Tumorigenicity and metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol enantiomers and metabolites in the A/J mouse

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a major metabolite of the tobacco-specific pulmonary carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), has a chiral center but the tumorigenicity of the NNAL enantiomers has not been previously examined. In this study, we assessed the relative tumorigenic activities in the A/J mouse of NNK, racemic NNAL, (R)-NNAL, (S)-NNAL and several NNAL metabolites, including [4-(methylnitrosamino)-1-(3-pyridyl)but-(S)-1-yl] β-O-D-glucosiduronic acid [(S)-NNAL-Gluc], 4-(methylnitrosamino)-1- (3-pyridyl N-oxide)-1-butanol, 5-(3-pyridyl)-2-hydroxytetrahydrofuran, 4-(3-pyridyl)butane-1,4.diol and 2-(3-pyridyl) tetrahydrofuran. We also quantified urinary metabolites of racemic NNAL and its enantiomers and investigated their metabolism with A/J mouse liver and lung microsomes. Groups of female A/J mice were given a single i.p. injection of 20 μmol of each compound and killed 16 weeks later. Based on lung tumor multiplicity, (R)-NNAL (25.6 ± 7.5 lung tumors/mouse) was as tumorigenic as NNK (25.3 ± 9.8) and significantly more tumorigenic than racemic NNAL (12.1 ± 5.6) or (S)-NNAL (8.2 ± 3.3) (P < 0.0001). None of the NNAL metabolites was tumorigenic. The major urinary metabolites of racemic NNAL and the NNAL enantiomers were 4-hydroxy-4-(3 pyridyl)butanoic acid (hydroxy acid), NNAL-N-oxide and NNAL-Gluc, in addition to unchanged NNAL. Treatment with (R)-NNAL or (S)-NNAL gave predominantly (R)-hydroxy acid or (S)-hydroxy acid, respectively, as urinary metabolites. While treatment of mice with racemic or (S)-NNAL resulted in urinary excretion of (S)-NNAL-Gluc, treatment with (R)-NNAL gave both (R)-NNAL-Gluc and (S)-NNAL-Gluc in urine, apparently through the metabolic intermediacy of NNK. (S)-NNAL appeared to be a better substrate for glucuronidation than (R)-NNAL in the A/J mouse. Mouse liver and lung microsomes converted NNAL to products of α-hydroxylation, to NNAL-N-oxide, to adenosine dinucleotide phosphate adducts and to NNK. In lung microsomes, metabolic activation by α-hydroxylation of (R)-NNAL was significantly greater than that of (S)-NNAL. The results of this study provide a metabolic basis for the higher tumorigenicity of (R)-NNAL than (S)-NNAL in A/J mouse lung, namely preferential metabolic activation of (R)-NNAL in lung and preferential glucuronidation of (S)-NNAL.