A novel C-3-substituted oleanolic acid benzyl amide derivative exhibits therapeutic potential against influenza A by targeting PA–PB1 interactions and modulating host macrophage inflammation

The influenza A virus (IAV), renowned for its high contagiousness and potential to catalyze global pandemics, poses significant challenges due to the emergence of drug-resistant strains. Given the critical role of RNA polymerase in IAV replication, it stands out as a promising target for anti-IAV therapies. In this study, we identified a novel C-3-substituted oleanolic acid benzyl amide derivative, <b>A5</b>, as a potent inhibitor of the PA_C-PB1_N polymerase subunit interaction, with an IC₅₀ value of 0.96 ± 0.21 μmol/L. <b>A5</b> specifically targets the highly conserved PA_C domain and demonstrates remarkable efficacy against both laboratory-adapted and clinically isolated IAV strains, including multidrug-resistant strains, with EC₅₀ values ranging from 0.60 to 1.83 μmol/L. Notably, when combined with oseltamivir, <b>A5</b> exhibits synergistic effects both <i>in vitro</i> and <i>in vivo</i>. In a murine model, dose-dependent administration of <b>A5</b> leads to a significant reduction in IAV titers, resulting in a high survival rate among treated mice. Additionally, <b>A5</b> treatment inhibits virus-induced Toll-like receptor 4 activation, attenuates cytokine responses, and protects against IAV-induced inflammatory responses in macrophages. In summary, <b>A5</b> emerges as a novel inhibitor with high efficiency and broad-spectrum anti-influenza activity.