A high-throughput cell-based reporter gene system for measurement of CYP1A1 induction.
BACKGROUND
Enzyme induction is undesirable in new drug discovery process, with consequences spanning from auto-induction to toxicity. Cytochrome P450 (CYP) 1A1 has long been known to be one of the metabolic enzymes involved in activating many procarcinogens, the first step toward tumor formation during chemical carcinogenesis. Induction of CYP1A1 during drug treatment may predispose the patients to some risk of chemical carcinogenesis.


METHODS
Based on the signal-transduction mechanism of CYP1A1 induction, a high-throughput reporter-gene system was established by stable transformation of H4IIE cells to incorporate the luciferase gene under control of CYP1A1 promoter. This stable cell line was validated with known CYP1A1 inducers, such as 3-methylcholanthrene (3-MC), beta-naphthoflavone (beta-NF), alpha-naphthoflavone (alpha-NF) and 3-indocarbinol. Thirty in-house new chemical entities (NCEs) were then screened with this reporter-gene system, and also administered to rats to evaluate in vivo CYP1A1 induction.


RESULTS
CYP1A1 reporter gene system can be used to identify strong inducers, such as 3-MC, beta-NF and alpha-NF, and weak inducers, such as 3-indocarbinol. In vitro induction of 30 in-house compounds in reporter gene system did not correlate with in vivo induction in rat liver microsome measured by ethoxyresorufin-O-dealkylation (EROD) activity, but had a reasonable correlation with Western blot signals.


CONCLUSIONS
This reporter-gene system may be useful in eliminating compounds that can cause CYP1A1 induction at an early stage of drug discovery.
Cui
Xiaoming
X
Department of Exploratory Drug Discovery, Schering-Plough Research Institute, D209, K15-2700, Kenilworth, NJ 07033, USA. xioming.cui@spcorp.com
Palamanda
Jairam
J
Norton
Laura
L
Thomas
Ann
A
Lau
Yau Yi
YY
White
Ronald E
RE
Cheng
K-C
KC
D000818:Animals; D001571:Benzoflavones / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000201:enzymology; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis* / Q000235:genetics; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D015507:Drugs, Investigational / Q000494:pharmacology; D004790:Enzyme Induction; D017930:Genes, Reporter; D007211:Indoles / Q000494:pharmacology; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D008748:Methylcholanthrene / Q000494:pharmacology; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014407:Tumor Cells, Cultured; D019324:beta-Naphthoflavone / Q000494:pharmacology
D001571:Benzoflavones; D015507:Drugs, Investigational; D007211:Indoles; D008748:Methylcholanthrene; C011512:alpha-naphthoflavone; D019324:beta-Naphthoflavone; C016517:indole-3-carbinol; D008156:Luciferases; D019363:Cytochrome P-450 CYP1A1