The Effect of Genetic Diversity on Mechanism-Based Inactivation of CYP2D6

遗传多样性对 CYP2D6 机制失活的影响

基本信息

批准号：
8433693
负责人：
LAURA L FURGE
金额：
$ 34.23万
依托单位：
KALAMAZOO COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-01-01 至 2016-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8433693
关键词：
Accounting Affect Age Amino Acids Apoproteins Biological Assay CYP2D6 gene Carcinogens Complex Computer Simulation Cytochrome P450 Development Digestion Drug Design Drug Evaluation Drug Interactions Elderly Enzymes Event Family Genetic Polymorphism Genetic Variation Goals Half-Life Health Heme High Pressure Liquid Chromatography Human Individual Individual Differences Investigation Kinetics Laboratories Lead Ligand Binding Liver Medical Medicine Metabolism Mind Molecular Biology Techniques Molecular Models Mutagenesis Paroxetine Pharmaceutical Preparations Pharmacologic Substance Pharmacotherapy Plant alkaloid Plasma Proteins Publications Reaction Research Role Site Structure Structure-Activity Relationship Students Substrate Specificity Techniques Testing Training Variant Vitamins Xenobiotics adduct base clinically relevant design drug metabolism ecstasy enzyme activity interest liquid chromatography mass spectrometry meetings molecular modeling mutant public health relevance research study response small molecule steroid metabolism success tool undergraduate research

项目摘要

DESCRIPTION (provided by applicant): Cytochrome P450 enzymes (CYPs) in humans are a family of heme-containing enzymes involved in metabolism of steroids, vitamins, eiconsanoids, and xenobiotics such as drugs, complex plant alkaloids, carcinogens, and other small molecules. The role of CYPs in metabolism of nearly all drugs has made this family of enzymes of considerable interest in medicine and for human health. CYP2D6 is one of the significant drug metabolizing enzymes responsible for metabolism of ~25% of pharmaceutical compounds while only accounting for a fraction of liver CYP content. The enzyme also displays multiple polymorphic forms that contribute to inter-individual difference in responses to drugs metabolized by CYP2D6. One caveat of such broad substrate specificity among drug metabolizing CYPs is that the enzymes are susceptible to inhibition by products of their own reactions. This type of inhibition is called mechanism-based inactivation and leads to irreversible inactivation of the enzyme. Compounds that act as mechanism-based inactivators are important tools for understanding structure/function of these enzymes and there are only a few known for CYP2D6. The project proposed here will contribute to understanding mechanisms inactivation of CYP2D6 and select allelic variants by two classes of inactivators - protein adductors and heme modifiers. These complimentary lines of investigation will be pursued through two aims: Aim 1: To determine the amino acid(s) of CYP2D6 targeted by mechanism-based protein adduct inactivators. In silico and LC/MS predictions of amino acids involved in apoprotein adduction will be tested by mutagenesis of postulated nucleophiles to non-nucleophilic amino acids. The expressed mutant enzyme(s) will be characterized in terms of kinetics with normal and inactivator substrates including two protein adductors (SCH66712 and EMTPP) and two heme modifiers (paroxetine and MDMA). Aim 2: To determine the interaction of the most common polymorphisms of CYP2D6 with inactivators. Almost 20% of drug therapies are affected by polymorphisms. Polymorphisms lead to different plasma concentrations and half-life of drugs that lead to adverse drug reactions. CYP2D6 is highly polymorphic and the role of polymorphisms in inactivation will be characterized. Together, these studies will provide an understanding of how in activators interact with CYP2D6 and lead to new opportunities in rational drug design and evaluation of inter-individual differences in drug metabolism in the age of personalized medicine.

描述（由申请人提供）：人类细胞色素 P450 酶（CYP）是一类含血红素的酶，参与类固醇、维生素、类二十烷酸和异生物质（如药物、复杂植物生物碱、致癌物和其他小分子）的代谢。 CYP 在几乎所有药物代谢中的作用使得该酶家族在医学和人类健康领域引起了极大的兴趣。 CYP2D6 是重要的药物代谢酶之一，负责约 25% 的药物化合物的代谢，但仅占肝脏 CYP 含量的一小部分。该酶还表现出多种多态性，导致个体间对 CYP2D6 代谢药物的反应存在差异。药物代谢 CYP 中如此广泛的底物特异性的一个警告是，这些酶容易受到其自身反应产物的抑制。这种类型的抑制称为基于机制的失活，并导致不可逆的酶的失活。作为基于机制的灭活剂的化合物是理解这些酶的结构/功能的重要工具，而 CYP2D6 已知的化合物只有少数。这里提出的项目将有助于理解 CYP2D6 失活机制，并通过两类失活剂（蛋白质加合剂和血红素修饰剂）选择等位基因变体。这些互补的研究路线将通过两个目标进行：目标 1：确定基于机制的蛋白质加合物灭活剂靶向的 CYP2D6 氨基酸。在计算机和 LC/MS 中，参与脱辅基蛋白加合的氨基酸预测将通过假定的亲核试剂诱变为非亲核氨基酸来测试。表达的突变酶将根据正常底物和失活底物的动力学进行表征，包括两种蛋白质加合物（SCH66712 和 EMTPP）和两种血红素修饰剂（帕罗西汀和 MDMA）。目标 2：确定 CYP2D6 最常见的多态性与灭活剂的相互作用。近 20% 的药物疗法受到多态性的影响。多态性导致药物的血浆浓度和半衰期不同，从而导致药物不良反应。 CYP2D6 具有高度多态性，将表征多态性在失活中的作用。总之，这些研究将有助于了解激活剂如何与 CYP2D6 相互作用，并为合理的药物设计和个性化医疗时代药物代谢的个体间差异评估带来新的机遇。