Structural Biology of Alpha-Synuclein in Lewy Body Dementia

摘要 帕金森病(PD)的病理定义是α-突触核蛋白(Asyn)纤维在 神经元胞浆和神经元包涵体称为路易小体和路易神经突起。ASYN在网络中的作用 编码Asyn基因的显性突变支持帕金森病的发病机制 (SNCA)在罕见的家族性帕金森病中。痴呆症在帕金森病患者中高发。它有时开始于 与运动症状大致相同的时间(通常称为路易小体痴呆或DLB)，或 运动症状开始后长达20年(帕金森病伴痴呆症或PDD)。路易体痴呆(LBD)一词 包括这种临床表现的谱系，并与广泛的不同步沉积有关 纤维遍布大脑，特别是新大脑皮层。针对Asyn的多种治疗方法 人们正在追求积累。另一个优先事项是开发正电子发射断层扫描(PET) 用于量化Asyn在活体个体中的沉积的显像剂，作为靶点接触和 疾病的发展。了解LBD的Asyn原纤维结构可指导Asyn靶向疾病的发展 治疗和显像剂。在这个项目中，我们将使用固态核磁共振(SS核磁共振)和低温核磁共振的组合。 电子显微镜(Cryo-EM)确定LBD中Asyn原纤维的原子分辨结构。我们开发了 从组织中分离纤维并在标记的情况下生长的多种互补方法 用于SS核磁共振和冷冻-EM研究的单体Asyn蛋白质，使更全面的结构分析成为可能。 我们将分析和比较从LBD尸检病例的多个亚组分离的Asyn纤维的结构 定义为早发和晚发痴呆症，以及是否存在共同出现的淀粉样蛋白β 积累，并确定结构变异是否与疾病表型有关。为了促进 这些结构研究的翻译我们将使用SS核磁共振和冷冻-EM来确定结构特征 主要的PET成像配体的结合位点，可以指导进一步的PET配体的优化。 最后，我们将利用来自LBD组织的Asyn纤维在细胞培养中积累Asyn纤维。 和老鼠模型。我们将利用SS核磁共振、低温电子显微镜和冷冻电子断层扫描技术来分析异步纤维 这些模型系统中的结构，将指导未来疾病机制的研究，PET配体 发展和治疗发展。

Abstract Parkinson 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 Positron Emission Tomography (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 a combination of solid-state NMR (SSNMR) and cryo- electron microscopy (cryo-EM) to determine atomic resolution structures of Asyn fibrils in LBD. We developed multiple complimentary approaches to isolate fibrils from tissue and grow them in the presence of labeled monomeric Asyn protein for SSNMR and cryo-EM studies, enabling more comprehensive analysis of structure. 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, and determine whether structural variations relate to disease phenotype. To promote the translation of these structural studies we will utilize SSNMR and cryo-EM to determine the structural features of binding sites for leading PET imaging ligand candidates, which can guide further optimization of PET ligands. Finally, we will utilize Asyn fibrils derived from LBD tissue to seed accumulation of Asyn fibrils in cell culture and mouse models. We will utilize SSNMR, cryo-EM and cryo-electron tomography to analyze Asyn fibril structure in these model systems, which will guide future studies of disease mechanisms, PET ligand development and therapeutic development.