曲妥珠单抗联合NSC 74859 对曲妥珠耐药的乳腺癌细胞的抑制作用研究

目的:观察曲妥珠单抗(Trastuzumab)与转录信号转导子与激活子3蛋白(STAT3)抑制剂NSC 74859联用对曲妥珠耐药细胞株SK-BR-3R的生长抑制作用及机理研究。方法:采用四甲基偶氮唑蓝(MTT)法鉴定曲妥珠耐药的SK-BR-3R细胞株并检测曲妥珠单药处理、NSC 74859单药处理以及两药联用处理对SK-BR-3R细胞的生长抑制程度。建立SK-BR-3R的皮下肿瘤模型,观察两药联用对肿瘤生长的抑制效果;通过免疫印迹(Western Blot)实验检测SK-BR-3R细胞中磷酸化HER2(p-HER2),磷酸化STAT3(p-STAT3)及磷酸化AKT(p-AKT)的水平。结果:当曲妥珠浓度在50 nmol/L及NSC 74859的浓度在50μmol/L联用时,较之两药单用显示了显著的抑制效果,其差异具有统计学意义;进一步在建立的SK-BR-3R小鼠肿瘤模型中观察到了曲妥珠联合NSC74859治疗组显示了比曲妥珠或NSC 74859单独使用时更显著的抑瘤效果。最后,免疫印迹实验显示了曲妥珠和NSC74859联合处理显著降低了SK-BR-3R细胞的HER2,STAT3及AKT的磷酸化水平。结论:曲妥珠单抗联合NSC 74859使用可显著抑制曲妥珠耐药的乳腺癌细胞SK-BR-3R的生长,其机制可能是药物协同抑制了对肿瘤生长重要的PI3K/AKT信号通路。本研究可为临床上治疗曲妥珠耐药的乳腺癌提供参考。

Study in the Effects of Trastuzumab and NSC78459 on Inhibiting Trastuzumab-resistant Human Breast Cancer Cells

Objective:To evaluate the growth inhibitory effects of of trastuzumab-resistant SK-BR-3R cells with combination of trastuzumab and NSC 74859 and further explore the mechanisminvolved.Methods:Trastuzumab-resistant SK-BR-3 cells was characterized and cell proliferation of SK-BR-3R upon treatment with trastuzumab, NSC 74859 or trastuzumab plus NSC 74859 were evaluated by MTT assay. Subsequently, nude mice bearing subcutaneous SK-BR-3R cells xenograft tumor model was set up to evaluate the tumor growth inhibition upon combinational treatment. Additionally, the levels of phosphorylated HER2, STAT3 and AKT were detected by western blot assay.Results:Trastuzumab-resistant SK-BR-3 cell line were obtained and characterized by MTT assay. Next, Trastuzumab (50 nmol/L) plus NSC 74859 (50 umol/L) exhibited significant inhibitory effect compared with either agent alone by MTT assay, showing statistical difference; Furthermore, combinatorial treatment with trastuzumab plus NSC 74859 resulted in a significant benefit over either agent alone in SK-BR-3R xenograft models. Finally, combined treatment of trastuzumab and NSC 74859 suppressed HER2, STAT3 and AKT phosphorylation in SK-BR-3R cells.Conclusion:Combination treatment of trastuzumab with NSC 74859 markedly inhibited the growth of SK-BR-3R cells. Suppression of PI3K/AKT pathway in SK-BR-3R cells could explain the inhibitory effects upon combinational treatment. To conclude, this study will provide a new regimen for treating the trastuzumab-resistant breast cancer in clinics.