In Vivo Responses of the Human and Murine Pregnane X Receptor to Dexamethasone in Mice

Abstract

ABSTRACT:Dexamethasone (DEX) is a potent and widely used anti-inflammatory and immunosuppressant glucocorticoid. It can bind and activate the pregnane X receptor (PXR), which plays a critical role as xenobiotic sensor in mammals to induce the expression of many enzymes, including cytochromes P450 in the CYP3A family. This induction results in its own metabolism. We have used a series of transgenic mouse lines, including a novel, improved humanized PXR line, to compare the induction profile of PXR-regulated drugmetabolizing enzymes after DEX administration, as well as looking at hepatic responses to rifampicin (RIF). The new humanized PXR model has uncovered further intriguing differences between the human and mouse receptors in that RIF only induced Cyp2b10 in the new humanized model. DEX was found to be a much more potent inducer of Cyp3a proteins in wild-type mice than in mice humanized for PXR. To assess whether PXR is involved in the detoxification of DEX in the liver, we analyzed the consequences of high doses of the glucocorticoid on hepatotoxicity on different PXR genetic backgrounds. We also studied these effects in an additional mouse model in which functional mouse Cyp3a genes have been deleted. These strains exhibited different sensitivities to DEX, indicating a protective role of the PXR and CYP3A proteins against the hepatotoxicity of this compound.Dexamethasone (DEX) is a synthetic glucocorticoid that is used to treat a wide variety of medical conditions. It is a potent anti-inflammatory and immunosuppressant, which is metabolized primarily by the liver and undergoes renal excretion (Minagawa et al., 1986).DEX induces cytochrome P450 (P450) genes, such as Cyp3a11 and Cyp2b10 in mice and CYP3A4 in humans Wrighton et al., 1985;Strom et al., 1996;Yanagimoto et al., 1997). The mechanism of this induction is complex and is mediated by a number of nuclear receptors. For example, the glucocorticoid receptor (GR) is essential for the DEX-mediated induction of Cyp2b proteins in mouse liver, but at high doses the induction of Cyp3a enzymes is mediated by other transcription factors (Schuetz et al., 2000). In addition, it has been reported that DEX induction of CYP3A4 in human hepatocytes is mediated by several transcription factors (Pascussi et al., 2001). At low (nanomolar) concentrations, CYP3A4 induction is mediated by a GR-dependent elevation of pregnane X receptor (PXR) expression, whereas at high (supramicromolar) concentrations, DEX directly binds to and activates PXR. In addition, submicromolar concentrations of DEX also enhance the expression of the constitutive androstane receptor (CAR) and the retinoid X receptor-␣ in human hepatocytes (Pascussi et al., 2000a,b). It has been shown that DEX is a more potent ligand for mouse PXR than the human receptor (Meehan et al., 1988;Moore et al., 2000), and hepatic induction of Cyp3a, but not Cyp2b, expression in mice is PXRdependent (Schuetz et al., 2000;Scheer et al., 2008).CYP3A plays a central role in DEX disposition in human liver because it catalyzes th...

Quantitative Prediction of Human Pregnane X Receptor and Cytochrome P450 3A4 Mediated Drug-Drug Interaction in a Novel Multiple Humanized Mouse Line

“…Mice with a deletion of the seven closely linked mouse Cyp3a genes on chromosome 5 have been previously reported (Scheer et al, 2010). To summarize, these Cyp3a(Ϫ/Ϫ)/3a13(ϩ/ϩ) mice were created from a C57BL/6 mouse ES cell line that was double-targeted with loxP sites at the Cyp3a57 and Cyp3a59 loci.…”

“…huPXR/huCAR/huCYP3A4/3A7 mice were obtained by breeding of huCYP3A4/3A7 mice with the previously described huCAR and huPXR mice (Scheer et al, 2008(Scheer et al, , 2010. Homozygous Cyp3a(Ϫ/Ϫ)/3a13(ϩ/ϩ), huCYP3A4/3A7, and huPXR/huCAR/huCYP3A4/3A7 mice appeared normal, could not be distinguished from WT mice, and had normal survival rates and fertility (data not shown).…”

“…Whereas the conventional approach of crossing a random insertion of a human transgene with a knockout of the corresponding mouse gene(s) requires the combination of two independently segregating genetic loci, the Cyp3a knockout and CYP3A4 humanization are linked in the huCYP3A4/3A7 model. Because PXR and CAR humanized models were generated by a targeted insertion strategy as well (Scheer et al, 2008(Scheer et al, , 2010, the combination of these different modifications in the multiple humanized huPXR/huCAR/huCYP3A4/3A7 model described in the present work was straightforward.…”

“…Expression in the intestine was constitutively high and was also inducible, although compared with the liver, the magnitude of the intestinal induction response was lower. When crossed with the previously described humanized mouse models for PXR (huPXR) (Scheer et al, 2010) and CAR (huCAR) (Scheer et al, 2008), the induction response with the above inducers in the huPXR/huCAR/huCYP3A4/3A7 mouse line was reversed. We show that the huPXR/huCAR/huCYP3A4/3A7 model allows the ranking of different PXR activators according to their potency to induce CYP3A4 expression in humans and that this model might be a useful tool to quantitatively predict PXR/CYP3A4-mediated drug-drug interactions in the clinic.…”

“…Many such models have become available over the last few years (for review, see Cheung and Gonzalez, 2008). For example, humanized mouse models for PXR (Xie et al, 2000;Ma et al, 2007;Scheer et al, 2008Scheer et al, , 2010, CAR (Zhang et al, 2002;Scheer et al, 2008), and CYP3A4 (Granvil et al, 2003;Yu et al, 2005;van Herwaarden et al, 2007) have been described by various groups. By combining two of these modifications, a PXR/CYP3A4 double humanized model was successfully generated (Ma et al, 2008).…”