Analysis of CYP2A Contributions to Metabolism of 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone in Human Peripheral Lung Microsomes

Abstract

ABSTRACT:The objectives of this study were to determine the contributions of CYP2A13 and CYP2A6 to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) metabolism in human peripheral lung microsomes and to determine the influence of the genetic polymorphism, CYP2A13 Arg257Cys, on NNK metabolism. 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), the keto-reduced metabolite of NNK, was the major metabolite produced, ranging from 0.28 to 0.9%/mg protein/min. Based on total bioactivation of NNK and NNAL by ␣-carbon hydroxylation, subjects could be classified as either high (17 subjects) or low (12 subjects) bioactivators [(5.26 ؎ 1.23) ؋ 10 ؊2 and (6.49 ؎ 5.90) ؋ 10 ؊3% total ␣-hydroxylation/mg protein/min, P < 0.05]. Similarly, for detoxification, subjects could be grouped into high (9 subjects) and low (20 subjects) categories [(2.03 ؎ 1.65) ؋ 10 ؊3 and (2.50 ؎ 3.04) ؋ 10 ؊4% total N-oxidation/mg protein/min, P < 0.05]. When examining data from all individuals, no significant correlations were found between levels of CYP2A mRNA, CYP2A enzyme activity, or CYP2A immunoinhibition and the degree of total NNK bioactivation or detoxification (P > 0.05). However, subgroups of individuals were identified for whom CYP2A13 mRNA correlated with total NNK and NNAL ␣-hydroxylation and NNAL-N-oxide formation (P < 0.05). The degree of NNAL formation and CYP2A13 mRNA was also correlated (P < 0.05). Subjects (n ‫؍‬ 84) were genotyped for the CYP2A13 Arg257Cys polymorphism, and NNK metabolism for the one variant (Arg/Cys) was similar to that for other subjects. Although results do not support CYP2A13 or CYP2A6 as predominant contributors to NNK bioactivation and detoxification in peripheral lung of all individuals, CYP2A13 may be important in some.Lung cancer is the leading cause of cancer-related death in the world, and it is estimated that cigarette smoking accounts for approximately 90% of lung cancer cases (Hecht, 2003). The tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is thought to play a major role in human tobacco-related cancers (Hecht, 2003). NNK is the most prevalent pulmonary carcinogen in tobacco smoke and the most potent cancer-causing tobacco-specific nitrosamine in all animal species tested (Hecht, 1998). NNK selectively induces lung adenocarcinoma in animals (Hecht, 1998) and is believed to be a causal agent in the induction of human lung adenocarcinoma, which is now the leading form of lung cancer (Hoffmann et al., 1996;Thun et al., 1997).To induce carcinogenesis, NNK and its keto-reduced metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), require metabolic activation via ␣-carbon hydroxylation (Fig. 1). Hydroxylation of the ␣-methylene carbons of NNK and NNAL leads to the formation of DNA-methylating species, whereas hydroxylation of the ␣-methyl carbons of NNK and NNAL results in the formation of DNA-pyridyloxobutylating and -pyridylhydroxybutylating species, respectively. The formation of both types of adducts is believed to be important in the induction of ...

Differential Distribution of CYP2A6 and CYP2A13 in the Human Respiratory Tract

“…Moreover, the interindividual variation in bronchiolar CYP2A6 expression shown in table 1 is consistent in some previous studies in human peripheral lung tissue with qPCR [29] and immunoblot assay [30]. The interindividual variation in CYP2A6 may contribute to the large variation in NNK metabolism by the peripheral lung microsomes from individuals with different levels of CYP2A6 [33] and thus may significantly influence the susceptibility of individuals to TSNAs. Because of the heterogeneity of cell types in the respiratory tract, by using protein or mRNA samples from whole tissue it is extremely difficult to precisely define those protein molecules with specific expression in certain types of cells, such as epithelial or alveolar cells.…”

A Bioinformatics Approach for the Phenotype Prediction of Nonsynonymous Single Nucleotide Polymorphisms in Human Cytochromes P450

“…CYP2A13 is active in the metabolism of a number of procarcinogens. CYP2A13 is the most efficient enzyme in the metabolic activation of the tobacco-specific procarcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, a tobacco-specific lung carcinogen (Brown et al, 2007). CYP2A13, but not CYP2A6, is also highly efficient in metabolizing the mycotoxin aflatoxin B 1 to its carcinogenic metabolites 8,9-epoxide and 1-8,9-epoxide.…”

Targeted omics analyses, and metabolic enzyme activity assays demonstrate maintenance of key mucociliary characteristics in long term cultures of reconstituted human airway epithelia

“…(i) diol-epoxides from CYP1A1-dependent oxidation of benzo[a]pyrene (B[a]P) found in tobacco smoke and exhaust fumes (Uppstad et al, 2010), (ii) diazo groups resulting from the a-hydroxylation of tobacco nitrosamines by the CYP2A enzyme family (Brown et al, 2007), (iii) formaldehyde hydrate as a by-product from the glutathione conjugation through inhalation of dichloromethane vapors (Hashmi et al, 1994).…”