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CYP2C19 and CYP2C9 : new aspects of pharmacogenetics and transcriptional regulation

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posted on 2024-09-02, 21:11 authored by Jessica Mwinyi

Cytochrome P450s (CYPs) are responsible for approximately 75% of the phase I-dependent drug metabolism. Several important polymorphisms in these enzymes are known to significantly affect the individual drug response. CYP2C9 and CYP2C19 are polymorphically expressed CYP family members which are responsible for the metabolism of many different clinically important drugs, e.g. anticoagulants, antidepressants and antiulcer drugs. This thesis focuses on novel aspects with regard to the regulation of CYP2C9 and CYP2C19 gene expression.

The influence of the recently found common allele variant CYP2C19*17 on CYP2C19 enzyme activity towards two CYP2C19 substrates was investigated in the frame of two pharmacokinetic in vivo studies. The studies compared the single-dose pharmacokinetics of omeprazole and the steady-state kinetics of escitalopram in healthy CYP2C19*17/*17 carriers with the pharmacokinetic outcome obtained for CYP2C19 wild-type subjects. While no significant differences in the metabolic efficacy were observed for escitalopram, CYP2C19*17/*17 subjects showed significantly lower plasma levels of omeprazole compared to CYP2C19 wild type carriers. This observation suggests that CYP2C19*17/*17 carriers might be at a higher risk for therapy failure during treatment with proton pump inhibitors.

The transcriptional regulation of CYP2C9 and CYP2C19 gene expression by GATA transcription factors and by estrogen receptor α (ERα) was investigated in vitro. In the proximal promoter regions of both the CYP2C9 and the CYP2C19 gene, two adjacent putative GATA-binding sites with an ER-binding half-site in their vicinity were predicted in silico and initially studied by luciferase gene reporter assay. HepG2 and Huh-7 hepatoma cells were transfected with CYP2C9 or CYP2C19 promoter fragment carrying pGL3basic-constructs along with expression vectors for the transcription factors GATA-2, GATA-4, or ERα. Luciferase activities driven by wild-type CYP2C19 or CYP2C9 promoter were highly increased by GATA-4 and GATA-2 in both cell lines, whereas mutations introduced into the GATA binding sites or the co-transfection of the GATA-4 antagonist FOG-2 caused a significant loss of luciferase activity. In contrast, treatment with estradiol derivatives of ERα-transfected cells caused a significant inhibition of CYP2C19 and CYP2C9 promoter activity that was antagonized by site-directed mutagenesis of the putative ERα-binding half-sites. Additionally, estradiol derivatives significantly suppressed both CYP2C9 and CYP2C19 mRNA expression in human hepatocytes, as measured by real time PCR. Electrophoretic mobility shift assays revealed sequence-specific binding of GATA-4, GATA-6, and ERα to the two adjacent GATA binding sites and to the predicted ER binding half sites, respectively. ChIP assay in the cultured cells furthermore confirmed the association of both GATA-4 and ERα with CYP2C9 and CYP2C19 gene promoter. In conclusion, we have established novel mechanisms of CYP2C9 and CYP2C19 transcriptional regulation that involve transcription factors from the GATA family and estrogen receptor 􀀀. The estrogen mediated regulation may explain the clinically observed inhibitory effects of oral contraceptives on CYP2C19 and CYP2C9 activity.

History

Defence date

2010-10-15

Department

  • Department of Physiology and Pharmacology

Publication year

2010

Thesis type

  • Doctoral thesis

ISBN

978-91-7457-049-6

Language

  • eng

Original publication date

2010-09-24

Author name in thesis

Mwinyi, Jessica

Original department name

Department of Physiology and Pharmacology

Place of publication

Stockholm

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