miR-137/YB-1调控肿瘤细胞干性参与卵巢癌耐药调节的机制研究

The residual cells after treatment were thought to be the root of recurrence and drug resistance for ovarian cancer. In our previous studies, we found that ovarian cancer cells reprogrammed to stem-like cells and downregulated miR-137 during the process of drug resistance inducement, moreover, the elevated miR-137 could induce the cancer stem cells (CSCs) to lose their characteristics and sensitive ovarian cancer cells to a variety of chemotherapy drugs, suggesting that miR-137 involved in the regulation of multidrug resistance (MDR) and biological characteristics of CSCs in ovarian cancer. On this basis, the current study is divided into the following four parts, ①In first part, expression of miR-137 will be assessed in ovarian carcinoma tissues, the relationship between the expression of miR-137 and chemotherapy resistance or clinicopathological factors will be studied. ②In the second part, the OCSCs will be isolated and identitied from the human ovarian cancer cell lines and specimens, the potential targets of miR-137 will be analyzed and identified, the role of miR-137 in the regulation of MDR and biological characteristics of CSCs will be explored by the algorithms for target gene prediction，the gene rescue experiments and so on. ③Furthermore, the molecular mechanism of the chemosensitivity and biological characteristics of CSCs modulation by miR-137 in ovarian cancer cells will be studied from the aspect of transcriptional regulation. ④Finally, to analysis the impact of miR-137 on drug resistance of ovarian cancer cells in vivo by establishment of ovarian carcinoma in nude mice tumor models. The study purposes to find effective targets to improve the efficacy of chemotherapy by overcoming MDR of ovarian cancer is of great significance.

治疗后残存的肿瘤干细胞是卵巢癌耐药、复发的根源。我们的前期研究发现，卵巢癌细胞诱导性耐药过程中伴有miR-137表达的下调及肿瘤细胞干性化，且过表达的miR-137可诱导肿瘤干细胞去干性化并增加卵巢癌细胞对多种化疗药物的敏感性，提示miR-137参与卵巢癌干性及耐药性调节。在此基础上，本课题拟进行以下四个方面的研究：一是从临床入手，分析miR-137表达水平与卵巢癌化疗疗效的相关性；二是从卵巢癌临床标本及细胞中分选并鉴定出卵巢癌干细胞，利用靶基因分析、基因拯救试验等从细胞水平上探讨miR-137对卵巢癌细胞干性及耐药性的影响；三是从基因转录调控环节研究miR-137调节卵巢癌细胞干性及耐药性的分子机制；四是建立卵巢癌裸鼠荷瘤模型，体内实验观察miR-137对卵巢癌细胞干性及化疗敏感性的调节作用。这一研究对于寻找有效靶点克服卵巢癌耐药，提高卵巢癌化疗疗效具有重要意义。

项目的背景：. 卵巢癌的病死率高居妇科恶性肿瘤之首，化疗过程中多药耐药的发生是其治疗失败的重要原因，化疗后残存的肿瘤干细胞是肿瘤耐药的始作俑者，由此，探索新的miRNA在卵巢癌细胞干性及耐药性调节中的作用与机制对寻找有效靶点逆转耐药、提高化疗疗效具有重要价值。.主要研究内容：.1. 筛选卵巢癌耐药相关miRNAs，评估miRNAs与化疗疗效、临床病理特征及预后的相关性；.2. 提取、分离卵巢癌干细胞，检测miRNA差异性表达对细胞干性与耐药性靶控点的调节作用与机制；.3. 建立体内耐药模型，探讨特异性miRNAs递送对卵巢癌耐药的预防与逆转作用。..重要结果与关键数据：.1. miRNAs与卵巢癌耐药的相关性分析. 收集卵巢癌组织标本65例，筛选耐药与敏感组织中显著差异表达的miRNAs（miR-137、miR-145、miR-186、miR-204），结合临床分析显示：这些miRNAs表达水平与卵巢癌化疗疗效、远期生存率正相关，与复发时间间隔负相关。.2. miRNAs对卵巢癌细胞耐药的调节作用与机制.1) miR-137在卵巢癌干细胞中显著下调，可通过miR-137/YB-1/MDR1/CD44通路介导卵巢癌细胞干性维护受阻、药物外排泵功能失活，进而增加肿瘤细胞对多种化疗药物的敏感性；.2) miR-145的表达下调与表观遗传修饰相关，外源性的miR-145可通过miR-145/CDK6/P-RB将肿瘤细胞阻滞于G0/G1期，亦可通过miR-145/SP1/P-gp将化疗药物富集胞内，增加药物的杀伤效应；.3) 卵巢癌细胞在体外诱导耐药过程中发生上皮间质转化，伴随miR-186表达下调，过表达的miR-186可靶向Twist1，逆转上皮间质化，并通过对p53的蛋白修饰作用促进卵巢癌细胞凋亡。.4) 由化疗损伤刺激分泌的IL-6与膜表面受体IL-6R结合，触发IL6-IL-6R/STAT3/miR-204正反馈环路，递进式活化卵巢癌细胞增殖通路，显著降低化疗药物的体内外药物敏感性。..科学意义：. 课题组探讨了miR-137、miR-186、miR-145、miR-204与卵巢癌耐药的相关性及其调节肿瘤耐药的作用与机制。这一研究丰富了卵巢癌耐药形成的机制，为寻求新的行之有效的耐药逆转与预防的策略奠定了理论与实验基础。

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