Rac1b-Snail1表达差异与上皮性卵巢癌顺铂获得性耐药的关系及机制研究

The main reason for failure in treating epithelial ovarian cancer is drug resistance. The impacts of chemotherapy drugs on EMT key regulatory factors are closely related with the acquired drug resistance.From our preliminary experiments,we found that, after neoadjuvant chemotherapy of cisplatin plus paclitaxel,the Rac1b expression was increased in surgical resection specimen of epithelial ovarian cancer. Cisplatin-resistant SKOV3 cells strengthened their clone formation efficiency,and consequently the ratio of CD44+CD117+ cells in nude mice transplant tumor was increased. Gene chip results showed that Rac1b-Snail1 expression was up-regulated in cisplatin-resistant SKOV3 cells. Taking into account the effect of Rac1b-Snail1 on the epithelial-mesenchymal transition (EMT) and proportion of cancer stem cells, we hypothesis that the direct connection between the Rac1b-snail1 signaling pathway and EMT,cancer stem cell,acquired resistance of epithelial ovarian cancer.We'd like to carry out the followed experiments to testify the hypothesis. We will explore the relationship between the variant expression of Rac1b and the EMT,the ratio of cancer stem cells.The relationship between expression of Rac1b of nude mice transplant tumor and the effect of cisplatin was observed.The relationship between the Rac1b-Snail1 and prognosis of patients with epithelial ovarian cancer treated by combination of cisplatin and paclitaxel was investigated.The results will provide a theoretical support for explanation of cisplatin resistance of patients with epithelial ovarian cancer and the therapy strategy of the disease.

耐药是上皮性卵巢癌治疗失败的最主要原因，化疗药物对EMT关键调控因子的影响与获得性耐药密切相关。我们预实验发现：TP方案新辅助化疗后上皮性卵巢癌标本Rac1b表达上调；耐顺铂SKOV3细胞克隆能力增强，其裸鼠移植瘤CD44+/CD117+细胞比例升高，基因芯片结果显示该细胞Rac1b-Snail1表达上调。结合Rac1b-Snail1通路与EMT发生、肿瘤干细胞比例的关系，我们推测：Rac1b-Snail1介导EMT发生、肿瘤干细胞形成，进而导致上皮性卵巢癌获得性耐药。为证实该推测，拟考察：①SKOV3细胞在顺铂诱导与shRNA干预条件下，Rac1b表达差异与EMT发生、肿瘤干细胞比例的关系；②裸鼠移植瘤中Rac1b表达水平与顺铂疗效关系；③临床样本Rac1b-Snail1与TP方案治疗上皮性卵巢癌患者预后的关系。该研究为揭示上皮性卵巢癌顺铂获得性耐药机制及临床方案的选择提供理论依据。

在临床上，上皮性卵巢癌的复发、转移和耐药高度相关。过往研究报道，化疗药物对 EMT 的关键调控因子的影响可能是耐药性产生的机制之一。Rac1作为Rho超家族的一员，被认为是增殖、迁移、粘附等多项细胞重要生命过程的分子开关，其下游转录因子Snail1是EMT的关键调控因子。因此，我们围绕Rac1-Snail1、EMT、顺铂获得性耐药开展研究。我们构建了耐顺铂SKOV3细胞株和Rac1基因敲降细胞株，并检测了它们在细胞粘附、侵袭、迁移、以及对顺铂的敏感性的差异。分子标记检测显示，上皮性卵巢癌细胞在产生顺铂耐药性的过程中，RAC1活化增多，激活了下游的Snail1，使癌细胞向间质样细胞转化。敲降RAC1后发现，耐药性的上皮癌细胞对铂类药物的敏感性在一定程度上得到了恢复，且癌细胞的由间质样向上皮样细胞转化，转移能力下降。克隆形成实验显示耐药性细胞增殖增强，且耐药细胞裸鼠移植瘤 CD44+/CD117+细胞比例升高。结合以上结果，我们推断：Rac1-Snail1 信号的激活介导 EMT 发生、肿瘤干细胞形成，进而导致上 皮性卵巢癌获得性耐药。此外，临床样本研究显示Rac1的活化水平与患者的耐药与预后存在相关性，本研究为揭示上皮性卵巢癌顺铂获得性耐药机制及临床方案的选择提供了理论依据。