PXR: Regulation and Pharmacogenomics

描述(申请人提供)：由配体激活的PXR诱导药物解毒基因继发的不良药物相互作用是导致药物疗效降低的重要临床问题。尽管我们能够使用快速体外筛选来识别激活PXR的化合物，但仍然没有办法预测个别患者的药物相互作用的程度。我们认为，人类PXR表达的差异是导致细胞色素P3A诱导的个体间差异的原因。此外，由PXR诱导的药物解毒基因的不断增加进一步证明了确定PXR可变表达的原因的必要性。我们先前的研究表明，PXR编码区的序列变异(SNPs，单核苷酸多态)并不显著地影响人类PXR表达的变异，也不显著影响人类诱导CYP3A靶基因的变异。我们目前的研究旨在进一步阐明调控PXR表达的细胞和分子机制。我们先前的研究表明，PXR有多种不同的亚型，它们的氨基末端和配体结合域都不同。我们目前在AIM 1的研究旨在发现具有不同氨基末端和不同配体结合结构域的PXR在人体组织中的表达程度，以及这些PXR剪接变异体是否改变了功能。我们假设，这些变异的PXR微调PXR激活，创造了一层额外的调节，并最终影响CYP3A诱导表型。为了了解调控PXR表达的因素，目标2和4将确定肝脏富集型转录因子在调节人PXR转录和人肝脏PXR表达中的作用。为了可视化PXR在小鼠组织中的转录活性，在目标3中，我们将开发一种PXR启动子驱动荧光素酶报告基因的转基因小鼠。通过小鼠皮肤的生物发光成像将在最具信息量的生物条件下实时监测PXR转录--即在活的完整动物中。这种独特而强大的全身成像方法将使我们能够在不同的生理和药物治疗环境下可视化PXR在多个组织中的转录。总之，这项提案描述了一套全面的实验，以确定PXR是如何受到监管的。

DESCRIPTION (provided by applicant): Adverse drug interactions secondary to induction of drug detoxification genes by ligand activated PXR is an important clinical problem leading to decreased drug efficacy. Despite our ability to use rapid in vitro screens to identify compounds that activate PXR, there is still no way to predict the extent of drug interaction in individual patients. We believe that human variation in PXR expression contributes to the interindividual differences in induction of CYP3A. Moreover, the growing list of drug detoxification genes induced by PXR further justifies the need to determine what is responsible for variable PXR expression. Our prior studies have revealed that sequence variation (SNPs, single nucleotide polymorphisms) in the PXR coding region is not significantly contributing to human variation in PXR expression or human variation in induction of the CYP3A target gene. Our current research is designed to further elucidate the cellular and molecular mechanisms regulating PXR expression. Our prior studies have revealed that there are multiple isoforms of PXR that differ in their amino terminus and in the ligand binding domain. Our current research in Aim 1 is designed to discover the extent to which PXR with different amino termini and different ligand binding domains are expressed in human tissues and whether these PXR splice variants have altered function. We hypothesize that these variant PXRs fine tune PXR activation creating an additional layer of regulation, and ultimately influence the CYP3A inductive phenotype. Towards the goal of understanding factors regulating PXR expression Aims 2 and 4 will determine the role of liver enriched transcriptional factors in regulation of human PXR transcription and PXR expression in human liver. In order to visualize PXR transcriptional activity in mouse tissues, in Aim 3 we will develop a transgenic mouse with the PXR-promoter-driving a luciferase reporter. Bioluminescent imaging through the skin of the mouse will monitor in real-time PXR transcription under the most informative biological conditions-i.e., in a living intact animal. This unique and powerful whole body imaging approach will allow us to visualize PXR transcription in multiple tissues under different physiological and drug treated environments. In total this proposal describes a comprehensive set of experiments to determine how PXR is regulated.