Tetrandrine: a potent abrogator of G2 checkpoint function in tumor cells and its mechanism.
OBJECTIVE
To assess the ability of tetrandrine (Tet) to enhance the sensitivity to irradiation and its mechanism in cell lines of human breast cancer p53-mutant MCF-7/ADR, p53-wild-type MCF-7 and human colon carcinoma p53-mutant HT-29 as well as in C26 colorectal carcinoma-bearing BALB/c mice.


METHODS
MCF-7/ADR, HT-29 and MCF-7 cells were exposed to irradiation in the absence or presence of tetrandrine. The effect of Tet on the cytotoxicity of X-irradiation in these three cells was determined and the effect of tetrandrine on cell cycle arrest induced by irradiation in its absence or presence was studied by flow cytometry. Moreover, mitotic index measurement determined mitosis of cells to enter mitosis. Western blotting was employed to detect cyclin B 1 and Cdc2 proteins in extracts from irradiated or non-irradiated cells of MCF-7/ADR, HT-29 and MCF-7 treated with tetrandrine at various concentrations. Tumor growth delay assay was conducted to determine the radio-sensitization of tetrandrine in vivo.


RESULTS
Clonogenic assay showed that tetrandrine markedly enhanced the lethal effect of X-rays on p53-mutant MCF-7/ADR and HT-29 cells and the sensitization enhancement ratio (SER) of tetrandrine was 1.51 and 1.63, but its SER was only 1.1 in p53-wt MCF-7 cells. Irradiated p53-mutant MCF-7/ADR and HT-29 cells were only arrested in G2/M phase while MCF-7 cells were arrested in G1 and G2/M phases. Radiation-induced G2 phase arrests were abrogated by tetrandrine in a concentration-dependent manner in MCF-7/ADR and HT-29 cells, whereas redistribution within MCF-7 cell cycle changed slightly. The proportion of cells in M phase increased from 1.3% to 14.7% in MCF-7/ADR cells, and from 1.5% to 13.2% in HT-29 cells, but 2.4% to 7.1% in MCF-7 cells. Furthermore, the levels of cyclin B 1 and Cdc2 expression decreased after X-irradiation in MCF-7/ADR and HT-29 cells, and the mitotic index was also lower. Tet could reverse the decrease and induce the irradiated cells to enter mitosis (M phase). Endosomatic experiment showed that tetrandrine caused tumor growth delay in irradiated mice.


CONCLUSIONS
Tetrandrine boosts the cell killing activity of irradiation both in vitro and in vivo. Tetrandrine is a potent abrogator for G2 checkpoint control and can sensitize the cells to radiation.
Sun
Xin-Chen
XC
Cancer Center of Zhongda Hospital, Southeast University, Nanjing 210009, Jiangsu, China. xunxch505@yahoo.com.cn
Cheng
Hong-Yan
HY
Deng
Yu-Xia
YX
Shao
Rong-Guang
RG
Ma
Jun
J
D000818:Animals; D044182:Benzylisoquinolines / Q000494:pharmacology*; D042846:CDC2-CDC28 Kinases / Q000378:metabolism; D045744:Cell Line, Tumor; D019926:Cyclin B / Q000378:metabolism; D056744:Cyclin B1; D004354:Drug Screening Assays, Antitumor; D016195:G2 Phase / Q000187:drug effects*; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011836:Radiation Tolerance
D044182:Benzylisoquinolines; C534699:CCNB1 protein, human; C534700:Ccnb1 protein, mouse; D019926:Cyclin B; D056744:Cyclin B1; C009438:tetrandrine; D042846:CDC2-CDC28 Kinases