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.
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