PXR cross-talks with internal and external signals in physiological and pathophysiological responses

PXR interaction with p53: a meeting of two masters

Pregnane X receptor (PXR) is a xenobiotic receptor that can be activated by numerous chemicals including endogenous hormones, dietary steroids, pharmaceutical agents, and environmental chemicals. PXR has been established to function as a xenobiotic sensor to coordinately regulate xenobiotic metabolism by regulating the expression of many enzymes and transporters required for xenobiotic metabolism. Recent studies have implicated a potentially important role for PXR in obesity and metabolic disease beyond xenobiotic metabolism, but how PXR action in different tissues or cell types contributes to obesity and metabolic disorders remains elusive. To investigate the role of adipocyte PXR in obesity, we generated a novel adipocyte-specific PXR deficient mouse model (PXRΔAd). Notably, we found that loss of adipocyte PXR did not affect food intake, energy expenditure, and obesity in high-fat diet-fed male mice. PXRΔAd mice also had similar obesity-associated metabolic disorders including insulin resistance and hepatic steatosis as control littermates. PXR deficiency in adipocytes did not affect expression of key adipose genes in PXRΔAd mice. Our findings suggest that adipocyte PXR signaling may be dispensable in diet-induced obesity and metabolic disorders in mice. Further studies are needed to understand the role of PXR signaling in obesity and metabolic disorders in the future. SIGNIFICANCE STATEMENT: The authors demonstrate that deficiency of adipocyte pregnane X receptor (PXR) does not affect diet-induced obesity or metabolic disorders in mice and infers that adipocyte PXR signaling may not play a key role in diet-induced obesity. More studies are needed to understand the tissue-specific role of PXR in obesity.

Hepatic Fatty Acid Transporter Cd36 Is a Common Target of LXR, PXR, and PPARγ in Promoting Steatosis

The pregnane X receptor (PXR) is an orphan nuclear receptor that regulates the expression of phase I and phase II drug metabolizing enzymes and transporters involved in the absorption, distribution, metabolism, and elimination of xenobiotics. PXR is expressed predominantly in the liver and intestine and resembles cytochrome P450s (CYPs), which is a phase I drug metabolizing enzyme. It is estimated that CYP 3As and CYP2Cs metabolize > 50% of all prescription drugs. PXR upregulates gene expression of these CYPs. Therefore, PXR plays a crucial role detoxifying xenobiotics and could potentially have effects on drug-drug interactions. PXR is reportedly responsible for activating a variety of target genes through cross-talk with other nuclear receptors and coactivators at transcriptional and translation levels. Recent findings have demonstrated the regulatory role of PXR and show the potential use of a PXR antagonist during drug therapy. In addition, genetic variations in the PXR gene are associated with the pharmacological effects of several drugs, and inter-individual differences in the clinical response are likely to be understood through these PXR polymorphisms. Many approaches have been used to explain the PXR regulatory mechanisms, such as microRNA-mediated PXR post-translational regulation and diverse PXR haplotype analysis. Understanding these PXR polymorphisms may lead to improving personalized therapeutic treatments.

The pregnane X receptor (PXR), a ligand-activated nuclear receptor, plays a central role in regulating the metabolism of both endogenous substances and xenobiotics. In recent years, increasing evidence has highlighted its involvement in chronic diseases, particularly metabolic disorders and cancer. PXR modulates drug-metabolizing enzymes, transporters, inflammatory factors, lipid metabolism, and immune-related pathways, contributing to the maintenance of hepatic–intestinal barrier homeostasis, energy metabolism, and inflammatory responses. Specifically, in type 2 diabetes mellitus (T2DM), PXR influences disease progression by regulating glucose metabolism and insulin sensitivity. In obesity, it affects adipogenesis and inflammatory processes. In atherosclerosis (AS), PXR exerts protective effects through cholesterol metabolism and anti-inflammatory actions. In metabolic dysfunction-associated steatotic liver disease (MASLD), it is closely associated with lipid synthesis, oxidative stress, and gut microbiota balance. Moreover, PXR plays dual roles in various cancers, including hepatocellular carcinoma, colorectal cancer, and breast cancer. Currently, PXR-targeted strategies, such as small molecule agonists and antagonists, represent promising therapeutic avenues for treating metabolic diseases and cancer. This review comprehensively summarizes the structural features, signaling pathways, and gene regulatory functions of PXR, as well as its role in metabolic diseases and cancer, providing insights into its therapeutic potential and future drug development challenges.

Background: Stevioside is used as a sweetener in food additives and pharmaceutical products. There is a growing concern about the adverse effects posed by stevioside usage. Stevioside is hydrolyzed into the aglycone steviol that is absorbed into the body. Recent studies suggest that steviol, but not stevioside, activates Pregnane X receptor (PXR) in human hepatic cells. In addition to the role in xenobiotic metabolism, the atherogenic and dyslipidemic effects of PXR have been revealed. Controversially, steviol not only increases the mRNA expression of PXR direct downstream gene CYP3A4, but also inhibits the enzymatic activities of CYP3A4. Thus, it is urgent to elucidate the molecular mechanisms by which steviol activates PXR signaling and to assess the possible adverse effects of steviol on pro-atherosclerotic events, such as dyslipidemia. Objective: Our study aims to explore the molecular mechanisms by which exposure to steviol activates human PXR and increases the risk of dyslipidemia. Methods: Both human hepatic and intestinal cells were used to test if steviol was a PXR agonist via transfection assay. The key residues within PXR’s ligand-binding pocket that steviol interacted with were investigated using a computational docking study with site-directed mutagenesis assay. Human intestinal cells were treated with steviol and/or PXR antagonist resveratrol (RES) to estimate the function of steviol in cholesterol uptake. Individual pairwise comparisons and other comparisons were analyzed with Student’s t-test and two-way ANOVA, respectively. Results: Steviol was a more potent agonist of human PXR than mouse PXR. Steviol promoted PXR to dissociate from its nuclear co-repressors. The key amino acids that were essential for the agonistic effects of steviol within PXR’s ligand binding pocket were established. Mechanistically, steviol induced gene expression of key intestinal cholesterol transporters, which led to increased cholesterol uptake by intestinal cells. Future studies in mice are needed to explore if exposure to steviol alters in vivo lipid profiles via PXR signaling. Our study provides potential evidence for the future risk assessment of steviol on cardiovascular disease. Keywords: Steviol, PXR, cholesterol uptake, dyslipidemia

Species differences in the induction and activation of the pregnane X receptor (PXR)

Preface. Abbreviations. Contributors. Chapter 1. Drug Metabolism: Significance and Challenges (Chandra Prakash and Alfin D.N. Vaz). 1.1. Introduction. 1.2. Phase I Drug Metabolizing Enzymes. 1.3. Phase II Conjugative Enzymes. 1.4. Drug Efflux Transporters. 1.5. Drug Uptake Transporters. 1.6. Challenges in Drug Metabolism. 1.7. Summary. 1.8. References. Chapter 2. Establishing Orphan Nuclear Receptors PXR and CAR as Xenobiotic Receptors (Tao Li, Junichiro Sonoda, and Ronald M. Evans). 2.1. Introduction. 2.2. Nuclear Receptor and Orphan Nuclear Receptor Superfamily. 2.3. Orphan Nuclear Receptors as Xenobiotic Receptors and Their Implications in Phase I Enzyme Regulation. 2.4. Perspectives. 2.5. References. Chapter 3. Nuclear Receptor-Mediated Regulation of Phase II Conjugating Enzymes (Olivier Barbier). 3.1. Introduction. 3.2. Phase II Drug Metabolizing Enzymes. 3.3. The Xenosensors CAR and PXR: 2 Masters Regulators of Phase II Metabolism. 3.4. AhR And Nrf2, Two Important Regulators of Phase II Enzymes. 3.5. PPARS and Phase II XMEs Regulation. 3.6. FXR/LXR and Phase II XMEs Regulation. 3.7. HNF and Phase II XMEs Regulation. 3.8. Regulation of Phase II Conjugating Enzymes by Steroid and Thyroid Receptors. 3.9. Concluding remarks and perspectives. 3.10. References. Chapter 4. Nuclear Receptor-Mediated Regulation of Drug Transporters (Oliver Burk). 4.1. Introduction. 4.2. Drug Transporters. 4.3. Induction of Drug Transporters by Activation of PXR and CAR. 4.4. Induction of Drug Transporters by Activation of PPARa. 4.5. Molecular Mechanism of PXR- and CAR-Dependent Drug Transporter Regulation. 4.6. Induction of Drug Transporter Expression and Drug Disposition. 4.7. Conclusions and Future Perspectives. 4.8. References. Chapter 5. Structure and Function of PXR and CAR (X. Edward Zhou and H. Eric Xu). 5.1. Introduction. 5.2. Structure and Function of PXR. 5.3. Structure and Function of CAR. 5.4. Concluding Remarks. 5.5. References. Chapter 6. Xenobiotic Receptor CoFactors and Coregulators (John Y. L. Chiang). 6.1. Regulation of PXR and CAR Nuclear Translocation. 6.2. Nuclear Receptor Coregulators and Epigenetic Regulation of Gene Transcription. 6.3. PXR and CAR Crosstalk with other Nuclear Receptors and Transcription Factors. 6.4. PXR and CAR Regulation of Lipid and Glucose Homeostasis. 6.5. Conclusion. 6.6. References. Chapter 7. Animal Models of Xenobiotic Nuclear Receptors and Their Utility in Drug Development (Haibiao Gong and Wen Xie). 7.1. Introduction. 7.2. PXR and CAR Loss-of-Function (Knock Out) Mouse Models. 7.3. PXR and CAR Gain-of-Function (Transgenic) Mouse Models. 7.4. Humanized Mouse Models. 7.5. Utility of Xenobiotic Mouse Models in Pharmaceutical Development. 7.6. Closing Remarks. 7.7. References. Chapter 8. Nuclear Receptors and Drug-Drug Interactions with Prescription Drugs and Herbal Medicines (Rommel G. Tirona and Richard B. Kim). 8.1. Introduction. 8.2. Prescription Drugs/Drug Classes Commonly Involved in Inductive Interactions. 8.3. Herbal Drug Medicines Commonly Involved in Inductive Interactions. 8.4. Pharmacology of Induction. 8.5. Clinical Aspects of Induction-Type Drug Interactions. 8.6. Inhibition of Nuclear Receptors in Clinical Drug Interactions. 8.7. Nuclear Receptor-Mediated Drug Side-Effects. 8.8. Perspectives. 8.9. References. Chapter 9. Genetic Variants of Xenobiotic Receptors and Their Implications in Drug Metabolism and Pharmacogenetics (Jatinder Lamba and Erin G. Schuetz). 9.1. PXR (Pregnane X Receptor) Background. 9.2. PXR Gene Structure. 9.3. PXR Alternative mRNAs. 9.4. Genetic Variants in PXR's Exons and their Functional Consequences. 9.5. Genetic Variants In Introns 2-8 and the 3'-UTR of PXR and their Functional Consequences. 9.6. Resequencing Strategy for the PXR Promoter and Intron 1. 9.7. Genetic Variation in the PXR Promoter and 5'-UTR and its Functional Relevance. 9.8. Genetic Variation in PXR's Intron 1 and its Functional Relevance. 9.9. In Silico Analysis for Functional Effect of SNPs in PXR's Promoter, 5'-UTR and Intron 1. 9.10. SNPs in PXR's Promoter and Intron 1 Affect Putative HNF Binding Sites. 9.11. PXR SNPs Have Been Associated with Intestinal and Hepatic Inflammation and Diseases. 9.12. PXR Structural Variation and other Genomic Features. 9.13. PXR Summary. 9.14. CAR-Background. 9.15. CAR Gene Structure. 9.16. CAR Alternatively Spliced RNAs. 9.17. CAR Genetic Variants (SNPs) and their Functional Consequences. 9.18. CAR Summary. 9.19. References. Chapter 10. Beyond PXR and CAR, Regulation of Xenobiotic Metabolism by Other Nuclear Receptors (Martin Wagner, Gernot Zollner, and Michael Trauner). 10.1. Introduction. 10.2. Farnesoid X Receptor. 10.3. Hepatocyte Nuclear Factor 4. 10.4. Vitamin D receptor. 10.5. Glucocorticoid Receptor. 10.6. Peroxisome Proliferator Activated Receptors. 10.7. Aryl Hydrocarbon Receptor (AhR). 10.8. Conclusions. 10.9. References. Chapter 11. Emerging Role of Retinoid-Related Orphan Receptor (ROR) and Its Crosst alk With LXR(Liver X Receptor) in the Regulation Of Drug-Metabolizing Enzymes (Taira Wada and Wen Xie). 11.1. Introduction. 11.2. Orphan Nuclear Receptor RORalpha. 11.3. A Potential Role of RORs in Xeno- and Endobiotic Gene Regulation. 11.4. LXR and its Regulation of Drug Metabolizing Enzymes. 11.5. A Functional Cross-Talk Between RORa and LXR in the Regulation of Xeno- and Endobiotic Genes. 11.6. Closing Remarks. Index.

Cannabidiol (CBD), a non-psychoactive phytocannabinoid of cannabis, is therapeutically used as an analgesic, anti-convulsant, anti-inflammatory, and anti-psychotic drug. There is a growing concern about the adverse side effects posed by CBD usage. Pregnane X receptor (PXR) is a nuclear receptor activated by a variety of dietary steroids, pharmaceutical agents, and environmental chemicals. In addition to the role in xenobiotic metabolism, the atherogenic and dyslipidemic effects of PXR have been revealed in animal models. CBD has a low affinity for cannabinoid receptors, thus it is important to elucidate the molecular mechanisms by which CBD activates cellular signaling and to assess the possible adverse impacts of CBD on pro-atherosclerotic events in cardiovascular system, such as dyslipidemia. Our study aims to explore the cellular and molecular mechanisms by which exposure to CBD activates human PXR and increases the risk of dyslipidemia. Both human hepatic and intestinal cells were used to test if CBD was a PXR agonist via cell-based transfection assay. The key residues within PXR's ligand-binding pocket that CBD interacted with were investigated using computational docking study together with site-directed mutagenesis assay. The C57BL/6 wildtype mice were orally fed CBD in the presence of PXR antagonist resveratrol (RES) to determine how CBD exposure could change the plasma lipid profiles in a PXR-dependent manner. Human intestinal cells were treated with CBD and/or RES to estimate the functions of CBD in cholesterol uptake. CBD was a selective agonist of PXR with higher activities on human PXR than rodents PXRs and promoted the dissociation of human PXR from nuclear co-repressors. The key amino acid residues Met246, Ser247, Phe251, Phe288, Trp299, and Tyr306 within PXR's ligand binding pocket were identified to be necessary for the agonistic effects of CBD. Exposure to CBD increased the circulating total cholesterol levels in mice which was partially caused by the induced expression levels of the key intestinal PXR-regulated lipogenic genes. Mechanistically, CBD induced the gene expression of key intestinal cholesterol transporters, which led to the increased cholesterol uptake by intestinal cells. CBD was identified as a selective PXR agonist. Exposure to CBD activated PXR signaling and increased the atherogenic cholesterol levels in plasma, which partially resulted from the ascended cholesterol uptake by intestinal cells. Our study provides potential evidence for the future risk assessment of CBD on cardiovascular disease, such as dyslipidemia.

Unique Transcription Start Sites and Distinct Promoter Regions Differentiate the Pregnane X Receptor (PXR) Isoforms PXR 1 and PXR 2

As a devastating neurological disease, spinal cord injury (SCI) results in severe tissue loss and neurological dysfunction. Pregnane X receptor (PXR) is a ligand-activated nuclear receptor with a major regulatory role in xenobiotic and endobiotic metabolism and recently has been implicated in the central nervous system. In the present study, we aimed to investigate the role and mechanism of PXR in SCI. The clip-compressive SCI model was performed in male wild-type C57BL/6 (PXR+/+ ) and PXR-knockout (PXR-/- ) mice. The N2a H2 O2 -induced injury model mimicked the pathological process of SCI in vitro. Pregnenolone 16α-carbonitrile (PCN), a mouse-specific PXR agonist, was used to activate PXR in vivo and in vitro. The siRNA was applied to knock down the PXR expression in vitro. Transcriptome sequencing analysis was performed to discover the relevant mechanism, and the NRF2 inhibitor ML385 was used to validate the involvement of PXR in influencing the NRF2/HO-1 pathway in the SCI process. The expression of PXR decreased after SCI and reached a minimum on the third day. In vivo, PXR knockout significantly improved the motor function of mice after SCI, meanwhile, inhibited apoptosis, inflammation, and oxidative stress induced by SCI. On the contrary, activation of PXR by PCN negatively influenced the recovery of SCI. Mechanistically, transcriptome sequencing analysis revealed that PXR activation downregulated the mRNA level of heme oxygenase-1 (HO-1) after SCI. We further verified that PXR deficiency activated the NRF2/HO-1 pathway and PXR activation inhibited this pathway in vitro. PXR is involved in the recovery of motor function after SCI by regulating NRF2/HO-1 pathway.

SUN-042 PREGNANE X RECEPTOR (PXR) IS A NOVEL THERAPEUTIC TARGET FOR THE TREATMENT OF CISPLATIN-INDUCED NEPHROTOXICITY IN MICE

RNA-Seq reveals common and unique PXR- and CAR-target gene signatures in the mouse liver transcriptome

The cellular response to DNA damage comprises of DNA repair, cell-cycle checkpoint control and DNA damage-induced apoptosis to promote genomic integrity and suppress tumorigenesis. However, cancer cells can modulate the DNA damage response by suppressing DNA repair, altering cell cycle checkpoint control and ultimately downregulating tumor suppressors that contribute to apoptosis promoting genomic instability and avoidance of apoptosis. Although PXR is well established as a xenobiotic nuclear receptor that plays a central role in xenobiotic metabolism and disposition, emerging evidence suggest PXR as a regulator of apoptosis, promoting a malignant phenotype, both in vitro and in vivo. The tumor suppressor p53 can be activated in the presence of DNA damage and can induce cell cycle arrest to allow for DNA repair, or ultimately apoptosis suppressing tumor formation. We hypothesized that the novel protein-protein interaction between PXR and p53 contributes to the oncogenic functions of PXR enhancing colon carcinogenesis. Using immunoprecipitation, we found that PXR forms a protein-protein interaction with p53, resulting in reduced p53 activation and expression of downstream pro-apoptotic genes. In addition, utilizing the soft agar assay, we found that PXR overexpression leads to increased malignant transformation in colon cancer cells. Our findings show for the first time that PXR can physically bind to p53, resulting in downregulation of p53 activity and increased malignant transformation in colon cancer cells. Thus, these results provide insight into the biological significance of novel protein-protein interactions with PXR and how the PXR-p53 protein-protein interaction influences p53-mediated mechanism of tumor suppression. Citation Format: Delira F. Robbins, Jing Wu, Taosheng Chen. Regulation of cellular apoptosis via a novel protein-protein interaction of tumor suppressor p53 with the xenobiotic pregnane X receptor (PXR) in colon cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1556. doi:10.1158/1538-7445.AM2014-1556

The pregnane X receptor (PXR), a ligand-activated transcription factor, regulates the expression of genes involved in endobiotic and xenobiotic metabolism, inflammation, and fibrosis. Disruption of PXR functions can affect processes critical to metabolic dysfunction-associated steatohepatitis (MASH) progression. Although ligand-dependent PXR functions are well studied, its regulation by post-translational modification, particularly phosphorylation, remains unclear. PXR has a conserved phosphorylation motif within its ligand binding domain (Ser347 in mice; Ser350 in humans). In vitro studies showed that this site mutation impairs human PXR transcriptional activity; however, the mechanism remains elusive. To investigate this phosphorylation site role in MASH development, wild-type and PXR Ser347Ala knock-in mutation (PXR-KI) mice were fed either a high-fat diet or a control chow diet for 16 weeks. On control chow diet, PXR-KI mice exhibited decreased expression of alternative bile acid (BA) synthesis genes compared with wild-type mice. On a high-fat diet, PXR-KI mice manifested more severe hepatic steatosis, revealed by elevated serum total cholesterol, and increased expression of genes involved in lipid metabolism. In addition, changes in BA metabolism and transporter genes suggested a cholestatic pattern in this group of mice. BA profiling showed higher levels of conjugated, hydrophilic, primary BA in the serum and liver, and increased unconjugated BA in the intestine. The data suggest that PXR Ser347 phosphorylation motif is essential for regulating PXR functions to maintain endobiotic metabolism and alleviate hepatotoxicity during MASH progression. SIGNIFICANT STATEMENT: The ligand-independent role of pregnane X receptor (PXR) is unclear. In phosphodeficient PXR knock-in mice, loss of Ser347 phosphorylation worsened hepatic steatosis and altered bile acid homeostasis under high-fat diet feeding, uncovering a novel role and therapeutic potential of PXR phosphorylation in fatty liver diseases.