Xenoreceptors of the nuclear receptor superfamily, such as pregnane X receptor (PXR), are liver-enriched ligand-activated transcription factors regarded as crucial sensors in xenobiotic exposure and detoxification. PXR controls transcription of many drug-handling genes and influx/efflux transporters, thus playing a crucial role in drug metabolism and excretion. Liver functions have been studied using primary human hepatocytes (PHHs), which, when conventionally cultured, undergo rapid de-differentiation, leaving them unsuitable for long-term studies. Recently, 3D PHHs called spheroids have emerged as an in vitro model that is similar to in vivo hepatocytes regarding phenotype and function and represents the first in vitro model to study the long-term regulation of drug-handling genes by PXR. In this study, we used mathematical modelling to analyze the long-term activation of PXR in 3D PHHs through expression kinetics of three key PXR-regulated drug-metabolizing enzymes, CYP3A4, CYP2C9, and CYP2B6 and the P-glycoprotein efflux transporter encoding gene, MDR1. PXR action in 3D PHHs was induced by the antibiotic rifampicin at two clinically relevant concentrations. The results confirmed that high rifampicin concentrations activated PXR nearly to its full capacity. The analysis indicated the highest PXR-induced transcription rate constant for CYP2B6. The rate constant dictating mRNA degradation associated with activated PXR was highest for CYP3A4. Moreover, we measured the metabolic activity of CYP3A4, CYP2C9, and CYP2B6 and quantified their metabolic rate constants. Metabolic activity rate constant of CYP3A4 was found to be the highest whereas that of CYP2B6 was found to be the lowest among the studied enzymes. Our results provide important insight into the regulation of PXR-target genes in 3D PHHs and show that mRNA expression and metabolic activity data can be combined with quantitative analysis to reveal the long-term action of PXR and its effects on drug-handling genes.

• MeSH
• Models, Biological MeSH
• Spheroids, Cellular * metabolism   MeSH
• Cytochrome P-450 CYP3A metabolism   genetics   MeSH
• Hepatocytes * metabolism   MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Pregnane X Receptor * metabolism   MeSH
• Gene Expression Regulation * MeSH
• Rifampin pharmacology   MeSH
• Receptors, Steroid * metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Cytochrome P-450 CYP3A MeSH
• Pregnane X Receptor * MeSH
• Rifampin MeSH
• Receptors, Steroid * MeSH

Xenobiotic receptors, such as the pregnane X receptor, regulate multiple host physiologic pathways including xenobiotic metabolism, certain aspects of cellular metabolism, and innate immunity. These ligand-dependent nuclear factors regulate gene expression via genomic recognition of specific promoters and transcriptional activation of the gene. Natural or endogenous ligands are not commonly associated with this class of receptors; however, since these receptors are expressed in a cell-type specific manner in the liver and intestines, there has been significant recent effort to characterize microbially derived metabolites as ligands for these receptors. In general, these metabolites are thought to be weak micromolar affinity ligands. This journal anniversary minireview focuses on recent efforts to derive potentially nontoxic microbial metabolite chemical mimics that could one day be developed as drugs combating xenobiotic receptor-modifying pathophysiology. The review will include our perspective on the field and recommend certain directions for future research. SIGNIFICANCE STATEMENT: Xenobiotic receptors (XRs) regulate host drug metabolism, cellular metabolism, and immunity. Their presence in host intestines allows them to function not only as xenosensors but also as a response to the complex metabolic environment present in the intestines. Specifically, this review focuses on describing microbial metabolite-XR interactions and the translation of these findings toward discovery of novel chemical mimics as potential drugs of the future for diseases such as inflammatory bowel disease.

• MeSH
• Ligands MeSH
• Receptors, Steroid * metabolism   MeSH
• Intestines MeSH
• Carrier Proteins MeSH
• Xenobiotics metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Research Support, N.I.H., Extramural MeSH
• Names of Substances
• Ligands MeSH
• Receptors, Steroid * MeSH
• Carrier Proteins MeSH
• Xenobiotics MeSH

The pregnane X receptor (PXR, NR1I2) is a xenobiotic-activated transcription factor with high levels of expression in the liver. It not only plays a key role in drug metabolism and elimination, but also promotes tumor growth, drug resistance, and metabolic diseases. It has been proposed as a therapeutic target for type II diabetes, metabolic syndrome, and inflammatory bowel disease, and PXR antagonists have recently been considered as a therapy for colon cancer. There are currently no PXR antagonists that can be used in a clinical setting. Nevertheless, due to the large and complex ligand-binding pocket (LBP) of the PXR, it is challenging to discover PXR antagonists at the orthosteric site. Alternative ligand binding sites of the PXR have also been proposed and are currently being studied. Recently, the AF-2 allosteric binding site of the PXR has been identified, with several compounds modulating the site discovered. Herein, we aimed to summarize our current knowledge of allosteric modulation of the PXR as well as our attempt to unlock novel allosteric sites. We describe the novel binding function 3 (BF-3) site of PXR, which is also common for other nuclear receptors. In addition, we also mention a novel allosteric site III based on in silico prediction. The identified allosteric sites of the PXR provide new insights into the development of safe and efficient allosteric modulators of the PXR receptor. We therefore propose that novel PXR allosteric sites might be promising targets for treating chronic metabolic diseases and some cancers.

• Keywords
• AF-2 site, BF-3 site, CAR, PAM-antagonist, PXR, allosteric site, pregnane X receptor,
• MeSH
• Allosteric Site MeSH
• Diabetes Mellitus, Type 2 * MeSH
• Furylfuramide MeSH
• Humans MeSH
• Ligands MeSH
• Pregnane X Receptor MeSH
• Receptors, Cytoplasmic and Nuclear MeSH
• Receptors, Steroid * metabolism   MeSH
• Xenobiotics MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Furylfuramide MeSH
• Ligands MeSH
• Pregnane X Receptor MeSH
• Receptors, Cytoplasmic and Nuclear MeSH
• Receptors, Steroid * MeSH
• Xenobiotics MeSH

The pregnane X receptor (PXR, encoded by the NR1I2 gene) is a ligand-regulated transcription factor originally described as a master regulator of xenobiotic detoxification. Later, however, PXR was also shown to interact with endogenous metabolism and to be further associated with various pathological states. This review focuses predominantly on such aspects, currently less covered in literature, as the control of PXR expression per se in the context of inter-individual differences in drug metabolism. There is growing evidence that non-coding RNAs post-transcriptionally regulate PXR. Effects on PXR have especially been reported for microRNAs (miRNAs), which include miR-148a, miR-18a-5p, miR-140-3p, miR-30c-1-3p and miR-877-5p. Likewise, miRNAs control the expression of both transcription factors involved in PXR expression and regulators of PXR function. The impact of NR1I2 genetic polymorphisms on miRNA-mediated PXR regulation is also discussed. As revealed recently, long non-coding RNAs (lncRNAs) appear to interfere with PXR expression. Reciprocally, PXR activation regulates non-coding RNA expression, thus comprising another level of PXR action in addition to the direct transactivation of protein-coding genes. PXR expression is further controlled by several transcription factors (cross-regulation) giving rise to different PXR transcript variants. Controversies remain regarding the suggested role of feedback regulation (auto-regulation) of PXR expression. In this review, we comprehensively summarize the miRNA-mediated, lncRNA-mediated and transcriptional regulation of PXR expression, and we propose that deciphering the precise mechanisms of PXR expression may bridge our knowledge gap in inter-individual differences in drug metabolism and toxicity.

• Keywords
• CYP3A4, Gene expression, Non-coding RNA, Post-transcriptional regulation, Pregnane X receptor, microRNA,
• MeSH
• Biological Variation, Population * MeSH
• Biotransformation MeSH
• Pharmacogenetics MeSH
• Pharmacogenomic Variants * MeSH
• Phenotype MeSH
• Transcription, Genetic * MeSH
• Genotype MeSH
• Humans MeSH
• MicroRNAs genetics   metabolism   MeSH
• RNA Processing, Post-Transcriptional * MeSH
• Pregnane X Receptor genetics   metabolism   MeSH
• RNA, Long Noncoding genetics   metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• MicroRNAs MeSH
• NR1I2 protein, human MeSH Browser
• Pregnane X Receptor MeSH
• RNA, Long Noncoding MeSH

Pregnane X receptor (PXR) is the major regulator of xenobiotic metabolism. PXR itself is controlled by various signaling molecules including glucocorticoids. Moreover, negative feed-back regulation has been proposed at the transcriptional level. We examined the involvement of the 3'-untranslated region (3'-UTR) of NR1I2 mRNA and microRNAs in PXR- and glucocorticoid receptor (GR)-mediated regulation of NR1I2 gene expression. PXR ligands were found to significantly downregulate NR1I2 mRNA expression in a set of 14 human hepatocyte cultures. Similarly, PXR was downregulated by PCN in the C57/BL6 mice liver. In mechanistic studies with the full-length 3'-UTR cloned into luciferase reporter or expression vectors, we showed that the 3'-UTR reduces PXR expression. From the miRNAs tested, miR-18a-5p inhibited both NR1I2 expression and CYP3A4 gene induction. Importantly, we observed significant upregulation of miR-18a-5p expression 6 h after treatment with the PXR ligand rifampicin, which indicates a putative mechanism underlying NR1I2 negative feed-back regulation in hepatic cells. Additionally, glucocorticoids upregulated NR1I2 expression not only through the promoter region but also via 3'-UTR regulation, which likely involves downregulation of miR-18a-5p. We conclude that miR-18a-5p is involved in the down-regulation of NR1I2 expression by its ligands and in the upregulation of NR1I2 mRNA expression by glucocorticoids in hepatic cells.

• Keywords
• 3′-UTR, 3′-untranslated region, CAR, constitutive androstane receptor, CYP3A4, cytochrome P450 3A4, Cytochrome P450 3A4, DEX, dexamethasone, DMEs, drug metabolizing enzymes, DMSO, dimethyl sulfoxide, ER, estrogen receptor, GRα, glucocorticoid receptor α, Gene expression, Gluc, Gaussia luciferase, Glucocorticoid, LBD, ligand binding domain, MRE, miRNA-response element, MicroRNA, NR, nuclear receptor, PB, phenobarbital, PCN, pregnenolone 16α-carbonitrile, PHHs, primary human hepatocytes, PPARα, peroxisome proliferator-activated receptor α, PXR, pregnane X receptor, Pregnane X receptor, RXRα, retinoid X receptor α, Regulation, Rif, rifampicin, SEAP, secreted alkaline phosphatase, miRNA, microRNA,
• Publication type
• Journal Article MeSH

The pregnane X receptor (PXR) is a drug/xenobiotic-activated transcription factor of crucial importance for major cytochrome P450 xenobiotic-metabolizing enzymes (CYP) expression and regulation in the liver and the intestine. One of the major target genes regulated by PXR is the cytochrome P450 enzyme (CYP3A4), which is the most important human drug-metabolizing enzyme. In addition, PXR is supposed to be involved both in basal and/or inducible expression of many other CYPs, such as CYP2B6, CYP2C8, 2C9 and 2C19, CYP3A5, CYP3A7, and CYP2A6. Interestingly, the dynamics of PXR-mediated target genes regulation has not been systematically studied and we have only a few mechanistic mathematical and biologically based models describing gene expression dynamics after PXR activation in cellular models. Furthermore, few indirect mathematical PKPD models for prediction of CYP3A metabolic activity in vivo have been built based on compartmental models with respect to drug⁻drug interactions or hormonal crosstalk. Importantly, several negative feedback loops have been described in PXR regulation. Although current mathematical models propose these adaptive mechanisms, a comprehensive mathematical model based on sufficient experimental data is still missing. In the current review, we summarize and compare these models and address some issues that should be considered for the improvement of PXR-mediated gene regulation modelling as well as for our better understanding of the quantitative and spatial dynamics of CYPs expression.

• Keywords
• Pregnane X receptor, gene regulation, mathematical models, simulation,
• MeSH
• Gene Regulatory Networks * MeSH
• Humans MeSH
• Pregnane X Receptor MeSH
• Receptors, Steroid genetics   metabolism   MeSH
• Cytochrome P-450 Enzyme System genetics   metabolism   MeSH
• Models, Theoretical * MeSH
• Feedback, Physiological MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH
• Names of Substances
• Pregnane X Receptor MeSH
• Receptors, Steroid MeSH
• Cytochrome P-450 Enzyme System MeSH