Immunomodulation targeting abnormal conformation and the influence of apoE

摘要 阿尔茨海默病的神经病理包括斑块中的纤维淀粉样蛋白(Fibillary Amyloidβ，Aβ)，脑淀粉样血管病变， 神经原纤维缠结(NFT)中过度磷酸化的tau纤维。然而，毒性最大的物种是Aβ 以普通结构β-Sheet二级结构为特征的tau齐聚物。这些错误折叠了一个β和 尽管到目前为止进行了许多临床试验，但tau构象是免疫干预的合适靶点。 失败的原因是：1)自身免疫毒性；2)缺乏针对寡聚体Aβ和tau的特异性伴随靶向 淀粉样蛋白相关成像异常(ARIA)，特别是载脂蛋白(Apo)E4 携带者，被认为与纤维血管淀粉样蛋白清除有关。为了克服这些限制，我们 开发了创新的抗体结合位点(β联合抗体副表位)，仅识别主要的β- 错误折叠的蛋白质的片状二级结构，这是所有病理性低聚物的普遍特征 神经退行性疾病。我们的初步结果表明，某些aβ结合抗体结合在两种抗体的任一种上 或者免疫球蛋白类可以无副作用地穿透预先存在Aβ的AD TG小鼠模型的血脑屏障，并 Tau病理，实现显著的认知挽救，降低Aβ和tau病理寡聚体水平，并在 CAA AD TG小鼠模型(TgSwDI)无ARIA样毒性(两种模型均以小鼠载脂蛋白E为模型 背景)。在这个项目中，我们将测试β组合抗体(IgM或Ig G)的假设，而不是mAbs定向的 针对Aβ序列(6E10)，可以安全输注并产生修改的疗效而不需要 3xTg的血管ARIA样毒性与TgSwDI或APP/PS1dE9小鼠模型的杂交 人类APOE2、E3或E4背景。此外，我们预计血管淀粉样蛋白 这些小鼠的蛋白质组将类似于与不同载脂蛋白E相关的投影1和2 背景、治疗和有无微出血。这一信息是至关重要的 需要阐明与ARIA相关的机制并选择用于未来临床的β复合抗体表位 审判。具体目标是： 1)产生并鉴定特异性结合位点的免疫球蛋白M和免疫球蛋白G形式(Aβ联合抗体副表位) 识别有毒低聚物的β-Sheet二级结构。 2)测定来自Aim 1的4个a-β单抗与脑的生化和组织化学相互作用 外周输注的TgSwDI小鼠的血管系统与KI人APOE2，E3和E4背景杂交。 3)检测被动免疫治疗后的行为、组织化学和生化变化 β复合抗体在3xTg、Tg APP/PS1dE9和TgSwDI小鼠中的Ig M和Ig G上的对位 在Ki人类APOE2、E3和E4背景上杂交。

SUMMARY The neuropathology of AD includes fibrillary amyloid β (Aβ) in plaques, cerebral amyloid angiopathy (CAA), and hyperphosphorylated tau fibrils in neurofibrillary tangles (NFT). However, the most toxic species are Aβ and tau oligomers characterized by generic structural β-sheet secondary structure. These misfolded Aβ and tau conformers are suitable targets for immunological intervention, although numerous clinical trials thus far have failed due to: 1) autoimmune toxicity; 2) lack of specific concomitant targeting of oligomeric Aβ and tau species; and 3) Amyloid Related Imaging Abnormalities (ARIA), particularly among apolipoprotein (apo)E4 carriers, thought to be associated with fibrillar vessel amyloid clearance. To overcome these limitations, we developed innovative antibody combining sites (aβComAb paratopes) that only recognize the dominant β- sheet secondary structure of misfolded proteins, a generic characteristic of all pathologic oligomers found in neurodegenerative diseases. Our preliminary results show that some aβComAb combining sites on either IgM or IgG class can, without side effects, penetrate the BBB of an AD Tg mouse model with pre-existing Aβ and tau pathology, achieve significant cognitive rescue, reduce levels of Aβ and tau pathological oligomers, and in a CAA AD Tg mouse model (TgSwDI) act without ARIA-like toxicity (both models on murine apoE background). In this project, we will test the hypothesis that aβComAbs (IgM or IgG), unlike mAbs directed against Aβ sequences (6E10), can be infused safely and produce modifying therapeutic effects without vascular ARIA-like toxicity in 3xTg and the vascular TgSwDI or APP/PS1dE9 mouse models cross-bred to human apoE2, E3, or E4 backgrounds. Furthermore, we anticipate that the vascular amyloid proteomes of these mice will resemble those that Projects 1 and 2 associate with different apoE backgrounds, treatments, and presence or absence of microhemorrhages. This information is critically needed to elucidate the mechanism associated with ARIA and select aβComAbs paratopes for future clinical trials. The specific aims are to: 1) Produce and characterize IgM and IgG forms of combining sites (AβComAb paratopes) that specifically recognize the β-sheet secondary structures of toxic oligomers. 2) Determine the biochemical and histochemical interaction of the four aβComAbs from Aim 1 with the brain vasculature of peripherally infused TgSwDI mice crossed on KI human ApoE2, E3, and E4 backgrounds. 3) Determine behavioral, histochemical, and biochemical changes after passive immunotherapy with two AβComAb paratopes on an IgM and an IgG selected from Aim 2, in 3xTg, Tg APP/PS1dE9, and TgSwDI mice crossed on KI human ApoE2, E3, and E4 backgrounds.