Discovery of two novel small-molecule series with potent SARS-CoV-2 inhibition via putative nsp13 open-state trapping

Abstract

We report the discovery of 31 SARS-CoV-2 inhibitors identified across two computational-to-experimental screening campaigns, with average cell-based antiviral potency satisfying 〈EC50〉 ≤ CC50/3 in A549-hACE2 cells. These compounds were selected from 60 successfully tested molecules evaluated using nanoluciferase reporter assay (nLuc), cytopathic effect (CPE), and host-cell cytotoxicity (CC50) assays. We used a funnel-like computational framework where candidate compounds were progressively prioritized through different scores, improving robustness against the limitations of individual metrics. However, none of the docking-based scoring metrics, including MM/GBSA and Glide scores, showed statistically significant correlation with experimental activity, illustrating the limitations of docking-only approaches and supporting a major contribution of the machine-learning predictions to the high hit-identification rate. Although direct target validation is still lacking, the machine-learning (ML) workflow was grounded in experimental nsp13 inhibition labels, and molecular dynamics (MD) simulations suggest that these compounds can act by stabilizing apo-like open conformations of nsp13, with differential interactions involving the 1B domain contributing to potency differences. Among the compounds evaluated, the phenoxypropanol (PP) and bipiperidine (BPP) series stood out as the most promising SARS-CoV-2 antivirals. Integrated analysis of structure–activity relationships, physicochemical parameters, metabolic stability, and interdomain dynamics delineated structural optimization paths for both series.