Chemotherapy of malignant brain tumors is currently severely limited by both inherent and acquired resistance of the brain tumor cells to the chloroethylnitrosoureas (CENU)s, the most effective agents available for the treatment of brain tumors. In this project, the role of glutathione (GSH) and its related enzymes, GSH-transferase, glutathione reductase, gamma-glutamylcysteine synthetase and glutathione synthetase in mechanisms of resistance of human brain tumor cells to chloroethylnitrosoureas will be investigated. Primary brain tumor specimens will be used to establish early passage cell lines with graded inherent resistance to bischloroethylnitrosourea (BCNU), a clinically active CENU. An acquired resistance model will be developed by treating a BCNU-sensitive cell line in vitro with increasing doses of BCNU over several passages. GSH levels and the activities of GSH-metabolizing enzymes in these cells will be determined and correlated with the degree of development resistance to BCNU. Agents that are capable of depleting intracellular GSH and/or inhibiting GSH-relatined enzymes will be used to cause GSH-depletion and enzyme inhibition in BCNU- resistant cells. Dose-effect and kinetic studies will be performed to establish the optimum inhibitor concentration and treatment times to achieve maximum GSH-depletion and enzyme inhibition in brain tumor cells of graded degrees of BCNU-resistance. Based on these results, strategies will be developed with which resistant brain tumor cells will be """"""""re-sensitized"""""""" to killing by BCNU. The degree of conversion of resistant cells to sensitive ones will be estimated in a clonogenic cell assay and the increase in DNA interstrand crosslinking will be measured by alkaline elution techniques. Although the glutathione system is one of the most important cellular pathways to detoxify carcinogens, free radicals and electrophiles, its role in the CENU-resistance of brain tumor cells has until now been little investigated. The findings in this project, therefore, promise to contribute substantially to our understanding of GSH-dependent mechanisms involved in both the sensitivity and resistance of brain tumor cells to the chloroethylnitrosoureas, and thus, provide potential guidelines with which clinical strategies for overcoming this resistance may be developed.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29CA046410-02
Application #
3458585
Study Section
Experimental Therapeutics Subcommittee 2 (ET)
Project Start
1988-02-01
Project End
1993-01-31
Budget Start
1989-02-01
Budget End
1990-01-31
Support Year
2
Fiscal Year
1989
Total Cost
Indirect Cost
Name
University of Washington
Department
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Sriram, R; Ali-Osman, F (1993) Purification and biochemical characterization of gamma-glutamylcysteine synthetase from a human malignant astrocytoma cell line. Biochem Mol Biol Int 30:1053-60
Ali-Osman, F; Berger, M S; Rajagopal, S et al. (1993) Topoisomerase II inhibition and altered kinetics of formation and repair of nitrosourea and cisplatin-induced DNA interstrand cross-links and cytotoxicity in human glioblastoma cells. Cancer Res 53:5663-8
Silbergeld, D L; Ali-Osman, F; Winn, H R (1991) Induction of transformational changes in normal endothelial cells by cultured human astrocytoma cells. J Neurosurg 75:604-12
Ali-Osman, F; Srivenugopal, K; Berger, M S et al. (1990) DNA interstrand crosslinking and strand break repair in human glioma cell lines of varying [1,3-bis(2-chloroethyl)-1-nitrosourea] resistance. Anticancer Res 10:677-82
Sriram, R; Ali-Osman, F (1990) S1-nuclease enhancement of the ethidium bromide binding assay of drug-induced DNA interstrand crosslinking in human brain tumor cells. Anal Biochem 187:345-8
Srivenugopal, K S; Ali-Osman, F (1990) Stimulation and inhibition of 1,3-bis(2-chloroethyl)-1-nitrosourea-induced strand breaks and interstrand cross-linking in Col E1 plasmid deoxyribonucleic acid by polyamines and inorganic cations. Biochem Pharmacol 40:473-9
Ali-Osman, F; Stein, D E; Renwick, A (1990) Glutathione content and glutathione-S-transferase expression in 1,3-bis(2-chloroethyl)-1-nitrosourea-resistant human malignant astrocytoma cell lines. Cancer Res 50:6976-80
Ali-Osman, F; Caughlan, J; Gray, G S (1989) Decreased DNA interstrand cross-linking and cytotoxicity induced in human brain tumor cells by 1,3-bis(2-chloroethyl)-1-nitrosourea after in vitro reaction with glutathione. Cancer Res 49:5954-8
Ali-Osman, F (1989) Quenching of DNA cross-link precursors of chloroethylnitrosoureas and attenuation of DNA interstrand cross-linking by glutathione. Cancer Res 49:5258-61