Ju Lixia, Han Mingquan, Li Xuefei, Zhao Chao
Department of Integrative Medicine, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Tongji University, Shanghai, People's Republic of China.
Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University Medical School Cancer Institute, Tongji University, Shanghai, People's Republic of China.
J Cancer. 2017 May 11;8(7):1311-1318. doi: 10.7150/jca.17817. eCollection 2017.
The findings of EGFR mutations and the development of targeted therapies have significantly improved the overall survival of lung cancer patients. Still, the prognosis remains poor, so we need to know more about the genetic alterations in lung cancer. MicroRNAs are dysregulated in lung cancer, and some of them can regulate EGFR. So it is very important to predict the candidate microRNAs that target mutated EGFR and to investigate the role of these candidate microRNAs in lung cancer. In this study, we investigated the difference of microRNAs expression between lung adenocarcinoma cell lines with EGFR exon 19 deletion (H1650 and PC9) and wild-type (H1299 and A549) using the Phalanx Human Whole Genome Microarray. Then the expression of individual microRNAs was validated by qRT-PCR assays. Moreover, we detected the microRNAs expression in plasma of lung adenocarcinoma patients with EGFR exon 19 deletion and wild-type. Lastly, we explored the function of the positive microRNA in EGFR tyrosine kinase inhibitors (EGFR-TKIs ) resistance using MTT and Annexin V-APC assays. The expression of 1,732 microRNAs was evaluated, and we found that microRNAs expression was different between these two groups. Hsa-miR-141-3p, hsa-miR-200c-3p, hsa-miR-203, hsa-miR-3182, hsa-miR-934 were up-regulated and hsa-miR-3196 was down-regulated in the EGFR exon 19 deletion group compared with wild-type group. The detection of circulating microRNAs showed that miR-3196 was down-regulated in lung adenocarcinoma patients with EGFR exon 19 deletion compared with wild-type. And then the MTT assay results showed that miR-3196 had no effect on the sensitivity of erlotinib. The results of apoptosis analysis showed that inhibition of miR-3196 and erlotinib induced more apoptosis in H1299 cells than erlotinib alone, and overexpressed miR-3196 and erlotinib induced less apoptosis in PC9 cells than erlotinib alone (P<0.05). It is suggested that microRNAs associate with EGFR exon 19 deletion and miR-3196 may be further explored as a potential predictor and targeted biomarker when it is difficult to get the tumors.
表皮生长因子受体（EGFR）突变的发现以及靶向治疗的发展显著提高了肺癌患者的总生存率。尽管如此，其预后仍然较差，因此我们需要更多地了解肺癌中的基因改变。微小RNA（miRNA）在肺癌中表达失调，其中一些可以调节EGFR。因此，预测靶向突变型EGFR的候选miRNA并研究这些候选miRNA在肺癌中的作用非常重要。在本研究中，我们使用方阵人全基因组微阵列研究了具有EGFR外显子19缺失的肺腺癌细胞系（H1650和PC9）与野生型细胞系（H1299和A549）之间miRNA表达的差异。然后通过qRT-PCR分析验证了单个miRNA的表达。此外，我们检测了具有EGFR外显子19缺失和野生型的肺腺癌患者血浆中的miRNA表达。最后，我们使用MTT和膜联蛋白V-APC分析探讨了阳性miRNA在EGFR酪氨酸激酶抑制剂（EGFR-TKIs）耐药中的作用。评估了1732种miRNA的表达，我们发现这两组之间miRNA表达存在差异。与野生型组相比，EGFR外显子19缺失组中hsa-miR-141-3p、hsa-miR-200c-3p、hsa-miR-203、hsa-miR-3182、hsa-miR-934上调，hsa-miR-3196下调。循环miRNA检测显示，与野生型相比，具有EGFR外显子19缺失的肺腺癌患者中miR-3196下调。然后MTT分析结果显示，miR-3196对厄洛替尼的敏感性没有影响。细胞凋亡分析结果显示，抑制miR-3196和厄洛替尼诱导H1299细胞凋亡比单独使用厄洛替尼更多，而过表达miR-3196和厄洛替尼诱导PC9细胞凋亡比单独使用厄洛替尼更少（P<0.05）。这表明miRNA与EGFR外显子19缺失有关，当难以获取肿瘤时，miR-3196可能作为一种潜在的预测指标和靶向生物标志物被进一步探索。
