PI3K/Akt信号通路活化在曲妥珠单抗耐药机制意义的研究

目的探讨磷脂酰肌醇-3-激酶/丝苏氨酸蛋白激酶(PI3K/Akt)信号转导通路激活是曲妥珠单抗耐药的重要靶点之一,为乳腺癌曲妥珠单抗耐药的靶向治疗提供理论基础。方法对人乳腺癌细胞株BT474连续处理建立了耐曲妥珠单抗的耐药亚株BT-HerR,FISH法对耐药细胞株BT-HerR做Her-2表型分析,MTT法检测曲妥珠单抗对BT474和BT-HerR细胞的体外增殖抑制情况,流式细胞仪检测曲妥珠单抗干预后细胞的凋亡变化,PI3K/Akt抑制剂LY294002干预细胞后Western blot检测p-Akt表达。结果耐药细胞株BT-HerR Her-2基因表达为强阳性;曲妥珠单抗干预细胞72 h后,细胞的体外增殖受到抑制且随着浓度的升高而增强;经曲妥珠单抗处理后比较BT474与BT-HerR细胞凋亡率,差异具有显著性(P<0.01);曲妥珠单抗仅能抑制BT474的Akt蛋白磷酸化,LY294002则能同时抑制BT474的BT-HerR Akt蛋白磷酸化。结论曲妥珠单抗耐药细胞Akt蛋白磷酸化活化,PI3K/Akt抑制剂LY294002能明显抑制曲妥珠单抗耐药细胞Akt蛋白磷酸化,PI3K/Akt信号转导通路与曲妥珠单抗耐药存在明确相关性。

Research on significance of PI3K/Akt signaling pathway activation in drug resistance mechanism of trastuzumab

Aim T discuss the activation of signal transduction pathway of phosphatidylinositol 3′-kinase /serine-threonine kinase(PI3K/Akt) as one of the important targets of drug resistance of trastuzumab,which provides theoretical basis for the targeted therapy of the drug resistance of trastuzumab in breast cancer.Methods The drug-resistance sub-strain BT-HerR of trastuzumab was established for the continuous treatment of human breast cancer cell strain BT474,Her-2 phenotype analysis was conducted on the drug-resistance cell strain BT-HerR with FISH method,the proliferation inhibition(in vitro) of trastuzumab to BT474 and BT-HerR cells was deteced with MTT method,the apoptosis variation after in terference of trastuzumab was detected with flow cytometry and p-Akt and apoptosis-related protein expression was detected with Western blot after PI3K/Akt inhibitor LY294002 interfered the cells.Results The gene expression of drug-resistance cell strain BT-HerR Her-2 was strongly positive;Seventy-two hours after interference of trastuzumab,the proliferation in vitro of cells was inhibited,which was strengthened with the increase of concentration;after treatment of trastuzumab,in terms of the cell apoptotic rate of BT474 and BT-HerR,there was a significant difference(P<0.01);trastuzumab could only inhibit the Akt protein phosphorylation of BT474, while LY294002 could inhibit the BT-HerR Akt protein phosphorylation of BT474 simultaneously.Conclusion Akt protein phosphorylation of trastuzumab drug-resistance cell is activated;LY294002,PI3K/Akt inhibitor,can obviously inhibit Akt protein phosphorylation of trastuzumab drug-resistance cell and there exists a clear relevancy between PI3K/Akt signal transduction pathway and drug resistance of trastuzumab.