miR-181b/Notch2调控肿瘤干细胞样特性介导非小细胞肺癌化疗耐药机制研究

Drug resistance is the main reason for the failure of chemotherapy in cancer. The drug-resistant tumor cells have cancer stem cell-like properties. It has been confirmed that miR181b enhanced chemosensitivity, inhibited the forming of tumor spheres and reduced the percentage of CD133+ cells in non-small cell lung cancer. Luciferase report experiments have proved that miR-181b can bind Notch2, which is one of the stem cell signal molecule. Therefore, we propose that miR-181b/Notch2 may mediate chemoresistance through regulating cancer stem cell-like properties. But its regulating mechanism has not been fully elucidated. In cancer cell line, nude mice and non-small cell lung cancer tissues, we’ll further reveal its effect on chemosensitivity by knock-down or over-expression miR-181b/Notch2, and detect its effect on cancer stem cell-like properties by using stem cell suspension culture and flow cytometry. qPCR and Western blot will be used to identify the molecule of Notch2 signaling pathway regulated by miR-181b. This project will provide new theory and molecular target for clinical reversing chemoresistance in non-small cell lung cancer.

耐药是肿瘤化疗失败的主要原因之一，且肿瘤耐药细胞具有肿瘤干细胞样特性。我们前期实验证实miR-181b能增加非小细胞肺癌化疗药物敏感性；抑制细胞成球能力；降低CD133+细胞比例；且miR-181b能靶向结合干细胞信号分子Notch2。因此我们推测miR-181b/Notch2在调控肿瘤干细胞样特性介导非小细胞肺癌化疗耐药中发挥重要作用。但其作用机制尚待阐明。本项目拟从细胞、裸鼠及人体组织标本着手，通过沉默或过表达miR-181b/Notch2等关键分子，观察其对非小细胞肺癌化疗药物敏感性的影响；利用干细胞悬浮培养，流式细胞技术等实验证明miR-181b/Notch2等关键分子对肿瘤干细胞样特性的调控作用；通过qPCR、Western blot等实验进一步证实 miR-181b对Notch2下游信号传导通路分子活性的调控。本项目将为临床逆转非小细胞肺癌化疗耐药提供新的理论依据和分子靶点。

肿瘤干细胞样特性与肿瘤耐药密切相关。miRNA通过靶向调控靶基因的表达及功能，参与调控肿瘤的发生、发展等生物学过程。然而，miRNA在调控肿瘤干细胞样特性及药物耐药方面的机制还有待进一步阐明。miR-181b是一个抑制性miRNA，我们前期研究证实其与非小细胞肺癌的耐药有关。在本研究中，体外细胞实验证实miR-181b可以抑制NSCLC细胞系肿瘤干细胞样特性，增加细胞对顺铂的药物敏感性。荧光素酶实验证实miR-181b与Notch2有靶向结合作用。同时miR-181b进一步调控Notch2/Hes1信号通路。此外，裸鼠实验证实，外源性注射miR-181b显著抑制肿瘤的生长。临床组织标本实验证实，低miR-181b表达及高Notch2的患者OS较低，与NSCLC患者的肿瘤干细胞样特性有关。本研究证实miR-181b具有调控肿瘤干细胞样特性，通过调控Notch2及其下游信号转导通路调控肿瘤对化疗药物的敏感性。