Toxicological importance of CYP3A4 catalysis and inhibition

 描述（由申请方提供）：人细胞色素P450 3A 4（CYP 3A 4）是人体中含量最高且临床相关的异源代谢P450，因其在底物特异性和变构行为方面的极端混杂性而闻名。除了药物和内源性化合物的代谢外，CYP 3A 4还参与杀虫剂、杀虫剂、除草剂、致癌物和环境污染物的氧化和生物活化。CYP 3A 4也可被外源性物质抑制和激活，这可能导致不期望的药物-药物相互作用、毒性和人类死亡。尽管CYP 3A 4在药物代谢中起着中心作用，但其抑制和激活机制仍知之甚少。该提案旨在解决CYP 3A 4研究两个领域的关键问题，并集中于我们在CYP 3A 4-配体结合过程的结构和功能研究方面的进展。目的1的目标是优化CYP 3A 4抑制剂药效团模型，该模型是基于我们对利托那韦类似物的研究而建立的，利托那韦是迄今为止上市的最有效的药效增强剂。我们将使用新的，合理设计的化合物，这将进一步优化的基础上结构和功能的解释准确地确定药效决定因素。基于结构的CYP 3A 4特异性抑制剂药效团可以帮助在开发用于农业和公共卫生的药物和其他化学品期间识别和早期消除潜在的CYP 3A 4灭活剂，以及设计更有效的药物增强剂。该提案的第二部分集中在CYP 3A 4的变构行为，认为是由多底物结合引起的。CYP 3A 4可以被各种外源性物质和天然膳食化合物（如植物黄酮）激活，但其潜在机制仍在争论中。我们最近的研究结果表明，底物通道，而不是以前确定的周边地区作为一个高亲和力的效应结合区。目的2的目的是通过比较类黄酮样分子激活CYP 3A 4的外周和通道内位点的相对重要性来检验这一假设。使用一个综合的结构，生物化学和生物物理的方法，我们将评估这些领域的破坏如何影响蛋白质动力学，底物结合化学计量，动力学和代谢。这些研究将提供新的见解CYP 3A 4的激活机制，底物协同性和药物-药物相互作用，这是必不可少的预测/预防化学毒性和了解药物，膳食化合物和环境化学品的代谢的个体差异。

 DESCRIPTION (provided by applicant): Human cytochrome P450 3A4 (CYP3A4) is the most abundant and clinically relevant xenobiotic-metabolizing P450 in humans and is well known for its extreme promiscuity in substrate specificity and allosteric behavior. In addition to the metabolism of drugs and endogenous compounds, CYP3A4 is implicated in oxidation and bioactivation of pesticides, insecticides, herbicides, carcinogens and environmental pollutants. CYP3A4 can also be inhibited and activated by xenobiotics, which may lead to undesired drug-drug interactions, toxicity and human death. Despite the central role of CYP3A4 in drug metabolism, its mechanism of inhibition and activation are still poorly understood. This proposal is designed to address key issues in both areas of the CYP3A4 research and centers on our advances in the structural and functional investigation of the CYP3A4- ligand binding process. The goal of Aim 1 is to optimize the CYP3A4 inhibitor pharmacophore model that was built based on our studies of analogs of ritonavir, the most potent pharmacoenhancer marketed to date. We will accurately identify the pharmacophoric determinants using novel, rationally designed compounds, which will be further optimized based on structural and functional interpretations. The structure-based CYP3A4-specific inhibitor pharmacophore could assist in identification and early elimination of potential CYP3A4 inactivators during development of drugs and other chemicals used in agriculture and public health, as well as designing more potent pharmacoenhancers. The second part of the proposal focuses on the allosteric behavior of CYP3A4, thought to arise from the multiple substrate binding. CYP3A4 can be activated by various xenobiotics and natural dietary compounds, such as plant flavonoids, but the underlying mechanism is still under debate. Our recent findings suggest that the substrate-access channel rather than the previously identified peripheral area serves as a high affinity effector binding sie. The goal of Aim 2 is to test this hypothesis by comparing the relative importance of the peripheral and intra-channel sites in the CYP3A4 activation by flavonoid-like molecules. Using an integrated structural, biochemical and biophysical approach, we will assess how disruption of these areas affects protein dynamics, substrate binding stoichiometry, kinetics and metabolism. These studies will provide novel insights into the mechanism of CYP3A4 activation, substrate cooperativity and drug-drug interactions, which is essential for prediction/prevention of chemical toxicity and understanding interindividual differences in metabolism of drugs, dietary compounds and environmental chemicals.