In vitro CYP Induction 

Cytochrome P450 (CYP) induction by a new chemical entity results in the increased amount and activity of relevant drug-metabolizing CYPs, which may lead to increase the metabolism of co-medicated drugs, or the metabolism of itself (autoinduction). That will reduce plasma levels, resulting in a decrease in efficacy. Induction of cytochrome P450 enzymes can also lead to toxicity by increasing reactive metabolite formation. Because CYP induction could pose a significant risk to patients, induction-mediated drug-drug interaction (DDI) needs to be carefully evaluated to determine and/or predict their safety risk. The newly FDA guidance is a strong suggestion that the in vitro DDI studies should be done earlier in the drug discovery.

MB Biosciences offer CYP induction assays to understand the potential drug-drug interaction liabilities of your compounds. We deliver reliable, high quality data with study designs based on the criteria and recommendations of the FDA or EMA guidelines.

At MB Biosciences, we use human 3D hepatocyte spheroids culture and fully characterized human hepatocytes in sandwich culture for CYP450 enzyme induction assays. Our assay ensures similar composition and architecture to primary liver tissue, uptake and efflux hepatic transporters are functioning in the model. Our model has better in vitro – in vivo correlations and is superior at predicting clinical outcomes.

Our CYP induction studies includes the following programs: 

  • Cytochrome P450 induction Screening assays

  • Regulatory agency-driven cytochrome P450 (CYP) induction services

  • PXR, CAR and AhR Nuclear Receptor Activation Assay

Our CYP induction studies

  • 2-week data turnaround

  • Best Price Guarantee 

  • Better in vitro – in vivo correlations

Regulatory Compliant Cytochrome P450 Induction Assay Protocol

Test System 3D human hepatocytes spheroids or Cryopreserved human hepatocytes (3 donors recommended). Fresh human hepatocytes are available on request
Test Compound Concentration 6 concentrations (dependent upon unbound Cmax, dose, solubility and cytotoxicity) plus vehicle control, in triplicate
CYP Isoforms Assessed CYP1A2, CYP2B6 and CYP3A4; For CYP2C, UGT or transporter studies, please contact directly for information
Number of Replicates 3
Negative Control Vehicle (0.1% DMSO)
Positive Controls Omeprazole (CYP1A2) Phenobarbital (CYP2B6) or CITCO (CYP2B6) Rifampin (CYP3A4)
Compound Requirements Dependent on top concentration (recommend 0.1% DMSO in incubation)
Exposure Period 72 hours (media changed every 24 hours)
Analysis Method qRT-PCR for relative mRNA expression levels
Data Delivery Full submission quality report, EC50 / Emax data, mRNA levels Fold induction above vehicle control, % of positive control induction


We also provide regulatory agency-driven cytochrome P450 (CYP) induction services in all the major CYP isoforms including CYP1A2, CYP2B6, CYP3A4, CYP2C8, CYP2C9, CYP2C19 at enzyme activity endpoints. Data include EC50 and Emax. The assay protocols are can be modified based on specific customer requirements.

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Cytochrome P450 induction Screening assays using 3D hepatocyte spheroids

Spheroids showed a significantly higher baseline level of CYP450 activity and induction over 2D monolayers. At MB Biosciences, we use human 3D hepatocyte spheroids culture for CYP450 enzyme induction assays. Our assay ensures that 3D environment that replicates native tissue, uptake and efflux hepatic transporters are functioning in the model, our model has better in vitro – in vivo correlations and is superior at predicting clinical outcomes.

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Benefits 

  • 3D hepatocyte spheroids more closely mimic normal liver tissue micro-anatomy

  • 3D hepatocyte spheroids are more phenotypically accurate than conventional 2D plated culture of cells

  • 3D hepatocyte spheroids maintenance of CYP450, uptake and efflux hepatic transporters activity in 3D in comparison with 2D culture

  • Our highly characterized 3D hepatocyte spheroids allow you to interrogate mechanisms of action of your compound.


Cytochrome P450 Induction Screening Protocol

Test System Human 3D primary hepatocyte spheroids culture ( one donor)
Test Article Concentration 1 or 3 plus vehicle control (or custom)
CYP Isoforms CYP1A2, CYP2B6 and CYP3A4 For CYP2C, UGT or transporter studies, please contact directly for information
Negative Control Non-inducer
Positive Control Omeprazole (CYP1A2) Phenobarbital (CYP2B6) or CITCO (CYP2B6) Rifampicin (CYP3A4)
Test Article Requirements Dependent on top concentration (recommend 0.1% DMSO in incubation)
Analysis Method qPCR of mRNA expression (CYP1A2, CYP2B6 and CYP3A4)
Data Delivery Assay methods mRNA levels, Fold induction above vehicle control % of positive control induction

Nuclear Receptor Interaction Screening

Screening for nuclear receptor activation is a widely used and accepted practice to predict adverse drug effects and drug-drug interactions. The pregnane X receptor (PXR), the aryl hydrocarbon receptor (AhR), and the constitutive androstane receptor (CAR) which are known to regulate CYP3A4, CYP1A2 and CYP2B6, respectively. The nuclear receptor interaction screening assay using human cell line co-transfected with DNA-binding fusion protein containing human PXR, AhR and CAR ligand binding domain together with luciferase reporter construct. After 24 hours exposure to the study compound, the luciferase activities are measured and compared to control incubations to deliver fold activation.

Screening assay design includes 6 concentrations in duplicate that can be used to calculate EC50 values. Also included in the study results are positive and negative (solvent) controls and cell viability of the test compound.

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