Identification of potential COVID-19 treatment compounds which inhibit SARS Cov2 prototypic, Delta and Omicron variant infection

Abstract

Recurrent waves of COVID19 remain a major global health concern. Repurposing either FDA-approved or clinically advanced drug candidates can save time and effort required for validating the safety profile and FDA approval. However, the selection of appropriate screening approaches is key to identifying novel candidate drugs with a higher probability of clinical success. Here, we report a rapid, stratified two-step screening approach using pseudovirus entry inhibition assay followed by an infectious prototypic SARS CoV2 cytotoxic effect inhibition assay in multiple cell lines. Using this approach, we screened a library of FDA-approved and clinical-stage drugs and identified four compounds, apilimod, berbamine, cepharanthine and (S)-crizotinib which potently inhibited SARS CoV2-induced cell death. Importantly, these drugs exerted similar inhibitory effect on the delta and omicron variants although they replicated less efficiently than the prototypic strain. Apilimod is currently under clinical trial (NCT04446377) for COVID19 supporting the validity and robustness of our screening approach.

Keywords: Anti-viral drug repurposing; Delta; Omicron; SARS CoV2.

Published by Elsevier Inc.

Figures

Fig. 1

Pseudovirus entry inhibition screening of drug library. A-D) 293T, 293T-ACE2, A549-ACE2 and Vero TMPRSS2 cells were transduced with SARS CoV2 pseudotyped virus particles containing luciferase reporter gene and luminescence was measured after 72 h. Bar graph shows respective Relative Light Units (RLUs) from Mock (Blue) without envelope and SARS CoV2 spike pseudotyped virus particle (Red) transduced cells. E) RLU values from 293T-ACE2 cells transduced with serially diluted SARS CoV2 spike pseudotyped virus particles. F–I) Bar graphs show RLU values from pseudovirus entry inhibition assay where 293T ACE2 cells were pre-treated with 1 μM of indicated drugs for 1 h and then transduced with SARS CoV2 spike pseudotyped virus particles. Cathepsin-L inhibitor CAS 108005943 was used as a positive control. J) Bar graph shows summary of percent pseudovirus entry inhibition values for each drug. Values represent Mean ± SEM, n = 3 and two independent experiments.

Fig. 2

Validation of cell death inhibition assay. A) Cells were infected with serial dilutions of SARS CoV2 (10^-2 to 10^-8) and cell death was quantified by Cell-Titer-Glo cell viability assay and TCID50 for each cell line was determined at 72 h. B) Cells were pre-treated with different concentrations of Remdesivir followed by infection with 10 TCID50 of SARS CoV2 and the effect on cell death inhibition was quantified. C-E) Dose response effect of candidate drugs apilimod, berbamine, cepharanthine and (S)-crizotinib on USA/WA1 SARS CoV2 induced cell death in 293T-ACE2, Vero E6 and Vero-TMPRSS2 cells was analyzed. F) Vero TMPRSS2 cells were infected with 10TCID50 SARS CoV2 and the nucleocapsid positive (NP+) cells were enumerated by flow cytometry 24 h post infection. Numbers in the histograms show the percentage of NP + cells. G) Bar graph shows the % inhibition in NP + cells in the presence of candidate drugs. Values represent Mean ± SEM, n = 3 and two independent experiments.

Fig. 3

A-B) Representative histograms show the frequency of NP + cells in the Vero-TMPRSS2 (A) and Vero-ACE2-TMPRSS2 (B) cells infected with different doses of the USA-WA1, delta and omicron strains of SARS CoV2. C-D) Graphs show the TCID50 values for each strain of the virus in Vero TMPRSS2 (C) and Vero ACE2-TMPRSS2 cells (D). E-H) Dose response effect of the drugs on delta and omicron variant induced cell death in Vero TMPRSS2 and Vero-ACE2-TMPRSS2 cells. Values represent Mean ± SEM, n = 3 and two independent experiments.