Preferential Metabolic Activation of N-Nitrosopiperidine as Compared to Its Structural Homologue N-Nitrosopyrrolidine by Rat Nasal Mucosal Microsomes

N-Nitrosopiperidine (NPIP) is a potent rat nasal carcinogen whereas N-nitrosopyrrolidine (NPYR), a hepatic carcinogen, is weakly carcinogenic in the nose. NPIP and NPYR may be causative agents in human cancer. P450-catalyzed α-hydroxylation is the key activation pathway by which these nitrosamines elicit their carcinogenic effects. We hypothesize that the differences in NPIP and NPYR metabolic activation in the nasal cavity contribute to their differing carcinogenic activities. In this study, the kinetics of tritium-labeled NPIP or NPYR α-hydroxylation mediated by Sprague-Dawley rat nasal olfactory or respiratory microsomes were investigated. To compare α-hydroxylation rates of the two nitrosamines, tritiated 2-hydroxytetrahydro-2H-pyran and 2-hydroxy-5-methyltetrahydrofuran, the major NPIP α-hydroxylation products, and tritiated 2-hydroxytetrahydrofuran, the major NPYR α-hydroxylation product, were quantitated by HPLC with UV absorbance and radioflow detection. These microsomes catalyzed the α-hydroxylation of NPIP more efficiently than that of NPYR. K_M values for NPIP were lower as compared to those for NPYR (13.9-34.7 vs 484-7660 μM). Furthermore, catalytic efficiencies (V_max/K_M) of NPIP were 20-37-fold higher than those of NPYR. Previous studies showed that P450 2A3, present in the rat nose, also exhibited this difference in catalytic efficiency. For both types of nasal microsomes, coumarin (100 μM), a P450 2A inhibitor, inhibited NPIP and NPYR α-hydroxylation from 63.8 to 98.5%. Furthermore, antibodies toward P450 2A6 inhibited nitrosamine α-hydroxylation in these microsomes from 68.8 to 78.4% whereas antibodies toward P450 2E1 did not inhibit these reactions. Further immunoinhibition studies suggest some role for P450 2G1 in NPIP metabolism by olfactory microsomes. In conclusion, olfactory and respiratory microsomes from rat nasal mucosa preferentially activate NPIP over NPYR with P450 2A3 likely playing a key role. These results are consistent with local metabolic activation of nitrosamines as a contributing factor in their tissue-specific carcinogenicity.