Project 3: Development of multiplex assays for clinical monitoring of disease

项目 3：开发用于疾病临床监测的多重检测方法

• 批准号: 10573266
• 负责人: ANDREW N HOOFNAGLE
• 金额: $ 60.82万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10573266
• 关键词: Alternative Splicing; Alzheimer' s Disease; Alzheimer' s disease diagnosis; Alzheimer' s disease patient; Alzheimer' s disease related dementia; Amyloid; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Bathing; Behavior; Biochemical; Biological; Biological Assay; Biological Markers; Cause of Death; Cell Death; Cell Separation; Central Nervous System; Cerebral cortex; Cerebrospinal Fluid; Cessation of life; Clinical; Computer software; Data; Dementia; Deposition; Development; Disease; Disease Management; Ensure; Event; Goals; Guidelines; Hand; Impaired cognition; Injury; Laboratories; Lipoproteins; Liquid Chromatography; Liquid substance; Mediator; Methods; Molecular Conformation; Molecular Weight; Monitor; Neurodegenerative Disorders; Neurofibrillary Tangles; Neuronal Dysfunction; Neurons; Outcome; Pathologic; Patients; Phosphorylation Site; Post-Translational Protein Processing; Procedures; Process; Proteins; Proteomics; Reproducibility; Research Personnel; Sampling; Senile Plaques; Speed; Synapses; Tissue imaging; Validation; abeta deposition; amyloid formation; biomarker discovery; brain tissue; clinical translation; differential expression; experience; extracellular vesicles; gene product; gray matter; interest; mild cognitive impairment; multiplex assay; neuron loss; next generation; novel; novel marker; protein aggregation; regional atrophy; success; tandem mass spectrometry; tau Proteins; tau-1; translational proteomics; verification and validation

Abstract The pathological hallmarks of Alzheimer’s disease are the neurofibrillary tangles and amyloid plaques that form in gray matter regions of the cerebral cortex. Misfolded and aggregated proteins reside within those pathological features, which may to be central to the neuronal death that causes regional atrophy, dementia, and ultimately death. The cerebrospinal fluid that bathes the central nervous system is the fluid most proximal to the disease and has been the focus of biomarker discovery for more than 30 years. There are two important gene products that are present in cerebrospinal fluid and have been shown to be predictive of disease activity and cognitive decline in patients that present with mild cognitive impairment: (1) the amyloid precursor protein, which gives rise to Aβ(1-42) and other fragments, and (2) tau, which has many phosphorylation sites. The deposition of Aβ(1-42) as amyloid aggregates appears to lead to the death of adjacent neurons. The resulting injury increases the amount of tau protein released into the cerebrospinal fluid. From previous biomarker studies of Aβ proteins and tau, it seems likely that the entire disease process, from Aβ(1-42) deposition to cell death, is orchestrated differently than in other neurodegenerative diseases. As a result, it is a compelling hypothesis that novel biomarkers, which may be mediators of disease, are present in the cerebrospinal fluid and could add more information over the current biomarkers used in the diagnosis of Alzheimer’s disease. There have been previous attempts to use discovery proteomics to identify proteins that are differentially expressed in cerebrospinal fluid in patients with Alzheimer’s disease compared with controls and there have been few efforts to expand upon these studies in a clinically meaningful way. However, some of these efforts have identified the presence of additional fragments of amyloid precursor protein [i.e., in addition to Aβ(1-42)] and other proteins, which suggests that post-translational changes may be important in the formation of amyloid, the dysfunction of neuronal synapses, and the subsequent death of neurons. In this project entitled, “Identification of mass spectrometric targets and development of multiplex assays for disease management,” our objective is to identify the post-translational modifications that lead to altered LP/EV concentration, molecular weight, and stability in cerebrospinal fluid. Part of the reason that the clinical translation of Aβ(1-42), tau, and phosphorylated tau as cerebrospinal fluid biomarkers took decades was that reproducible assays were not available early on. As a result, another overarching goal of this project is to develop precise, transferable, validated targeted proteomic assays to quantify proteins in cerebrospinal fluid that can be used to investigate disease mechanism and predict poor outcomes.

抽象的 阿尔茨海默病的病理特征是形成神经原纤维缠结和淀粉样斑块 错误折叠和聚集的蛋白质存在于大脑皮层的灰质区域中。 病理特征，这可能是局部导致萎缩、痴呆的神经元死亡的核心， 沐浴中枢神经系统的脑脊液是最接近的液体。 30多年来，有两个重要的生物标志物一直是该疾病发现的焦点。 存在于脑脊液中并已被证明可以预测疾病活动的基因产物 轻度认知障碍患者的认知能力下降：（1）淀粉样前体蛋白， 它产生 Aβ(1-42) 和其他片段，以及 (2) tau，它具有许多磷酸化位点。 Aβ(1-42) 作为淀粉样蛋白聚集体的沉积似乎会导致邻近神经元的死亡。 损伤会增加先前生物标志物释放到脑脊液中的 tau 蛋白量。 对 Aβ 蛋白和 tau 蛋白的研究表明，从 Aβ(1-42) 沉积到细胞内的整个疾病过程似乎 死亡的发生方式与其他神经退行性疾病不同，因此，它是一种引人注目的疾病。 假设脑脊液中存在可能是疾病介质的新生物标志物 并可以为当前用于诊断阿尔茨海默病的生物标志物添加更多信息。 之前曾尝试使用发现蛋白质组学来识别差异蛋白质 与对照组相比，阿尔茨海默病患者的脑脊液中表达 很少有人努力以具有临床意义的方式扩展这些研究，但是其中一些努力。 已鉴定出淀粉样前体蛋白的其他片段的存在[即，除了 Aβ(1-42) 之外] 和其他蛋白质，这表明翻译后变化可能对形成 淀粉样蛋白、神经元突触功能障碍以及随后的神经元死亡。 在这个题为“质谱目标的识别和多重分析的开发”的项目中 疾病管理”，我们的目标是确定导致疾病管理的翻译后修饰 LP/EV在脑脊液中的浓度、分子量和稳定性的部分原因。 Aβ(1-42)、tau 和磷酸化 tau 作为脑脊液生物标志物的临床转化花费了数十年 因此，早期无法进行可重复的检测，这是该项目的另一个总体目标。 的目标是开发精确的、可转移的、经过验证的靶向蛋白质组测定法，以量化蛋白质组学中的蛋白质 脑脊液可用于研究疾病机制并预测不良结果。