Dynamic interactions and inhibitory mechanisms of Artemisia annua terpenoids with carbonic anhydrase IX.

This study evaluates the inhibitory potential of terpenoids isolated from Artemisia annua against carbonic anhydrase IX (CAIX), a crucial enzyme overexpressed in hypoxic tumor environments. Employing a multidisciplinary approach, we utilized in vitro assays, enzyme kinetics, molecular docking, and molecular dynamics (MD) simulations to comprehensively assess the efficacy of these compounds. Among the terpenoids tested, manool emerged as the most potent inhibitor, exhibiting the lowest IC value of 160.2 ± 15.2 nM. This was followed by labda-8(17),12-diene-15,16-dial with an IC of 297.9 ± 8.84 nM. Enzyme kinetics revealed a mixed inhibition mode for both compounds. Molecular docking aligned well with in vitro data, showing extensive polar and hydrophobic interactions within the CAIX binding site. Further insights were gained through 300 ns MD simulations, which highlighted the dynamic interactions and stability of these complexes. Manool demonstrated the most significant stabilization of CAIX, as evidenced by favorable RMSD, Rg, SASA profiles, and the strongest hydrogen bonding interactions. Additionally, MM/PBSA calculations confirmed manool's superior binding affinity. These findings underscore the therapeutic potential of manool as a potent CAIX inhibitor, providing a foundation for the development of effective anticancer agents targeting hypoxic tumor environments. ADMET analysis revealed favorable pharmacokinetic profiles for the terpenoids, with manool demonstrating high lipophilicity and BBB permeability, though potential CYP-mediated interactions were noted.