Characterization of a caspase-cleavage resistant tau knock-in mouse

 描述（由申请人提供）：阿尔茨海默病（AD）是世界上年龄依赖性痴呆的最常见原因，与大脑淀粉样斑块和神经纤维缠结（NFT）相关。淀粉样蛋白斑块主要由A β肽组成，而NFT由细胞骨架蛋白tau的过度磷酸化形式的异常聚集体组成。淀粉样前体蛋白（APP）的双切割产生脱乙酰肽。BACE 1裂解产生C-末端片段，β-CTF，其然后被β-分泌酶加工成几种腺苷酸同工型。遗传数据表明，APP加工的调节有助于AD。目前AD的药物发现方法集中在预防淀粉样蛋白形成或去除现有的淀粉样蛋白沉积。靶向淀粉样蛋白的化合物/生物制剂在临床试验中的反复失败可归因于纳入了疾病进展太快而无法从治疗干预中获益的AD患者，或者归因于阿尔茨海默病不是AD病理生理学的主要原因。AD的最先进的替代假设是tau毒性。与这一假设一致，降低内源性tau蛋白表达可防止表达具有家族性AD突变的人APP的转基因小鼠的行为缺陷，而不改变A β水平。我们的初步数据表明，tau还介导家族性丹麦痴呆（FDDKI小鼠）中的行为缺陷，这是一种与APP处理BRI 2/ITM 2B的调节因子突变相关的AD样痴呆，也以tau蛋白病为特征。一些证据表明，半胱天冬酶在AD进展的早期被激活，并且可能在神经元损失和NFT病理中起作用。Tau在D421处被执行人半胱天冬酶切割（Tau）。在半胱天冬酶切割后，Tau促进成核依赖性丝的形成，并采用由早期病理性tau标志物MC 1识别的构象变化。 NFT Tau可以被磷酸化并被NFT抗体PHF-1识别。在AD大脑中，Tau与NFT的标志物相关，并与认知下降相关。此外，Tau启动NFT形成并可发挥毒性作用。这些发现导致了以下假设：Tau是神经变性的关键毒性部分。为了直接测试该理论，我们已经产生了敲入小鼠，其中编码D421的外显子12中的内源性tau密码子GAC已经突变为AAC，其现在编码天冬酰胺（N）。这些被称为TauD 421 N的敲入小鼠表达tau突变体，taucas/res，其不能被半胱天冬酶切割，因此不能产生Tau Tau。TauD 421 N小鼠将用于确定在AD和FDD的动物模型中，Tau是否介导突触可塑性和记忆缺陷，AD和FDD是两种神经退行性疾病，其中tau起致病作用。我们的研究将讨论调节β-Tau形成是否是治疗AD和由神经毒性tau形式介导的其他神经退行性疾病的潜在策略。

 DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is the most common cause of ageing-dependent dementia in the world and is associated with cerebral amyloid plaques and neurofibrillary tangles (NFTs). Amyloid plaques are mostly composed of Aß peptides while NFTs are composed of abnormal aggregates of hyperphosphorylated forms of the cytoskeletal protein tau. Aß peptides are produced by a double cleavage of the amyloid precursor protein (APP). BACE1 cleavage produces the C-terminal fragment, ß-CTF, which is then processed into several Aß isoforms by -secretase. Genetic data suggest that regulation of APP processing contributes to AD. Current drug discovery approaches in AD have focused on preventing Aß formation or removing existing amyloid deposits. The repeated failures of compounds/biologics targeting amyloid in clinical trials can either be attributed to the inclusion of AD patients who were too advanced in their disease progression to benefit from therapeutic intervention, or to the fact that Aß is not the main cause of AD patho-physiology. The most advanced alternative hypothesis of AD is that of tau toxicity. Consistent with this hypothesis, reducing endogenous tau expression prevents behavioral deficits in transgenic mice expressing human APP with familial AD mutations, without altering Aß levels. Our preliminary data indicate that tau also mediates behavioral deficits in Familial Danish Dementia (FDDKI mice), an AD-like dementia associated with mutations in the regulator of APP processing BRI2/ITM2B, also characterized by tauopathy. Some evidence suggests that caspases are activated early in the progression of AD and may play a role in neuronal loss and NFT pathology. Tau is cleaved at D421 (Tau) by executioner caspases. Following caspase-cleavage, Tau facilitates nucleation-dependent filament formation and adopts a conformational change recognized by MC1, an early pathological tau marker. Tau can be phosphorylated and recognized by the NFT antibody PHF-1. In AD brains, Tau associates with markers of NFTs and correlates with cognitive decline. Furthermore, Tau initiates NFT formation and can exert toxic effects. These findings have led to the hypothesis that Tau is a critical toxic moiety underlying neurodegeneration. To directly test this theory, we have generated knock-in mice in which the endogenous tau codon GAC in exon 12, encoding for D421, has been mutated into AAC, which now encodes for an asparagine (N). These knock-in mice, called TauD421N, express a tau mutant, taucas/res that cannot be cleaved by caspases and therefore cannot generate Tau. TauD421N mice will be used to determine whether Tau mediates synaptic plasticity and memory deficits in animal models of AD and FDD, two neurodegenerative disorders in which tau plays a pathogenic role. Our studies will speak to whether modulation of Tau formation is a potential strategy for the treatment of AD and other neurodegenerative disorders mediated by neuro-toxic tau forms.