Cell-based high-throughput bioassays to assess induction and inhibition of CYP1A enzymes

Toxicol In Vitro. 2005 Mar;19(2):275-87. doi: 10.1016/j.tiv.2004.10.003.

Abstract

CYP1A is a subfamily of cytochrome P450 enzymes involved in the metabolism of numerous therapeutic drugs and in the bioactivation of procarcinogens to mutagens. Because of their diverse metabolic capacities, differences in expression of CYP1A enzymes may profoundly influence drug-drug interactions and drug or carcinogen activation and detoxification. Here, we demonstrate that cell-based bioassays are capable of identifying xenobiotics that either alter aryl hydrocarbon receptor (AhR)-mediated CYP1A levels or produce inhibition of enzyme activity. To assess induction, a stable cell line harboring a luciferase reporter driven by multiple dioxin response elements (DREs) was developed. Using this cell line, AhR agonists and antagonists were identified among drugs, dietary agents, and environmental compounds. Of the chemicals examined, the therapeutic agent omeprazole induced reporter gene activity 12.5+/-0.41 fold above control, whereas the phytochemical, chrysin and environmental pollutant, benzanthracene enhanced luciferase activity 3.3+/-0.03 and 28.7+/-1.7 fold above control, respectively. Several natural products, polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAHs) prevented TCDD-mediated increases in luciferase expression. For example, the botanical kava inhibited TCDD-mediated induction by 88%. Northern blot analyses of CYP1A1 in HepG2 cells treated with similar agents validated results generated in the stable cell line. The stable cells were further used to identify inhibitors of CYP1A-mediated metabolism. Resveratrol and furafylline exhibited dose-dependent decreases in CYP1A1 and CYP1A2 enzyme activities with IC50 values of 1.89 and 0.79 microM, respectively. In summary, chemicals that possess the ability to alter CYP1A expression or inhibit CYP1A enzyme activities can be rapidly identified with the cell-based bioassays described here.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Biological Assay / methods*
  • Cell Line, Tumor*
  • Cytochrome P-450 CYP1A1 / biosynthesis*
  • Cytochrome P-450 CYP1A1 / genetics
  • Cytochrome P-450 CYP1A2 / biosynthesis*
  • Cytochrome P-450 CYP1A2 / genetics
  • Dose-Response Relationship, Drug
  • Enzyme Induction / drug effects
  • Enzyme Inhibitors / classification
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation, Enzymologic
  • Genes, Reporter / drug effects
  • Hepatocytes / drug effects
  • Hepatocytes / enzymology*
  • Humans
  • Luciferases / genetics
  • Receptors, Aryl Hydrocarbon / agonists
  • Receptors, Aryl Hydrocarbon / antagonists & inhibitors
  • Xenobiotics / classification
  • Xenobiotics / pharmacology

Substances

  • Enzyme Inhibitors
  • Receptors, Aryl Hydrocarbon
  • Xenobiotics
  • Luciferases
  • Cytochrome P-450 CYP1A1
  • Cytochrome P-450 CYP1A2