Pyruvate Dehydrogenase Complex Activation as a Strategy to Ameliorate Metabolic Disease

代谢重塑是2型心脏病等疾病的一个潜在主题 糖尿病和癌症，这些疾病的细胞偏离了它们典型的燃料利用情况。丙酮酸 脱氢酶复合体(PDC)是葡萄糖有氧利用的守门人。 下调监管与这些疾病有很大关联。因此，最近的努力集中在 关于激活PDC作为一种治疗方法。目前，努力只集中在抑制 丙酮酸脱氢酶激酶(PDK)，通过磷酸化抑制PDC。然而， 激活丙酮酸脱氢酶磷酸酶(PDP)作为一种治疗手段，它可以降低 磷酸化PDC以恢复活性，已被忽视。此外，众所周知， PDC活性被NADH抑制，但从治疗上针对这种调节机制的努力 已经被忽视了。作为更全面的PDC激活策略，我们希望扩展 PDK抑制剂，确定PDP小分子激活剂，并优化可 已被证明从NADH抑制中恢复PDC作为一种新的治疗策略 代谢性疾病。其次，线粒体产生的活性氧(ROS) 电子传输链(METC)被认为被PDC激活而增强，但其影响 PDC作为ROS的来源，以及NADH/NAD和ATP等特定条件如何需求， 就像在运动中一样，对ROS动态平衡的影响尚不清楚。ROS的现场特定量化 生成需要在各种条件下进行详细的酶模拟才能阐明。这个 该项目的目标是1)虚拟筛选和实验验证PDC激活剂和2) 应用数学模型确定PDC激活对特定部位mETC ROS的影响 一代。我们从锌数据库中获得了1500万种类似药物的虚拟化合物 单独筛选位于三个已知抑制位点的所有PDK同工酶(1-4)：硫胺，ATP/ADP， 和pzf3，寻找同工酶特异性和PAN抑制剂。该虚拟复合集合也将被使用 通过虚拟筛选混合晶体/计算衍生的PDPC来鉴定PDP激活剂- PDPr复合体。我们将在引导的结构中使用PDC(E3)的NADH结合结构域 方法，以确定PDC的最佳苯二酚类似物。详细的酶动力学模型 将集成PDC、TCA循环和mETC来模拟各种氧化状态，包括 减少应激和运动条件以量化特定部位的线粒体ROS 制作。我们认为，我们的建议解决了激活战略方面的重大差距 PDC作为一种治疗手段，将提供对PDC影响的定量了解 线粒体ROS产生的活性。

Metabolic remodeling is an underlying theme in diseases such as heart disease, type 2 diabetes, and cancer, where cells deviate from their typical fuel utilization profile. The pyruvate dehydrogenase complex (PDC) is the gatekeeper for aerobic glucose utilization and its downregulation is greatly associated with these diseases. As such, recent efforts have focused on the activation of PDC as a therapeutic. Currently, efforts have focused only on inhibition of the pyruvate dehydrogenase kinase (PDK), which inhibits PDC by phosphorylation. However, activation of the pyruvate dehydrogenase phosphatase (PDP) as a therapeutic, which de- phosphorylates PDC to recover activity, has been ignored. Furthermore, it is well-known that PDC activity is inhibited by NADH but efforts to therapeutically target this regulatory mechanism have been overlooked. As a more comprehensive PDC activation strategy, we look to expand PDK inhibitors, identify PDP small molecule activators, and optimize quinone compounds that have been shown to recover PDC from NADH inhibition as a new therapeutic strategy for metabolic disease. Secondly, reactive oxygen species (ROS) generated from the mitochondrial electron transport chain (mETC) are thought to be enhanced by PDC activation but the impact of PDC as a source of ROS and how specific conditions such as NADH/NAD and ATP demand, as in exercise, influence ROS homeostasis is unclear. Site-specific quantification of ROS generation requires detailed enzymatic simulations in various conditions to be elucidated. The aims of this project are to 1) virtually screen and experimentally validate PDC activators and 2) apply mathematical modeling to determine effects of PDC activation on site-specific mETC ROS generation. We obtained 15M drug-like virtual compounds from the ZINC database to individually screen all PDK isozymes (1-4) at three known inhibition sites: lipoamide, ATP/ADP, and pzf3 to find isozyme specific and pan inhibitors. This virtual compound set will also be used to identify PDP activators by virtually screening a hybrid crystal/computationally derived PDPc- PDPr complex. We will use the NADH binding domain of PDC (E3), in a structure guided approach, to identify optimal quinone analogs for PDC. Detailed enzyme kinetics models of PDC, TCA cycle, and mETC will be integrated to simulate various oxidative states including reductive stress and exercising conditions to quantify site-specific mitochondrial ROS production. We believe that our proposal addresses significant gaps in strategies to activate PDC as a therapeutic and will provide a quantitative understanding of the influence of PDC activity on mitochondrial ROS production.