Salicylamide Derivatives Inhibit Multiple Picornavirus Infections through Early Endosome Neutralization

Abstract The global resurgence of picornaviruses highlights an urgent need for broad-spectrum antivirals. Here, we screened 102 salicylamide derivatives, identifying HJW1107 and CZY1111 as inhibitors with submicromolar IC50 values. Time-of-addition and RNA transfection assays confirmed that these derivatives target early infection stages rather than virus replication or assembly. Mechanistically, HJW1107 and CZY1111 exhibit protonophore-like activity that neutralizes endosomal acidification, trapping internalized virions within EEA1-positive early endosomal compartments. This arrest prevents pH-dependent viruses from undergoing uncoating after endocytosis required for virus replication. These results demonstrate that optimizing the salicylamide scaffold to disrupt endosomal maturation offers a therapeutic strategy against acidification-dependent picornaviruses, potentially imposing a higher barrier to resistance than direct targeting of highly variable viral proteins.