Redox Regulation of Multidrug Resistance Gene Expression

描述（由申请人提供）：本续期申请要求支持扩展氧化还原条件对耐药和肿瘤进展的调节研究。这项应用的长期目标是开发有效的化疗药物用于治疗人类恶性肿瘤，从而提高人类癌症的治愈率。这与国家癌症研究所的使命是一致的。先前资助年份的研究已经确定γ -谷氨酰半胱氨酸合成（γ - gcs）是谷胱甘肽（GSH）生物合成的限速酶，是细胞内氧化还原条件的重要调节因子。γ - gcs的一个重亚基（γ - gcsh）的过表达减少了细胞内氧化还原条件，因此，改变了一系列参与耐药（MDR1、MRP1、MRP3和HCtr1）和肿瘤进展（MMP1、MMP3和MMP10）的基因的表达谱。因此，了解调节γ - gcs表达的机制在癌症药理学和肿瘤生物学中具有重要意义。尽管细胞毒性药物对γ - gcsh的转录调控已被广泛研究，但转录后调控尚未得到详细研究。我们最近观察到一种新的mRNA稳定性增强的转录后调控机制对各种氧化应激条件下γ - gcsh的上调很重要。这一更新应用在Specific Aim 1中提出，以阐明氧化还原调节的γ - gcsh mRNA稳定性是由MAPK途径介导的假设。特异性目标2中提出了验证MAPK通路也通过过表达γ - gcsh参与MMP表达下调的假设的方法。引人注目的是，γ - gcsh的表达升高与铜转运体hCtr1的上调有关，hCtr1将顺铂转运到培养细胞中。因此，我们建议在Specific Aim 3中研究不同氧化还原条件下hCtr1的调控机制。我们希望从这些研究中了解氧化还原条件影响药物敏感性的分子基础，通过调控各自的耐药基因表达。

DESCRIPTION (provided by applicant): This renewal application requests support to extend studies of the regulation of drug resistance and tumor progression by redox conditions. The long-term objectives of this application are to develop effective use of chemotherapeutic agents for the treatment of human malignancies, thereby improving the cure rates of human cancers. This is consistent with the mission of National Cancer Institute. Studies from the previous funding years have established that gamma-glutamylcysteine synthesis (gamma-GCS), which is the rate-limiting enzyme for the biosynthesis of glutathione (GSH), is an important regulator of intracellular redox conditions. Overexpression of a heavy subunit of gamma-GCS (gamma-GCSh) reduces intracellular redox conditions, as a consequence, changes the expression profile of a whole host of genes involved in drug resistance (MDR1, MRP1, MRP3, and HCtr1) and tumor progression (MMP1, MMP3, and MMP10). Thus, understanding mechanisms that regulate gamma-GCS expression is important in cancer pharmacology and tumor biology. Whereas transcriptional regulation of gamma-GCSh by cytotoxic agents has been studied quite extensively, posttranscriptional regulation has not been investigated in detail. We recently observed that a novel posttranscriptional regulation mechanism of enhancement of mRNA stability is important for the upregulation of gamma-GCSh under various oxidative stress conditions. This renewal application proposes in Specific Aim 1 to elucidate the hypothesis that redox-regulated gamma-GCSh mRNA stability is mediated by the MAPK pathway. Approaches to testing the hypothesis that the MAPK pathway is also involved in the down regulation of MMP expression by overexpression of gamma-GCSh are proposed in Specific Aim 2. Strikingly, elevated expression of gamma-GCSh is associated with upregulation of the copper transporter hCtr1, which transport cisplatin into cultured cells. We therefore propose in Specific Aim 3 to investigate the regulation mechanism of hCtr1 under various redox conditions. We hope from these studies to learn the molecular bases of redox conditions that affect drug sensitivity through the regulation of their respective drug resistance gene expression.