This proposal will explore depletion of intracellular reduced glutathione (GSH) as a chemotherapeutic strategy for the treatment of human solid tumors. As an example, the role of GSH depletion in the antineoplastic action of N-methylformamide (NMF) is examined. NMF, a polar solvent currently in clinical trials, 1) is active against experimental human solid tumors in vivo, 2) can suppress the malignant phenotype (e.g. clonigenicity, tumorgenicity) of human colon carcinoma cells and 3) can decrease the expression of certain proto-oncogenes e.g. c-myc in these cells. The action of NMF involves a depletion of intracellular GSH as 1) the GSH precursor, 1-cysteine, restores the GSH content, growth, level of proto-oncogene expression, and malignant behavior of NMF-treated colon carcinoma cells and 2) other known GSH depleting agents e.g. buthionine sulfoximine, can duplicate the actions of NMF. This proposal will investigate the mechanism by which NMF depletes GSH, how this GSH depletion results in a decrease in proto-oncogene expression, and whether this suppression of proto-oncogene expression is responsible for the growth inhibition and loss of the malignant phenotype in NMF-treated human colon carcinoma cells, both in vitro and in vivo. It will also be examined if other, more potent, GSH depleting agents can be found, such as phorone (disopropylidene acetone) or other polar solvents structurally related to NMF, that will have the same actions as NMF but greater efficacy. Finally, the exploitability of the above-mentioned findings in the chemotherapy of colon carcinoma will be examined, including a) can glutathione repletion therapy using cysteine precursors reverse NMF-induced hepatotoxicity, b) can one take advantage of defects in cysteine metabolism that occur in certain colon tumors and selectively rescue normal tissues form the glutathione-depleting effects of NMF and c) can these polar solvents be used in combination with other antineoplastic agents, e.g., cis-platin, whose action may be compromised by high intracellular levels of glutathione. The proposed studies will provide 1) information on the mechanism of action of the experimental antitumor agent NMF, 2) insights into better ways of clinically utilizing NMF in the treatment of solid tumors, 3) a better understanding of colonic cell proliferation and differentiation and 4) a broader view of the biological roles of glutathione.
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