Preventing early events in Aβ-driven pathology in vivo

预防体内 Aβ 驱动病理学的早期事件

• 批准号: 9913438
• 负责人: EFRAT LEVY
• 金额: $ 40.43万
• 依托单位: NATHAN S. KLINE INSTITUTE FOR PSYCH RES
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9913438
• 关键词: Age; Alzheimer' s Disease; Amino Acid Substitution; Amino Acids; Amyloid; Amyloid Fibrils; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Amyloid deposition; Autologous; Behavior; Behavioral; Brain; Brain Diseases; Cell Line; Cells; Culture Media; Data; Deposition; Development; Early treatment; Elderly; Event; Extracellular Space; Family; Family member; Fibroblasts; Foundations; Harvest; Hippocampus (Brain); Human; In Vitro; Individual; Intranasal Administration; Investigation; Messenger RNA; Mus; Mutate; Mutation; Neurons; Nomenclature; Nose; Pathology; Pathway interactions; Peptides; Peripheral; Positioning Attribute; Presenile Alzheimer Dementia; Prevention; Promoter Regions; Property; Proteins; RNA delivery; Research; Role; Senile Plaques; Source; Suggestion; Techniques; Testing; Therapeutic; Therapeutic Uses; Time; Toxic effect; Transgenic Mice; Wild Type Mouse; abeta accumulation; abeta deposition; amyloid formation; amyloidogenesis; base; beta amyloid pathology; exosome; experimental study; extracellular vesicles; in vivo; interest; mouse model; mutant; mutation carrier; neuron loss; neurotoxicity; novel; novel therapeutic intervention; overexpression; polymerization; prevent; protective effect; protein metabolite; tool

PROJECT SUMMARY / ABSTRACT An APP mutation identified in an Italian family that results in an amino acid substitution at position 2 of amyloid β (AβIt) protects heterozygous carriers from AD even in advanced age. In vitro, AβIt prevents wild-type Aβ (Aβwt) from forming Aβ amyloid fibrils and oligomerization, suggesting that co-expression of AβIt interferes with Aβ nucleation and/or polymerization, a mechanism that is hypothesized to inhibit Aβ oligomerization, amyloidogenesis and neurotoxicity in the heterozygous Italian APP (APPIt) carriers. In order to directly explore the in vivo effect of this mutation we have generated a transgenic mouse overexpressing human APP with the A673V Italian mutation under the control of Thy1.2 promoter sequences (TgAPPIt mice). We will investigate the anti-amyloidogenic effect of AβIt when co-expressed with the human Aβwt in vivo using amyloid-depositing mice (APP23) crossed with the TgAPPIt mice (Aim 1). Examining mechanism within the brain, we will determine whether AβIt protection is due to the inhibition of Aβ nucleation or due to enhanced Aβ degradation, decreased oligomer/early aggregate stability, and/or enhanced clearance of the AβIt/Aβwt interacting peptides, testing the hypothesis that the amino-terminally mutated AβIt interferes with the earliest events necessary for the initiation of β amyloid-driven pathology. In an initial therapeutic strategy, the anti-amyloidogenic function of AβIt will be investigated by administering to amyloid depositing mice exosomes-enriched extracellular vesicles (EV) as a source of AβIt (Aim 2). In part because of their stability, autologous EV have many appealing properties as peptide, protein, and RNA delivery tools, and we have shown that brain EV contain APP, are highly enriched with APP carboxyl-terminal fragments (APP-CTFs) and are therefore a source of EV-generated Aβ. In preliminary studies we used our novel technique to isolate EV from the brain extracellular space of TgAPPIt mice and administered these intranasally to amyloid depositing mice. Amyloid deposition was reduced in mice treated with brain-derived EV from TgAPPIt mice compared to mice treated with EV isolated from the brain of wild-type mice, supporting our hypothesis that AβIt interferes with developing Aβ pathology and supporting our idea that EV can be used to deliver protective constituents to the brain. TgAPPIt brain-derived EV will be delivered prior to amyloid deposition and at an age when amyloid deposition is abundant, and recipient mice will be examined for Aβ oligomerization, amyloid deposition, neuronal loss and behavioral deficits. We will further examine the anti-amyloidogenic potential and protective effects of AβIt using EV loaded with APP-A673V mRNA or AβIt peptides, as well as a non-fibrillogenic peptide comprised of the 6 amino- terminal residues of AβIt. These studies are timely, given the growing interest in developing EV as therapeutic vehicles for both systemic and brain disorders, and our investigations of EV as candidates to act as vehicles to deliver protective AβIt within the brain as a foundation for developing novel EV-based treatments for AD.

项目摘要 /摘要 在意大利家族中鉴定出的APP突变，该突变在淀粉样蛋白的位置2导致氨基酸取代 β（AβIT）即使在高龄，也可以保护杂合载体免受AD的侵害。在体外，AβIT可防止野生型Aβ （AβWT）从形成Aβ淀粉样蛋白原纤维和低聚，表明AβIT的共表达会干扰 Aβ核化和/或聚合化，该机制假设抑制Aβ寡聚化的机制， 杂合意大利应用（APPIT）载体中的淀粉样生成和神经毒性。为了直接探索 该突变的体内效应我们已经产生了一种过表达人类应用的转基因小鼠 在THY1.2启动子序列（TGAPPIT小鼠）控制下的A673V意大利突变。我们将调查 当使用淀粉样蛋白抑制与人AβWT共表达时，AβIT的抗淀粉样生成作用 小鼠（APP23）与TGAPPIT小鼠（AIM 1）交叉。检查大脑内的机制，我们将 确定AβIT保护是由于抑制Aβ核还是由于Aβ降解的抑制作用引起的 降低寡聚物/早期骨料稳定性和/或增强AβIT/AβWT相互作用肽的清除率， 测试氨基末端突变的AβIT的假设会干扰最早的事件 β淀粉样蛋白驱动病理的倡议。在最初的治疗策略中，抗淀粉样生殖功能 AβIT将通过对富含细胞外蔬菜的淀粉样蛋白沉积进行研究 （EV）作为AβIT的来源（AIM 2）。自体电动汽车的稳定性部分是由于它们的稳定性 作为肽，蛋白质和RNA递送工具的特性，我们已经表明含有大脑EV的应用是 高度富含App羧基末端片段（APP-CTF），因此是EV生成的来源 Aβ。在初步研究中，我们使用新技术将EV与大脑外部空间分离 TGAPPIT小鼠并将其内经局局部用于淀粉样蛋白沉积小鼠。减少淀粉样蛋白沉积 在用Tgappit小鼠治疗的小鼠中，与用从eV处理的小鼠相比 野生型小鼠的大脑，支持我们的假设，即AβIT会干扰发展Aβ病理学和 支持我们的想法，即EV可用于向大脑提供受保护的构成。 tgappit脑源性 EV将在淀粉样沉积之前和淀粉样沉积丰富的年龄之前交付，并且 将检查受体小鼠的Aβ寡聚，淀粉样蛋白沉积，神经元丧失和行为 缺陷。我们将进一步检查使用EV载的AβIT的抗淀粉样生殖潜力和受保护的作用 用App-A673V mRNA或AβITpetides，以及6个氨基的非原纤维胡椒 AβIT的末端残基。鉴于人们对开发EV作为治疗的兴趣日益增加，这些研究是及时的 系统性和脑部疾病的车辆，以及我们对EV作为候选人的调查 在大脑内提供受保护的AβIT，作为开发基于EV的新型AD处理的基础。