High potency of bioactivation of 2-amino-1-methyl-6-phenylimidazo[4,5- b]pyridine (PhIP) in mouse colon epithelial cells with ApcMin mutation

Abstract

2-Amino-1-methyl-6-phenylimidazo[4,5- b]pyridine (PhIP) is a prominent heterocyclic aromatic amine (HAA) found in meat and fish cooked at moderate to high temperature. It is considered as a potent dietary factor promoting colon carcinogenesis. However, the role of intestinal cells in PhIP bioactivation has not been fully explained, particularly when cells are pre-malignant. Loss of function of the adenomatous polyposis coli ( APC) gene product is an early and frequent event in human colorectal carcinogenesis. Normal ( Apc +/+) and pre-malignant ( Apc Min/+ , where Min = multiple intestinal neoplasia) colonic epithelial cells of mice can be used to study promotion of carcinogenesis, but these cells have not been characterized for bio-activation of HAA. We investigated the metabolism of 14C-PhIP in these two murine cell lines. Cells induced by 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) metabolized PhIP into 4′-OH-PhIP as the main metabolite in PhiP detoxification. Besides, 5-OH-PhIP was identified, revealing the formation of intermediary reactive metabolites, since it results from a degradation of conjugates of N-acetoxy-PhIP. Apc Min/+ cells produce significantly higher amounts of these metabolites. Demethylated metabolites are also observed, indicating that the colon contains a significant CYP1 family dependent metabolic activity. A minor hydroxy-glucuronide-PhIP metabolite is observed in Apc Min/+ cells, the glucuronidation being known as an important step in the detoxification pathway. Quantitative real-time reverse transcription polymerase chain reaction experiments demonstrate that induction by TCDD has prevailing effects in gene expression of CYP1A1, CYP1A2 and CYP1B1 in Apc Min/+ cells. In these cells, N-acetyltransferase-2 is also expressed at higher levels. So, the more important potency to metabolically bio-activate PhIP, as measured in Apc Min/+ cells, can be linked to a higher probability to generate new in situ mutations.