Cholinesterase Inhibitors in Alzheimer's Disease

描述（由申请人提供）：阿尔茨海默病（AD）的特征是胆碱能功能障碍和源自AB前体蛋白（APP）的淀粉样β肽（AB）沉积。目前fda批准的四种AD药物属于胆碱酯酶抑制剂（ChEI）的范畴。这些药物对轻度至中度阿尔茨海默病的治疗有用，但在晚期使用有限。我们的目标是测试新型乙酰胆碱酯酶抑制剂（AChEIs）或丁基胆碱酯酶抑制剂（BChEIs）在AD病理生理中的关键步骤，以更好地了解药物的作用。我们的重点是测试新的治疗策略，以验证和新的疾病目标。我们的假设是，由于某些chei具有降低ab的特性，因此具有独立于胆碱能活性的神经保护活性。我们提出研究ChEI药物对淀粉样变途径的机制是基于我们的研究结果，即某些ChEI显著降低了培养细胞和动物的AB水平。具体目的是：1]研究新型AChEIs和BChEIs对APP通路步骤的影响。我们将研究i)新型ChEIs对APP通路：AB和BACE水平的影响，ii)取代官能团的影响，iii) APPmRNA 5'-UTR, iv)突触蛋白。[2]研究一类新型ChEIs对AB在体内沉积的影响。我们将研究不同剂量的选择性AChEIs和BChEIs对双转基因appsw - tau淀粉样斑块加缠结小鼠APP和AB的影响。[3]探讨ChE酶在AChE基因敲除小鼠中的作用。我们将研究AChE酶对新型AChE敲除小鼠脑组织样本中APP、AB肽和突触蛋白标记物的作用。[4]探讨ChEIs对人（存档）样本分子标记物的影响。为了验证动物研究，我们将测试选定的ChEIs处理对人血浆和/或CSF样本中APP和AB水平的影响。主要终点是定量和功能性的：细胞存活、酶分析和APP通路蛋白/肽和突触蛋白标记物的水平。我们将从机制上选择与ChE的外周变构结合域以及与酯和阴离子结合域相互作用的chei，并在细胞培养和动物模型中进行测试。这项工作将表明ChEIs对AB和突触蛋白的独特作用，独立于它们对酶的选择性。这一特性将有助于最大限度地发挥其对淀粉样蛋白和突触蛋白的有益作用，从而可用于设计更好的阿尔茨海默病治疗剂。公共卫生相关性：阿尔茨海默病（AD）的特点是胆碱能系统突触前标记物减少，特别是在与记忆和学习相关的大脑区域，以及淀粉样蛋白β肽（AB）的沉积，AB来源于AB前体蛋白（APP）。目前fda批准的治疗AD的五种药物中有四种是胆碱酯酶抑制剂（ChEI），它通过抑制胆碱酯酶（ChE）来增加大脑中乙酰胆碱（ACh）的供应，从而保持胆碱能回路，这种回路被认为是调节记忆途径的。更令人惊讶的是，这些药物对记忆并没有实质性的影响，但却有一个意想不到的、受欢迎的结果，即在一段时间内保持认知。本研究试图描述ChEIs的神经保存和保护机制，并微调这一途径以改善ChEIs的这一有益特性。我们的目标是测试新型乙酰胆碱酯酶抑制剂（AChEIs）或丁基胆碱酯酶抑制剂（BChEIs）在AD病理生理中的关键步骤，以更好地了解药物的作用。我们建议研究ChEI药物对淀粉样变途径的作用机制，将APP加工成潜在毒性的AB。我们的研究最初集中在这些药物对AB产生的影响上，最近已扩展到突触蛋白标记物和其他神经保护作用。该建议的结果是确定ChEIs阻断潜在毒性AB水平的机制，并利用这一特性开发新的治疗剂。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is characterized by cholinergic dysfunction and depositions of the amyloid beta-peptide (AB) derived from AB precursor protein (APP). Four current FDA-approved drugs for AD belong to the category of cholinesterase inhibitor (ChEI). These drugs are useful for treatment of mild to moderate AD but limited use in advanced stages. Our goal is to test novel acetylcholinesterase inhibitors (AChEIs) or butyrylcholinesterase inhibitors (BChEIs) against pivotal steps in the pathophysiology of AD to better understand the drugs' effects. Our focus is to test new therapeutic strategies to both validated and novel disease targets. Our hypothesis is that certain ChEIs have neuroprotective activity independent of their cholinergic activity due to their AB-lowering property. Our proposal to study the mechanism of ChEI drugs on amyloidogenic pathways is based on our results that certain ChEIs significantly reduced AB levels in cultured cells and animals. The specific aims are: 1] To study the effect of novel AChEIs and BChEIs on APP pathway steps. We will examine i) the effect of novel ChEIs on APP pathway: AB and BACE levels, ii) the effect of substitution of functional groups, iii) APPmRNA 5'-UTR, and iv) synaptic proteins. 2] To test the effect of a novel group of ChEIs on AB deposition in vivo. We will study the effects of different doses of selective AChEIs and BChEIs on APP and AB in double transgenic APPSWE-Tau amyloid plaque plus tangle producing mice. 3] To examine the role of ChE enzyme in AChE knockout mice. We will study the role of the AChE enzyme on APP, AB peptides and synaptic protein markers in the brain tissue samples from novel AChE knockout mice. 4] To investigate the effect of ChEIs on molecular markers in human (archived) samples. To validate the animal studies, we will test the effect of treatment of selected ChEIs on APP and AB levels in human plasma and/or CSF samples. The primary end-points are quantitative and functional: Cell survival, enzyme assay and levels of APP pathway protein/peptides and synaptic protein markers. We will mechanistically select ChEIs that interact with the peripheral allosteric binding domain of ChE and with the esteratic and anionic binding domains and test them in cell culture and animal models. This work will indicate unique effect of ChEIs on AB and synaptic proteins, independent of their selectivity for the enzyme. This property will help maximize their beneficial effects on amyloid and synaptic proteins, which can be utilized to design better therapeutic agents for AD. PUBLIC HEALTH RELEVANCE: Alzheimer's disease (AD) is characterized by a reduction in the presynaptic markers of the cholinergic system, particularly in areas of the brain related to memory and learning, and by depositions of the amyloid beta peptide (AB), which is derived from the AB precursor protein (APP). Four of five current FDA-approved drugs for AD are cholinesterase inhibitors (ChEI), which increase the brain's supply of acetylcholine (ACh) by inhibiting cholinesterases (ChE) enzyme and thus preserve cholinergic circuits, which are believed to mediate memory pathways. Rather surprisingly, these drugs did not have a substantial effect on memory, but had an unexpected and welcome outcome of preserving cognition for a slightly extended period of time. The present proposal attempts to characterize the mechanisms of neuropreservation and protection by the ChEIs and fine tune this pathway to improve this beneficial property of ChEIs. Our goal is to test novel acetylcholinesterase inhibitors (AChEIs) or butyrylcholinesterase inhibitors (BChEIs) against pivotal steps in the pathophysiology of AD to better understand the effects of the drugs. We propose to study the mechanism of ChEI drugs on amyloidogenic pathways that process APP to potentially toxic AB. Our studies initially focused on the effects of these drugs on AB production and have recently been extended to synaptic protein markers and other neuroprotective effects. The outcome of the proposal is to identify mechanisms by which ChEIs block potentially toxic AB levels and to utilize this property in developing novel therapeutic agents.