Discovery of quinoxaline-chalcone derivatives as covalent colchicine binding site inhibitors for breast and fibrosarcoma treatment.

Targeting the colchicine binding site is an attractive pharmacological strategy to overcome the multidrug resistance microtubule-targeting agents are faced with. Herein, a series of novel covalent quinoxaline-chalcone derivatives were rationally designed using pharmacophore fusion with S-Ⅳ-25 as lead compound. All obtained target compounds were evaluated for their inhibitory activities and the structure-activity relationships were summarized, leading to the identification of a potent colchicine binding site inhibitor 10v. 10v showed the most potent antiproliferative activity against three cancer cell lines (HT-1080, MCF-7, MDA-MB-231), and further inhibited colony formation and migration. Mechanistic studies demonstrated that 10v inhibited tubulin polymerization and disrupted microtubule network via covalent binding to the colchicine binding site of tubulin. Further mechanistic assays revealed 10v arrested cell cycle at G2/M phase and induced cell apoptosis through mitochondrial-mediated pathway. More importantly, 10v exhibited antiproliferative activity against MCF-7/ADM cells via covalent binding and P-gp inhibition. Meanwhile, molecular docking result suggested 10v occupied colchicine binding site and formed covalent binding with β-Cys239. Finally, in vivo study indicated 10v showed better in vivo antitumor efficacy than paclitaxel with acceptable safety.