Measurement of cytochrome P450 2A6 and 2E1 gene expression in primary human bronchial epithelial cells.

Bronchogenic carcinomas arise from bronchial epithelial cells (BECs). Inhalation exposure of BECs to nitrosamines in cigarette smoke is an important exogenous risk factor for malignant transformation of BECs. Thus, an important endogenous risk factor is likely to be the capacity of BECs to metabolize nitrosamines. Among the cytochrome P450 enzymes capable of metabolizing nitrosamines, CYP2A6, CYP2E1 and CYP2B6 are expressed in BECs. In this study, we used quantitative RT-PCR to evaluate expression of CYP2A6 and CYP2E1 in primary human BECs from 12 non-smokers and eight smokers. CYP2A6 was expressed in 20/20 cases and quantifiable in 18/20 cases, with a mean level of 580 mRNA/10(6) beta-actin mRNA. CYP2E1 expression was observed in 9/20 cases, but in all cases it was expressed at levels below our limit of quantification (10 mRNA/10(6) beta-actin mRNA). There was significant (P < 0.05) 20-fold inter-individual variation in expression of CYP2A6. Further, the mean level of CYP2A6 among smokers (260 mRNA/10(6) beta-actin mRNA) was significantly lower than among non-smokers (740 mRNA/10(6) beta-actin mRNA). It is hypothesized that: (i) inter-individual variation in CYP2A6 gene expression may contribute to inter-individual variation in risk for bronchogenic carcinoma; (ii) smoking may reduce the level of expression of CYP2A6 in the BECs of some individuals; and (iii) CYP2A6 is more important than CYP2E1 for metabolic activation of nitrosamines in bronchial epithelial cells.

[1]  J. Willey,et al.  Expression measurement of many genes simultaneously by quantitative RT-PCR using standardized mixtures of competitive templates. , 1998, American journal of respiratory cell and molecular biology.

[2]  W. Thilly,et al.  Quantitative RT-PCR measurement of cytochromes p450 1A1, 1B1, and 2B7, microsomal epoxide hydrolase, and NADPH oxidoreductase expression in lung cells of smokers and nonsmokers. , 1997, American journal of respiratory cell and molecular biology.

[3]  M. Grever,et al.  Reductase enzyme expression across the National Cancer Institute Tumor cell line panel: correlation with sensitivity to mitomycin C and EO9. , 1996, Journal of the National Cancer Institute.

[4]  W. Thilly,et al.  Xenobiotic metabolism enzyme gene expression in human bronchial epithelial and alveolar macrophage cells. , 1996, American journal of respiratory cell and molecular biology.

[5]  D W Nebert,et al.  P450 superfamily: update on new sequences, gene mapping, accession numbers and nomenclature. , 1996, Pharmacogenetics.

[6]  T. Massey,et al.  Biochemical and Molecular Aspects of Mammalian Susceptibility to Aflatoxin B1 Carcinogenicity , 1995, Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine.

[7]  B. Lake,et al.  Expression and alternative splicing of the cytochrome P-450 CYP2A7. , 1995, The Biochemical journal.

[8]  J. Willey,et al.  Measurement of gene expression by multiplex competitive polymerase chain reaction. , 1993, Analytical biochemistry.

[9]  R. Hines,et al.  Specific nuclear protein binding to a negative regulatory element on the human CYP1A1 gene. , 1993, The Journal of biological chemistry.

[10]  M. Zenilman,et al.  A rapid and versatile method to synthesize internal standards for competitive PCR. , 1993, Nucleic acids research.

[11]  F. Guengerich,et al.  Metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in human lung and liver microsomes and cytochromes P-450 expressed in hepatoma cells. , 1992, Cancer research.

[12]  T. Shimada,et al.  Activation of Amino-α-carboline, 2-Amino-1-methyl-6-phenylimidazo[4,5- b ]pyridine, and a Copper Phthalocyanine Cellulose Extract of Cigarette Smoke Condensate by Cytochrome P-450 Enzymes in Rat and Human Liver Microsomes , 1991 .

[13]  S. Perrin,et al.  Analysis of cytokine mRNA and DNA: detection and quantitation by competitive polymerase chain reaction. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[14]  F. Gonzalez,et al.  The CYP2A3 gene product catalyzes coumarin 7-hydroxylation in human liver microsomes. , 1990, Biochemistry.

[15]  O. Mcbride,et al.  Human ethanol-inducible P450IIE1: complete gene sequence, promoter characterization, chromosome mapping, and cDNA-directed expression. , 1988, Biochemistry.

[16]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.

[17]  L. Kedes,et al.  Evolutionary conservation in the untranslated regions of actin mRNAs: DNA sequence of a human beta-actin cDNA. , 1984, Nucleic acids research.

[18]  S. Hecht,et al.  Recent studies on mechanisms of bioactivation and detoxification of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco-specific lung carcinogen. , 1996, Critical reviews in toxicology.

[19]  G. Clark,et al.  Dioxin-responsive genes: examination of dose-response relationships using quantitative reverse transcriptase-polymerase chain reaction. , 1994, Cancer research.

[20]  D. Hoffmann,et al.  Studies in tobacco carcinogenesis. , 1991, IARC scientific publications.