Design,Synthesis And Bioactivity Evaluation Of 1,2-diarylbenzimidazole Derivatives As Potential Tubulin Inhibitors

Microtubules were composed of tubulin and microtubule-binding proteins,which were the major components of the cytoskeleton.Microtubules had the kinetic properties of polymerization and depolymerization,and played a role in maintaining cell morphology,cell division and proliferation,organelle composition and transport,and signal transduction.Utilizing the kinetic properties of tubulin,anti-cancer drugs targeting tubulin promoted depolymerization or inhibited their polymerization.Studies had shown that there are mainly three different inhibitory binding sites on tubulin:paclitaxel site,vincristine site and colchicine site.Due to the small volume of the colchicine cavity and the simple structure of the corresponding inhibitors,research on its inhibitors had received much attention in recent years.CA-4 was originally a product isolated from Combretum Caffrum in South Africa.The structure-activity relationship study showed that the cis configuration of the two aromatic rings and the trimethoxy group of the A ring are important conditions for the anti-tumor activity of CA-4.However,the cis-double bond in the CA-4 structure was easily isomerized to a trans-double bond during storage and use,which is thermodynamically more stable,thereby reducing activity.Therefore,many studies have attempted to replace the cis double bond of CA-4 with different linkers,including different heterocyclic structures,to avoid structural isomerization of CA-4.At the same time,the substituents of the B ring of CA-4 were altered to obtain CA-4 analogs with better pharmacodynamic and pharmacokinetic characteristics.The structure of the designed compound was modified to replace the double bond of CA-4 and a heterocyclic ring,which not only stabilized the cis configuration but also improved the stability of the compound.Benzimidazole was one of the earliest nitrogen-containing heterocyclic compounds,and its structure had been widely studied in the pharmaceutical field as an advantageous skeleton.Benzimidazole-containing compounds had a wide range of biological activities,such as anti-inflammatory,anti-hypertensive,anti-clotting,anti-cancer,etc.,especially as anti-cancer drugs have good application prospects.It had been reported in the literature that a novel compound containing a benzimidazole structure can bind to tubulin at the colchicine site and inhibit tubulin polymerization.Based on our previous research,we designed and synthesized a series of 1,2-diarylbenzimidazole derivatives and evaluated their biological activities.The benzimidazole structure was added to the microtubule inhibitor structure to increase structural stability and compound activity.The structure of the positive drug CA-4 was modified to retain the same 3,4,5-trimethoxy group as the A ring in the CA-4 structure,and the B ring was substituted with a different substituent.Only benzimidazole was used as a bridge between the A ring and the B ring,and the two benzene rings were substituted at the 1 and 2 positions of the benzimidazole,respectively.Bioactivity evaluation results show that most of them are potential anti-tumor drugs.Among them,Compound c24 showed the biggest anti-tumor activity against HeLa,HepG2 and MCF-7 cells,GI50=0.71-2.41 μM.It was less toxic to normal cells and had a CC50>100 μM against 293T cells.In the tubulin binding assay,it showed excellent ability to inhibit tubulin polymerization(IC50=8.47 μM).The binding ability of compound c24 to tubulin crystals was verified by molecular dynamics simulation docking experiments.Further studies on HepG2 and HeLa cells showed that Compound c24 caused mitotic arrest of tumor cells to the G2/M phase and induced apoptosis.In summary,Compound c24 was a promising inhibitor of tubulin assembly.