RT Journal Article SR Electronic T1 Cytochrome P450 Induction in Rat Hepatocytes Assessed by Quantitative Real-Time Reverse-Transcription Polymerase Chain Reaction and the RNA Invasive Cleavage Assay JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 1243 OP 1250 VO 29 IS 9 A1 Michael E. Burczynski A1 Michael McMillian A1 James B. Parker A1 Stewart Bryant A1 Angelique Leone A1 Elfrida R. Grant A1 Jacqueline M. Thorne A1 Zhong Zhong A1 Robert A. Zivin A1 Mark D. Johnson YR 2001 UL http://dmd.aspetjournals.org/content/29/9/1243.abstract AB The acceleration of drug discovery due to combinatorial chemistry and high-throughput screening methods has increased the numbers of candidate pharmaceuticals entering the drug development phase, and the capability to accurately predict whether drug candidates will induce various members of the drug-metabolizing cytochrome P450 (CYP) enzyme superfamily is currently of great interest to the pharmaceutical industry. In the present study, we describe the rapid and reliable analysis of CYP induction in a readily obtained model system (cultured rat hepatocytes) using both real-time quantitative reverse transcription-polymerase chain reaction (real-time RT-PCR) and the RNA invasive cleavage assay. The levels of members in the three primary inducible rat CYP subfamilies (CYP1A1, CYP2B1/2, and CYP3A1) were analyzed in untreated and induced (β-naphthoflavone, phenobarbital, and hydrocortisone) hepatocyte cultures under various media conditions to screen for optimal CYP induction profiles. The fold inductions measured by real-time RT-PCR and the RNA invasive cleavage assay were also compared with enzyme activity measurements in parallel cultures using liquid chromatography/double mass spectrometry-based assays, and the sensitivity and the specificity of the two RNA analysis methods were compared. Using these techniques, various culture conditions were examined for optimizing induction of the three CYP subfamily members. Both real-time RT-PCR and the RNA invasive cleavage assay prove to be effective methods for determining the effects of drugs on specific CYPs in primary rat hepatocytes. The American Society for Pharmacology and Experimental Therapeutics