Arce, R.; Andreu-Moreno, I.; Dufloo, J.; Sanjuan, R.

Emerging viruses pose a major threat to global health, underscoring the need to improve antiviral preparedness. Here, we developed a phylogenetically-informed framework to identify entry inhibitors with potential activity against yet-to-emerge viruses. First, using pseudotypes displaying the receptor-binding proteins (RBPs) of 8 enveloped RNA viruses from different families, we screened a library of 2,320 FDA-approved compounds to identify candidate entry inhibitors. Hits were tested for potency, selectivity, consistency across cell types and pseudotyping vectors and, in some cases, were validated using authentic viruses. Then, to define the breadth of antiviral activity, 25 selected drugs were assayed against an expanded panel of 68 RBPs from 13 families, as well as against pairs of closely related RBPs. This revealed both narrow and broad-range inhibitors, including selective estrogen receptor modulators, alkaloids, aminoquinolines, and anidulafungin, which showed particularly broad activity. Importantly, RBPs from the same phylogenetic cluster frequently displayed correlated drug-sensitivity profiles, indicating that antiviral effects are predictable across closely related viruses. Our findings provide a proof of concept for anticipatory antiviral discovery, showing that phylogenetic relationships can guide the identification of entry inhibitors against potential future zoonotic threats.