Discovery of novel microtubule destabilizing agents via virtual screening methods and antitumor evaluation.

Background And Objective: Agents targeting to Colchicine-site of tubulin have become a focus of attention in the development of new anti-cancer drugs due to their ability to overcome multi-drug resistant and anti-angiogenesis effects against tumor endothelium.

Methods: In this investigation, the machine learning methods and molecular docking approach were combined to identify promising destabilizing agents targeting for colchicine-site of tubulin.

Results: Among these screened agents, hit22 represented excellent anti-tumor activity, which exhibited anti-proliferative activity toward H1299 cell with IC value of 3.93 μM, and significantly inhibited the colony formation in a dose-dependent manner. Furthermore, studies elucidated hit22 suppressed tubulin polymerization assembly in vitro, disrupted intracellular microtubule network, and caused G2/M phase cell cycle arrest and apoptosis in H1299 cells. Additionally, hit22 exhibited effective anti-migration abilities against H1299 cells, and suppressed angiogenesis of HUVECs. Importantly, hit22 could significantly inhibit tumor growth in the H1299 xenograft tumor model with a tumor growth inhibition rate of 70.30 %. The molecular dynamics simulations revealed hit22 could stably bind to colchicine site, interacting with some key residues, and inducing βT7 region to produce larger fluctuation. The average binding free energy for the Tubulin-hit22 was -90.9 kJ·mol.

Conclusions: Overall, these results illustrated that hit22 with novel scaffold might be a potential microtubule destabilizing agent targeting colchicine-site and deserves further investigation.