Discovery of novel microtubule stabilizers targeting taxane binding site by applying molecular docking, molecular dynamics simulation, and anticancer activity testing.

Disruption of the dynamic equilibrium of microtubules can induce cell cycle arrest in G2/M phase and apoptosis. Hence, discovery of novel tubulin polymerization inhibitors is very necessary and an important task in drug research and development for treatment of various tumors. In this investigation, 50 compounds were screened as microtubule stabilizers targeting the taxane site by combination of molecular docking methods. Among these hits, hits 19 and 38 with novel scaffolds exhibited the highest anti-proliferative activity with IC ranging from 9.50 to 13.81 μM in four cancer cell lines. The molecular dynamics simulations confirmed that tubulin and two hits could form stable systems. Meanwhile, the mechanism of the interactions between tubulin and two hits at simulated physiological conditions were probed. The in vitro tubulin polymerization assay revealed hits 19 and 38 were able to promote tubulin polymerization in a dose-dependent manner. Further, the immunofluorescence assay suggested that hits 19 and 38 could accelerate microtubule assembly in A549 and HeLa cells. Finally, studies on antitumor activity indicated that hits 19 and 38 induced G2/M phase cell cycle arrest and apoptosis, and inhibited cancer cell motility and migration in A549 and HeLa cells. Importantly, hit38 exhibited better anti-tubulin and anti-cancer activity than hit19 in A549 and HeLa cells. Therefore, these results suggest that hit38 represents a promising microtubule stabilizer for treating cancer and deserves further investigation.