CYP2C9

Cytochrome P450 2C9 (CYP2C9) is an important enzyme in the human body that is responsible for the metabolism of a large number of drugs, including nonsteroidal anti-inflammatory drugs (NSAIDs) such as ibuprofen and diclofenac, as well as anticoagulants like warfarin. CYP2C9 plays a crucial role in the liver's ability to break down and eliminate these drugs from the body. CYP2C9 is primarily found in the liver and accounts for approximately 18% of the total cytochrome P450 protein present in liver microsomes. CYP2C9 is known for its genetic polymorphisms, which can significantly impact its enzymatic activity and drug metabolism.

Table 1. Common CYP2C9 substrates, inducers and inhibitors.

CYP2C9 Induction Assay Service

At Creative Biolabs, our CYP2C9 induction assay using hepatocytes involves the detection of mRNA levels via qPCR or assessment of enzyme activity via HPLC-MS/MS.

• CYP2C9 mRNA Induction

Hepatocytes are treated with various test compounds or control inducers at different concentrations for 72 hours at 37°C, with refreshed media every 24 hours. After incubation, the hepatocytes are harvested and mRNA is extracted using a commercially available kit following the manufacturer's instructions. The mRNA levels of the CYP2C9 enzyme are measured using qRT-PCR. The mRNA levels can indicate the induction or suppression of CYP enzymes by the test compounds. A significant increase in CYP2C9 mRNA levels in treated hepatocytes compared to untreated controls indicates the induction of CYP2C9 activity by the test compound. The inducer control helps verify that the test compound is capable of upregulating the expression of the target CYP enzyme, and serves as a reference point for evaluating the extent of induction in other treatment groups within the assay.

• CYP2C9 Enzyme Induction

Hepatocytes are treated with various test compounds or control inducers at different concentrations for 72 hours at 37°C, with refreshed media every 24 hours. After 72 hours, a selective substrate specific to CYP2C9 is added to the reaction mixture for 2 hours. The reaction mixture is then analyzed using HPLC-MS/MS to detect and quantify the metabolites produced by the CYP2C9 enzyme.

Key features of our CYP2C9 induction assay using hepatocytes by qPCR include:

• Utilization of primary human hepatocytes to closely mimic in vivo conditions
• Robust quantification of mRNA levels using qPCR technology
• Functional assessment of CYP2C9 enzyme activity by HPLC-MS/MS
• Reliable assessment of CYP2C9 induction potential and inducer specificity
• Detailed analysis and interpretation of the results provided by our experienced team

Contact Us

Creative Biolabs is a leading provider of ADME services for drug development. We are committed to offering in vitro ADME tests, including CYP induction assay, CYP inhibition assay, and ADME screening tests. Our team of experts helps you assess the pharmacokinetic properties of your compounds to guide drug design and development. If you are interested in performing a CYP2C9 induction assay, we will provide you with a comprehensive and tailored experimental approach. Please let us know if you would like to learn more about this assay or have any specific requirements.

Support Data

Data 1: The experiment is used to determine if RG108 affects human CYP enzymes. The result showed that RG108 upregulated the expression level of the CYP2C9 gene.

Host cell: HepG2 cells

Incubation time: 72h

Analysis method: qRT-PCR

Fig.1 Expression levels of CYP2C9.^1,2

References

1. Nakamura, Kazuaki, et al. "Zebularine upregulates expression of CYP genes through inhibition of DNMT1 and PKR in HepG2 cells."Scientific Reports 7.1 (2017): 41093.
2. Distributed under Open Access license CC BY 4.0, without modification.

For Research Use Only | Not For Clinical Use