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BIOTRANSFORMATION AND GENOTOXICITY OF THE DRINKING WATER DISINFECTION BYPRODUCT BROMODICHLOROMETHANE: DNA BINDING MEDIATED BY GLUTATHIONE TRANSFERASE THETA 1-1
Citation:
Ross, M. K. AND R A. Pegram. BIOTRANSFORMATION AND GENOTOXICITY OF THE DRINKING WATER DISINFECTION BYPRODUCT BROMODICHLOROMETHANE: DNA BINDING MEDIATED BY GLUTATHIONE TRANSFERASE THETA 1-1. TOXICOLOGICAL AND APPLIED PHARMICOLOGY. Elsevier Science BV, Amsterdam, Netherlands, 195(2):166-81, (2004).
Impact/Purpose:
to investigate the DNA covalent binding potential of reactive intermediates generated by GSTT1-1-mediated metabolism of CHBrCl2
Description:
The drinking water disinfection byproduct bromodichloromethane (CHBrCl2) was
previously shown to be mutagenic in Salmonella typhimurium that overexpress rat glutathione
transferase theta 1-1 (GSTT1-1). Several experimental approaches were undertaken in this study
to investigate the DNA covalent binding potential of reactive intermediates generated by
GSTT1-1-mediated metabolism of CHBrCl2. First, rodent hepatic cytosol incubations
containing [14C]CHBrCl2, supplemented glutathione (GSH), and calf thymus DNA resulted in
~3-fold (rat liver cytosol) and 7-fold (mouse liver cytosol) greater amounts of total radioactivity
associated with the purified DNA as compared to a control (absence of rodent cytosol) following
liquid scintillation counting of isolated DNA. The increase in DNA labeling is consistent with
the conjugation activity of these rodent cytosols toward CHBrCl2. Second, exposure of GSTT1-
1-expressing S. typhimurium to [14C]CHBrCl2 resulted in a concentration-dependent increase of
bacterial DNA-associated total radioactivity. Characterization of DNA-associated radioactivity
could not be assigned to a specific deoxynucleoside adduct(s) following enzymatic hydrolysis of
DNA and subsequent HPLC analysis. A possible explanation for this observation was formation
of a ?transient' adduct that was unstable in the DNA isolation and hydrolysis procedures
employed. To circumvent problems of adduct instability, reactions of [14C]CHBrCl2 with GSH
catalyzed by recombinant rat GSTT1-1 were performed in the presence of calf thymus DNA or,
alternatively, the model nucleophile deoxyguanosine. Hydroxyapatite chromatography of [14C]-labeled DNA or HPLC chromatography of [14C]-labeled deoxyguanosine derivatives
demonstrated the covalent binding of [14C]CHBrCl2-derived metabolites to DNA and
deoxyguanosine in low yield (~0.02% of [14C]CHBrCl2 biotransformed by GSTT1-1 resulted in
DNA adducts). Cytochrome P450 (CYP)- and GST-catalyzed biotransformation of CHBrCl2 in
rat tissues (kidney and large intestine) that develop tumors following chronic CHBrCl2 exposure
were compared with rat liver (a non-target tissue). Rat liver had a significant capacity to
detoxify CHBrCl2 (to carbon dioxide) compared with kidney and large intestine as a result of
CYP-catalyzed oxidation; liver was ~16-fold more efficient than kidney and large intestine when
intrinsic clearance values (Vmax/Km) were compared. In contrast, the efficiency of GST-mediated
GSH conjugation of CHBrCl2 in kidney and large intestine was only slightly lower than liver
(~2?4-fold lower), thus the relative amounts of reactive intermediates that are produced with the
capacity to covalently modify DNA may be enhanced in these extrahepatic tissues. The
significance of these findings is that conjugation of CHBrCl2 with GSH can result in the covalent
modification of DNA and that cancer target tissues in rats have a much reduced detoxification
capacity, but only a modest decrease in bioactivation capacity, as compared to the liver (a non-target tissue in rats).