The role of CYP3A7 in the disposition and toxicity of HIV inhibitors in the developing infant

摘要 每年，感染艾滋病毒的患病婴儿和早产儿都要接受各种药物治疗， 事实上，这些药物从未在这一弱势群体中进行过安全性和有效性测试。主 许多这些药物的药代动力学（PK）和肝毒性特征的决定因素是细胞色素P450 新陈代谢. CYP 3A 7是发育中婴儿（≤6个月）的主要P450酶。后 地质年代）。最近发表的报告和我们自己的初步结果表明，CYP 3A 7 可以产生与成人CYP 3A 4显著不同的代谢谱。的这些差异 代谢可以改变儿科患者使用的许多HIV药物的PK参数和安全性， 导致效力降低或毒性增加。因此，迫切需要确定PK， 临床相关CYP 3A 7 HIV药物底物的代谢特征，以提高这些药物的安全性和有效性 婴儿的药物。本提案的目的是确定功能性后果和机械性 基于CYP 3A 7和成人CYP 3A 4之间HIV药物代谢的差异。我们的假设是 HIV药物代谢的差异是由于酶结构的变化， 药物清除、药物间相互作用和反应性代谢产物的产生的实质性改变。我们 第一个目的是开发一个基于生理学的PK（PBPK）模型，用于四种常见的 处方的HIV抑制剂：利托那韦、洛匹那韦、奈韦拉平和依法韦仑。为了实现这一目标，我们将监测药物 在体外用重组CYP 3A 7和CYP 3A 4酶以及人CYP 3A 7和人CYP 3A 4酶进行代谢并鉴定代谢产物 来自不同发育阶段的成人和婴儿的肝微粒体（HLM）。我们将利用 这些研究的结果，以产生发育中婴儿的预测PK模型。我们的第二个目标是 确定CYP 3A 7抑制能力、潜在的药物相互作用和每种药物的反应性代谢产物 这些抑制剂。最后，我们的第三个目标是确定HIV药物代谢差异的结构基础 在CYP 3A 7和CYP 3A 4之间观察到。为了做到这一点，我们建议获得X射线晶体结构， CYP 3A 7与底物利托那韦和奈韦拉平，并进行饱和转移差NMR， 鉴定对于酶-配体相互作用重要的氨基酸残基和配体官能团。我们 预计这些研究将使我们能够确定结构差异如何影响观察到的 代谢差异。拟议的研究是创新的，因为它试图改变目前的概念， 通过使用HLM产生用于HIV药物处置的改进的婴儿PBPK模型， 这种重要酶的结构。这项研究将通过提高我们的能力来改善健康结果， 准确地预测药物动力学和药物相互作用的一些重要的艾滋病毒药物处方， 发育中的婴儿此外，它将积极影响我们了解CYP 3A 7 HIV药物代谢的能力 这将有助于开发专门针对婴儿的新药。

ABSTRACT Each year sick and premature infants infected with HIV are treated with a variety of drugs, despite the fact that these drugs have never been tested for safety and efficacy in this vulnerable population. A primary determinant for the pharmacokinetic (PK) and hepatotoxicity profile of many of these drugs is cytochrome P450 metabolism. CYP3A7 is the predominant P450 enzyme in the in the developing infant (≤6 mos. Post- Gestational Age). Recently published reports and our own preliminary results have demonstrated that CYP3A7 can produce a metabolic profile significantly different than that of adult CYP3A4. These differences in metabolism can alter the PK parameters and safety profiles of many HIV drugs used in pediatric patients, leading to reduced efficacy or increased toxicity. Therefore, there exists a critical need to determine the PK and metabolic profiles of clinically relevant CYP3A7 HIV drug substrates to the improve safety and efficacy of these drugs in infants. The objective of this proposal is to determine the functional consequences and mechanistic basis of the differences in HIV drug metabolism between CYP3A7 and adult CYP3A4. Our specific hypothesis is that the differences in HIV drug metabolism are due to changes in enzyme structure which lead to substantial alterations in drug clearance, drug-drug interactions, and production of reactive metabolites. Our first aim is to develop a physiologically based PK (PBPK) model for the disposition of four commonly prescribed HIV inhibitors: ritonavir, lopinavir, nevirapine, and efavirenz. To achieve this, we will monitor drug metabolism and identify metabolites in vitro with both recombinant CYP3A7 and CYP3A4 enzymes and human liver microsomes (HLMs) from the adult and the infant at different stages of development. We will utilize the results from these studies to produce predictive PK models for the developing infant. Our second aim is to identify the CYP3A7 inhibitory capacity, potential drug-drug interactions, and reactive metabolites for each of these inhibitors. Finally, our third aim is to define the structural basis of the differences in HIV drug metabolism observed between CYP3A7 and CYP3A4. To do this, we propose to obtain an X-ray crystal structure for CYP3A7 with the substrates ritonavir and nevirapine, and perform Saturation Transfer Difference NMR to identify amino acid residues and ligand functional groups that are important for enzyme-ligand interaction. We expect that these studies will allow us to determine how structural differences contribute to the observed metabolic differences. The proposed research is innovative because it seeks to change the current conceptual framework by producing improved infant PBPK models for HIV drug disposition using HLMs, and novel crystal structures for this important enzyme. This research will improve health outcomes by improving our ability to accurately predict pharmacokinetics and drug-drug interactions of some important HIV drugs prescribed to the developing infant. Additionally, it will positively impact our ability to understand CYP3A7 HIV drug metabolism based on structure, which will be useful in the development of new drugs specifically targeted to infants.