Axl调控的miRNA表达在NSCLC吉非替尼获得性耐药中的意义

Axl-modulated miRNA regulates gefitinib-resistance in lung cancer

目的:探讨酪氨酸激酶Axl调控的miRNA在非小细胞肺癌(NSCLC)吉非替尼获得性耐药中的意义。方法:比较表皮生长因子受体(EGFR)突变型NSCLC细胞株HCC827及其吉非替尼耐药株HCC827-Gef中Axl mRNA[实时荧光定量PCR(qRT-PCR)]和蛋白(Western blot)的表达,再利用微阵列分析筛选出20个差异表达miRNA,其中miR-374a、miR-548b敏感度最高;采用qRT-PCR检测Axl沉默对HCC827-Gef细胞miR-374a、miR-548b表达的影响;对不同Axl mRNA表达水平的NSCLC患者(共40例)miR-374a、miR-548b表达进行比较并进行Kaplan-Meier法生存分析。结果:HCC827-Gef细胞中Axl mRNA及蛋白的表达均高于HCC827细胞(t=9.393,24.038;P<0.001)。与转染空质粒的HCC827-Gef细胞比较,HCC827-Gef-esiA XL细胞中Axl mRNA、miR-374a表达下降,miR-548b表达则上调(t=22.155,56.000,16.000;P<0.001)。与Axl低表达的NSCLC患者比较,Axl mRNA高表达者癌组织中miR-374a表达升高,miR-548b表达降低(t=5.231,2.473;P<0.05),无病生存期缩短(χ2=6.550,P=0.011)。结论:Axl及其调控的miR-374a和miR-548b在NSCLC吉非替尼获得性耐药中起重要作用,可作为NSCLC预后标记或潜在的治疗靶点。

Aim：To explore the significance of tyrosine kinase Axl-modulated miRNA in gefitinib-resistance for non-small cell lung cancer（NSCLC）.Methods：The expressions of Axl mRNA and protein in NSCLC cell lines HCC 827 with epidermal growth factor receptor mutation , and its gefitinib-resistant lung cancer cell HCC 827-Gef were detected by qRT-PCR and Western blot .Twenty differential expression miRNA were scaned by microarray analysis , and among which ,miR-374a and miR-548b were proved the most sensitive .qRT-PCR was used to detected the miR-374a and miR-548b expres-sions in HCC827-Gef silenced by esiAxl .The expressions of miR-374 a and miR-548 b in NSCLC patients with different Axl mRNA expression levels were compared , and survival analysis was performed by Kaplan-Meier method .Results：The ex-pressions of Axl mRNA and protein were significantly higher in HCC 827-Gef than those in HCC827（t=9.393,24.038;P〈0.001）.Down-regulated expression of miR-374a and up-regulated expression of miR-548b were detected in HCC827-Gef after Axl gene was knocked down （t=56.000,16.000;P〈0.001）.The patients with high Axl mRNA expression level had significantly higher expression of miR-374a and lower miR-548b（t=5.231,2.473;P〈0.05）,and shorter disease-free sur-vival time compared with patients with low Axl mRNA expression level （χ2 =6.550,P=0.011）.Conclusion：Axl and its modulated miR-374 a and miR-548 b may play important roles in gefitinib-resistance and may serve as a prognostic biomarker or potential therapeutic target for NSCLC .