Publisher Summary This chapter discusses the role of glutathione (GSH) and glutathione s-transferases in metabolism of chemical carcinogens and other electrophilic agents. GSH is a tripeptide (I) that is present in nearly all living cells and is the most abundant sulfhydryl compound present in animal tissues, mainly in the cytosol. The chapter illustrates the wide range of electrophilic agents, including several known mutagens and carcinogens, which conjugate with GSH, a process usually catalyzed by the GSH S-transferases. This conjugation is probably a protective mechanism and is the initial stage in mercapturic acid biosynthesis for the elimination of foreign compounds from the body. GSH S-transferases provide protection not only by catalyzing the conjugation of a potential toxicant with GSH, but also by preferentially binding, even covalently, that toxicant. The reactive electrophiles that conjugate with GSH also bind to DNA, RNA, and protein and identification of GSH conjugates provide information on the nature of these biologically active intermediates or even their immediate precursors. Thus, the knowledge of the way in which mutagens and carcinogens are metabolized is essential to a better understanding of their mode of action and of the processes for their detoxication.

The role of glutathione and glutathione S-transferases in the metabolism of chemical carcinogens and other electrophilic agents.

Previous studies reported that metabolic activation of certain polychlorinated biphenyls (PCBs) resulted in binding to protein, RNA and DNA fractions. However, the formation of DNA adducts has not been demonstrated nor have methods been optimized for the detection of such adducts. In the present study we investigated activation and binding to DNA of lower chlorinated biphenyls using 32P-postlabeling. The incubation of 2-chloro-, 3-chloro-, 3,4-dichloro- and 3,4,5-trichlorobiphenyl with calf thymus DNA and liver microsomes from rats treated with phenobarbital and 3-methylcholanthrene, followed by oxidation with a peroxidase, produced 1-3 major adducts. Reaction of deoxyguanosine 3'-monophosphate with metabolites of the congeneric chlorinated biphenyls produced adducts with similar chromatographic mobility as those with DNA, suggesting that guanine was the preferential site of attack. Furthermore butanol and nuclease P1 enrichments showed different adduct recoveries, depending upon the the chlorobiphenyl. Adducts derived from incubations with monochlorobiphenyls were recovered 2- to 3-fold higher with butanol, while the recovery of di- and tri-chlorobiphenyl adducts was 5- to 7-fold higher with nuclease P1. DNA adducts formed during the metabolism of 3,4-dichlorobiphenyl were reduced by the sulfur nucleophiles, glutathione and N-acetyl-L-cysteine, suggesting that reactive semiquinone(s) or quinone(s) are involved. In contrast, the addition of superoxide dismutase increased adduct formation, suggesting that the quinone metabolites are responsible for the major adducts formed. Our results are consistent with the hypothesis that lower chlorinated biphenyls are metabolically activated to electrophilic quinoid species which bind to DNA.

Analysis of polychlorinated biphenyl-DNA adducts by 32P-postlabeling

Reactions of 2,2',5,5'-tetrachlorobiphenyl 3,4-oxide with methionine, cysteine and glutathione in relation to the formation of methylthio-metabolites of 2,2',5,5'-tetrachlorobiphenyl in the rat and mouse

In vivo binding of 2,3,6,2′,3′,6′-hexachlorobiphenyl and 2,4,5,2′,4′,5′-hexachlorobiphenyl to mouse liver macromolecules

Time and dose responses of the reduction in retinoid concentrations in C57BL/Rij and DBA/2 mice induced by 3,4,3',4'-tetrachlorobiphenyl.

SummaryIncubation of tritiated 2,5,-2′,5′-tetrachlorobiphenyl with normal monkey liver microsomes in a NADPH-generating system results in the formation of active metabolite(s) of the [3H]2,5,2′,5′-tetrachlorobiphenyl capable of covalently binding to RNA and protein isolated from the incubation mixture. The metabolite is not formed when the control microsomes are held at 100° for 10 min prior to incubation. The addition of microsomal supernate to the solution causes an increase in the binding of the active metabolite to macromolecules while the addition of glutathione to the incubation medium significantly inhibits this increase.This investigation was supported in part by U.S. Public Health Service Grants ES-00472, ES-00958, and RR-00167 from the National Institutes of Health. This is Primate Center Publication No. 15-019.

S. P. Schmidt

Papers

In Vitro Generation of a Chemically Reactive Metabolite of 2,5,2′,5′-Tetrachlorobiphenyl by Rhesus Monkey Liver Microsomes