EZH2通过调控CHEK1信号通路促进卵巢癌干细胞对顺铂耐药的机制研究

The chemotherapy resistance of ovarian cancer is the main reason of its recurrences and treatment failure. Recent research shows that cancer stem cells play a critical role in chemo-resistance and tumor recurrence, however, the specific mechanism is not clear. Our preliminary data showed that histone methyltransferase EZH2 could promote the cisplatin resistance of ovarian cancer stem cells. Moreover, we have successfully screened out the cell cycle checkpoint kinase 1 (CHEK1) as the potential downstream target gene of EZH2 in ovarian cancer stem cells by PCR array. Accordingly, we propose that EZH2 contributes to the chemo-resistance of ovarian cancer stem cells by regulating CHEK1. In this study, we will further explore the expression and function of EZH2 and CHEK1 in ovarian cancer stem cells under the cisplatin treatment. Then, we will clarify the specific mechanism in which EZH2 regulates CHEK1 pathway to result in cisplatin resistance. In addition, we will identify the clinical significance of EZH2 and CHEK1 in ovarian cancer and further explore whether the inhibitors of EZH2 and CHK1 combined with cisplatin were able to reverse the cisplatin resistance of ovarian cancer stem cells. The implementation of this study can elucidate the molecular mechanism of EZH2 in cisplatin resistance of ovarian cancer stem cells, and also provide a new strategy and theoretical basis for clinical treatment of platinum-resistant ovarian cancer.

卵巢癌的化疗耐药是其复发及治疗失败的主要原因。最新研究显示,肿瘤干细胞是肿瘤耐药及复发的根源，但其具体机制尚不明确。本研究前期结果发现：组蛋白甲基转移酶EZH2能够促进卵巢癌干细胞对顺铂的耐药性，进一步通过PCR芯片技术筛选出卵巢癌干细胞中EZH2发挥作用的潜在下游靶基因——细胞周期检查点激酶CHEK1。据此我们提出：EZH2很可能通过调控CHEK1促进卵巢癌干细胞的耐药。本研究将进一步从分子水平探讨顺铂作用下卵巢癌干细胞中EZH2和CHEK1的表达及功能关系，并阐明EZH2调控CHEK1通路影响耐药的具体机制。同时，研究EZH2及CHEK1在卵巢癌中的临床意义并探讨其抑制剂联合顺铂用药能否逆转卵巢癌干细胞的耐药。本研究的实施可以阐明EZH2 参与卵巢癌干细胞耐药的分子机制，并为临床治疗顺铂耐药性卵巢癌提供新的策略和理论依据。

背景：卵巢癌是死亡率最高的妇科恶性肿瘤。卵巢癌的化疗耐药是其复发及治疗失败的主要原因。最新研究显示：卵巢癌干细胞 （OCSC）是肿瘤耐药及复发的根源。组蛋白甲基转移酶 EZH2在维持CSC中起着关键作用。本研究旨在探讨EZH2调控CSCs导致卵巢癌的耐药及复发的具体机制。 .方法：我们采用裸鼠模型获得了富含OCSC的细胞系，命名为SK-3rd细胞。 使用CRISPR和Cas9核酸内切酶系统建立敲除EZH2基因的SK-3rd卵巢癌细胞系。通过高通量PCR芯片和生物信息学方法筛选出EZH2的下游靶基因。荧光素酶报告基因测定和染色质免疫沉淀测定证实了EZH2对CHK1的激活作用。最后我们通过免疫组化的方法检测了 EZH2 / CHK1在卵巢癌患者组织中的表达与卵巢癌患者的化学耐药性和预后的相关性。 .结果：本研究证实了EZH2在维持卵巢癌干细胞干性和化学抗性中起着至关重要的作用。 CHK1是EZH2调控卵巢癌干细胞的下游靶基因。卵巢癌干细胞中EZH2的下调能够降低CHK1的表达，而CHK1的过表达能够逆转下调EZH2对肿瘤干细胞球形体形成能力以及耐药性的抑制作用。此外，本研究还证实了EZH2 通过与CHK1启动子结合激活而不是抑制CHK1信号的独特作用。在上皮性卵巢癌患者在临床样本中，EZH2和CHK1高表达患者不仅对铂类化疗具有更强的耐药性，而且预后也更差。.结论：本研究首次证实了EZH2对卵巢癌干细胞调控的新机制，即通过激活CHK1信号通路，促进卵巢癌干细胞的耐药，从而促进卵巢癌患者的化疗耐药和复发。为临床治疗顺铂耐药性卵巢癌提供新的策略和理论依据。