Adenovirus vector-mediated reporter system for in vivo analyses of human CYP3A4 gene activation.
The use of cultured mammalian cells and artificial promoters for analyses of gene regulation gives results that are sometimes inconsistent with in vivo events and thus inconclusive. To understand the in vivo mechanism of chemically mediated CYP3A4 gene activation, we have used a natural promoter of the CYP3A4 gene and an adenovirus as a reporter vector. The adenovirus reporter vector (AdCYP3A4-362) was constructed with a proximal promoter region (-362 to +11 nt) of the CYP3A4 gene and a luciferase-reporter gene. AdCYP3A4-362 was then infected into mice, and both the reporter and mouse CYP3A activities were measured. Clear increases in the reporter activity were observed in livers of all mice treated with chemicals. The profile of the CYP3A4 gene activation with chemicals was in good agreement with that of endogenous mouse CYP3A-mediated testosterone 6beta-hydroxylase. Introduction of nucleotide mutations in the receptor-binding region (ER-6) of the CYP3A4 promoter resulted in diminished reporter activity. These results indicate the advantage of the adenovirus-mediated in vivo system over the currently available in vitro systems for gene transcriptional activation.
Furukawa
Masayuki
M
Division of Drug Metabolism and Molecular Toxicology, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578, Japan.
Okubo
Takayuki
T
Ogino
Makoto
M
Yamazaki
Toru
T
Shimada
Miki
M
Nagata
Kiyoshi
K
Yamazoe
Yasushi
Y
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D003022:Clotrimazole / Q000494:pharmacology; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D003907:Dexamethasone / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005822:Genetic Vectors; D005938:Glucocorticoids / Q000494:pharmacology; D006131:Growth Inhibitors / Q000494:pharmacology; D006801:Humans; D008099:Liver / Q000378:metabolism; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D006899:Mixed Function Oxygenases / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012293:Rifampin / Q000494:pharmacology; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D005938:Glucocorticoids; D006131:Growth Inhibitors; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D008156:Luciferases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D003022:Clotrimazole; D012293:Rifampin