Amyloid Beta CAR Macrophages: a cell engineering strategy to clear pathogenic proteins

小胶质细胞的吞噬作用是阿尔茨海默病(AD)风险的核心，小胶质细胞是大脑中常驻的巨噬细胞 发病机制。目前的β-淀粉样蛋白(A-β)清除免疫疗法使用单抗间接诱导 吞噬作用，试图逆转AD的表现。这种形式的被动免疫会诱导小胶质细胞 反应性和神经炎症，这两个特征在AD神经病理标本中非常丰富。由于缺乏 工具，先前对小胶质细胞-Aβ相互作用的机制研究使用了功能丧失策略，如TREM2KO或 小胶质细胞耗尽，不确定小胶质细胞Aβ吞噬本身是否足以改变 神经病理学。为了解决这一知识差距，我们 净空 嫁接 测试 改造巨噬细胞获得特异性淀粉样蛋白 用抗原受体(AβCar)，并开发了高效巨噬细胞的方法 在大脑里。这一双重PI建议将免疫细胞疗法和小胶质细胞生物学的专家团结在一起 最重要的假设是 β嵌合体 β基因工程巨噬细胞体内有效清除淀粉样蛋白的研究 而相关的神经炎症可以通过操纵炎症小体来预防。在……里面 初步数据我们成功设计了一辆β汽车 吞没 vt.进入，进入 确定 嫁接 神经病理学 和 一辆β汽车 暴露 NLRP3 在……上面 移植 确定 在……里面 目标 炎性小体 关 至 表达巨噬细胞，发现它们特异性地 淀粉样蛋白化疗依赖和非依赖方法移植巨噬细胞 小鼠大脑以前所未有的效率，确保了这一方案的可行性。在目标1中，我们将 活性细胞为基础的Aβ靶向如何影响小鼠模型中的淀粉样蛋白病理。要做到这一点，我们将 β车巨噬细胞进入β病理5xFAD模型并检测其作用 和行为。目的1将澄清小胶质细胞Aβ吞噬之间的有争议的关系 使用细胞工程策略的病理学具有很高的治疗潜力。目标1的重点是如何 巨噬细胞影响大脑Aβ，Aim 2阐述了大脑Aβ如何反过来影响巨噬细胞。淀粉样蛋白 诱导小胶质细胞状态改变和神经炎症，具有很强的机械性数据暗示 炎性小体信号是主要驱动因素。因此，目标2将阐明β汽车目标的影响 NLRP3基因缺失对小胶质细胞反应性、神经炎症及其挽救程度的影响。要做到这一点，我们将 将Nlrp3KO或Nlrp3WT供体小鼠的βCAR巨噬细胞移植到5xFAD模型中，并 A)CAR增强的Aβ摄取增加巨噬细胞炎症特征和 神经炎症b)Nlrp3缺失是否挽救了这一疾病，以及c)Aβ病理的后果。 2将提高我们对免疫疗法如何导致神经炎症的理解，并确定 抑制是加强神经退行性变的细胞治疗的一种策略。这些目标加在一起，将 关于小胶质细胞吞噬在AD治疗中的作用的知识空白和创建基于细胞的策略 利用我们的多PI团队已经在人类疾病中测试的原理，专门针对CNS中的蛋白质。 ，并开发了 D

Phagocytosis by microglia, the brain's resident macrophages, is central to Alzheimer Disease (AD) risk and pathogenesis. Current β-amyloid (Aβ) clearing immunotherapies use monoclonal antibodies to indirectly elicit phagocytosis, in an attempt to reverse AD manifestations. This form of passive immunization induces microglial reactivity and neuroinflammation, two features abundant in AD neuropathologic specimens. Due to a dearth of tools, prior mechanistic study of microglia-Aβ interactions used loss of function strategies such as Trem2 KO or microglial depletion, leaving uncertain whether microglial Aβ phagocytosis itself is sufficient to alter neuropathology. To address this knowledge gap, we clearance engraftment test engineered macrophages to achieve specific amyloid with antigen receptors ( AβCAR) , and developed methods for efficient macrophage in the brain. This dual PI proposal unites experts in immune cell therapies and microglial biology to the overarching hypothesis that Aβ-chimeric AβCAR-engineered macrophages effectively clear amyloid in vivo and that associated neuroinflammation is prevented by manipulation of the inflammasome. In preliminary data we successfully engineered AβCAR engulf into determine engraft neuropathology and AβCAR exposure NLRP3 on transplant determine in Aim inflammasome close to expressing macrophages, found that they specifically amyloid chemotherapy-dependent and - independent methods to transplant macrophages the mouse brain at unprecedented efficiency, ensuring the feasibility of this proposal. In Aim 1 we will how active cell-based Aβ targeting affects amyloid pathology in murine models. To do so, we will AβCAR macrophages into the 5xFAD model of Aβ pathology and measure their effects on and behavior. Aim 1 will clarify the controversial relationship between microglial Aβ phagocytosis pathology using a cell engineering strategy with high t herapeutic potential. Where Aim 1 focuses on how macrophages affect the brain Aβ, Aim 2 addresses how brain Aβ in turn affects macrophages. Amyloid induces microglial state changes and neuroinflammation, with strong mechanistic data implicating inflammasome signaling as a major driver. Aim 2 will therefore elucidate the effects of AβCAR targeting microglial reactivity, neuroinflammation, and their degree of rescue by NLRP3 eficiency. To do so, we will AβCAR macrophages engineered from Nlrp3 KO or Nlrp3 WT donor mice into the 5xFAD model and a) the degree to which CAR-augmented Aβ uptake increases macrophage inflammatory profiles and turn neuroinflammation b) whether this is rescued by Nlrp3 loss, and c) the consequences on Aβ pathology. 2 will improve our understanding of how immunotherapies lead to neuroinflammation and determine whether inhibition is a strategy to enhance cell therapies for neurodegeneration. Together, these aims will knowledge gaps about the role of microglial phagocytosis in AD treatment and create a cell-based strategy specifically target proteins in the CNS, using principles our multi-PI team has already tested in human disease. , and developed d