Cytochrome C Oxidase in Malignant Gliomas

描述（申请人提供）：多形性胶质母细胞瘤（GBM）是最常见的原发性颅内肿瘤，其致死率几乎一致。替莫唑胺（TMZ）是治疗GBM患者的标准护理，其中位生存期虽小但显著增加，为4周。然而，获得tmz化疗耐药是GBM患者有效治疗的主要障碍之一。尽管采用积极的治疗方法，90%的GBM患者仍会复发。造成这种不良结果的一个原因是耐药性的发展。耐药性的潜在机制仍未被很好地理解，因此挑战了开发更有效的策略和/或新的治疗方法来克服这种耐药性。线粒体（mt）处于能量产生和凋亡途径的十字路口。它们在癌症病因学中的作用是显著的；然而，关于mt功能和化学耐药的信息仍然不明确。细胞色素c氧化酶（CcO） （EC 1.9.3.1）是一种大型的mt跨膜蛋白复合物。它是呼吸电子传递链（ETC）的最后一个酶，将电子从细胞色素c （Cyt c）转移到分子氧（O2）。Cyt c和O2是CcO的主要底物，它们都直接参与内源性mt凋亡途径和缺氧。虽然迄今为止的大多数研究都集中在CcO参与mt氧化磷酸化，但CcO活性、CcO核编码亚基与GBM中凋亡抗性发展之间的因果关系尚不清楚，这也是本提案的主题。最近，我们证明了tmz耐药性的获得与培养胶质瘤细胞以及配对的原发性复发性GBM患者活检中CcO活性的显著增加相关。在接受TMZ放射治疗的原发性和复发性GBM患者标本中，CcO活性的相似改变的发现强调了该发现的临床意义以及TMZ在介导这些影响中的主要作用。此外，在tmz耐药细胞中，药理或遗传抑制CcO可恢复tmz诱导的细胞凋亡。本提案的目标是通过实现三个特定目标来验证CcO亚基4异构体1 （COX4-1）通过改变1)CcO组装/功能和2)细胞氧化还原稳态来驱动GBM化学耐药的假设：(1)确定COX4-1和2异构体对CcO组装/功能和对TMZ抗性的影响；(2)确定COX4-1和2异构体对细胞氧化还原状态和对TMZ抗性的影响；(3)在脑肿瘤患者活检中测定CcO活性和COX4-1和2异构体的表达。在完成这项研究后，我们期望对CcO在TMZ的凋亡反应中的作用有一个实质性的基础知识。我们的长期目标是将这些信息应用于开发基于cco的治疗方法，以及开发化疗耐药和GBM复发的预后标志物。

DESCRIPTION (provided by applicant): Glioblastoma multiforme (GBM) is the most common primary intracranial neoplasm and its almost uniform lethality. Temozolomide (TMZ) is the standard of care for the treatment of GBM patients showing small but significant increases in median survival of 4 weeks. However, acquisition of TMZ-chemoresistance is one of the major obstacles to efficient therapy for GBM patients. Despite aggressive treatment approaches, recurrence occurs in 90% of GBM patients. One cause of this poor outcome is development of a drug resistance. The mechanism underlying drug resistance is still not well understood thus challenging the development of more effective strategies and/or novel therapeutics to overcome this resistance. Mitochondria (mt) are at the cross road of energy production and apoptotic pathways. Their role in cancer etiology is significant; however, information regarding mt function and chemoresistance remains poorly defined. The enzyme cytochrome c oxidase (CcO) (EC 1.9.3.1) is a large mt transmembrane protein complex. It is the last enzyme in the respiratory electron transport chain (ETC) that transfers electrons from cytochrome c (Cyt c) to molecular oxygen (O2). Cyt c and O2 are the main substrates of CcO and both are directly involved in the intrinsic mt apoptotic pathway and hypoxia. While most of the studies to date have focus on CcO involvement in mt oxidative phosphorylation, any casual relation between CcO activity, CcO-nuclear-encoded subunits and the development of resistance to apoptosis in GBM is unclear and is the subject of this proposal. Recently, we demonstrated that acquisition of TMZ-resistance correlates with a significant increase of CcO activity in culture glioma cells as well a paired primary-recurrent GBM patient biopsies. The discovery of similar alterations in CcO activity between primary and recurrent human GBM specimens in patients subjected to TMZ-radiotherapy emphasizes the clinical significance of the findings and the primary role of TMZ in mediating these effects. Moreover, pharmacological or genetic inhibition of CcO in TMZ-resistant cells restores TMZ-induced apoptosis. The goal of this proposal is to test the hypothesis that CcO subunit 4 isoform 1 (COX4-1) drives chemoresistance in GBM through changes in 1) CcO assembly/ function and 2) cellular redox homeostasis by achieving three specific aims: We will determine whether: (1) Determine the effects of COX4-1 and 2 isoforms on the assembly/ function of CcO and on resistance to TMZ, (2) Determine the effects of COX4-1 and 2 isoforms on cellular redox status and on resistance to TMZ and 3) Determine the activity of CcO and expression of COX4-1 and 2 isoforms in brain tumor patient biopsies. Upon completion of this research, we expect to have developed a substantial base of knowledge on the role of CcO in the apoptotic response to TMZ. Our long term goal is to apply this information for the development of CcO-based therapies as well as for the development of prognostic markers for chemoresistance and GBM recurrence.