Buthionine sulfoximine induction of gamma-L-glutamyl-L-cysteine synthetase gene expression, kinetics of glutathione depletion and resynthesis, and modulation of carmustine-induced DNA-DNA cross-linking and cytotoxicity in human glioma cells.
Glutathione (GSH) depletion by buthioninine sulfoximine (BSO) is being explored clinically as a means of enhancing the efficacy of cancer chemotherapy. We investigated the kinetics of GSH depletion and altered gamma-L-glutamyl-L-cysteine synthetase (gamma-GC-S) gene expression in two human malignant glioma cell lines, HBT5 and HBT28, and examined how these relate to GSH resynthesis and changes in DNA interstrand cross-link induction and cytotoxicity of 1,3-bis(2-chloroethyl)-nitrosourea (BCNU). GSH content was 54 and 126 nmol/mg/protein in HBT 5 and HBT 28, respectively, and after a 24-hr exposure to 100 microM BSO was decreased by 95% in HBT 5 and 91% in HBT 28. Basal gamma-GC-S enzyme activity in HBT 28 was twice that in HBT 5, and steady state gamma-GC-S gene transcripts were 2.6-fold higher in HBT 28 than in HBT 5, with no apparent amplification or rearrangement of the gene in either cell line. BSO exposure (100 microM) for 24 hr increased gamma-GC-S gene transcripts by 1.7-fold in HBT 5 and 2.8-fold in HBT 28. After BSO removal, the rate of GSH resynthesis in HBT 28 was twice that in HBT 5. Continuous BSO exposure increased the level of BCNU-induced DNA interstrand cross-links, and cytotoxicity was significantly higher in cells exposed continuously to BSO than in cells with only a 24-hr BSO preexposure. This increase was, however, greater in HBT 28 than in HBT 5. These findings indicate significant heterogeneity in the effects of BSO on gamma-GC-S gene expression and in the ability of BSO to sensitize tumors and cell lines to BCNU. The data also suggest that by preventing GSH resynthesis, a greater level of cytotoxicity is achieved with continuous BSO exposure than with BSO preexposure alone.
Ali-Osman
F
F
Department of Experimental Pediatrics, University of Texas M.D. Anderson Cancer Center, Houston, 77030, USA. ali-osman@utmdacc.mda.uth.tmc.edu
Antoun
G
G
Wang
H
H
Rajagopal
S
S
Gagucas
E
E
D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D001483:Base Sequence; D019328:Buthionine Sulfoximine; D002330:Carmustine / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D005910:Glioma / Q000473:pathology; D005721:Glutamate-Cysteine Ligase / Q000235:genetics*; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008717:Methionine Sulfoximine / Q000031:analogs & derivatives* / Q000494:pharmacology; D008969:Molecular Sequence Data
D018906:Antineoplastic Agents, Alkylating; D008717:Methionine Sulfoximine; D019328:Buthionine Sulfoximine; D004247:DNA; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D002330:Carmustine