Rosmarinic Acid as a Potent Influenza Neuraminidase Inhibitor: In Vitro and In Silico Study

Background: Neuraminidase (NA), a major glycoprotein found on the surface of the influenza virus, is an important target for the prophylaxis and treatment of influenza virus infections. Recently, several plant-derived polyphenols, especially caffeic acid analogs, have been reported to exert the inhibitory activity against NA. Objective: Herein, we aimed to investigate the anti-influenza NA activity of caffeic acid and its hydroxycinnamate analogues, rosmarinic acid and salvianolic acid A, in comparison to a known NA inhibitor, oseltamivir. Methods: In vitro MUNANA-based NA inhibitory assay was used to evaluate the inhibitory activity of the three interested hydroxycinnamic compounds towards the influenza NA enzyme. Subsequently, all-atom molecular dynamics (MD) simulations and binding free energy calculations were employed to elucidate the structural insights into the protein-ligand complexations. Results: Rosmarinic acid showed the highest inhibitory activity against NA with the IC50 of 0.40 mu M compared to caffeic acid (IC50 of 0.81 mu M) and salvianolic acid A (IC50 of >1 mu M). From 100-ns MD simulations, the binding affinity, hot-spot residues, and II-bond formations of rosmarinic acid/NA complex were higher than those of caffeic acid/NA model, in which their molecular complexations was driven mainly by electrostatic attractions and H-bond formations from several charged residues (R118, E119, D151, R152, E227, E277, and R371). Notably, the two hydroxyl groups on both phenyl and phenylacetic rings of rosmarinic acid play a crucial role in stabilizing NA through a strongly formed H-bond(s). Conclusion: Our findings shed light on the potentiality of rosmarinic acid as a lead compound for further development of a potential influenza NA inhibitor.