Fluorescent probes for detection of misfolded protein oligomers in Alzheimer's Disease and related disorders

用于检测阿尔茨海默病和相关疾病中错误折叠蛋白寡聚体的荧光探针

• 批准号: 10604908
• 负责人: David E Kang
• 金额: $ 20.04万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10604908
• 关键词: APP-PS1; Address; Alzheimer' s Disease; Alzheimer' s disease brain; Alzheimer' s disease model; Alzheimer' s disease related dementia; Amyloid; Amyloid beta-Protein; Animal Model; Antibodies; Binding; Biological; Biological Assay; Biological Markers; Categories; Characteristics; Chemicals; Clinical; Data; Dependence; Deposition; Detection; Development; Disease; Dyes; Evolution; Exhibits; Fluorescence; Fluorescent Dyes; Fluorescent Probes; Future; Gentian Violet; Goals; Growth; Human; Image; In Vitro; Kinetics; Label; Laboratories; Link; Monitor; Muramidase; Mus; Normal tissue morphology; Oranges; Outcome; Pathogenicity; Pathologic; Pathology; Patients; Phase; Population; Positron-Emission Tomography; Proliferating; Protocols documentation; Public Health; Role; Sampling; Series; Solid; Solvents; Specificity; Stains; Symptoms; Techniques; Testing; Thioflavin S; Tissues; Transgenic Mice; Transgenic Organisms; Validation; abeta oligomer; amyloid formation; animal tissue; brain tissue; cohort; detection assay; detection method; experimental study; imaging probe; improved; in vivo; in vivo imaging; in vivo monitoring; indexing; innovation; insight; mild cognitive impairment; misfolded protein; monomer; novel; protein oligomer; response; scaffold; screening; success; tau Proteins; tau aggregation

Fluorescent Probes for Detection of Misfolded Protein Oligomers in Alzheimer's Disease and Related Disorder There is significant evidence that the clinical symptoms of Alzheimer’s Disease (AD) and related disorders are closely linked to the formation and proliferation of small oligomers that precede the emergence of the prominent late-stage fibrils and plaques. Therefore, amyloid beta oligomers (AβOs) are the most direct biomarkers for monitoring the onset and progression of AD. Attempts at utilizing this biomarker, however, have been severely hampered by the dearth of techniques for the reliable detection of AβOs in biological samples and tissues. While oligomer-selective antibodies have provided important insights into AβOs, their use doesn't extend to detecting oligomer populations in vivo, let alone monitoring their temporal evolution. The overall objectives of this proposal are therefore to identify small oligomer-selective dyes for the detection of AβOs and to validate their specificity for AβOs, and potentially related oligomers, in tissues of animal models of AD and patient tissue. Multiple laboratories have observed that in vivo Aβ assembly displays not only purely sigmoidal but also biphasic ThT kinetics. We have shown that the onset of biphasic ThT kinetics directly correlates with the onset and rapid increase in prefibrillar oligomer populations with increases in monomer concentrations. Here we propose to use this transition from essentially oligomer-free sigmoidal to oligomer-dominated biphasic kinetics to screen a selection of readily available fluorescent dyes for their selectivity for AβOs over AβFs and monomers. An initial test of this approach already yielded a highly promising dye candidate. Our preliminary data also indicate that this dye specifically stains oligomer deposits in animal models of AD. While very encouraging, the utility of our current oligomer-selective dye requires further validation. In addition, we seek to identify multiple chemically and structurally distinct oligomer-selective dyes to improve the chances to develop one of them into a PET probe for in vivo imaging of oligomers. We will therefore extend our current screen for AβO-selective dyes to a larger set of fluorescent dyes selected from different dye categories. In parallel, we will scrutinize whether the current dye reliably detects AβOs at various stages of the disease, and does so in animal models as well as patient tissues. Promising novel AβO-selective dyes identified through our screening assay will be subjected to the same ex vivo validation of their specificity in tissues. We anticipate that these experiments will yield multiple promising AβO-selective dyes with application for fundamental studies of oligomer formation, for the development of new assays for detecting AβOs ex vivo, and, most importantly, as the detection moiety for a future oligomer-selective PET probe for antemortem in vivo oligomer imaging in patients.

用于检测阿尔茨海默病中错误折叠蛋白寡聚体的荧光探针 疾病及相关病症 有显著的证据表明，阿尔茨海默病（AD）和相关疾病的临床症状 与小寡聚体的形成和增殖密切相关，这些小寡聚体先于 后期纤维和斑块突出。因此，淀粉样蛋白β寡聚体（AβOs）是最直接的 用于监测AD的发作和进展的生物标志物。然而，利用这种生物标志物的尝试， 由于缺乏可靠检测生物样品中Aβ O的技术， 组织中虽然寡聚体选择性抗体为AβOs提供了重要的见解，但它们的应用并没有扩大到 更不用说监测它们的时间演变了。总体目标 因此，该提案的目的是确定用于检测Aβ O的小寡聚体选择性染料，并验证 它们对AD动物模型组织和患者组织中的Aβ 0和潜在相关寡聚体的特异性。 多个实验室已经观察到，体内Aβ组装不仅表现出纯粹的S形， 双相ThT动力学。我们已经表明，双相ThT动力学的开始与ThT的开始直接相关。 以及随着单体浓度的增加，预纤维状低聚物群体迅速增加。这里我们 我建议使用这种从基本上无低聚物的S形到低聚物主导的两相动力学的转变 筛选一系列易于获得的荧光染料，以确定其对Aβ O的选择性优于Aβ F和单体。 对这种方法的初步测试已经产生了一种非常有前途的染料候选物。我们的初步数据还 表明该染料特异性染色AD动物模型中的寡聚体沉积物。 虽然非常令人鼓舞，但我们目前的寡聚体选择性染料的实用性需要进一步验证。在 此外，我们试图确定多种化学和结构上不同的寡聚体选择性染料，以改善其性能。 有机会将其中之一开发成PET探针，用于寡聚体的体内成像。因此，我们将扩大 目前筛选Aβ O-选择性染料的方法是从不同的染料类别中选出一组更大的荧光染料。在 与此同时，我们将仔细检查目前的染料是否能可靠地检测疾病不同阶段的Aβ O， 在动物模型和患者组织中也是如此。通过我们的研究发现了有前途的新型Aβ O选择性染料 筛选试验将进行相同的组织特异性离体验证。 我们预计，这些实验将产生多种有前途的Aβ O选择性染料， 寡聚体形成的基础研究，用于开发体外检测AβOs的新方法，以及， 最重要的是，作为未来用于死前体内的寡聚体选择性PET探针的检测部分， 寡聚体成像。