CYP2A13: Variable Expression and Role in Human Lung Microsomal Metabolic Activation of the Tobacco-Specific Carcinogen 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone

Abstract

CYP2A13 is the most efficient cytochrome P450 enzyme in the metabolic activation of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco-specific lung carcinogen. The aims of this study were to determine the levels of CYP2A13 protein in human lung microsomes and to ascertain whether CYP2A13 plays any role in lung microsomal NNK metabolic activation. The expression of CYP2A6 and CYP2A13 was examined using a high-resolution immunoblotting method, following immunopurification with an anti-CYP2A5 antibody. We found that, of 116 human lung microsomal samples analyzed, ϳ90% had detectable CYP2A6, whereas only 12% had detectable CYP2A13 with a detection limit of ϳ2 fmol of CYP2A/mg protein. For the majority of microsomal samples analyzed, the level of CYP2A13 was found to be lower than the level of CYP2A6; overall, the highest level of CYP2A13 found (ϳ20 fmol/mg protein) was ϳ10-fold lower than the highest level of CYP2A6 detected. Quantitative RNA-polymerase chain reaction analysis confirmed that the highly variable expression of the CYP2A proteins was consistent with variations in the levels of the corresponding CYP2A mRNAs in the same tissue samples. It is noteworthy that the level of CYP2A13, but not CYP2A6, was correlated with lung microsomal NNK metabolic activation activity. Furthermore, the addition of 8-methoxypsoralen, a CYP2A inhibitor, led to greater inhibition of NNK metabolic activation in microsomes containing relatively high levels of CYP2A13 than in samples containing no detectable CYP2A13. Taken together, these data indicate that human lung microsomal CYP2A13 is active in NNK metabolic activation. Therefore, individuals having relatively high levels of CYP2A13 expression will likely have an increased risk of developing smoking-related lung cancer.The human cytochrome P450 (P450) CYP2A gene subfamily is known to have two functional members, CYP2A6 and CYP2A13 (Fernandez-Salguero et al., 1995;Su et al., 2000). Previous studies on CYP2A mRNA expression in human tissues indicated that CYP2A6 is expressed in the lung at much lower levels than in the liver, whereas CYP2A13 is selectively expressed in the respiratory tract (Koskela et al., 1999;Su et al., 2000). A more recent study indicated that CYP2A13 is also expressed in the bladder (Nakajima et al., 2006). Heterologously expressed CYP2A6 and CYP2A13 enzymes are both active in the metabolic activation of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a lung procarcinogen, which is found in cigarettes (Hecht, 1998) and can also be produced endogenously from nicotine metabolites (Hecht et al., 2000). NNK metabolic activation involves P450-catalyzed hydroxylation at one of the two ␣ carbons to the N-nitroso group, leading to the formation of reactive intermediates that can form either DNA adducts or else stable metabolites, including 4-hydroxy-1-(3-pyridyl)-1-butanone (HPB) and 4-oxo-1-(3-pyridyl)-1-butanone (OPB) (Hecht, 1998). Although multiple human P450 enzymes, including CYP1A1, CYP1A2, CYP2A6, CYP2A13, CYP2B6, CYP2E1, and CYP...

Immunohistochemical analysis of CYP2A13 in various types of human lung cancers

“…Zhang et al detected CYP2A13 protein in lung microsomes by immunoblotting using the anti-Cyp2a5 antibody, in which immunoprecipitants from lung microsomes using the anti-Cyp2a5 antibody were separated. (19) Thus, since a specific antibody to CYP2A13 has not been available, great effort has been required to detect CYP2A13 protein. This background prompted us to prepare a specific antibody against human CYP2A13.…”

HapMap-based study on the association between MPO and GSTP1 gene polymorphisms and lung cancer susceptibility in Chinese Han population

“…Phase I enzymes such as myeloperoxidase (MPO) and cytochrome P450 can metabolically activate a wide range of tobacco mutagens to DNAdamaging metabolites [3,4] . After phase I bioactivation, phase II enzymes (such as glutathione S-transferase) can detoxify the DNA-damaging metabolites via conjugation with endogenous molecules to form hydrophilic conjugates [5,6] .…”

The Effects of Single Nucleotide Polymorphisms in CYP2A13 on Metabolism of 5-Methoxypsoralen

“…Although the furanoepoxide and ␥-ketoenal species of 5-MOP were not directly detected in this study, these intermediates were probably generated during 5-MOP metabolism by CYP2A13 and were hydrolyzed to dihydrodiol spontaneously. The tissue distribution of CYP2A13 in vivo and its metabolic activity led us to the hypothesis that the human CYP2A13 is a major player in the production of mutagenic intermediates from NNK (Jalas et al, 2003;He et al, 2004;Zhang et al, 2007). Our findings indicate the possibility that reactive intermediates from 5-MOP are produced in respiratory organs by CYP2A13, as well as in liver by other P450s.…”