Hormonal Regulation of Human CYP3A

描述(由申请人提供)：在小肠中，细胞色素P3A4活性的个体间差异导致了作为细胞色素P3A底物的许多药物的口服生物利用度低且可变。这种差异似乎是由于十二指肠肠上皮细胞中细胞色素P3A4的特定含量存在很大差异的结果。从治疗的角度来看，首次通过的肠道提取效率的巨大差异可能导致不同的全身性药物暴露和不同的药理作用，在口服适合“一般”患者的剂量后，增加治疗失败和不良毒性的风险。肠道细胞色素P3A表达变异的原因在很大程度上尚不清楚，但被认为与遗传和环境因素有关。重要的是，我们以前已经证明了最具生物活性的维生素D形式，1，25-二羟基维生素D3(1，25(OH)2D3)，以VDR依赖的方式增强CYP3A4基因的转录，而CYP3A4反过来可以催化1，25(OH)2D3的代谢清除。这项赠款提案的总体目标是确定在体内是否通过以下顺序过程来调节肠道中的CYP3A4：25(OH)D3-葡萄糖醛酸脂结合物的形成和胆汁排泄，该结合物在近端肠腔中被水解为25(OH)D3，并被初级肠道细胞吸收，在那里它被CYP27B1转化为1，25(OH)2D3。肠道细胞以这种方式产生的活性激素可以调节CYP3A4和其他VDR靶基因的表达，包括钙转运蛋白TRPV6和Calbindin D9k。我们将通过确定和表征参与人体胆汁中维生素D结合物排泄的肝脏转运蛋白，并测试这些结合物是否可以影响培养的人肠道细胞中VDR靶基因的表达和功能来验证这一假设。由于CYP3A4可以催化1，25(OH)2D3的氧化代谢，我们还提出，已知的受体激动剂激活小肠中的hPXR可以促进肠道1，25(OH)2D3的清除，导致钙转运体的形成减少，并可能改变全身钙稳态指标。我们将使用培养的人肝细胞、肠细胞和原代肾小管上皮细胞来验证这一假设，这是一种新的人体肠道粘膜的微流体三维模型，并在健康志愿者中进行了体内CYP3A4相互作用研究。阐明细胞色素P3A依赖的药物代谢个体间差异的分子基础，可以通过更清楚地了解其他药物、环境和疾病状态可能如何影响作为肠道细胞色素P3A底物的新药候选药物的处置，从而提高制药业开发安全有效药物的能力。此外，如果我们关于CYP3A4参与小肠内1，25(OH)2D3基因组效应的负反馈控制的假设被证明是正确的，那么它可能指向相对简单的方法(例如，饮用西柚汁)来防止有效的hPXR激动剂对“高危”患者的骨健康的不利影响。

DESCRIPTION (provided by applicant): Inter-individual differences in the activity of CYP3A4 in the small intestine contribute to the low and variable oral bioavailability observed for many drugs that are CYP3A substrates. This variability appears to be the result of large differences in the specific content of CYP3A4 in duodenal enterocytes. From a therapeutic perspective, large differences in first-pass intestinal extraction efficiency can lead to variable systemic drug exposure and variable pharmacological effects following oral administration of doses that are appropriate for the "average" patient, increasing the risk of therapeutic failure and adverse toxicity. The cause of variable intestinal CYP3A expression is largely unknown, but thought to involve both genetic and environmental factors. Importantly, we have shown previously that the most biologically active form of vitamin D, 1,25-dihydroxy vitamin D3 (1,25(OH)2D3), enhances transcription of the CYP3A4 gene in a VDR-dependent manner and that CYP3A4 in turn can catalyze the metabolic clearance of 1,25(OH)2D3. The overall objectives of this grant proposal are to determine whether or not intestinal CYP3A4 is regulated in vivo by the following sequential process: formation and biliary excretion of a 25(OH)D3-glucuronide conjugate, hydrolysis of the conjugate to 25(OH)D3 in the proximal intestinal lumen and its absorption into the primary enterocytes, where it is converted to 1,25(OH)2D3 by CYP27B1. Active hormone produced in the enterocyte in this manner can regulate the expression of CYP3A4 and other VDR target genes, including the calcium transport proteins TRPV6 and calbindin D9k. We will test this hypothesis by identifying and characterizing the hepatic transporters involved in the biliary excretion of vitamin D conjugates in humans and testing whether or not these conjugates can affect the expression and function of VDR target genes in cultured human enterocytes. Because CYP3A4 can catalyze the oxidative metabolism of 1,25(OH)2D3, we also propose that activation of hPXR in the small intestine by known receptor agonists enhances intestinal 1,25(OH)2D3 clearance, resulting in a decrease in the formation of calcium transporters, and a potential change in systemic indices of calcium homeostasis. We will test this hypothesis with the use of cultured human hepatocytes, enterocytes and primary tubular epithelial cells, a novel microfluidic, 3-dimensional model of the human intestinal mucosa and the conduct of an in vivo CYP3A4 interaction study in healthy volunteers. Elucidating the molecular basis of inter-individual differences in CYP3A-dependent drug metabolism could enhance the ability of the drug industry to develop safe and efficacious drugs through a clearer understanding of how other medications, the environment, and disease states might impinge on the disposition of new drug candidates that are intestinal CYP3A substrates. In addition, if our hypothesis about the participation of CYP3A4 in negative feedback control of 1,25(OH)2D3 genomic effects within the small intestine proves correct, it could point to relatively simple ways (e.g., grapefruit juice consumption) to prevent the adverse effects of potent hPXR agonists on bone health in "at-risk" patients.