DataSheet_1_Evaluation of mitochondrial biogenesis and ROS generation in high-grade serous ovarian cancer.pdf

IntroductionOvarian cancer is one of the leading causes of death for women with cancer worldwide. Energy requirements for tumor growth in epithelial high-grade serous ovarian cancer (HGSOC) are fulfilled by a combination of aerobic glycolysis and oxidative phosphorylation (OXPHOS). Although reduced OXPHOS activity has emerged as one of the significant contributors to tumor aggressiveness and chemoresistance, up-regulation of mitochondrial antioxidant capacity is required for matrix detachment and colonization into the peritoneal cavity to form malignant ascites in HGSOC patients. However, limited information is available about the mitochondrial biogenesis regulating OXPHOS capacity and generation of mitochondrial reactive oxygen species (mtROS) in HGSOC.
MethodsTo evaluate the modulation of OXPHOS in HGSOC tumor samples and ovarian cancer cell lines, we performed proteomic analyses of proteins involved in mitochondrial energy metabolism and biogenesis and formation of mtROS by immunoblotting and flow cytometry, respectively.
Results and discussionWe determined that the increased steady-state expression levels of mitochondrial- and nuclear-encoded OXPHOS subunits were associated with increased mitochondrial biogenesis in HGSOC tumors and ovarian cancer cell lines. The more prominent increase in MT-COII expression was in agreement with significant increase in mitochondrial translation factors, TUFM and DARS2. On the other hand, the ovarian cancer cell lines with reduced OXPHOS subunit expression and mitochondrial translation generated the highest levels of mtROS and significantly reduced SOD2 expression. Evaluation of mitochondrial biogenesis suggested that therapies directed against mitochondrial targets, such as those involved in transcription and translation machineries, should be considered in addition to the conventional chemotherapies in HGSOC treatment.

引言
卵巢癌是全球女性癌症相关死亡的主要病因之一。上皮性高级别浆液性卵巢癌（epithelial high-grade serous ovarian cancer, HGSOC）的肿瘤生长能量需求需通过有氧糖酵解（aerobic glycolysis）与氧化磷酸化（oxidative phosphorylation, OXPHOS）协同满足。尽管OXPHOS活性降低已被证实是肿瘤侵袭性增强与化疗耐药的重要诱因之一，但HGSOC患者发生基质脱落并定植于腹膜腔形成恶性腹水时，需要上调线粒体抗氧化能力。然而目前关于调控OXPHOS能力的线粒体生物发生（mitochondrial biogenesis），以及HGSOC中线粒体活性氧（mitochondrial reactive oxygen species, mtROS）生成的相关研究信息仍较为有限。
方法
为评估HGSOC肿瘤样本与卵巢癌细胞系中OXPHOS的调控模式，本研究分别通过免疫印迹法（immunoblotting）对线粒体能量代谢与生物发生相关蛋白进行蛋白质组学分析，并借助流式细胞术（flow cytometry）检测mtROS的生成情况。
结果与讨论
本研究发现，线粒体编码与细胞核编码的OXPHOS亚基稳态表达水平上调，与HGSOC肿瘤及卵巢癌细胞系中线粒体生物发生增强呈显著正相关。其中MT-COII（线粒体编码细胞色素c氧化酶亚基II）的表达上调更为显著，这与线粒体翻译因子TUFM和DARS2的表达水平显著升高相一致。另一方面，OXPHOS亚基表达与线粒体翻译能力均降低的卵巢癌细胞系，其mtROS生成水平最高，同时超氧化物歧化酶2（superoxide dismutase 2, SOD2）的表达显著下调。对线粒体生物发生的评估表明，除常规化疗方案外，靶向线粒体靶点（如参与线粒体转录与翻译机器的靶点）的治疗策略，有望应用于HGSOC的临床治疗中。