Pregnane X receptor (PXR), extensively expressed in human tissues related to digestion and metabolism, is responsible for recognizing and detoxifying diverse xenobiotics encountered by humans. To comprehend the promiscuous nature of PXR and its ability to bind a variety of ligands, computational approaches, viz., quantitative structure-activity relationship (QSAR) models, aid in the rapid dereplication of potential toxicological agents and mitigate the number of animals used to establish a meaningful regulatory decision. Recent advancements in machine learning techniques accommodating larger datasets are expected to aid in developing effective predictive models for complex mixtures (viz., dietary supplements) before undertaking in-depth experiments. Five hundred structurally diverse PXR ligands were used to develop traditional two-dimensional (2D) QSAR, machine-learning-based 2D-QSAR, field-based three-dimensional (3D) QSAR, and machine-learning-based 3D-QSAR models to establish the utility of predictive machine learning methods. Additionally, the applicability domain of the agonists was established to ensure the generation of robust QSAR models. A prediction set of dietary PXR agonists was used to externally-validate generated QSAR models. QSAR data analysis revealed that machine-learning 3D-QSAR techniques were more accurate in predicting the activity of external terpenes with an external validation squared correlation coefficient (R2) of 0.70 versus an R2 of 0.52 in machine-learning 2D-QSAR. Additionally, a visual summary of the binding pocket of PXR was assembled from the field 3D-QSAR models. By developing multiple QSAR models in this study, a robust groundwork for assessing PXR agonism from various chemical backbones has been established in anticipation of the identification of potential causative agents in complex mixtures.

Anti-tuberculosis drug-induced hepatotoxicity (ATDH) is a serious adverse drug reaction, and its pathogenic mechanism is still largely unknown. Pregnane X receptor (PXR, encoded by the NR1I2 gene) is a ligand-dependent transcription factor, and rifampicin is a human PXR-specific activator. Rifampicin and isoniazid co-therapy targets porphyrin biosynthesis via PXR and results in hepatic protoporphyrin IX accumulation and subsequent liver injury. The present study aimed to investigate the associations between genetic polymorphisms in NR1I2 and ATDH in an Eastern Chinese Han population.
A 1:4 matched case-control study was conducted using 146 ATDH cases and 584 controls. Seven single nucleotide polymorphisms (SNPs) were detected and analysed. Multivariate conditional logistic regression analysis was used to estimate the association between genotypes and risk of ATDH by the odds ratios (ORs) with 95% confidence intervals (CIs), with liver disease history, hepatoprotectant use, smoking history and drinking history as covariates.
Patients carrying the GG genotype of rs7643645 were at a higher risk of ATDH than those carrying the AA genotype (OR = 1.864, 95% CI: 1.106-3.141, P = .020), and significant differences were also found under the recessive model (P = .029) and additive model (P = .021). Patients with a polymorphism at rs2276707 were at a reduced risk of ATDH under the recessive model (OR = 0.600, 95% CI: 0.364-0.988, P = .045). Subgroup analysis confirmed the relationship in mild hepatotoxicity cases under the additive model (rs7643645, OR = 1.429, 95% CI: 1.027-1.988, P = .034) and recessive model (rs2276707, OR = 0.478, 95% CI: 0.253-0.902, P = .023). Functional annotation using ENCODE data also indicated that rs2276707 and rs7643645 were located in the histone modification regions targeting enhancers or promoter (H3K4Me1, H3K4Me3 and H3K27Ac).
Based on this case-control study, SNPs rs7643645 and rs2276707 in NR1I2 may contribute to susceptibility to ATDH in Eastern Chinese Han anti-TB treatment patients. Further studies in larger varied populations are needed to validate our findings.

Time-resolved fluorescence resonance energy transfer (TR-FRET) protein-protein interaction assays, especially in the format of receptor coregulator (coactivator and corepressor) recruitment/repression assays, have been widely used in nuclear receptor research to characterize the modes of action, efficacies, and binding affinities of ligands (including their properties as agonists, antagonists, and inverse agonists). However, there has been only limited progress in using this assay format for pregnane X receptor (PXR). In this chapter, we discuss TR-FRET protein-protein interaction assays and focus on a novel PXR TR-FRET coactivator interaction assay that we have developed based on a PXR coactivator cocrystal study. This new PXR TR-FRET coactivator interaction assay can characterize the binding affinities of PXR ligands and also differentiate antagonists from agonists. This assay is very robust, with the signal remaining stable over a long incubation time (up to 300min has been tested). It can tolerate high concentrations of DMSO (up to 5%) and has a high signal-to-noise ratio (six under typical assay conditions). This newly developed PXR TR-FRET coactivator interaction assay has potential application in high-throughput screening to identify and characterize novel PXR agonists and antagonists.

The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are important nuclear receptors involved in the regulation of cellular responses from exposure to many xenobiotics and various physiological processes. Phenobarbital (PB) is a non-genotoxic indirect CAR activator, which induces cytochrome P450 (CYP) and other xenobiotic metabolizing enzymes and is known to produce liver foci/tumors in mice and rats. From literature data, a mode of action (MOA) for PB-induced rodent liver tumor formation was developed. A MOA for PXR activators was not established owing to a lack of suitable data. The key events in the PB-induced liver tumor MOA comprise activation of CAR followed by altered gene expression specific to CAR activation, increased cell proliferation, formation of altered hepatic foci and ultimately the development of liver tumors. Associative events in the MOA include altered epigenetic changes, induction of hepatic CYP2B enzymes, liver hypertrophy and decreased apoptosis; with inhibition of gap junctional intercellular communication being an associative event or modulating factor. The MOA was evaluated using the modified Bradford Hill criteria for causality and other possible MOAs were excluded. While PB produces liver tumors in rodents, important species differences were identified including a lack of cell proliferation in cultured human hepatocytes. The MOA for PB-induced rodent liver tumor formation was considered to be qualitatively not plausible for humans. This conclusion is supported by data from a number of epidemiological studies conducted in human populations chronically exposed to PB in which there is no clear evidence for increased liver tumor risk.

To assess potential PCB153-associated human health effects and risks, it is necessary to model past exposure. PCB153 blood concentrations, obtained from the AMAP biomonitoring programme, in Inuit women covering the years 1994-2006 at Disko Bay, 1999-2005 at Nuuk, and 1992-2007 at Nunavik were used to extrapolate body burden and exposure to the whole lifespan of the population by the one-compartment toxicokinetic model. By using risk characterisation modelling, calculated Hazard Quotients were higher than 1 between the years 1955 and 1987 for the 90th population percentile and during 1956-1984 for the 50th population percentile. Cancer risk for overall exposure of PCB153 ranged from 4.6×10(-5) to 1.8×10(-6) for the 90th percentile and 3.6×10(-5) to 1.4×10(-10) for the 50th percentile between 1930 and 2049, when central estimates or upper-bound slope factors were applied. Cancer risk was below 1×10(-6) for the same time period when a lower slope factor was applied. Significant future research requirements to improve health risk characterisation include, among others, larger sample sizes, better analytical accuracy, fewer assumptions in exposure assessment, and consequently, a better choice of the toxicity benchmark used to develop the hazard quotient.

The exposure to pesticides and toxic compounds in xenobiotic transport and metabolism genes has been shown to affect risk of developing multiple myeloma (MM). Therefore, we hypothesized that genetic variations in xenobiotic transport and metabolism regulator genes PXR (NR1I2) and CAR (NR1I3) could determine a difference in MM susceptibility. Ten tagging single-nucleotide polymorphisms (SNPs) for PXR and seven for the CAR genes were selected and genotyped in 627 MM cases and 883 controls collected in the context of the International Multiple Myeloma rESEarch (IMMEnSE) consortium. None of the 17 SNPs investigated showed significant association with MM risk either alone or when combined in haplotypes. Significant SNP-SNP interactions were not found, neither with 58 previously genotyped polymorphisms in ABC transporters. We can therefore exclude that common genetic variants in the xenobiotic transport and metabolism regulator genes PXR and CAR affect MM risk.

The pregnane X receptor belongs to the nuclear hormone receptor superfamily and is involved in the transcriptional control of numerous genes. It was originally thought that it was a xenobiotic sensor controlling detoxification pathways. Recent studies have shown an increasingly important role in inflammation and cancer, supporting its function in abrogating tissue damage. PXR orthologs and PXR-like pathways have been identified in several non-mammalian species which corroborate a conserved role for PXR in cellular detoxification. In summary, PXR has a multiplicity of roles in vivo and is being revealed as behaving like a \"Jekyll and Hyde\" nuclear hormone receptor. The importance of this review is to elucidate the need for discovery of antagonists of PXR to further probe its biology and therapeutic applications. Although several PXR agonists are already reported, virtually nothing is known about PXR antagonists. Here, we propose the development of PXR antagonists through chemical, genetic and molecular modeling approaches. Based on this review it will be clear that antagonists of PXR and PXR-like pathways will have widespread utility in PXR biology and therapeutics.

The human pregnane X receptor (hPXR) is a nuclear receptor that regulates the expression of phase I and II drug-metabolizing enzymes as well as that of drug transporters. In addition, this receptor plays a critical role in cholesterol homeostasis and in protecting tissues from potentially toxic endobiotics. hPXR is activated by a broad spectrum of low-affinity compounds including xenobiotics and endobiotics such as bile acids and their precursors. Crystallographic studies revealed a ligand binding domain (LBD) with a large and conformable binding pocket that is likely to contribute to the ability of hPXR to respond to compounds of varying size and shape. Here, we describe an in silico method that allowed the identification of nine novel hPXR agonists. We further characterize the compound 1-(2-chlorophenyl)-N-[1-(1-phenylethyl)-1H-benzimidazol-5-yl]methanesulfonamide (C2BA-4), a methanesulfonamide that activates PXR specifically and more potently than does the reference compound 4-[2,2-bis(diethoxyphosphoryl)ethenyl]-2,6-ditert-butyl-phenol (SR12813) in our stable cell line expressing a Gal4-PXR and a GAL4 driven luciferase reporter gene. Furthermore treatment of primary human hepatocytes with C2BA-4 results in a marked induction of the mRNA expression of hPXR target genes, such as cytochromes P450 3A4 and 2B6. Finally, C2BA-4 is also able to induce hPXR-mediated in vivo luciferase expression in HGPXR stable bioluminescent cells implanted in mice. The study suggests new directions for the rational design of selective hPXR agonists and antagonists.

The use of herbals/botanicals has been gaining wide popularity in recent years in the United States as well as in other parts of the world. The mechanism of action of most of these herbals/botanicals has not been subjected to thorough scientific investigations. St. John\'s wort (Hypericum perforatum) represents a useful case study in this sense. Traditionally, it is used as a natural treatment for depression; however, in recent years its molecular mechanism of action has been elucidated by a number of laboratories across the world. Such studies have helped understand potential interactions of St. John\'s wort with drugs and other xenobiotics. St. John\'s wort activates a nuclear receptor called pregnane X receptor (PXR). PXR is a ligand-activated transcription factor that induces a number of xenobiotic-metabolizing enzymes and transporters including cytochrome P4503A4 (CYP3A4) in humans. Because CYP3A4 alone metabolizes about 60% of all clinically relevant drugs, induction of CYP3A4 may result in the rapid elimination of these drugs and a consequent reduction in drug efficacy. Ironically, such enzyme-inducing effects may not produce any immediate adverse symptomatology in the person taking St. John\'s wort. Therefore, the case of St. John\'s wort should serve as a good example of the usefulness and importance of studies on the mechanism of action of the herbals/botanicals, particularly those with widespread use. Scientists, physicians, and other health professionals can make use of the knowledge from such studies as an additional risk management tool.