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. 1995 May 15;308(Pt 1):283–290. doi: 10.1042/bj3080283

Glutathione analogues as novel inhibitors of rat and human glutathione S-transferase isoenzymes, as well as of glutathione conjugation in isolated rat hepatocytes and in the rat in vivo.

S Ouwerkerk-Mahadevan 1, J H van Boom 1, M C Dreef-Tromp 1, J H Ploemen 1, D J Meyer 1, G J Mulder 1
PMCID: PMC1136874  PMID: 7755575

Abstract

Inhibitors of rat and human Alpha- and Mu-class glutathione S-transferases that effectively inhibit the glutathione (GSH) conjugation of bromosulphophthalein in the rat liver cytosolic fraction, isolated rat hepatocytes and in the rat liver in vivo have been developed. The GSH analogue (R)-5-carboxy-2-gamma-(S)-glutamylamino-N-hexylpentamide [Adang, Brussee, van der Gen and Mulder (1991) J. Biol. Chem. 266, 830-836] was used as the lead compound. To obtain more potent inhibitors, it was modified by replacement of the N-hexyl moiety by N-2-heptyl and by esterification of the 5-carboxy group with ethyl and dodecyl groups. In isolated hepatocytes, the branched N-2-heptyl derivatives were stronger inhibitors of GSH conjugation of bromosulphophthalein than the N-hexyl derivatives. The ethyl ester compounds were more efficient than the corresponding unesterified derivatives. The dodecyl ester of the N-2-heptyl analogue was the most effective inhibitor in isolated hepatocytes, but was relatively toxic in vivo. However, the corresponding ethyl ester was a potent in vivo inhibitor: GSH conjugation of bromosulphophthalein (as assessed by biliary excretion of the conjugate) was decreased by 70% after administration of a dose of 200 mumol/kg. The isoenzyme specificity of the inhibitors towards purified rat and human glutathione S-transferases was also examined. The unesterified compounds were more potent than the esterified analogues, and inhibited Alpha- and Mu-class isoenzymes of both rat and human glutathione S-transferase (Ki range 1-40 microM). Other GSH-dependent enzymes, i.e. GSH peroxidase, GSH reductase and gamma-glutamyltranspeptide, were not inhibited. Thus (R)-5-ethyloxycarbonyl-2-gamma-(S)-glutamylamino-N-2-hept ylpentamide, the in vivo inhibitor of GSH conjugation, may be useful in helping to assess the role of the Alpha and Mu classes of glutathione S-transferases in cellular biochemistry, physiology and pathology.

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Selected References

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  1. Adang A. E., Brussee J., van der Gen A., Mulder G. J. Inhibition of rat liver glutathione S-transferase isoenzymes by peptides stabilized against degradation by gamma-glutamyl transpeptidase. J Biol Chem. 1991 Jan 15;266(2):830–836. [PubMed] [Google Scholar]
  2. Adang A. E., Brussee J., van der Gen A., Mulder G. J. The glutathione-binding site in glutathione S-transferases. Investigation of the cysteinyl, glycyl and gamma-glutamyl domains. Biochem J. 1990 Jul 1;269(1):47–54. doi: 10.1042/bj2690047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Adang A. E., Moree W. J., Brussee J., Mulder G. J., van der Gen A. Inhibition of glutathione S-transferase 3-3 by glutathione derivatives that bind covalently to the active site. Biochem J. 1991 Aug 15;278(Pt 1):63–68. doi: 10.1042/bj2780063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Anderson M. E., Powrie F., Puri R. N., Meister A. Glutathione monoethyl ester: preparation, uptake by tissues, and conversion to glutathione. Arch Biochem Biophys. 1985 Jun;239(2):538–548. doi: 10.1016/0003-9861(85)90723-4. [DOI] [PubMed] [Google Scholar]
  5. Armstrong R. N. Glutathione S-transferases: reaction mechanism, structure, and function. Chem Res Toxicol. 1991 Mar-Apr;4(2):131–140. doi: 10.1021/tx00020a001. [DOI] [PubMed] [Google Scholar]
  6. Berk P. D., Potter B. J., Stremmel W. Role of plasma membrane ligand-binding proteins in the hepatocellular uptake of albumin-bound organic anions. Hepatology. 1987 Jan-Feb;7(1):165–176. doi: 10.1002/hep.1840070131. [DOI] [PubMed] [Google Scholar]
  7. Bilzer M., Krauth-Siegel R. L., Schirmer R. H., Akerboom T. P., Sies H., Schulz G. E. Interaction of a glutathione S-conjugate with glutathione reductase. Kinetic and X-ray crystallographic studies. Eur J Biochem. 1984 Jan 16;138(2):373–378. doi: 10.1111/j.1432-1033.1984.tb07925.x. [DOI] [PubMed] [Google Scholar]
  8. Bogaards J. J., van Ommen B., van Bladeren P. J. An improved method for the separation and quantification of glutathione S-transferase subunits in rat tissue using high-performance liquid chromatography. J Chromatogr. 1989 Jul 19;474(2):435–440. doi: 10.1016/s0021-9673(01)93940-8. [DOI] [PubMed] [Google Scholar]
  9. Cmarik J. L., Inskeep P. B., Meredith M. J., Meyer D. J., Ketterer B., Guengerich F. P. Selectivity of rat and human glutathione S-transferases in activation of ethylene dibromide by glutathione conjugation and DNA binding and induction of unscheduled DNA synthesis in human hepatocytes. Cancer Res. 1990 May 1;50(9):2747–2752. [PubMed] [Google Scholar]
  10. Flohé L., Günzler W. A. Assays of glutathione peroxidase. Methods Enzymol. 1984;105:114–121. doi: 10.1016/s0076-6879(84)05015-1. [DOI] [PubMed] [Google Scholar]
  11. Habig W. H., Pabst M. J., Jakoby W. B. Glutathione S-transferases. The first enzymatic step in mercapturic acid formation. J Biol Chem. 1974 Nov 25;249(22):7130–7139. [PubMed] [Google Scholar]
  12. Harrison D. J., May L., Hayes J. D., Neal G. E. Glutathione S-transferase localization in aflatoxin B1-treated rat livers. Carcinogenesis. 1990 Jun;11(6):927–931. doi: 10.1093/carcin/11.6.927. [DOI] [PubMed] [Google Scholar]
  13. Hussey A. J., Stockman P. K., Beckett G. J., Hayes J. D. Variations in the glutathione S-transferase subunits expressed in human livers. Biochim Biophys Acta. 1986 Nov 7;874(1):1–12. doi: 10.1016/0167-4838(86)90094-4. [DOI] [PubMed] [Google Scholar]
  14. Kitamura T., Jansen P., Hardenbrook C., Kamimoto Y., Gatmaitan Z., Arias I. M. Defective ATP-dependent bile canalicular transport of organic anions in mutant (TR-) rats with conjugated hyperbilirubinemia. Proc Natl Acad Sci U S A. 1990 May;87(9):3557–3561. doi: 10.1073/pnas.87.9.3557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Leung K. H. Selective inhibition of leukotriene C4 synthesis in human neutrophils by ethacrynic acid. Biochem Biophys Res Commun. 1986 May 29;137(1):195–200. doi: 10.1016/0006-291x(86)91195-2. [DOI] [PubMed] [Google Scholar]
  16. Lewis A. D., Hickson I. D., Robson C. N., Harris A. L., Hayes J. D., Griffiths S. A., Manson M. M., Hall A. E., Moss J. E., Wolf C. R. Amplification and increased expression of alpha class glutathione S-transferase-encoding genes associated with resistance to nitrogen mustards. Proc Natl Acad Sci U S A. 1988 Nov;85(22):8511–8515. doi: 10.1073/pnas.85.22.8511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lyttle M. H., Hocker M. D., Hui H. C., Caldwell C. G., Aaron D. T., Engqvist-Goldstein A., Flatgaard J. E., Bauer K. E. Isozyme-specific glutathione-S-transferase inhibitors: design and synthesis. J Med Chem. 1994 Jan 7;37(1):189–194. doi: 10.1021/jm00027a024. [DOI] [PubMed] [Google Scholar]
  18. Mannervik B., Danielson U. H. Glutathione transferases--structure and catalytic activity. CRC Crit Rev Biochem. 1988;23(3):283–337. doi: 10.3109/10409238809088226. [DOI] [PubMed] [Google Scholar]
  19. Meister A. Glutathione deficiency produced by inhibition of its synthesis, and its reversal; applications in research and therapy. Pharmacol Ther. 1991;51(2):155–194. doi: 10.1016/0163-7258(91)90076-x. [DOI] [PubMed] [Google Scholar]
  20. Meister A., Tate S. S., Griffith O. W. Gamma-glutamyl transpeptidase. Methods Enzymol. 1981;77:237–253. doi: 10.1016/s0076-6879(81)77032-0. [DOI] [PubMed] [Google Scholar]
  21. Meyer D. J., Coles B., Pemble S. E., Gilmore K. S., Fraser G. M., Ketterer B. Theta, a new class of glutathione transferases purified from rat and man. Biochem J. 1991 Mar 1;274(Pt 2):409–414. doi: 10.1042/bj2740409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Min A. D., Johansen K. L., Campbell C. G., Wolkoff A. W. Role of chloride and intracellular pH on the activity of the rat hepatocyte organic anion transporter. J Clin Invest. 1991 May;87(5):1496–1502. doi: 10.1172/JCI115159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mulder G. J., Scholtens E., Meijer D. K. Collection of metabolites in bile and urine from the rat. Methods Enzymol. 1981;77:21–30. doi: 10.1016/s0076-6879(81)77006-x. [DOI] [PubMed] [Google Scholar]
  24. Murren J. R., Hait W. N. Why haven't we cured multidrug resistant tumors? Oncol Res. 1992;4(1):1–6. [PubMed] [Google Scholar]
  25. Reed D. J., Babson J. R., Beatty P. W., Brodie A. E., Ellis W. W., Potter D. W. High-performance liquid chromatography analysis of nanomole levels of glutathione, glutathione disulfide, and related thiols and disulfides. Anal Biochem. 1980 Jul 15;106(1):55–62. doi: 10.1016/0003-2697(80)90118-9. [DOI] [PubMed] [Google Scholar]
  26. Seglen P. O. Preparation of isolated rat liver cells. Methods Cell Biol. 1976;13:29–83. doi: 10.1016/s0091-679x(08)61797-5. [DOI] [PubMed] [Google Scholar]
  27. Seidegård J., Pero R. W., Miller D. G., Beattie E. J. A glutathione transferase in human leukocytes as a marker for the susceptibility to lung cancer. Carcinogenesis. 1986 May;7(5):751–753. doi: 10.1093/carcin/7.5.751. [DOI] [PubMed] [Google Scholar]
  28. Snel C. A., Zhao Y., Mulder G. J., Pang K. S. Methods for the quantitation of bromosulfophthalein and its glutathione conjugate in biological fluids. Anal Biochem. 1993 Jul;212(1):28–34. doi: 10.1006/abio.1993.1286. [DOI] [PubMed] [Google Scholar]
  29. Tew K. D., Bomber A. M., Hoffman S. J. Ethacrynic acid and piriprost as enhancers of cytotoxicity in drug resistant and sensitive cell lines. Cancer Res. 1988 Jul 1;48(13):3622–3625. [PubMed] [Google Scholar]
  30. Vander Jagt D. L., Hunsaker L. A., Garcia K. B., Royer R. E. Isolation and characterization of the multiple glutathione S-transferases from human liver. Evidence for unique heme-binding sites. J Biol Chem. 1985 Sep 25;260(21):11603–11610. [PubMed] [Google Scholar]
  31. Vos R. M., Snoek M. C., van Berkel W. J., Müller F., van Bladeren P. J. Differential induction of rat hepatic glutathione S-transferase isoenzymes by hexachlorobenzene and benzyl isothiocyanate. Comparison with induction by phenobarbital and 3-methylcholanthrene. Biochem Pharmacol. 1988 Mar 15;37(6):1077–1082. doi: 10.1016/0006-2952(88)90513-8. [DOI] [PubMed] [Google Scholar]
  32. Warholm M., Guthenberg C., Mannervik B. Molecular and catalytic properties of glutathione transferase mu from human liver: an enzyme efficiently conjugating epoxides. Biochemistry. 1983 Jul 19;22(15):3610–3617. doi: 10.1021/bi00284a011. [DOI] [PubMed] [Google Scholar]
  33. Warholm M., Guthenberg C., Mannervik B., von Bahr C. Purification of a new glutathione S-transferase (transferase mu) from human liver having high activity with benzo(alpha)pyrene-4,5-oxide. Biochem Biophys Res Commun. 1981 Jan 30;98(2):512–519. doi: 10.1016/0006-291x(81)90870-6. [DOI] [PubMed] [Google Scholar]
  34. Waxman D. J. Glutathione S-transferases: role in alkylating agent resistance and possible target for modulation chemotherapy--a review. Cancer Res. 1990 Oct 15;50(20):6449–6454. [PubMed] [Google Scholar]
  35. Whelan G., Hoch J., Combes B. A direct assessment of the importance of conjugation for biliary transport of sulfobromophthalein sodium. J Lab Clin Med. 1970 Apr;75(4):542–557. [PubMed] [Google Scholar]

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