日蟾蜍它灵通过HSP90/突变p53途径抑制三阴性乳腺癌细胞增殖

Gamabufotalin Inhibits the Proliferation of Triple Negative Breast Cancer Cells Through HSP90/mutant p53 Pathway

摘要

三阴性乳腺癌(triple negative breast cancer, TNBC)恶性程度高, 预后差, 其p53基因的突变频率达80%以上。日蟾蜍它灵(gamabufotalin, CS-6)是日本蟾蜍的皮肤分泌物, 有研究表明CS-6具有抗癌作用, 但其对TNBC的作用和机制尚无报道。本文采用MTT法和细胞克隆形成实验检测了不同浓度CS-6对TNBC细胞系MDA-MB-468和MDA-MB-231细胞增殖的影响; 采用Western-blot检测了CS-6在外源和内源水平对p53和热休克蛋白90 (heat shock protein 90, HSP90)表达水平的影响; 采用联合指数及等效线分析法考察了CS-6和HSP90的抑制剂坦螺旋霉素(tanespimycin, 17-AAG)对MDA-MB-468和MDA-MB-231细胞增殖的协同作用。结果显示, CS-6明显抑制MDA-MB-468和MDA-MB-231细胞的增殖, 其抑制增殖机制有可能与降解HSP90/突变p53有关; 而且, CS-6和HSP90抑制剂17-AAG联合用药对抑制MDA-MB-468和MDA-MB-231细胞增殖具有协同作用。

Abstract

Triple negative breast cancer (TNBC) is a highly malignant tumor with a poor prognosis, and the mutation frequency of p53 gene in TNBC patients is more than 80%. Gamabufotalin (CS-6), a skin secretion from Japanese common toad (Bufo japonicus), has been shown to have anticancer effects. However, the role and underlying mechanism of CS-6 against TNBC have not been reported. Herein, the effects of different concentrations of CS-6 on the proliferation of TNBC cell lines MDA-MB-468 and MDA-MB-231 were de-tected by MTT assay and clone formation assay. The effects of CS-6 on the expression of p53 and heat shock protein 90 (HSP90) at exogenous and endogenous levels were detected by Western-blot. The synergistic effect of CS-6 and the HSP90 inhibitor tanespimycin (17-AAG) on the proliferation of MDA-MB-468 and MDA-MB-231 cells was investigated by combination index and equivalent linearization analysis. The results showed that CS-6 significantly inhibited the proliferation of MDA-MB-468 and MDA-MB-231 cells. The mechanism of inhibition of the tumor cell proliferation may be related to degradation of HSP90/mutant p53 protein. In addition, CS-6 and HSP90 inhibitor 17-AAG had a synergistic inhibitory effect on the prolifera-tion of MDA-MB-468 and MDA-MB-231 cells.

关键词

日蟾蜍它灵(CS-6) / 三阴性乳腺癌(TNBC) / p53蛋白 / 热休克蛋白90 (HSP90) / 联合用药

Key words

gamabufotalin (CS-6) / triple negative breast cancer (TNBC) / p53 protein / heat shock protein 90 (HSP90) / drug combination