Fig. 6: ISM3312 treatment reduces the risk of drug resistance.
From: A novel, covalent broad-spectrum inhibitor targeting human coronavirus Mpro
SARS-CoV-2 BA.2.3 was serially passaged 18 times in VeroE6 cells with escalating concentrations of ISM3312 and Nirmatrelvir and P-gp inhibitor Elacridar (n = 3 biological replicates) (Method). Inhibition of passage eighteen viruses by ISM3312 (a) and Nirmatrelvir (b). c EC50 of ISM3312 and Nirmatrelvir against passage 18 resistant SARS-CoV-2. Fold change of EC50 mean values relative to inhibition of Ctrl strain from three biologically independent experiments. Color coding for variant frequency: white, no resistance ( < threefold); light blue, low-level resistance (3- to 10-fold); Medium blue, moderate-level resistance (10- to 50-fold); dark blue, high-level resistance ( > 50-fold). d Mutations in Mpro of ISM3312 and Nirmatrelvir resistance from the indicated passages. Dots indicate Ctrl at that residue. Mutations are shaded according to frequency. e IC50 (Compounds are compared for IC50 values under identical incubation time and conditions in each mutant assay) of Nirmatrelvir, Ensitrelvir, and ISM3312 against a panel of Mpro mutants induced by Omicron (P132H) and Nirmatrelvir (Y54A, N133H, F140A, E166A, E166V, L167F, H172Y) those previous reported and ISM3312 (T21I, L50L, P252L were bold labeled) using biochemical FRET assay. Colorimetric mapping of the dAffinity value (kcal/mol) for ISM3312-Mpro complex (f) and Nirmatrelvir-Mpro complex (g) by virtual alanine scanning. Residues around the binding sites are shown. Colors range from blue (negative values, indicating increased protein-ligand affinity) to red (positive values, indicating decreased protein-ligand affinity). ISM3312 is shown in pink, and Nirmatrelvir is shown in orange. h The affinity values for four mutations, including F140A, E166A, E166V and L167F. dAffinity values of E166A and E166V mutations in the Nirmatrelvir-bound system (bold labeled) were significantly higher than those in the ISM3312-bound system. Colorimetric mapping of the dStability value (kcal/mol) for ISM3312-Mpro comple x (i) and Nirmatrelvir-Mpro complex ( j) by virtual alanine scanning. Colors range from blue (negative values, indicating increased protein stability) to red (positive values, indicating decreased protein stability). k The dStability values for 4 mutations, including F140A, E166A, E166V and L167F. Data are presented as mean values ± SEM. Source data are provided as a Source Data file.
