Linkage-specific ubiquitylation patterns as highly sensitive markers for neurodegenerative disease

虽然灵敏的成像工具可以检测β淀粉样蛋白和tau蛋白等生物标志物 阿尔茨海默病(AD)和其他神经退行性疾病是可用的，类似的工具可以检测到 这些疾病的症状性、病因性生物标志物不是。AD中最早的步骤之一是 泛素蛋白酶体途径通过使错误折叠的蛋白质降解来对其作出反应，从而促进 聚合。这一早期病理事件的部分标志是蛋白质泛素化模式的变化。 神经细胞。“泛素化模式”包括多个泛素链和赖氨酸位置，这 信息远比简单的泛素化事件复杂得多。此外，神经元的泛素化模式 阿尔茨海默病患者的细胞随着疾病的进展而变化。无先兆症状或病因的非侵入性血液检测 治疗阿尔茨海默病S病的药物已被美国食品和药物管理局批准。除了诊断外，识别分子的能力 AD的特征可以帮助将患者群体分层，泛素化模式的变化也可以使用 监测以泛素蛋白酶体系统为靶点的药物的临床疗效。这个项目的目的是 通过以下方式捕捉神经退行性疾病建立的初始步骤以及疾病进展的阶段 确定可在诊断试验中常规检测的适当泛素化模式特征。 为此，在第一阶段，生命传感器公司开发了一种高度敏感的方法来分离多泛素化的 基于使用管的Ub链选择性的蛋白质--一种可以选择性地与多种 独特的多Ub链--与质谱学(MS)相结合，根据 泛素化。在模拟神经变性的神经细胞中模拟条件，并标记蛋白质 此外，还开发了一些方法来鉴定低水平的泛素化蛋白并观察 正常和AD患者脑脊液泛素化模式的变化在第二阶段， 建议将这些研究扩展到检测脑脊液和血清泛素化蛋白分子特征 大约100名正常和AD患者。Lifesensors将与尼古拉斯·塞弗里德博士和艾伦博士合作 阿尔茨海默病研究中心的利维，AD蛋白质组学的领导者，翻译开发的工具 在第一阶段，监测正常人和AD患者脑脊液泛素化模式的变化，并建立 生物标记物面板，区分不同疾病阶段的AD患者和对照组，验证标记物 利用现有的数据库和预测性研究。第二阶段的成功将导致小说的发现 AD不同阶段的泛素化特征。Lifesensors将把稳定同位素标记版本商业化 通过在血清和/或脑脊液中的质谱仪对这些蛋白质中的标志物进行定量。这些简单的测试将分层 患者群体在临床试验中监测疗效，指导治疗AD的药物选择和剂量。

While sensitive imaging tools to detect β amyloid and tau proteins and other biomarkers for established Alzheimer’s disease (AD) and other neurodegenerative diseases are available, similar tools for detecting pre- symptomatic, etiologic biomarkers of these diseases are not. Among the earliest steps in AD is a failure of the ubiquitin proteasome pathway to respond to misfolded proteins by causing their degradation, thereby promoting aggregation. This early pathological event is marked in part by changes in protein ubiquitylation patterns in neuronal cells. “Ubiquitylation patterns” encompasses multiple ubiquitin chains and lysine positions, and this information is far more complex than a simple ubiquitylation event. In addition, ubiquitylation patterns in neuronal cells of AD patients change as the disease progresses. No non-invasive blood test for pre-symptoms or etiology of Alzheimer ’s disease has been approved by the FDA. In addition to diagnosis, the ability to identify molecular signatures of AD could help stratify the patient population, and changes in ubiquitylation patterns could be used to monitor the clinical efficacy of drugs that target the ubiquitin proteasome system. The aim of this project is to capture the initial step in neurodegenerative disease establishment as well as stages in disease progression by identifying appropriate ubiquitylation pattern signatures that can be detected routinely in a diagnostic assay. Toward this end, in Phase I, LifeSensors developed a highly sensitive method to isolate poly-ubiquitylated proteins based on Ub chain selectivity using TUBES – an affinity matrix that can bind selectively to various distinct poly-Ub chains -- in concert with mass spectrometry (MS) to identify molecular signatures based on ubiquitylation. Conditions were simulated in neuronal cells that mimic neurodegeneration, and marker proteins were identified; in addition, methods were developed to identify low levels of ubiquitylated proteins and observe changes in ubiquitylation patterns from cerebrospinal fluid (CSF) of normal and AD patients. In Phase II, it is proposed to extend these studies to examine CSF and serum ubiquitylated protein molecular signatures from approximately 100 normal and AD patients. LifeSensors will collaborate with Drs. Nicholas Seyfried and Allan Levey of the Alzheimer’s Disease Research Center, leaders in AD proteomics, to translate the tools developed in Phase I to monitor ubiquitylation pattern changes in CSF of normal controls and AD patients and to establish biomarker panels that distinguish controls from AD patients at various disease stages, validating the markers using existing databases and predictive studies. Success in Phase II will lead to the discovery of novel ubiquitylation signatures for various stages of AD. LifeSensors will commercialize stable isotope labeled versions of these proteins to quantify the markers by mass spec in the serum and/or CSF. These simple tests will stratify patient populations to monitor efficacy in clinical trials and guide the choice and dosage of drug to treat AD.