Mechanisms of Cholinesterase Inhibitors on Beta-amyloid

阿尔茨海默病（Alzheimer's disease，AD）是一种以胆碱能神经元严重缺失和淀粉样β肽（amyloid beta peptide，Abeta）沉积为特征的疾病。 用于治疗AD受试者的三种FDA批准的药物（他克林、多奈哌齐和卡巴拉汀）属于胆碱酯酶（ChE）抑制剂（ChEI）的类别，其通过增加乙酰胆碱的脑供应而起作用，乙酰胆碱是AD中缺乏的神经通讯化学物质。 这些药物被批准用于治疗轻度至中度AD，并且在更晚期阶段可能不那么有用。 我们的目标是研究ChEI药物对淀粉样蛋白生成途径的作用机制，该途径将β-淀粉样蛋白前体蛋白（APP）加工成潜在的神经毒性Abeta。 随着越来越多的证据表明Abeta在AD的发病机制中起重要作用，该研究具有重要意义。 这一提议是基于我们的发现，即用某些ChEI（如他克林和苯丝氨酸）处理培养的细胞，可显著降低分泌的APP（sAPP）和Abeta的水平，并可减缓AD的进展以及改善认知。 值得注意的是，降低Abeta的机制并没有增加已知的替代加工途径，因此可能损害较小。 我们感兴趣的是确定ChEI阻断Abeta分泌的机制，以利用Abeta降低特性开发新的治疗药物。具体目标：1.研究乙酰胆碱酯酶抑制剂（AChEI）和丁酰胆碱酯酶抑制剂（BchEI）对sAPP和Abeta水平的影响。 为了检查其作用的特异性，i）AchEI（例如，将测试ii）BChEI（例如cymserine）和iii）作为他克林衍生物的化合物（例如维那克林）以鉴定降低Abeta的结构方面。 2.研究ChEI对APP代谢的作用。 将测试ChEI对i）FAD-APP突变细胞系中APP加工和ii）APP羧基截短片段命运的影响。 3.以确定药物的可能靶点。 ChEI对AD转基因小鼠模型中i）APP裂解酶（BACE），ii）5' -非翻译区和iii）Abeta水平抑制的影响。 我们将机械地选择与ChE酶的外周变构结合结构域或与酯-乳酸和阴离子结合结构域（苯丝氨酸和cymserine）相互作用的ChEI，并在APP/PS1双转基因小鼠中测试它。 这些结果将表明ChEI对APP加工的独特作用，这与其对酶的选择性无关。这一特性将被进一步研究，以最大限度地发挥其在减少淀粉样蛋白沉积方面的潜在作用，并可用于设计更好的治疗AD的药物。

Alzheimer's disease (AD) is characterized by the severe loss of cholinergic neurons and depositions of amyloid beta peptide (Abeta). Three FDA- approved drugs (tacrine, donepezil and rivastigmine) for treating AD subjects belong to the category of cholinesterase (ChE) inhibitor (ChEI), which works by increasing the brain's supply of acetylcholine, a nerve communication chemical that is deficient in AD. These drugs are approved for treatment of mild to moderate AD and may not be as useful in more advanced stages. Our goal is to study the mechanism of ChEI drugs on amyloidogenic pathways that process beta-amyloid precursor protein (APP) to potentially neurotoxic Abeta. Such study is significant as there is increasing evidence that Abeta plays an important role in AD pathogenesis. This proposal is based on our discovery that treating cultured cells with certain ChEIs, such as tacrine and phenserine, significantly reduced levels of secreted APP (sAPP) and Abeta and may serve to slow the progression of AD as well as improve cognition. Notably, the mechanism of reduction of Abeta did not increase known alternative processing pathways and may therefore be less damaging. We are interested in identification of the mechanisms by which ChEIs block Abeta secretion to take advantage of the Abeta lowering property in developing novel therapeutic agents. SPEC. AIMS: The specific aims are: 1. To study the effects of acetyl- ChEI (AChEI) and butyrl-ChEI (BchEI) on sAPP and Abeta levels. To examine the specificity of their actions, effects of i) AchEI (e.g., pheneserine) ii)BChEI (e.g. cymserine), and iii) compounds that are tacrine-derivatives (e.g. velnacrine) will be tested to identify structural aspects that lower Abeta. 2. To investigate the role of ChEIs on APP metabolism. Effects of ChEIs on i)APP processing in FAD-APP mutant cell lines and ii) the fate of APP carboxyl-truncated fragments will be tested. 3. To determine the possible targets of the drugs. Effects of ChEIs on the i) APP-cleaving enzyme (BACE), ii) 5' -untranslated region and iii) inhibition of Abeta levels in transgenic mice model of AD. We will mechanically select ChEIs that interact with the peripheral allosteric binding domain of ChEenzyme, or with the esteractic and anionic binding domains (phenserine and cymserine) and test it in APP/PS1 double transgenic mice. These results will indicate a unique effect of ChEIs on APP processing, which is independent of their selectivity for the enzyme. This property will be further investigated to maximize their potential effects in decreasing amyloid depositions, and which can be utilized to design better drugs for the treatment of AD.