Cytochrome P450 (CYP) induction assay

Aim: To identify the potential of test compound to induce CYP1A2, CYP2B6 or CYP3A4 in cultured human hepatocytes by evaluating mRNA levels.

Background information: Cytochrome P450 (CYP) induction plays an important role in the pharmacokinetics of a drug and can potentially affect drug efficacy by reducing plasma half-life, or drug toxicity if elevated levels of toxic metabolites are formed. Induction of CYP1A2, CYP2B6 and CYP3A4 gene expression is a representative endpoint for activation of the nuclear receptors (AhR, PXR and CAR) that mediate drug-induced changes in the expression of phase I and II enzymes and transporters.

After plating for 3 days, human hepatocytes are exposed to test compound, vehicle or positive control inducers in the medium at selected concentrations. The cell culture medium is replaced every 24 hours and incubations are conducted for 48 hours before mRNA analysis. Additionally, hepatocytes are seeded and exposed to chlorpromazine (cytotoxicity control).

Compound requirement: 5 mg

Turnaround time: 4 weeks 

Experimental design:

  • Test compound concentrations: 3 (variables on request and dependent on solubility and cytotoxicity)
  • DMSO content in incubation: 0.5%
  • Number of replicates: 3
  • Cell model: Inducible cryopreserved pooled human hepatocytes in monolayer
  • CYP isoforms: CYP1A2, CYP2B6 and CYP3A4 (other isoforms available)
  • Duration of exposure: 48 h
  • Endpoint: relative mRNA expression levels
  • Control inducers: Phenobarbital (CYP1A2), Rifampicin (CYP2B6) and Omeprazole (CYP3A4)
  • Analytical method: qRT-PCR

Delivered results:

  • Fold induction
  • Percentage of positive control