020b ; Ngo et al, 2020b ; Chowdhury et al, 2020 ; Gupta et al, 2020 ; Gupta and Zhou, 2020 ; Jukic et al, 2020 ; Li et al, 2020 ; Milenković et al, 2020 ; Tejera et al, 2020 ; Aghaee et al, 2021 ; Bhardwaj et al, 2021 ) (3CL M pro ) that mediates replication and transcription, the spike protein ( Patil et al, 2021 ) involved in initiating infection by penetrating the host cell, S-adenosyl-methionine dependent methyltransferase ( Sk et al, 2020 ) (nsp16) that adds the 5′-cap to mRNA essential for stability, envelope protein ( Dey et al, 2020 ) that is involved in virion assembly and budding, Papain-like protease that functions in viral replication and immune response evasion ( Bosken et al, 2020 ), and the host serine protease TMPRSS2 ( Singh et al, 2020 ) that primes the spike protein. Alanine scanning is combined with MM-PBSA to identify the hot-spot binding residues GLU166 and GLN189 on M pro as critical sites for inhibitors to target ( Aghaee et al, 2021 ).…”

Combining Different Docking Engines and Consensus Strategies to Design and Validate Optimized Virtual Screening Protocols for the SARS-CoV-2 3CL Protease

“…Among the other 102 molecules, there are 37 compounds that are known inhibitors of SARS-CoV 3CL-Pro but with pIC50 < 5, and 17 known inhibitors of the main proteases of other viruses (such as norovirus and HIV), and these compounds represent a further confirmation of the efficacy of the performed VS campaigns, since some of the retrieved hits (rupintrivir [31], saquinavir [32], and lopinavir [33]) were experimentally confirmed as promising inhibitors of SARS-CoV-2 3CL-Pro. Finally, among the other common molecules, cobicistat [34] and galloyl analogues [35] were identified as SARS-CoV-2 3CL-Pro inhibitors.…”