SARS-related coronaviruses poses continual threat to humanity by rapidly mutating and emerging as severe pandemic outbreaks, including the current nCoV-19 pandemic. Hence a rapid drug repositioning and lead identification strategy are required to mitigate these outbreaks. We report a pharmacophore and molecular dynamics-based approach for drug repositioning and lead identification against dual targets (3CLp and PLp) of SARS-CoV-2. The pharmacophore model of 3CLp inhibitors was apolar with two aromatic and two H-bond acceptors, whereas that of PLp was relatively polar, bearing one aromatic and three H-bond acceptors. Pharmacophore-based virtual screening yielded six existing FDA-approved drugs and twelve natural products with both the pharmacophoric features. Among them are nelfinavir, tipranavir and licochalcone-D, which has shown better binding characteristics with both the proteases compared to lopinavir. The molecular dynamics revealed that the connecting loop (residues 176-199) of 3CLp is highly flexible, and hence, inhibitors should avoid high-affinity interactions with it. Lopinavir, due to its high affinity with the loop region, exhibited unstable binding. Further, the van der Waals size of the 3CLp inhibitors positively correlated with their binding affinity with 3CLp. However, the van der Waals size of a ligand should not cross a threshold of 572Å 3 , beyond which the ligands are likely to make high-affinity interaction with the loop and suffer unstable binding as observed in the case of lopinavir. Similarly, the total polar surface area of the ligands were found to be negatively correlated with their binding affinity with PLp.
Background
The Transmembrane Serine Protease 2 (TMPRSS2) of human cell plays a significant role in proteolytic cleavage of SARS-Cov-2 coronavirus spike protein and subsequent priming to the receptor ACE2. Approaching TMPRSS2 as a therapeutic target for the inhibition of SARS-Cov-2 infection is highly promising. Hence, in the present study, we docked the binding efficacy of ten naturally available phyto compounds with known anti-viral potential with TMPRSS2. The aim is to identify the best phyto compound with a high functional affinity towards the active site of the TMPRSS2 with the aid of two different docking software. Molecular Dynamic Simulations were performed to analyse the conformational space of the binding pocket of the target protein with selected molecules.
Results
Docking analysis using PyRx version 0.8 along with AutoDockVina reveals that among the screened phyto compounds, Genistein shows the maximum binding affinity towards the hydrophobic substrate-binding site of TMPRSS2 with three hydrogen bonds interaction ( − 7.5 kcal/mol). On the other hand, molecular docking analysis using Schrodinger identified Quercetin as the most potent phyto compound with a maximum binding affinity towards the hydrophilic catalytic site of TMPRSS2 ( − 7.847 kcal/mol) with three hydrogen bonds interaction. The molecular dynamics simulation reveals that the Quercetin-TMPRSS complex is stable until 50 ns and forms stable interaction with the protein ( − 22.37 kcal/mol of MM-PBSA binding free energy). Genistein creates a weak interaction with the loop residues and hence has an unstable binding and exits from the binding pocket.
Conclusion
The compounds, Quercetin and Genistein, can inhibit the TMPRSS2 guided priming of the spike protein. The compounds could reduce the interaction of the host cell with the type I transmembrane glycoprotein to prevent the entry of the virus. The critical finding is that compared to Genistein, Quercetin exhibits higher binding affinity with the catalytic unit of TMPRSS2 and forms a stable complex with the target. Thus, enhancing our innate immunity by consuming foods rich in Quercetin and Genistein or developing a novel drug in the combination of Quercetin and Genistein could be the brilliant choices to prevent SARS-Cov-2 infection when we consider the present chaos associated with vaccines and anti-viral medicines.
In
this study, a pH-induced self-assembly-based method has been
developed to form silk fibroin nanoparticles (SFN-2) with a higher
drug loading capacity (21.0 ± 2.1%) and cellular uptake than
that of silk fibroin particles produced by a conventional desolvation
method (SFN-1). Using the self-assembly method, rifampicin-encapsulated
silk fibroin nanoparticles (R-SFN-2) were prepared with a size of
165 ± 38 nm at an optimum pH of 3.8. In silico analysis reveals that at acidic pH, the amino acid side chain charge
neutralization of acidic residues, especially GLU64, promotes the
formation of additional favorable interactions between the silk fibroin
and the drug. The SFN-2 also possess a good aerosol property with
a mass median aerodynamic diameter of 3.82 ± 0.71 μm and
fine particle fraction of 64.0 ± 1.4%. These SFN-2 particles
were selectively endocytosed by macrophages through clathrin- and
caveolae-mediated endocytosis with a higher uptake efficiency (66.2
± 2.1%) and were found to exhibit a sustained drug release in
the presence of macrophage intracellular lysates. The cytokine and
biomarker expression analyses revealed that SFN-2 could exhibit an
immunomodulatory effect by polarizing the macrophages to an initial
M1 phase and later M2 phase. Further, R-SFN-2 also exhibited an enhanced
and sustained intracellular antibacterial activity against Mycobacterium smegmatis-infected macrophages than
free rifampicin. Thus, the self-assembled silk fibroin particles with
immunomodulatory action combined with a good aerosol and intracellular
drug release property can be an attractive choice as a carrier for
developing pulmonary drug delivery systems.
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