Design,synthesis And Antitumor Activity Of CA-4 Derivatives Coupled With Aminophosphonate Derivatives

Combretastatin A-4 can kill cancer cells by inhibiting the polymerization of tubulin in cells,and has good activity on a variety of cancer cells.However,the clinical application of CA-4 is limited because of low oral bioavailability and high toxicity.Studies have found that aminophosphonate and its derivatives have a variety of biological activities,such as antibacterial activity,enzyme inhibitory activity,anti-cancer and anti-virus.The introduction of aminophosphonate into anticancer drugs can increase the biological activity of drugs.Sixteen CA-4derivatives were designed and synthesized including 9a-9h and 10a-10h.Structures of molecules were confirmed by ¹H NMR and ¹³C NMR.The in vitro inhibitory effect of sixteen CA-4 derivatives were evaluated against Hep G-2,HT29,A549,MGC-803,LO2 normal cells line by MTT method.The results showed that all of the compounds exhibited good cytotoxicity and that the anticancer activity of compound 9e was better than that of CA-4;the toxicity of CA-4 to LO2 cells was higher(IC₅₀value was only 3.27μM)and the toxicity of 16 compounds to LO2 cells was lower(IC₅₀value was more than 10μM except 9f).The anticancer activities of compounds 9b,9c and 9d substituted by halogen at C-3 position of benzene ring were similar to those of compounds 9a,and the activities of compounds 9g and 9h substituted by halogen at C-4 position of benzene ring were significantly increased,especially the activities of compounds 9e and 9f substituted by-OCH₃and-CH₃at C-4 position of benzene ring were better within compounds 9a-9h.The same situation existed within compounds 10a-10h.it was notable that compound 9e exhibited the best antitumor activity with IC₅₀values respectively of 0.36μM,0.31μM,0.19μM and 0.42μM for four kinds of cancer cells,and had less toxicity to LO2 cells(IC₅₀value of10.45μM).Related mechanism of representative compound 9e was conducted by Molecular Docking,Cell cycle,Apoptosis,ROS determination and Western Blotting.CA-4 was used as the positive control and A549 cells as the target.Then we found that the compounds tested could induced apoptosis through endogenous pathway.Compound 9e can arrest mitosis in G2 phase by binding to tubulin in cancer cells and can induce mitochondrial apoptosis by regulating the increase of Bax and caspase-3 content and the decrease of Bcl-2 content in mitochondriait;The excessive ROS production of cells is caused by the stimulation of compound9e which leads to the increase of the contents of CHOP,phospho-e IF2a and phospho-PERK proteins,which affects the normal function of endoplasmic reticulum;In addition,the increase of p53 content in cells was stimulated by compound 9e and the apoptosis was induced finally.