Biology subjects

Alugubelli, Y. R.

Publications and source records attributed to Alugubelli, Y. R..

3 recordsLinked to original sources

bioRxiv2021

MPI8 is Potent Against SARS-CoV-2 by Inhibiting Dually and Selectively the SARS-CoV-2 Main Protease and the Host Cathepsin L

A number of inhibitors have been developed for the SARS-CoV-2 main protease (MPro) as potential COVID-19 medications but little is known about their selectivity. Using enzymatic assays, we characterized inhibition of TMPRSS2, furin, and cathepsins B/K/L by more than a dozen of previously developed MPro inhibitors including MPI1-9, GC376, 11a, 10-1, 10-2, and 10- 3. MPI1-9, GC376 and 11a all contain an aldehyde for the formation of a reversible covalent hemiacetal adduct with the MPro active site cysteine and 10-1, 10-2 and 10-3 contain a labile ester to exchange with the MPro active site cysteine for the formation of a thioester. Our data revealed that all these inhibitors are inert toward TMPRSS2 and furin. Diaryl esters also showed low inhibition of cathepsins. However, all aldehyde inhibitors displayed high potency in inhibiting three cathepsins. Their determined IC50 values vary from 4.1 to 380 nM for cathepsin B, 0.079 to 2.3 nM for cathepsin L, and 0.35 to 180 nM for cathepsin K. All aldehyde inhibitors showed similar inhibition levels toward cathepsin L. A cellular analysis indicated high potency of MPI5 and MPI8 in inhibiting lysosomal activity, which is probably attributed to their inhibition of cathepsins. Among all aldehyde inhibitors, MPI8 shows the best selectivity toward cathepsin L. With respect to cathepsins B and K, the selective indices are 192 and 150, respectively. MPI8 is the most potent compound among all aldehyde inhibitors in cellular MPro inhibition potency and anti-SARS-CoV-2 activity in Vero E6 cells. Cathepsin L has been demonstrated to play a critical role in the SARS-CoV-2 cell entry. By selectively inhibiting both SARS-CoV-2 MPro and the host cathepsin L, MPI8 potentiates dual inhibition effects to synergize its overall antiviral potency and efficacy. Due to its high selectivity toward cathepsin L that reduces potential toxicity toward host cells and high cellular and antiviral potency, we urge serious consideration of MPI8 for preclinical and clinical investigations for treating COVID-19.

pharmacology and toxicology↗

bioRxiv2021

Cellular Activities of SARS-CoV-2 Main Protease Inhibitors Reveal Their Unique Characteristics

As an essential enzyme of SARS-CoV-2, the pathogen of COVID-19, main protease (MPro) triggers acute toxicity to its human cell host, an effect that can be alleviated by an MPro inhibitor with cellular potency. By coupling this toxicity alleviation with the expression of an MPro-eGFP fusion protein in a human cell host for straightforward characterization with fluorescent flow cytometry, we developed an effective method that allows bulk analysis of cellular potency of MPro inhibitors. In comparison to an antiviral assay in which MPro inhibitors may target host proteases or other processes in the SARS-CoV-2 life cycle to convene strong antiviral effects, this novel assay is more advantageous in providing precise cellular MPro inhibition information for assessment and optimization of MPro inhibitors. We used this assay to analyze 30 literature reported MPro inhibitors including MPI1-9 that were newly developed aldehyde-based reversible covalent inhibitors of MPro, GC376 and 11a that are two investigational drugs undergoing clinical trials for the treatment of COVID-19 patients in United States, boceprevir, calpain inhibitor II, calpain inhibitor XII, ebselen, bepridil that is an antianginal drug with potent anti-SARS-CoV-2 activity, and chloroquine and hydroxychloroquine that were previously shown to inhibit MPro. Our results showed that most inhibitors displayed cellular potency much weaker than their potency in direct inhibition of the enzyme. Many inhibitors exhibited weak or undetectable cellular potency up to 10 M. On contrary to their strong antiviral effects, 11a, calpain inhibitor II, calpain XII, ebselen, and bepridil showed relatively weak to undetectable cellular MPro inhibition potency implicating their roles in interfering with key steps other than just the MPro catalysis in the SARS-CoV-2 life cycle to convene potent antiviral effects. characterization of these molecules on their antiviral mechanisms will likely reveal novel drug targets for COVID-19. Chloroquine and hydroxychloroquine showed close to undetectable cellular potency to inhibit MPro. Kinetic recharacterization of these two compounds rules out their possibility as MPro inhibitors. Our results also revealed that MPI5, 6, 7, and 8 have high cellular and antiviral potency with both IC50 and EC50 values respectively below 1 M. As the one with the highest cellular and antiviral potency among all tested compounds, MPI8 has a remarkable cellular MPro inhibition IC50 value of 31 nM that matches closely to its strong antiviral effect with an EC50 value of 30 nM. Given its strong cellular and antiviral potency, we cautiously suggest that MPI8 is ready for preclinical and clinical investigations for the treatment of COVID-19.

biochemistry↗

bioRxiv2020

A Speedy Route to Multiple Highly Potent SARS-CoV-2 Main Protease Inhibitors

The COVID-19 pathogen, SARS-CoV-2, requires its main protease (SC2MPro) to digest two of its translated polypeptides to form a number of mature proteins that are essential for viral replication and pathogenesis. Inhibition of this vital proteolytic process is effective in preventing the virus from replication in infected cells and therefore provides a potential COVID-19 treatment option. Guided by previous medicinal chemistry studies about SARS-CoV-1 main protease (SC1MPro), we have designed and synthesized a series of SC2MPro inhibitors that contain {beta}-(S-2-oxopyrrolidin-3-yl)-alaninal (Opal) for the formation of a reversible covalent bond with the SC2MPro active site cysteine C145. All inhibitors display high potency with IC50 values at or below 100 nM. The most potent compound MPI3 has as an IC50 value as 8.5 nM. Crystallographic analyses of SC2MPro bound to 7 inhibitors indicated both formation of a covalent bond with C145 and structural rearrangement from the apoenzyme to accommodate the inhibitors. Virus inhibition assays revealed that several inhibitors have high potency in inhibiting the SARS-CoV-2-induced cytopathogenic effect in both Vero E6 and A549 cells. Two inhibitors MP5 and MPI8 completely prevented the SARS-CoV-2-induced cytopathogenic effect in Vero E6 cells at 2.5-5 M and A549 cells at 0.16-0.31 M. Their virus inhibition potency is much higher than some existing molecules that are under preclinical and clinical investigations for the treatment of COVID-19. Our study indicates that there is a large chemical space that needs to be explored for the development of SC2MPro inhibitors with extreme potency. Due to the urgent matter of the COVID-19 pandemic, MPI5 and MPI8 may be quickly advanced to preclinical and clinical tests for COVID-19.

biochemistry↗