Differentiating Abeta40/42 in plaques with small molecule fluorescent probes

使用小分子荧光探针区分斑块中的 Abeta40/42

基本信息

批准号：
10507501
负责人：
Chongzhao Ran
金额：
$ 45.93万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10507501
关键词：
Age Alanine Alzheimer&apos s Disease Alzheimer&apos s disease brain Alzheimer&apos s disease pathology Amino Acid Sequence Amino Acids Amyloid beta-Protein Antibodies Area Aspartate Autopsy Brain C-terminal Cells Cryoelectron Microscopy Dendritic Spines Development Disease Progression Environment Enzyme-Linked Immunosorbent Assay Epitopes Fluorescence Fluorescent Probes Future Human Hydrophobicity Image Imaging technology Immune Impaired cognition In Vitro Isoleucine Knowledge Length Lysine Machine Learning Mission Molecular Probes Monitor Morphology Mus N-terminal Neurites Oral cavity Pathology Peptides Performance Play Property Roentgen Rays Role Senile Plaques Severities Shapes Site Slide Stains Structure Surface Technology Testing Tissue Extracts Tissue Stains base brain tissue design histological stains hydrophilicity imaging modality in vitro testing neuron loss neurotoxicity preclinical study protein aminoacid sequence small molecule

项目摘要

Amyloid beta (Aβ) plaques constitute one of the most distinctive morphological hallmarks of Alzheimer’s disease (AD). In the past decades, the contribution of Aβ plaques to the overall cognitive decline in AD has been debated extensively. Postmortem studies suggest that plaque abundance does not correlate strongly with the severity of sporadic AD. Conversely, preclinical studies provide strong evidence that plaques are clear sites of pathology and are associated with dystrophic neurites, the loss of dendritic spines and rapid neuron cell death in their surroundings. In the plaques, A40 and A42 peptides are the major constituents. Nonetheless, unlike the role of plaque, there is nearly no argument that A42 has a much higher neurotoxicity than A40 does. Conceivably, differentiating A40 and A42 can considerably clarify the role of plaque in AD pathology. Differentiating A40 and A42 has long been considered as an impossible mission with small-molecule probes, due to the small difference in the amino acid sequence of the peptides. It is obvious that the C-terminal of A peptide is the key for designing small-molecule probes to distinguish them, because the only difference of two amino acids (isoleucine-alanine) is within the C-terminals of A40/42 peptides. There is no prior knowledge to teach us how to design such probes. Nonetheless, it has been routinely performed with anti-A42 antibodies to determine the contents of A42 in cell media and brain extracts. These anti-A42 antibodies were designed based on the epitope of the C-terminal of the peptide. This fact has bolstered us to believe that the properties of the C-terminal can be relied on to design our small molecule probes. In the past, X-ray structures of full-length As were rare. However, recently the advanced Cryo-EM technology has impressively facilitated A structure studies. In our preliminary results, based on the unique environment of the C-terminal, we designed small- molecule fluorescence probe ICTAD-1 that has the capacity to spectrally differentiate A40/42 in vitro and brain slides. In this proposal, we plan to design new probes and validate their capacity for differentiating A40/42.

β 淀粉样蛋白 (Aβ) 斑块是阿尔茨海默病最独特的形态学标志之一 (AD) 在过去的几十年里，Aβ 斑块对 AD 整体认知能力下降的影响一直存在争议。尸检研究表明，斑块丰度与斑块的严重程度没有很强的相关性。离线、临床前研究提供了强有力的证据，证明斑块是明确的病理部位。并与神经突营养不良、树突棘丧失和神经元细胞快速死亡有关然而，与作用不同，斑块中的主要成分是 A40 和 A42 肽。可以想象，A42 比 A40 具有更高的神经毒性，这一点几乎没有争议。区分 A40 和 A42 可以在很大程度上阐明斑块在 AD 病理学中的作用。长期以来，区分 A40 和 A42 一直被认为是小分子探针不可能完成的任务，由于肽的氨基酸序列差异很小，很明显A的C端。肽是设计小分子探针来区分它们的关键，因为两者唯一的区别氨基酸（异亮氨酸-丙氨酸）位于 A40/42 肽的 C 末端，目前尚无相关知识。教我们如何设计这样的探针然而，它已经常规地使用抗A42抗体来进行。确定细胞培养基和脑提取物中 A42 的含量设计了这些抗 A42 抗体。基于肽 C 末端的表位这一事实支持了我们相信的特性。过去，我们可以依靠C端来设计全长的X射线结构。 A 很少见，然而，最近先进的冷冻电镜技术令人印象深刻地促进了 A 结构。在我们的初步研究结果中，基于C端的独特环境，我们设计了小型- 分子荧光探针 ICTAD-1，能够在体外和大脑中对 A40/42 进行光谱区分在本提案中，我们计划设计新的探针并验证其区分 A40/42 的能力。