整合素β_1信号通路参与非小细胞肺癌吉非替尼获得性耐药

Integrin β_1 participates in acquired resistance to gefitinib in non-small cell lung cancer

目的探讨整合素β1及其下游信号转导通路在非小细胞肺癌表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKI)吉非替尼获得性耐药中的作用。方法以人肺腺癌细胞株PC-9和吉非替尼耐药株PC-9/G作为研究对象,免疫印迹分析检测整合素β1、Akt、磷酸化Akt蛋白的表达;用MTT法检测吉非替尼和(或)磷脂酰肌醇3激酶(PI3K)抑制剂LY294002、细胞外调节蛋白激酶(ERK)抑制剂PD98059对细胞增殖的影响;用AnnexinⅤ/PI试剂盒和TUNEL试剂盒检测细胞凋亡。结果吉非替尼耐药株PC-9/G高表达整合素β1,RNA干扰抑制整合素β1表达能够抑制PC-9/G细胞的生长和促进凋亡。PC-9/G细胞中吉非替尼对Akt磷酸化的抑制作用弱于PC-9细胞,RNA干扰抑制整合素β1表达后Akt的磷酸化水平降低。ERK抑制剂PD98059不能恢复PC-9/G细胞对吉非替尼的敏感性,PI3K抑制剂LY294002能恢复PC-9/G细胞对吉非替尼的敏感性。结论整合素β1过表达可以通过PI3K途径激活下游信号分子,这可能是一种重要的EGFR-TKI耐药机制。

Objective To explore the role of integrin β1 and relevant signaling pathway in acquired resistance to epidermal growth factor receptor（EGFR） tyrosine kinase inhibitor（TKI） gefitinib in non-small cell lung cancer（NSCLC）. Methods Human lung adenocarcinoma cell line PC-9 and gefitinib-resistant PC-9/G cell lines were used in the present study.Western blotting analysis was used to examine the expression of integrin β1,Akt and phospho-Akt protein.The inhibitory effects of gefitinib and/or phosphoinositide 3-kinase（PI3K） inhibitor LY294002 and extracellular regulated protein kinases（ERK） inhibitor PD98059 on cellular proliferation were tested by MTT assay.Cell apoptosis was analyzed by Annexin Ⅴ/PI and TUNEL method.Results Overexpression of integrin β1 was observed in PC-9/G cell line.Silencing integrin β1 by RNAi method inhibited the proliferation and promoted apoptosis of PC-9/G cells.The inhibitory effect of gefitinib against Akt phosphorylation in PC-9/G cells was weaker than that in PC-9 cells.Knockdown of integrin β1 with RNAi decreased the phosphorylation level of Akt.ERK inhibitor PD98059 failed to restore the sensitivity of PC-9/G cells to gefitinib.PI3K inhibitor LY294002 could restore the sensitivity of PC-9/G cells to gefitinib.Conclusion It is suggested that overexpressed integrin β1 can activate the downstream signaling pathways through PI3K,which may be an important mechanism for resistance to EGFR-TKIs.