四逆汤联合Imatinib抑制乳腺癌耐药细胞生长的研究

The inhibitoty effect of Sini Decoction combined with imatinib on the growth of breast cancer resistant cells

目的以中医温阳法代表方四逆汤和c-Abl抑制剂Imatinib为研究对象,研究两药联用对氟维司群耐药乳腺癌细胞生长的影响,为临床中西医结合治疗氟维司群耐药乳腺癌提供实验依据.方法采用MTT法测定耐药细胞T47D/Ful对Fulvestrant的敏感性,用Western Blot测定ER、PR及HER2蛋白表达;用MTT法及CompuSyn软件确定四逆汤与Imatinib联合用药的最佳剂量及联合指数;用Annexin V-FITC/7-AAD双染法检测细胞凋亡,Western Blot测定Bid、Bcl-xl及Bcl-2蛋白表达,从分子水平研究细胞凋亡机制.结果氟维司群耐药细胞T47D/Ful对Fulvestrant敏感性显著下降,且ER、PR及HER2均为阴性表达.四逆汤与Imatinib能协同抑制耐药细胞T47D/Ful的生长(P<0.01,CI=0.865).流式细胞术检测凋亡细胞结果表明,四逆汤联合Imatinib相较于单独用药能显著诱导氟维司群耐药乳腺癌细胞凋亡.四逆汤联合Imatinib能上调促凋亡蛋白Bid、下调抗凋亡蛋白Bcl-xl和Bcl-2.结论四逆汤联合Imatinib能协同抑制氟维司群耐药乳腺癌细胞的生长,其机制可能是通过上调促凋亡蛋白Bid、下调抗凋亡蛋白Bcl-xl和Bcl-2诱导耐药细胞凋亡.

Objective To study the effect of the combination of Sini Decoction and imatinib,on the growth of fulvestrant-resistant breast cancer cells,and to provide experimental basis for clinical treatment of fulvestrant—resistant breast cancer with integrated traditional Chinese and Western medicine.Methods MTT assay was used to determine the sensibility of T47D/Ful to fulvestrant.Western Blot was used to detect the expression of ER,PR and HER2.The optimal dosage and combination index of Sini Decoction and imatinib in combination were determined by MTT assay and CompuSyn software.Apoptosis was detected by Annexin V—FITC/7—AAD double staining.Bid,Bel—xl and Bel—2 protein expression was determined by Western Blot in order to study the mechanism of apoptosis at the molecular level.Results The sensirivity of fulvestrant—resistant cells T47D/Ful to fulvestrant was significantly decreased,and ER,PR and HER2 were negative.The growth of drug—resistant cells T47D/Ful were synergistically inhibited by the dual treatment of Sini Decoction and imatinib(P<0.01,CI=0.865).Flow cytometry analysis of apoptotic cells showed that Sini Decoction combined with imatinib significantly induced apoptosis in fulvestrant—resistant breast cancer cells compared w让h the drug alone.The pro-apoptotic protein Bid was up—regulated and the anti—apoptotic proteins Bel—xl and Bel—2 were down—regulated with dual treatment of Sini decoction and imarinib.Conclusion The growth of fulvestrant—resistant breast cancer cells can be synergistically inhibited by Sini Decoction combined with imatinib.The mechanism may be to induce apoptosis of drug—resistant cells by up—regulating the pro—apoptotic protein Bid and down—regulating the anti—apoptotic proteins Bcl-xl and Bel—2.