Trivalent triterpenoids block SARS-CoV-2 and influenza A virus infection by directly targeting homotrimeric envelope proteins.

Both SARS-CoV-2 and influenza viruses are major causes of contagious respiratory disease, posing a significant challenge to global public health. In previous work, we identified a class of triterpenoids with broad-spectrum antiviral activity by manipulating virus-host fusion. Here we further report the design and synthesis of 24 trivalent triterpenoid derivatives by targeting the homotrimeric spike protein of SARS-CoV-2 and hemagglutinin (HA) of influenza virus. Anti-SARS-CoV-2 pseudovirus screening in HEK293T-ACE2 cells identified two derivatives 18d and 20c as potent entry inhibitors with IC50 values of 0.82 μM and 0.36 μM, respectively, and negligible cytotoxicity. Flow cytometry and surface plasmon resonance (SPR) assays confirmed that these compounds block viral entry by directly binding to the spike protein. Notably, compound 20c also exhibited potent anti-influenza A virus activity in MDCK cells with EC50 value of 0.61 μM. Time-of-addition and SPR assays revealed that 20c acts at the early stage of infection, significantly suppressing HA expression. Molecular docking simulations further elucidated that the binding mode of triterpenoid moiety within the binding pocket of HA and identified key residues involved in the interaction. Collectively, our findings suggest that trivalent triterpenoid derivatives represent a promising class of broad-spectrum antiviral entry agents that inhibit the entry of both SARS-CoV-2 and influenza virus by targeting the homotrimeric envelope proteins.