REDOX REGULATION OF MULTIDRUG RESISTANCE GENE EXPESSION

描述：（申请人的摘要）最近的研究已经证明， 由MRP编码的多药耐药相关蛋白参与了 含有谷胱甘肽（GSH）结合物的底物的转运。 这些 研究结果表明，细胞内GSH水平和γ- 谷氨酰半胱氨酸合成酶（γ-GCS），从头开始的限速酶 在MRP介导的耐药性中，谷胱甘肽生物合成的抑制剂是重要的。 的确， 申请人已经证明了MRP和γ-GCS的协调表达， 许多独立建立的人类耐药肿瘤细胞系， 培养的细胞短暂暴露于细胞毒性剂（重金属， 烷化剂、酚类促氧化剂和一氧化氮），以及在人类中， 结肠直肠肿瘤活检。 因为已知这些药剂中有许多会产生 氧化应激和他的初步研究结果表明，过度表达 抗氧化剂GSH下调MRP，本申请的中心主题是 氧化应激是控制基因表达的潜在机制 MRP和γ-GCS 为了检验这一假设，申请人在本申请中提出， 应用：（i）确定抗氧化剂GSH在协调 使用诱导型表达系统来上调这些基因的表达， GSH水平和抑制剂下调水平，（ii）识别和 表征DNA序列和转录因子，参与 通过促氧化剂和抗氧化剂上调和下调MRP，（iii） 结肠癌细胞MRP表达调控研究 由一氧化氮信号传导产生的亚硝化应激，以及（iv）建立 结肠直肠癌中MRP和γ-GCS表达的动物模型， 因为氧化应激与结直肠癌发生的关系 被牵连。 申请人期望从这些研究中了解 控制多药耐药基因表达的分子机制 癌细胞，从而提高癌症化疗的功效。

DESCRIPTION: (Applicant's Abstract) Recent studies have demonstrated that the multidrug resistance-associated protein encoded by the MRP is involved in the transport of substrates containing glutathione (GSH) conjugates. These findings suggest that intracellular GSH levels and activities of gamma- glutamylcysteine synthetase (gamma-GCS), the rate-limiting enzyme for de novo biosynthesis of GSH, are important in MRP-mediated drug resistance. Indeed, the applicant has demonstrated coordinated expression of MRP and gamma-GCS in many independently established human drug-resistant tumor cell lines, in cultured cells transiently exposed to cytotoxic agents (heavy metals, alkylating agents, phenolic prooxidants and nitric oxide), and in human colorectal tumor biopsies. Because many of these agents are known to generate oxidative stress and his preliminary findings showed that overexpression of antioxidant GSH downregulates MRP, the central theme of this application is that oxidative stress is the underlying mechanism that controls the expression of MRP and gamma-GCS. To test this hypothesis, the applicant proposes in this application: (i) to determine the roles of antioxidant GSH in the coordinated regulation of these genes using inducible expression system to up-regulate the GSH levels and depletors to down-regulate the levels, (ii) to identify and characterize DNA sequences and transcription factors that are involved in the up- and down-regulation of MRP by prooxidants and antioxidants, (iii) to investigate regulation of MRP expression in colon cancer cells under nitrosative stress generated by nitric oxide signaling, and (iv) to establish animal models for the expression of MRP and gamma-GCS in colorectal cancers, because association of oxidative stress with colorectal carcinogenesis has been implicated. The applicant anticipates from these studies to learn the molecular mechanisms that control multidrug resistance gene expression in cancer cells, and thus to improve the efficacy of cancer chemotherapy.