The impact of glucuronidation on the bioactivation and genotoxicity of the cooked-food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine.

摘要

UDP-glucuronosyltransferase 1A (UGT1A)-catalyzed glucuronidation is an important detoxification pathway for endogenous and xenobiotic compounds including carcinogenic heterocyclic amines and their hydroxylated metabolites. The biotransformation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), the most mass abundant heterocyclic amine found in cooked meats, is highly dependent on cytochrome P4501A2 hydroxylation followed by UGT-catalyzed glucuronidation of the N-hydroxy-PhIP reactive intermediate. To better understand the effect of glucuronidation on PhIP bioactivation, subcellular, cellular, and whole animals studies were done to assess the role of UGTs in PhIP metabolism and toxicity. To determine which UGT1A proteins contribute to N-hydroxy-PhIP glucuronide formation, microsomal preparations from baculovirus-infected insect cells containing recombinant human UGT1A proteins were used. Kinetic parameters were measured for each UGT1A isozyme to determine the catalytic efficiency of each protein. Differential selectivity for the N-hydroxy-PhIP substrate was observed between each of the UGT1A isozymes, with UGT1A1 having the highest affinity for glucuronidating N-hydroxy-PhIP. To assess the role of glucuronidation on PhIP bioactivation in intact mammalian cells, Chinese hamster ovary cells that are nucleotide excision repair-deficient, and express CYP1A2, were transfected with a cDNA plasmid of human UGT1A1 to establish the UGT1A1 expressing 5P3hUGT1A1 cell line. The cytotoxic effect from PhIP was decreased ∼350 fold in the UGT1A1 expressing cells compared to the control cells. In addition no significant increase in mutation frequency was observed in the transfected cells where exposed to PhIP compared to vehicle treated controls. The role of UGTs in the biotransformation of PhIP in whole animals was assessed using the UGT1A proficient and deficient Wistar and Gunn rats, respectively. Rats were treated orally with a single dose of [14C]-PhIP. Urinary metabolite profiles and PhIP-DNA adduct formation in liver and colon was compared between the two rat strains. A decrease in urinary PhIP glucuronide levels in the Gunn rats correlated with an increase in hepatic DNA adducts, compared to the Wistar rats. These results indicate that UGT1A1 provides a protective effect against increased PhIP bioactivation. Therefore, a decrease in UGT1A1 activity can potentially lead to an increase in susceptibility to the deleterious effects from PhIP because the capacity to form nontoxic N-hydroxy-PhIP glucuronide conjugates will be diminished.