Anticancer Drug Resistance by Bcl-2 Oncogene Expression

描述（由申请人提供）：本研究项目的目标一直是并仍然是确定Bcl-2癌基因表达异常的肿瘤细胞对癌症化疗药物耐药的机制。Bcl-2是蛋白质家族的原型成员，其主要功能似乎是调节细胞凋亡。Bcl-2在多种人类癌症中高度表达，并通过在肿瘤进展期间阻断细胞凋亡而为肿瘤提供生长优势。然而，这种阻断也使这些肿瘤细胞对抗癌疗法具有抗性，包括依赖于细胞凋亡作为其主要细胞杀伤模式的大多数癌症化疗药物。事实上，Bcl-2表达产生多药耐药（MDR）表型。Bcl-2调节细胞凋亡的机制尚不清楚。我们以前已经表明，Bcl-2表达细胞具有较高的细胞内谷胱甘肽（GSH）水平。正如我们在本申请中所证明的，来自表达Bcl-2的细胞的线粒体也具有更高的GSH水平，并且具有增强的将GSH从胞质溶胶主动转运到线粒体中的能力。这些新的发现与Bcl-2抑制线粒体凋亡的能力完全一致;我们的论点是，由于Bcl-2表达的MDR可以通过抑制或逆转Bcl-2对线粒体GSH水平的增强来逆转。因此，在本申请中，具体目的1和2旨在表征Bcl-2对线粒体GSH转运的这种作用，并测试线粒体GSH水平直接调节线粒体凋亡的假设。在目标3中，我们建议跟踪一个新的线索，并调查Bcl-2蛋白是否在细胞收到凋亡信号后重新分布到质膜。最后，我们在具体目标4中提出，通过使用能够使GSH从细胞或线粒体流出的药物来研究克服Bcl-2介导的MDR的策略。本课题的研究意义涉及肿瘤治疗中的MDR问题。我们的中心假设是，了解Bcl-2阻断抗癌药物诱导的细胞凋亡的能力的生物化学和分子基础将导致逆转这种耐药性和增强治疗效果的策略的发展。

DESCRIPTION (provided by applicant): The goal of this research project has been and remains to define the mechanism responsible for the resistance of tumor cells with abnormal expression of the Bcl-2 oncogene to cancer chemotherapy agents. Bcl-2 is the prototypic member of a family of proteins, whose main function appears to be the regulation of apoptosis. Bcl-2 is highly expressed in a variety of human cancers and provides a growth advantage to tumors by blocking apoptosis during tumor progression. However, this block also renders such tumor cells resistant to anticancer therapies including most cancer chemotherapy drugs that rely on apoptosis as their primary mode of cell killing. Indeed, Bcl-2 expression produces a multi-drug resistance (MDR) phenotype. The mechanism by which Bcl-2 regulates apoptosis is not understood. We have previously shown that Bcl-2 expressing cells have higher intracellular levels of glutathione (GSH). As we demonstrate in this application, the mitochondria from Bcl-2 expressing cells also have higher GSH levels and have enhanced capacity to actively transport GSH from the cytosol into mitochondria. These are novel findings that are entirely consistent with the ability of Bcl-2 to suppress mitochondrial apoptosis; it is our contention that MDR due to Bcl-2 expression could be reversed by inhibiting or reversing Bcl-2's enhancement of mitochondrial GSH levels. Therefore, in this application, Specific Aims 1 and 2 are directed at characterizing this effect of Bcl-2 on mitochondrial GSH transport and testing the hypothesis that the level of mitochondrial GSH directly regulates mitochondrial apoptosis. In Aim 3, we propose to follow up on a new lead and investigate whether Bcl-2 protein is redistributed to the plasma membrane after cells receive an apoptotic signal. Finally, we propose in Specific Aim 4 to investigate strategies for overcoming Bcl-2 mediated MDR through the use of drugs that enable GSH efflux from cells or mitochondria. The significance of this project relates to the problem of MDR in cancer therapeutics. It is our central hypothesis that understanding the biochemical and molecular basis for Bcl-2's ability to block anticancer-drug induced apoptosis will lead to the development of strategies for reversing this drug resistance and enhancing therapeutic effects.