STRUCTURAL BIOLOGY OF ALPHA-SYNUCLEIN IN LEWY BODY DEMENTIA

帕金森病（PD）在病理学上被定义为α-突触核蛋白（Asyn）原纤维在神经元细胞质和称为路易体和路易神经突的神经炎包涵体中的积累。Asyn在PD发病机制中的作用得到了罕见家族性PD中编码Asyn的基因（SNCA）中显性突变的鉴定的支持。PD常发生痴呆。它有时与运动症状（通常称为路易体痴呆或DLB）大约同时开始，或在运动症状开始后长达20年（PD痴呆或PDD）。术语路易体痴呆（LBD）包括这一系列临床表现，并与Asyn纤维在整个大脑，特别是新皮质中的广泛沉积有关。正在寻求多种针对Asyn积累的治疗方法。另一个优先事项是开发一种PET成像剂，以量化Asyn在活体中的沉积，作为靶点参与和疾病进展的生物标志物。了解LBD中的Asyn纤维结构可以指导Asyn靶向治疗和成像剂的开发。在这个项目中，我们将使用低温电子显微镜（cryo-EM）来确定LBD中Asyn纤维的原子分辨率结构，结合固态NMR（SSNMR）来细化结构。我们将分析和比较从LBD尸检病例的多个亚组中分离的Asyn原纤维的结构，这些亚组由早发性与迟发性痴呆定义，以及是否存在共同发生的淀粉样蛋白β积累。我们还将利用冷冻电子断层扫描，SSNMR光谱分析和新的单克隆抗体，以扩展对Asyn原纤维结构的分析，以增加尸检病例。为了促进这些结构研究的翻译，我们将利用cryo-EM来确定PET成像配体的主要候选物的结合位点，并使用原纤维生长测定来鉴定Asyn蛋白中对原纤维生长和稳定性重要的特定氨基酸残基。

Parkinson’s disease (PD) is defined pathologically by the accumulation of alpha-synuclein (Asyn) fibrils in neuronal cytoplasmic and neuritic inclusions known as Lewy bodies and Lewy neurites. The role of Asyn in the pathogenesis of PD is supported by the identification of dominant mutations in the gene encoding Asyn (SNCA) in rare familial versions of PD. Dementia occurs frequently in PD. It sometimes begins at approximately the same time as motor symptoms (often referred to as dementia with Lewy bodies or DLB), or up to 20 years after motor symptoms begin (PD with dementia or PDD). The term Lewy body dementia (LBD) encompasses this spectrum of clinical presentations and is associated with widespread deposition of Asyn fibrils throughout the brain, particularly neocortex. Multiple therapeutic approaches targeting Asyn accumulation are being pursued. A further priority is to develop a PET imaging agent to quantify the deposition of Asyn in living individuals, as a biomarker for target engagement and disease progression. Understanding Asyn fibril structure in LBD can guide the development of Asyn-targeted therapies and imaging agents. In this project, we will use cryo-electron microscopy (cryo-EM) to determine atomic resolution structures of Asyn fibrils in LBD, in conjunction with solid-state NMR (SSNMR) for refinement of structures. We will analyze and compare structures of Asyn fibrils isolated from multiple subgroups of LBD autopsy cases defined by early versus late onset of dementia, as well as the presence or absence of co-occurring amyloid β accumulation. We will also utilize cryo-electron tomography, SSNMR spectral analysis and new monoclonal antibodies to extend the analysis of Asyn fibril structure to additional autopsy cases. To promote the translation of these structural studies we will utilize cryo-EM to determine binding sites of leading candidates for PET imaging ligand and use fibril growth assays to identify specific amino acid residues in the Asyn protein that are important for fibril growth and stability.