Synthesis and anticancer activity of betulin-succinate derivatives in vitro and in the zebrafish xenograft tumor model.

Betulin is a naturally occurring triterpene holding great promise as a lead for developing new antitumor agents. Given that the design, synthesis and anticancer activity of betulin-succinate derivatives remain unexplored, we herein reported the synthesis and cytotoxicity evaluation of such 22 new derivatives against A172, HT29, SW579, and TCA8113 cancer cell lines in vitro using the CCK8 (Cell Counting Kit-8) assay. The results revealed that most compounds showed potent antitumor activity relative to the positive control and betulin. Particularly, compound 21 also exhibited superior activities in inducing the early apoptosis of HT29 cells and blocking the cell cycle in the G2 phase than the chemotherapeutic agent cisplatin. In vitro kinase screening results from the 207 tested kinases demonstrated that MARK2 (Microtubule Affinity-Regulating Kinase 2), significantly inhibited by compound 21, may be the potential target of betulin-succinate derivatives. Hence, the affinity and interactions between compound 21 and MARK2 were obtained by molecular docking simulations. Compound 21, which prevented tumor cell metastasis and induced tumor cell apoptosis, exhibited potent antitumor activity in a zebrafish tumor model with xenografted HT29 cells. Furthermore, the histopathological evaluation demonstrated the ability of compound 21 to reduce the proliferation and improve the atypia of tumor cells in the abdominal cavity of zebrafish.