Multiplexed immunoassay for building patient-specific molecular profiles of CSF amyloid beta and TAU

摘要 该项目将开发一种多重免疫测定法，用于测量这两种蛋白的患者特异性分子谱。 阿尔茨海默病（AD）的蛋白质标志物：来自人脑脊液（CSF）的淀粉样蛋白β（ABeta）和TAU。 由于存在多种差异切割形式（ABeta）和6，这两种蛋白质在结构上都很复杂。 含有50+磷酸化位点（TAU）的同种型。CSF提供了丰富的潜在生物标志物来源， 可以广泛挖掘，以建立AD和AD相关痴呆（ADRD）的诊断和预后特征 或者开发个性化的患者档案用于药物发现。目前，这些努力受到 缺乏解析多种ABeta和TAU蛋白质型的技术能力，更具体地说是缺乏产品 用于ABeta和TAU的多重捕获和富集，用于基于定量质谱的蛋白质组学。 本提案旨在建立在Adepponent早期成功执行的产品开发项目的基础上， 一个医药客户使用我们的BAMS™平台，我们能够鉴定出至少14种新的低丰度CSF ABeta肽，并通过添加两种新类型的探针， 其补充了用于总TAU（tTAU）和磷酸-TAU（pTAU）的现有常规探针。拟议 称为ABeta/TauScan™的免疫测定将满足生物学家、临床医生和制药公司当前的需求， 用于研究神经退行性变蛋白标志物的全面分子分析工具。此外，它将 为开发其他蛋白质靶点（如神经丝轻多肽（NFL））的类似检测方法开辟了道路 和α-突触核蛋白。虽然ABeta/TauScan™主要用于CSF，但在II期后将对试剂进行评价 用于血清/血浆。 在整个项目中，我们将与学术，临床和行业合作伙伴密切合作，以确保强大的 ABeta/TauScan™的分析性能。一旦建立了检测试剂盒，将通过筛选300 CSF进行验证 代表认知正常（CN）受试者、轻度认知障碍（MCI）和AD受试者的样品，以及 建立CN至MCI至AD进展的分子标记，其将包含至少一种新的标记物， 提高了测定灵敏度和特异性。

ABSTRACT This project will develop a multiplexed immunoassay for measuring patient-specific molecular profiles of the two protein markers of Alzheimer’s disease (AD): amyloid beta (ABeta) and TAU from human cerebrospinal fluid (CSF). Both proteins are structurally complex due to the existence of multiple differentially cleaved forms (ABeta) and 6 isoforms containing 50+ phosphorylation sites (TAU). CSF provides a rich source of potential biomarkers, which can be extensively mined to build diagnostic and prognostic signatures of AD and AD-related dementias (ADRD) or to develop personalized patient profiles for drug discovery. At present, those efforts are constrained by the lack of technical capability to resolve multiple ABeta and TAU proteoforms, more specifically by the lack of products for multiplexed capture and enrichment of ABeta and TAU for quantitative mass spectrometry-based proteomics. This proposal seeks to build upon an earlier successful product development project performed by Adeptrix for a pharma customer. Using our BAMS™ platform, we were able to identify at least 14 novel low abundance CSF ABeta peptides and dramatically expand the sequence coverage of CSF TAU by adding two new types of probes, which complement the existing conventional probes for total (tTAU) and phospho-TAU (pTAU). The proposed immunoassay, termed ABeta/TauScan™ will address the current need of biologists, clinicians, and pharma for comprehensive molecular profiling tools for studying protein markers of neurodegeneration. Furthermore, it will open a path to developing similar assays for other protein targets, such as neurofilament light polypeptide (NFL) and alpha-synuclein. While ABeta/TauScan™ is intended primarily for CSF, post Phase II the reagents will be evaluated for use in serum/plasma. Throughout this project we will work closely with academic, clinical and industry partners to ensure the robust analytical performance of ABeta/TauScan™. Once the assay is created, it will be validated by screening 300 CSF samples representing cognitively normal (CN) subjects, mild cognitive impairment (MCI) and AD subjects and establishing a molecular signature of CN to MCI to AD progression that will contain at least one novel marker for improved assay sensitivity and specificity.