CYP induction screening, pooled hepatocytes

The test compounds are incubated with plated cryopreserved human hepatocytes (pooled donor lot) after which the cells are lysed, and total RNA is isolated. Target mRNA-levels (CYP1A2, CYP2B6, CYP3A4) are then analyzed by real-time quantitative PCR. Possible toxicity of the study compounds towards the cells is monitored visually during the study. Separate cytotoxicity assessment (LDH release) can be included in the study.
check Protocol

Test concentrations: 3

Time points: 48h

Replicates: 3

Positive controls

RT-qPCR analysis

check Deliverables

Induction (xx-fold) based on CYP-mRNA levels compared to vehicle

check Reporting

Excel report
(full report available per request)

Background

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 enhanced metabolism of co-medicated drugs. CYP induction is a relatively slow phenomenon due to gene regulation events, such as nuclear receptor activation and synthesis of mRNA and enzyme. Exposure times for the assays are usually 24 – 72 hours, followed by PXR activation, mRNA and/or CYP activity assessment. As a part of our drug-drug interactions services, the CYP induction screening assays are tailored for cost-efficient simultaneous evaluation of higher number of compounds, whereas the regulatory compliant induction assay with human hepatocytes is a comprehensive package required by FDA and EMA. Each assay delivers CYP induction relative to vehicle control (x-fold) and / or EC50 values if feasible (concentration where 50 % of maximal induction is achieved). The induction assays are available for CYP1A2, CYP2B6 and CYP3A4. Assays for other CYPs are available upon request.

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