The Study On Antitumor Molecular Mechanism Of Novel Tubulin Inhibitors CA4Analog

Combretastatin A4is one of the most potent microtubule inhibitors. It can destroy the mitotic spindle of tumor cells with strong proliferate capacity which lead tumor cells to lagging in the G2/M phase of mitosis. Based on poor water solubility, low bioavailability, the nature of being turned into inactive trans-configuration easily and the restriction in clinical application of CA4, we modified the molecule of CA4and successfully obtained a number of noval CA4analogs. In this study, we study on the tubule targeted inhibitor in molecular mechanisms of CA4and40B.In this study, novel CA4analogs were detected to inhibit proliferation activity in vitro of tumor cells and normal cells using MTT assays. The results showed that the four novel CA4analogs have a broad spectrum antitumor activity and40B is the best which the IC50is8nmol/L. Followed research use MCF-7as model to explore the inhibitory microtubule mechanism of40B. The result of FCM showed that both CA4and40B can block MCF-7cells in G2/M phase, and the block rate increased in a concentration dependence pattern, with48hours is the highest. Immunofluorescence detected the MCF-7cell apoptosis and the change of tubulin in the nucleus and plasma distribution from morphology. The results showed that CA4and40B make the cell nucleus microtubules depolymerization obviously in a dose dependent manner. They all induced cell apoptosis. Western blotting assay was used to analyse the change of regulatory protein Cyclin B1, Cdc2, p-Cdc2and a-tubuliin、β-tubulin at the level of protein. The results showed that CA4and40B raised the expression of CyclinB1in MCF-7cells with concentration dependence and down regulated Cdc2and p-Cdc2, but there was no effects on the expression of α-tubulinin、β-tubulin.To sum up,40B and CA4can target tubule and inhibit tubulin polymerization. They up-regulated Cyclin B1and down-regulated Cdc2and p-Cdc2in MCF-7lead to retardation in G2/M phases, which result in inhibiting tumor cell proliferation and cell apoptosis.The reach provides an important basis for40B effective safety evaluation on animal anti-tumors experiment.