GnRH类似物/GnRHR调控糖酵解关键酶PK-M2抑制上皮性卵巢癌增殖的研究

Gonadotropin releasing hormone and its receptor (GnRH/GnRHR) are closely related to the proliferation of ovarian cancer. Our researches verified that GnRH analogues can inhibit ovarian cancer proliferation through PI3K/Akt pathway, GnRHR expression in stages I-II of ovarian cancer was obviously higher than in stages III-IV, and over expression of GnRHR can inhibit ovarian cancer proliferation. Glycolysis is the main path of energy acquisition of tumors. Pyruvate kinase M2 (PK-M2) is one of the key enzymes of glycolysis. The activity and acetylation of PK-M2 is closely related to tumorigenesis. Forkhead box O1 (FoxO1) can regulate differentiation, proliferation and DNA damage/repair of cells and influence glycometabolism. We suppose GnRH analogues/GnRHR can inhibit proliferation of ovarian cancer by PK-M2 through PI3K/Akt-FoxO1 pathway. We intend to construct GnRHRsiRNA and GnRHR over expression lentivirus transfection ovarian cancer cell lines and detect gene expression, promoter activity, isoenzyme activity and acetylation of PK-M2. Meanwhile, the proliferation of ovarian cancer cells will be detected. Second, we plan to use FoxO1siRNA and over expression techniques to further study the signal mechanism of GnRH analogues/GnRHR and PK-M2.

促性腺激素释放激素及其受体（GnRH/GnRHR）与卵巢癌增殖密切相关。课题组前期研究证明GnRH类似物通过PI3K/Akt通路抑制卵巢癌细胞增殖；GnRHR在I-II期上皮性卵巢癌中表达明显高于III-IV期；GnRHR过表达卵巢癌细胞增殖明显被抑制。糖酵解是肿瘤细胞获取能量的主要途径，糖酵解途径关键酶-丙酮酸激酶M2（PK-M2）的活性、乙酰化修饰与肿瘤发生发展密切相关。转录因子FoxO1调控细胞分化、增殖，参与DNA损伤/修复，并在糖代谢中起重要作用。推测GnRH类似物/GnRHR可能通过PI3K/Akt-FoxO1通路调控PK-M2，抑制卵巢癌细胞增殖。本项目通过构建GnRHR过表达和低表达卵巢癌细胞株，检测GnRHR对PK-M2基因表达、启动子活性、酶活性、乙酰化修饰以及卵巢癌细胞增殖的影响。通过干扰和过表达FoxO1,探讨GnRH类似物/GnRHR调控PK-M2的信号分子机制。

促性腺激素释放激素及其受体（GnRH/GnRHR）与卵巢癌增殖密切相关。课题组前期研究证明GnRH类似物通过PI3K/Akt通路抑制卵巢癌细胞增殖。糖酵解是肿瘤细胞获取能量的主要途径，糖酵解途径关键酶-丙酮酸激酶M2（PK-M2）与肿瘤发生发展密切相关。转录因子FoxO1调控细胞分化、增殖，参与DNA损伤/修复，并在糖代谢中起重要作用。推测GnRH/GnRHR可能通过PI3K/Akt-FoxO1通路调控PK-M2，影响卵巢癌细胞增殖等生物学行为。. 本项目通过cck-8实验及实时无标记细胞分析技术（RTCA）检测不同浓度GnRHa/GnRHant对上皮性卵巢癌株增殖的影响；通过流式细胞仪及凋亡小体实验检测GnRHa/GnRHant对上皮性卵巢癌株生长、增殖、凋亡的影响等细胞功能实验；以及western检测关键分子akt/p-akt,foxo1,pkm2的表达变化；并应用si-RNA及PCR-array芯片等实验进行机制探讨。另外进行了裸鼠动物实验，验证细胞学实验的结果。. 通过增殖相关RTCA实验，流式细胞仪及凋亡小体实验验证了GnRHa/GnRHant影响上皮性卵巢癌细胞株生长、增殖、凋亡。并通过western检测、人细胞凋亡PCR芯片证实了GnRHa/GnRHant对凋亡相关关键蛋白的影响。并在裸鼠皮下移植瘤实验中得到了证实。. 我们得出结论：促性腺激素释放激素及其受体通过PI3K/Akt-FoxO1途径影响上皮性卵巢癌生长、增殖、凋亡。本研究结果为GnRH／GnRHR、PK-M2、FOXO1做为评价卵巢癌生物学行为、治疗和预后的指标提供了线索。并对上皮性卵巢癌的激素相关发病机制、治疗机制进行了有意探索。