Novel Glutathione-Dependent Thiopurine Prodrugs: Evidence for Enhanced Cytotoxicity in Tumor Cells and for Decreased Bone Marrow Toxicity in Mice
- 1Environmental Toxicology Center (S.G., A.A.E.) and 2Department of Comparative Biosciences (S.G., M.R., A.A.E.), University of Wisconsin, Madison, Wisconsin
- Dr. Adnan A. Elfarra, Department of Comparative Biosciences, School of Veterinary Medicine, 2015 Linden Dr., University of Wisconsin—Madison, Madison, WI 53706. E-mail: elfarraa{at}svm.vetmed.wisc.edu
Abstract
Elevated glutathione (GSH) levels have been detected in many tumors compared with the healthy, surrounding tissue. Often, this GSH up-regulation is associated with drug resistance. The prodrugs 6-(2-acetylvinylthio)guanine (AVTG) and 6-(2-acetylvinylthio)purine (AVTP) contain a novel butenone moiety that allows the prodrugs to react selectively with sulfhydryl nucleophiles to release the chemotherapeutic drug 6-thioguanine (6-TG) or 6-mercaptopurine (6-MP), respectively. The cellular uptake and metabolism oftrans-AVTG in two human renal carcinoma cell lines that were used as models were rapid and associated with depletion of intracellular GSH. Formation of 6-TG from trans-AVTG correlated positively with intracellular GSH concentrations, and was significantly reduced by diethyl maleate pretreatment. Intracellular concentrations of 6-TG after incubations with trans-AVTG were significantly higher than the 6-TG concentrations obtained after incubations with equimolar concentrations of 6-TG; thus, the prodrug delivered more 6-TG to the cell than did 6-TG itself. Cytotoxicity studies demonstrated that AVTG and AVTP had similar IC50values that were comparable with those of 6-TG, but were significantly lower than those of 6-MP. Furthermore, after in vivo treatment of mice with the prodrugs, no reduction was observed in circulating white blood cell counts, whereas white blood cell counts of mice treated with equimolar or 60% lower doses of 6-TG were reduced by 50 to 60%. Collectively, the results show that AVTG and AVTP are novel potential chemotherapeutic agents that may target tumors with up-regulated levels of GSH, and may exhibit less systemic toxicity than the parent thiopurines.
Footnotes
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This work was supported in part by Grant DK44295 from the National Institute of Diabetes, Digestive, and Kidney Diseases. Preliminary results from this work were presented at the joint meeting of the American Society for Biochemistry and Molecular Biology, and the American Society for Pharmacology and Experimental Therapeutics in Boston, MA, June 2000.
- Abbreviations:
- GSH
- glutathione (reduced form)
- GST
- glutathione S-transferase
- GR
- glutathione reductase
- 6-MP
- 6-mercaptopurine
- PG
- S-(9H-purin-6-yl)glutathione
- 6-TG
- 6-thioguanine
- PTA
- 2-(9H-purin-6-ylthio)acrylic acid
- AVTG
- 6-(2-acetylvinylthio)guanine
- AVTP
- 6-(2-acetylvinylthio)purine
- RCC
- renal cell carcinoma
- AZA
- azathioprine, 6-[(1-methyl-4-nitro-5-imidazolyl)thio]purine
- PBO
- trans-4-phenyl-3-buten-2-one
- DMSO
- dimethyl sulfoxide
- DEM
- diethyl maleate
- GSSG
- glutathione (oxidized form)
- DTNB
- 5,5′-dithio-bis(2-nitrobenzoic acid)
- SSA
- sulfosalicylic acid
- TFA
- trifluoroacetic acid
- HPLC
- high-pressure liquid chromatography
- HBSS
- Hanks' balanced salt solution
- MTT
- 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide
- AUC
- area under the curve
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- Received August 31, 2001.
- Accepted December 6, 2001.
- The American Society for Pharmacology and Experimental Therapeutics
