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De Guzman, Joshua Alrenzo Evangelista

University of British Columbia

There is an urgent need for new antivirals against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus causing coronavirus disease 2019 (COVID-19), because of reduced vaccine effectiveness to emerging variants of concern (VOCs) along with resistance to antibody therapeutics, and limitations present in current SARS-CoV-2 antivirals. A promising antiviral target is the SARS-CoV-2 main protease, also known as the 3-chymotrypsin-like protease (3CLpro), which plays an essential role in viral polyprotein processing. In a prior study (Ton et al., 2020), a 1.3 billion compound library was screened for potential 3CLpro inhibitors by using a Deep Docking (DD) platform to predict which molecules bind to the 3CLpro catalytic site. I hypothesize that compounds predicted by DD to bind to the SARS-CoV-2 3CLpro will present SARS-CoV-2 antiviral activity through inhibition of the 3CLpro. 604 potential 3CLpro inhibitors identified through DD were screened using an in vitro fluorescence resonance energy transfer (FRET)-based enzymatic assay with recombinant SARS-CoV-2 3CLpro. 15 of the molecules presented an inhibitory concentration 50 (IC₅₀) between 7-75 μM against 3CLpro. The best compound, C230, had an IC₅₀ of 74 ± 20 μM. In cellulo studies in Calu-3 lung cells showed that C230 had a mean effective concentration 50 (EC₅₀) of 17 μM against mNeonGreen (mNG) reporter SARS-CoV-2, was even more potent against SARS-CoV-2 Omicron BA.5, with EC₅₀ of 5 μM and 6 μM using nucleocapsid and dsRNA staining, respectively, and had a cytotoxic concentration 50 (CC₅₀) of > 100 μM. Additionally, an in cellulo enzymatic assay showed inhibition of recombinantly expressed SARS-CoV-2 3CLpro proteolytic activity in Caco-2 intestinal cells by 3CLpro inhibitors nirmatrelvir and GC376, with EC₅₀ values of 4.8 ± 0.4 μM and 8.0 ± 3.1 μM, respectively. Major findings of this study include the identification of novel 3CLpro inhibitors that display SARS-CoV-2 antiviral activity, validated through in vitro and in cellulo assays.

Text

2024-01-31

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/83560

Microbiology and Immunology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/