CYTOTOXICITY, DNA ADDUCT FORMATION AND DNA-REPAIR INDUCED BY 2-HYDROXYAMINO-3-METHYLIMIDAZO[4,5-F]QUINOLINE AND DROXYAMINO-1-METHYL-6-PHENYLIMIDAZO[4,5-B]PYRIDINE IN CULTURED HUMAN MAMMARY EPITHELIAL-CELLS

2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-ph enylimidazo[4,5-b]pyridine (PhIP) are heterocyclic amines (HAs) found in cooked meats, Both compounds are mammary gland carcinogens in rats, The initiation of carcinogenesis by the HAs is believed to be associa ted with their DNA adduct formation that occurs after metabolic activa tion of the parent amines via cytochrome P-450-mediated N-hydroxylatio n and esterification, To assess the capacity of the human mammary epit helium to metabolically activate the HAs, we used the P-32-postlabelin g method to measure the levels of DNA adducts in a culture of human ma mmary epithelial cells exposed to IQ, PhIP or their N-hydroxylamine me tabolites. Whereas 50 mu M parent amines did not form detectable level s of DNA adducts ill cultured human mammary epithelial cells after 24 h incubations, concentrations as low as 1 mu M N-hydroxylamines produc ed detectable levels of adducts after 2 h incubations, N-Hydroxy-PhIP formed higher adduct levels than N-hydroxy-IQ at all concentrations te sted, For example, following a 2 h incubation at 50 mu M, adduct level s (per 10(7) nucleotides) were 674 and 16 for N-hydroxy-PhIP and N-hyd roxy-IQ, respectively, At similar initial adduct levels (10-11/10(7) n ucleotides), 60-80% of IQ- and PhIP-DNA adducts were removed after 24 h, indicating that the mammary epithelial cell culture showed efficien t repair of HA adducts, Whereas neither IQ nor PhIP was cytotoxic, bot h N-hydroxy-IQ and N-hydroxy-PhIP were cytotoxic as assessed by a dose -dependent inhibition of colony formation, After exposure to 0.1, 1, 1 0 or 50 mu M N-hydroxy-PhIP (or N-hydroxy-IQ), colony formation was 10 3 (94), 84 (74), 37 (29) and 3 (2)% of the control values, respectivel y, Only N-hydroxy-PhIP (at 10 and 50 mu M), however, was cytotoxic as assessed by the MTT cell survival assay (which measures the capacity o f mitochondria to metabolize a tetrazolium salt), The ability of the N -hydroxylamines to form DNA adducts and to be cytotoxic in a culture o f human mammary epithelial cells may implicate these metabolites as pr oximate carcinogenic forms of IQ and PhIP in the human mammary gland, However, whether there are inter-individual differences in N-hydroxyla mine metabolism, adduct formation and repair in human mammary epitheli al cells requires further study. The results from this study support t he usefulness of cultured human mammary epithelial cells for studies o n the genotoxicity and metabolism of the N-hydroxylamines of HA food m utagens.