二甲双胍通过激活AMPK通路增强PARP抑制剂治疗同源重组修复正常型卵巢癌的有效性及其分子机制研究

Epithelial ovarian cancer is one of the most lethal gynecological malignancies worldwide, largely due to the advanced stage at diagnosis. While PARP inhibitor has potent anti-tumor activity against advanced-stage ovarian cancer, it has little effect on homologous recombination repair (HRR) proficient tumors. We have previously shown that PI3K inhibition sensitizes ovarian cancer and endometrial cancer cells to PARP inhibitor Olaparib (Gynecological Oncology 2016, Oncogene 2018). More recently, we found that combined use of Metformin and Olaparib led to exacerbated DNA damage and synergistically inhibited the growth and survival of ovarian cancer cells regardless of BRCA1/2 status. Herein, we generated a genetically engineered mouse model of ovarian cancer with competent HRR ability. Metformin is able to activate AMPK, an upstream kinase for both TSC1/TSC2 (negative regulators of mTOR activity) and ACC. In the current project, we proposed to investigate if inactivation of mTOR or ACC signaling resulted from Metformin-induced AMPK activation may enhance the anti-tumor effect of PARP inhibitors in the treatment of HRR proficient ovarian cancer. We will also integrate additional preclinical models, including ovarian cancer cell lines, patient-derived xenografts, ex vivo explant culture model as well as organoid models, to evaluate the therapeutic effects of Metformin and PARP inhibitors in combination. The goal of our project is to provide a new therapeutic strategy for HRR proficient ovarian caners meanwhile expanding the utility of PARP inhibitors to a broader spectrum of ovarian cancer.

上皮性卵巢癌恶性程度高，预后差。PARP抑制剂对携带BRCA1/2基因突变或DNA同源重组修复（HRR）缺陷型卵巢癌疗效显著，而目前临床对HRR正常型卵巢癌尚无有效靶向治疗方案。申请人前期研究表明抑制PI3K/AKT/mTOR通路显著增强癌细胞对PARP抑制剂的敏感性；我们近期还发现：二甲双胍激活AMPK，与PARP抑制剂联用导致细胞内DNA损伤加剧并协同抑制卵巢癌细胞生长与存活。由此推测：二甲双胍可能通过激活AMPK抑制其底物ACC活性或下游mTOR通路，增强PARP抑制剂的抗癌作用。为此，本项目拟用HRR正常型卵巢癌细胞系与已成功构建的卵巢癌遗传工程小鼠模型，深入探寻二甲双胍联合PARP抑制剂产生协同致死效应的分子机制；结合活组织切片离体培养、卵巢癌类器官和PDX等临床前模型加以验证，同时探寻监测疗效的分子标记物，为建立基于PARP抑制剂治疗HRR正常型卵巢癌的联合靶向方案提供新思路。

随着PARP抑制剂的临床应用，卵巢癌患者预后得到极大改善，然而PARP抑制剂耐药频繁发生。因此深入探究影响PARP抑制剂疗效的关键因子，探寻有效联药方案，对于拓展PARP抑制剂的治疗应用、改善患者预后意义重大。本项目成功建立PtenLkb1共缺失型卵巢癌遗传工程小鼠模型，可产生中、低分化浆液性卵巢癌并伴有血样腹水及转移，较真实的模拟人卵巢癌病理特征，为卵巢癌治疗研究提供重要临床前模型。研究发现，二甲双胍通过抑制线粒体呼吸链复合物I降低NAD+水平，从而增强PARP抑制剂与PARP1催化结构域结合，提高其捕获PARP1的能力，加剧DNA断裂损伤和细胞凋亡，降低卵巢癌细胞克隆形成和3D成球生长能力，有效提高PARP抑制剂治疗卵巢癌的疗效；重要的是，联药显著抑制小鼠卵巢癌生长，减少血样腹水产生，并延长小鼠的生存期。研究还发现，PARP抑制剂与CDK4/6抑制剂联用有效杀伤c-MYC高表达型卵巢癌，且c-MYC水平可预测联药抗癌疗效，该研究有望为c-MYC高表达型肿瘤患者提供一种全新治疗策略，为拓展PARP抑制剂临床应用范围奠定理论依据(EbioMedicine, 2019)。研究还发现：靶向SGK1与褪黑素联用通过破坏癌细胞ROS稳态协同抗肿瘤，为SGK1作为抗癌靶点开辟新思路(Redox biology，2019)；EphB4 通过激活GAB1/SHP2/ERK信号轴介导HER2治疗耐药（Cancer Letters, 2020）；褪黑素可通过诱导内质网应激与未折叠蛋白反应或内吞介导的HER2蛋白降解，增强HER2靶向治疗敏感性（Oncogene, 2021; Cancer Letters, 2021），揭示褪黑素提高HER2靶向疗效及克服耐药中的潜在应用。本项目阐明多种肿瘤靶向治疗敏感性与耐药分子机制，提出多项基于PARP抑制剂的抗肿瘤联药方案，同时为拓展已批准药物的临床应用提供一定理论依据。