Projects
Transcriptional regulation of drug metabolizing enzymes
Drug metabolizing
enzymes are transcriptionally regulated by several steroid (glucocorticoid),
nuclear (retinoid, VDR), xeno- (PXR, CAR, AhR) receptors. However, exact
molecular mechanisms underlying the role of these receptors in regulation of
DMEs is not fully understood yet. In particular, there is increasing
indication from literature and from experiments, that not only the
receptors, but also their co-activators, co-represors and other
transcriptional factors are involved and that the orchestration of this
machinery is the limiting step in final transcriptional response.
We did significant piece of work in the investigation of glucocorticoid receptor role in transcriptional regulation of DMEs. There exist at least three basic mechanisms of GR action.
The classical approach was the search for glucocorticoid responsive elements in the gene promoters. First evidence was brought by Sabine Gerbal-Chaloin and colleagues, who identifies functional GRE in CYP2C9 promoter (Gerbal-Chaloin et al (2002) J Biol Chem 277:209-217). This was followed by other who found GRE in CYP2C8 and CYP2C19 promoter.
Second mechanism of GR action in DMEs regulation we call “regulationg the regulator”; i.e. we have demonstrated the existence of a cascade GR/PXR-CAR/P450 in human hepatocytes; i.e. the regulators of P450 – orphan receptors CAR and PXR are under transcriptional control of GR, since PXR and CAR contain GRE in their promoters.
Pascussi et al (2003) Mol Endocrinol 17:42-55
Pascussi et al (2000) Mol Pharmacol 58:361-372
Pascussi et al (2000) Mol Pharmacol 58:1441-1450
In ongoing study we have confirmed the existence of GR/PXR-CAR/P450, as we found that microtubule disruption leads to inhibition of GR transcriptional activity and consequently to down-regulation of PXR and CAR, and to impairment of P450 induction.
Dvorak et al (2003) Mol Pharmacol 64:160-169
Recently, we have
demonstrated that GR may indirectly transcriptionally activate nicotin-metabolizing
CYP2A6 in human hepatocytes, which represent non-conventional regulation via
GR.
Onica T., Nichols K., Larin M., Ng L., Maslen A., Dvorak Z., Pascussi J.M., Vilarem M.J., Maurel P., Kirby G. (2008) Dexamethasone-mediated up-regulation of human CYP2A6 involves the glucocorticoid receptor and increased binding of HNF4a to the proximal promoter. Mol Pharmacol 73(2):451-460.
We found that DEX induces CYP2A6 mRNA and protein in primary human hepatocytes. This process was GR-dependent (tested by RU486) and did not required proteosynthesis de novo (cycloheximide treatment).
Surprisingly, when we performed progressive truncation of CYP2A6 promoter, the DEX-responsivenes was delineated to basic promoter, and we did not find any GRE in CYP2A6 promoter. In addition, we found increased binding of HNF4a1 to HNF-RE in CYP2A6 in response to DEX, and demonstrated physical association between GR and HNF4a1. Hence we postulated non-classical transcriptional mechanism involving increased binding of GR-HNF4a1 to HNF-RE in CYP2A6 promoter.
Consistently, we have reported elsewhere that HNF4a and GR are essential factors for CYP2C9 and CYP3A4 inducibility in placental cell lines.
Pavek P., Cerveny L., Svecova L., Brysch M., Libra A., Vrzal R., Staud F., Ulrichova J., Fendrich Z., Dvorak Z. (2007) Examination of Glucocorticoid receptor α –mediated transcriptional regulation of P-glycoprotein, CYP3A4 and CYP2C9 genes in several placental trophoblast cell lines. Placenta 28(10):1004-1011.