Concept: The Use of Targeted Immunoaffinity Proteomics for Routine Assessment of In Vitro Enzyme Induction
| Autor: | Oliver Poetz, Thomas Ebner, Caroline MacLean, Frederik Weiß |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
Proteomics
CYP2B6 Pharmaceutical Science 030226 pharmacology & pharmacy 03 medical and health sciences Tissue culture 0302 clinical medicine Cytochrome P-450 Enzyme System Tandem Mass Spectrometry Humans Drug Interactions RNA Messenger Enzyme inducer Cells Cultured Messenger RNA biology CYP1A2 Cytochrome P450 Molecular biology In vitro Biochemistry 030220 oncology & carcinogenesis Enzyme Induction biology.protein Hepatocytes Chromatography Liquid |
| Zdroj: | Journal of pharmaceutical sciences. 106(12) |
| ISSN: | 1520-6017 |
| Popis: | In vitro investigations on enzyme induction are indispensable for assessing drug–drug interactions of drug candidates. Regulatory bodies require measurement of changes of mRNA in cultured human hepatocytes. However, such data provide only indirect assessments of effects of enzyme induction in vivo . We describe the quantification of cytochrome P450 (CYP) enzyme protein levels by liquid chromatography–mass spectrometry for the routine assessment of enzyme induction. Protein concentration of CYP1A2, 2B6, 3A4, and 2C8 were measured in human hepatocytes after incubation with prototypical enzyme inducers and drug candidate BI-X using an antibody-based capturing method. In addition, CYP mRNA levels and CYP enzyme activities were determined. Except for CYP2B6, mRNA levels consistently showed more pronounced induction effects than CYP activity or CYP protein concentration. Induction of CYP activities was better reflected on the level of CYP protein. The described method requires small sample amounts and can be integrated in routine in vitro enzyme induction studies using tissue culture in 48- and 96-well plates. Assessment of changes of enzyme protein levels adds valuable information to conventional measurements of enzyme induction and can improve the use of in vitro data for the prediction of clinical outcomes. |
| Databáze: | OpenAIRE |
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