Metabolic Control Analysis

摘要：代谢控制分析是动力学和热力学的正式应用，以确定代谢途径的通量控制在多大程度上是由该途径中特定的酶步骤决定的。对中心能量产生途径、柠檬酸循环和线粒体生物能量学的综述已经发表。通过进行正式的代谢控制分析所发现的是，与普遍认为存在“速率控制步骤”的观点相反，对途径中的通量的控制分布在途径中的许多酶步骤中，并且随着条件的不同而变化。正是代谢途径中的这种流量变化通常决定了疾病过程的表型。此外，这些见解表明，针对单个酶步骤的药物或基因疗法可能无法纠正疾病表型，而是单个或几个相关途径中的多个步骤必须在疾病表型中改变。 这导致了对几种退行性神经和心脏疾病的研究，这些疾病可以通过改变提供的底物来治疗，其中包括阿尔茨海默氏症和帕金森氏病，第一型和第二型糖尿病以及心力衰竭。随后在外部实验室的工作加强了治疗轻度酮症帕金森病的可能性，在这些实验室中，D-R-羟丁酸钠减少了MPTP诱导的C57黑鼠海马神经元的死亡。在一项初步试验中，给一小部分帕金森患者喂食高酮饮食可导致大约60%的症状减轻。这些概念在由NIH罕见疾病办公室主办的两次会议上进行了讨论，并更全面地概述了疾病表型目标。在目前还没有有效治疗方法的致命疾病中，杜兴氏肌营养不良症在美国影响到3000名男婴中的1/3。该实验室和牛津大学Kieran Clarke教授的实验室所做的工作发现，MDX小鼠(Duchenne的啮齿动物类似物)缺乏dystrophin，导致胰岛素未能诱导胰岛素反应GLUT4移位到质膜。因此，这些患者属于可能用酮症治疗的疾病表型。我们一直在检查轻度酮病可以提高MDX小鼠心脏灌流的水力效率的程度，这是本实验室独有的一种技术难度很大的制剂。在过去的两年里，这份材料一直是由美国国立卫生研究院罕见疾病办公室主办的两次会议的主题。 最近，我们研究了使用酮体改变暴露在低氧培养中的海马神经元存活的影响。我们发现，在低氧2小时内，可以防止海马细胞的急性死亡，并在较长时间内减少凋亡细胞的死亡。这增加了酮用于治疗急性中风或脑出血或治疗创伤性脑损伤的可能性。 我们目前计划开发D-R-羟基丁酸酯的口服制剂，用于军事应用。这个项目中没有关于在疾病状态下使用这些材料的规定。希望美国国立卫生研究院内的各个相关机构可以考虑在人体试验中使用这些材料。这里特别相关的将是帕金森氏病的可能治疗，根据最近来自其他实验室的出版物，跟踪我们在这一领域的工作。同样，鉴于目前缺乏任何有效的治疗方法，应考虑杜氏肌营养不良症的可能治疗方法。 对本研究所项目的意义 本节报告的研究，通过应用严格的代谢控制分析，可以定义疾病表型，这些表型可能适用于各种常见和罕见疾病的新治疗方法，这些疾病具有共同的病理生理基础。这种方法定义了一种新的治疗方法，旨在改变相互关联和相互依赖的代谢途径，而不是试图将一个关键的“限速步骤”定义为疾病状态下的致病因素。

Summary: Metabolic control analysis is the formal application of kinetics and thermodynamics to determine the extent to which the control of flux through a metabolic pathway is determined by a particular enzymatic step in that pathway. Reviews of the central energy producing pathway, the citric acid cycle, and of mitochondrial bioenergetics have been published. What is found by performing a formal metabolic control analysis is that, in contrast to the widely held view that there is a 'rate controlling step', control of flux in a pathway is distributed across many enzymatic steps in a pathway and varies depending upon conditions. It is this change in flux in a metabolic pathway which often defines the phenotype of a disease process. Further, these insights suggest that a drug or gene therapy directed at a single enzymatic step may not correct a disease phenotype, but rather a number of steps in a single or in several linked pathways would have to be altered in a disease phenotype. This lead to an investigation of several degenerative neurological and cardiac diseases which might be treated by alteration of the substrate provided, among them were Alzheimer's and Parkinson's disease, type I and II diabetes and heart failure. The possibility of treating Parkinson's disease with mild ketosis has been strengthened by subsequent work in outside laboratories where MPTP induced death in hippocampal neurons in C57 black mice was reduced by Na D-R-hydroxybutyrate. Feeding a hyperketogenic diet to a small group of Parkinsonian patients resulted in an approximately 60% reduction of symptoms in a preliminary trail. These concepts were discussed in two meetings sponsored by the NIH Office of Rare Diseases and the disease phenotype targets more fully outlined. Among the fatal diseases for which there is no current effective therapy is Duchenne's muscular dystrophy affecting 1/3000 male births in the United States. Work done in this laboratory and in the laboratory of Professor Kieran Clarke in Oxford have found that lack of dystrophin in the MDX mouse, the rodent analogue of Duchenne's, results in a failure of insulin to induce translocation the insulin responsive GLUT4 to the plasma membrane. As such, these patients fall into the disease phenotype potentially treatable by ketosis. We have been examining the degree to which mild ketosis can improve the hydraulic efficiency of the isolated working perfused MDX mouse heart, a technically difficult preparation which is unique to this laboratory. This material has been the subject of 2 meetings sponsored by the NIH Office of Rare Disease over the past 2 years. More recently, we have examined the effects of the use of ketone bodies on altering the survival of hippocampal neurons exposed to hypoxia in culture. We found that acute cell death in hippocampal cells could be prevented during 2 hours of hypoxia and that apoptotic cell death could also be decreased for a long period of time. This raises the possibility of the use of ketones in the treatment of acute stroke or cerebral hemorrhage or in the treatment of traumatic brain injury. We currently are scheduled to develop oral preparations of D-R-hydroxybutyrate for use in military applications. There is no provision in this project for the use of these materials in disease states. It is hoped that various relevant institutes within the NIH may wish to consider the use of these materials in clinical trials of human subjects. Of particular relevance here would be the possible treatment of Parkinson's disease in the light of the recent publications from other labs following our work in this area. Likewise the possible treatment of Duchenne's muscular dystrophy should be considered in view of the absence of any currently effective therapy. Significance to the Programs of this Institute The research reported in this section, by applying a rigorous metabolic control analysis, allows the definition of disease phenotypes which may be amenable to new therapeutic approaches in a variety of common and rare diseases, which share a common pathophysiological basis. This approach defines a new way of approaching therapy which aims at changing interlinked and interdependent metabolic pathways rather that attempting to define one key "rate limiting step" as causative in a disease state.