Inhibition of CD33-sialic acid binding in Late-Onset Alzheimer's Disease

迟发性阿尔茨海默病中 CD33-唾液酸结合的抑制

基本信息

批准号：
10752139
负责人：
Jennifer Leigh Green
金额：
$ 4.77万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10752139
关键词：
Affinity Alanine Alzheimer&apos s Disease Alzheimer&apos s disease model Alzheimer&apos s disease patient Amyloid Amyloid beta-Protein Binding Binding Proteins Biological Assay Brain Cells Development Disease Progression Gene Expression Genes Genetic study Genotype Glycoproteins Human Immune Immune signaling Immunoprecipitation Impaired cognition In Vitro Individual Inflammatory Investigation Late Onset Alzheimer Disease Length Ligands Measures Mediating Methods Microglia Molecular Morphology Myeloid Cells Neurodegenerative Disorders PTPRC gene Pathogenesis Pathogenicity Pathology Pathway interactions Peptides Phase Phosphoric Monoester Hydrolases Predisposition Process Property Protein Isoforms Proteins Risk Risk Factors Role Senile Plaques Sialic Acids Signal Pathway Signal Transduction Solid Specificity Structure Surface TREM2 gene Testing Thick Western Blotting abeta accumulation amyloid pathology design experimental study genetic association genetic risk factor genome wide association study glial activation human model human old age (65+)improved in vitro Model in vivo imaging inhibitor insight monocyte neuron loss protein expression receptor response risk variant sialic acid binding Ig-like lectin sialic acid receptor targeted treatment tau Proteins uptake

项目摘要

PROJECT SUMMARY Alzheimer’s disease (AD) is a debilitating neurodegenerative disease characterized by progressive cognitive decline and pre-symptomatic accumulation of amyloid-beta (Aβ) and tau pathology with older age. AD pathogenesis is poorly understood, however genetic studies have clearly indicated a causal role for microglia, the innate immune cells of the CNS. Indeed, GWAS established the innate immune gene CD33 as a risk factor for Late-Onset AD (LOAD) in 2011. The CD33 LOAD associated rs3865444CC risk genotype is associated with diminished internalization of amyloid-β1-42 peptide, accumulation of neuritic amyloid pathology and fibrillar amyloid on in vivo imaging, and increased numbers of human microglia with small, thick processes, and a rounded morphology compared to the rs3865444AA protective genotype. Individuals with the LOAD associated rs3865444CC risk genotype have increased expression of full-length CD33, the isoform containing the sialic acid binding domain, compared to those with the rs3865444AA protective genotype, suggesting that the sialic- acid binding domain may be critical to the genetic association to AD. Based on the known inhibitory function of CD33, my hypothesis is that sialic acid mediated CD33 activation leads to microglial suppression, resulting in the inability to resolve inflammatory processes and mitigate pathogenic amyloid plaques, which may heighten susceptibility to neuronal loss and contribute to AD progression. To test this hypothesis, our lab developed an alternative approach for inhibiting CD33 function, by using peptides designed to bind to sialic acid to act as “decoy” receptors for sialic acid to reduce it’s binding to CD33 and inhibit CD33 signaling. I hypothesize that competitive inhibition of CD33 with sialic acid will reduce CD33 activation, restore microglial responsiveness, and increase Aβ uptake. The overarching objective of this proposal is to characterize the binding properties of these peptides and their inhibitory effects on CD33 activation and signaling (Aim 1) and then examine the functional responses of these peptides on immune pathways in vitro (Aim 2). In Aim 1, I will determine the potency and selectivity of peptides 1 and 2 in inhibiting CD33-sialic acid binding in a solid-phase binding assay I have optimized (Aim1a), determine the critical residues for efficacy through an alanine screen (Aim1b), and investigate the effects of the peptides on CD33 activation and signaling (Aim1c). In Aim 2, I will look at how human microglia functionally respond to treatment with peptides 1 and 2 in the context of AD using primary microglia-like cells from healthy donor and AD patients expressing the CD33 risk and protective genotypes by measuring gene expression changes in markers of microglial activation and amyloid uptake (Aim2a) and expression changes in key LOAD-associated immune-related proteins (Aim 2b).These experiments will provide insights to not only the mechanism of action of these peptides, but also to further clarify the role of CD33 and Siglec receptors in microglial activation, immune signaling pathways, and the pathogenesis of Alzheimer’s disease.

项目概要阿尔茨海默病 (AD) 是一种使人衰弱的神经退行性疾病，其特征是进行性认知障碍随着年龄的增长，β-淀粉样蛋白 (Aβ) 和 tau 蛋白病理学下降和症状前积累。人们对发病机制知之甚少，但遗传学研究清楚地表明小胶质细胞的因果作用，事实上，GWAS 将先天免疫基因 CD33 确定为危险因素。 2011 年晚发 AD (LOAD)。CD33 LOAD 相关 rs3865444CC 风险基因型与淀粉样蛋白-β1-42 肽的内化减少，神经炎淀粉样蛋白病理和纤维的积累体内成像中的淀粉样蛋白，以及具有小而厚的突起的人类小胶质细胞数量的增加，以及与 rs3865444AA 保护性基因型相比，具有 LOAD 相关的个体具有圆形形态。 rs3865444CC 风险基因型增加了全长 CD33 的表达，CD33 是含有唾液酸的亚型与具有 rs3865444AA 保护性基因型的那些相比，酸结合域，表明唾液酸- 根据已知的 AD 抑制功能，酸结合域可能对 AD 的遗传关联至关重要。 CD33，我的假设是唾液酸介导的 CD33 激活导致小胶质细胞抑制，从而导致无法解决炎症过程和减轻致病性淀粉样斑块，该斑块可能会更高为了检验这一假设，我们的实验室开发了一种方法。抑制 CD33 功能的另一种方法是使用设计用于与唾液酸结合的肽作为唾液酸的“诱饵”受体，以减少其与 CD33 的结合并抑制 CD33 信号传导。用唾液酸竞争性抑制 CD33 将减少 CD33 活化，恢复小胶质细胞反应性，该提案的首要目标是表征 Aβ 的结合特性。这些肽及其对 CD33 激活和信号传导的抑制作用（目标 1），然后检查这些肽对体外免疫途径的功能反应（目标 2）。在目标 1 中，我将确定固相结合测定中肽 1 和 2 抑制 CD33-唾液酸结合的效力和选择性我已经优化 (Aim1a)，通过丙氨酸筛选 (Aim1b) 确定功效的关键残留物，并且研究肽对 CD33 激活和信号传导的影响 (Aim1c) 在目标 2 中，我将了解如何进行。人类小胶质细胞在 AD 背景下对肽 1 和 2 的治疗有功能性反应来自健康供体和 AD 患者的小胶质细胞样细胞表达 CD33 风险和保护基因型测量小胶质细胞激活和淀粉样蛋白摄取标记物的基因表达变化（Aim2a）和关键 LOAD 相关免疫相关蛋白的表达变化（目标 2b）。这些实验将提供不仅深入了解这些肽的作用机制，还进一步阐明 CD33 和 Siglec 受体在小胶质细胞激活、免疫信号通路和阿尔茨海默病发病机制中的作用疾病。