miR-21 过表达与非小细胞肺癌中 EGFR-TKI 的获得性耐药相关。
MiR-21 overexpression is associated with acquired resistance of EGFR-TKI in non-small cell lung cancer.
机构信息
Tongji University Medical School Cancer Institute, Shanghai 200433, People's Republic of China.
Department of Medical Oncology, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai 200433, People's Republic of China.
出版信息
Lung Cancer. 2014 Feb;83(2):146-53. doi: 10.1016/j.lungcan.2013.11.003. Epub 2013 Nov 13.
BACKGROUND AND PURPOSE
With the increasing use of epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI) in patients with advanced non-small cell lung cancer (NSCLC), its acquired resistance has become a major clinical problem. Recent studies revealed that miR-21 was involved into the resistance of cytotoxic agents. The aim of this study was to investigate its role in the acquired resistance of NSCLC to EGFR-TKI.
METHODS
EGFR-TKI-sensitive human lung adenocarcinoma cell line PC9 and the acquired resistant cell line, PC9R, were used. Lentiviral vectors were used to infect PC9 or PC9R to regulate the miR-21 expression. The expression of targeted proteins PTEN and PDCD4 was controlled by RNA interference. MicroRNA array, RT-PCR and TaqMan MicroRNA Assays were used to detect miR-21 expression. The MTT and Annexin V assays were used to determine proliferation and apoptosis. Western Blot and immunohistochemistry were used to analyze target protein expression (PTEN, PDCD4, Akt, p-Akt). We also constructed PC9R xenograft tumor model to observe the relationship between miR-21 and EGFR-TKI resistance in vivo and validated it in the clinical serum specimens of NSCLC patients treated with EGFR-TKI.
RESULT
MiR-21 was overexpressed in the EGFR-TKI resistant cell line PC9R relative to PC9. The level of miR-21 was reversely correlated with the expression of PTEN and PDCD4 and positive correlated with PI3K/Akt pathway. Inhibiting miR-21 with lentivirus vector induces apoptosis in PC9R cell line and inhibiting miR-21with ASO suppressed tumor growth in nude mice treated with EGFR-TKI. Furthermore, serum miR-21 expression in NSCLC patients treated with EGFR-TKI was significantly higher at the time of acquiring resistance than at baseline (p<0.01).
CONCLUSION
miR-21 is involved in acquired resistance of EGFR-TKI in NSCLC, which is mediated by down-regulating PTEN and PDCD4 and activating PI3K/Akt pathway.
背景与目的
随着表皮生长因子受体酪氨酸激酶抑制剂（EGFR TKI）在晚期非小细胞肺癌（NSCLC）患者中的应用日益增多，其获得性耐药已成为一个主要的临床问题。最近的研究表明，miR-21 参与了细胞毒药物的耐药性。本研究旨在探讨其在 NSCLC 对 EGFR-TKI 获得性耐药中的作用。
方法
使用 EGFR-TKI 敏感的人肺腺癌细胞系 PC9 和获得性耐药细胞系 PC9R。慢病毒载体用于感染 PC9 或 PC9R 以调节 miR-21 的表达。RNA 干扰控制靶向蛋白 PTEN 和 PDCD4 的表达。采用微阵列、RT-PCR 和 TaqMan 微 RNA 检测 miR-21 的表达。MTT 和 Annexin V 检测增殖和凋亡。Western Blot 和免疫组化分析靶蛋白表达（PTEN、PDCD4、Akt、p-Akt）。我们还构建了 PC9R 异种移植肿瘤模型，以观察体内 miR-21 与 EGFR-TKI 耐药的关系，并在接受 EGFR-TKI 治疗的 NSCLC 患者的临床血清标本中验证。
结果
在 EGFR-TKI 耐药细胞系 PC9R 中，miR-21 的表达高于 PC9。miR-21 的水平与 PTEN 和 PDCD4 的表达呈负相关，与 PI3K/Akt 通路呈正相关。用慢病毒载体抑制 miR-21 可诱导 PC9R 细胞系凋亡，用 ASO 抑制 miR-21 可抑制接受 EGFR-TKI 治疗的裸鼠肿瘤生长。此外，接受 EGFR-TKI 治疗的 NSCLC 患者在获得耐药时的血清 miR-21 表达明显高于基线时（p<0.01）。
结论
miR-21 参与了 NSCLC 中 EGFR-TKI 的获得性耐药，其机制是通过下调 PTEN 和 PDCD4 并激活 PI3K/Akt 通路。
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