Chemical Fingerprints of Cognitive Impairment-related alpha-Synuclein Strains using 3D Small Molecule Microarray and Related Therapeutic Application

使用 3D 小分子微阵列的认知障碍相关 α-突触核蛋白菌株的化学指纹及相关治疗应用

• 批准号: 10360139
• 负责人: Xiaobo Mao
• 金额: $ 45.03万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-15 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10360139
• 关键词: 3-Dimensional; Achievement; Affinity; Aging; Alzheimer' s Disease; Alzheimer' s disease patient; Autopsy; Big Data; Binding; Biochemical; Biological Assay; Biological Markers; Biophysics; Blinded; Brain; Cell model; Cells; Cerebrospinal Fluid; Cerebrum; Clinical; Cognition; Collaborations; Contracts; Data; Dementia; Dementia with Lewy Bodies; Deposition; Digestion; Dimensions; Double-Blind Method; Endopeptidase K; Enzyme-Linked Immunosorbent Assay; Exhibits; FK506; Fingerprint; Genetic; Goals; Heterogeneity; Impaired cognition; In Vitro; Lewy Bodies; Lewy Body Dementia; Libraries; Macrocyclic Compounds; Medical; Methods; Molecular; Molecular Conformation; Morphology; Multiple System Atrophy; Natural Products; Neurodegenerative Disorders; Neurologic Signs; Neurons; Oligopeptides; Onset of illness; Organic Synthesis; Parkinson' s Dementia; Pathogenicity; Pathologic; Pathology; Patients; Pattern; Periodicity; Pharmaceutical Chemistry; Phenotype; Population; Positioning Attribute; Prions; Property; Proteins; Recombinants; Role; S phase; Sampling; Seeds; Series; Sirolimus; Solid; Structure; Structure-Activity Relationship; Substantia nigra structure; Synthesis Chemistry; Techniques; Testing; Therapeutic; Therapeutic Agents; Time; alpha synuclein; analog; base; care costs; cell type; chemical fingerprinting; cognitive impairment in Parkinson' s; combinatorial chemistry; diagnostic tool; dopaminergic neuron; economic implication; efficacy evaluation; efficacy testing; expectation; experimental study; improved; in vitro activity; motor symptom; neurotoxicity; novel; novel diagnostics; prevent; prion-like; protein misfolding cyclic amplification; screening; small molecule; social; social implication; synucleinopathy; transmission process

PROJECT SUMMARY Dementia has significant social and economic implications in terms of direct medical and social care costs. Lewy bodies dementia (LBD) is one of the most common causes of dementia, including Parkinson's disease with dementia (PDD) and dementia with Lewy bodies (DLB). Approximately 30% of Alzheimer' disease (AD) patients also suffer from LBD resulting in a more rapid and severe cognition decline than AD alone. LBD is associated with abnormal deposits of a protein called α-synuclein (α-syn) in the brain. Substantial postmortem studies by Braak et al. show that α-syn pathology spreads in a stereotypical fashion in PDD, and the onset of motor symptoms occurs with loss of dopaminergic neurons in the substantia nigra (SN), and ~80% of patients finally progress to PDD with α-syn pathology in the cortex. In brief, the spread of pathogenic α-syn acts as a major driver of cognitive impairment (CI) in LBD. Recent studies support the notion that pathogenic α-syn may behave in a manner similar to strain-specific prions exhibiting distinct biochemical and pathologic phenotypes. Even recombinant α-syn aggregates (one strain) can convert to another strain after cyclic aggregation, and these two strains exhibited different neurotoxicity and immunoblot patterns after digestion with proteinase K. When injected intracerebrally, MSA (multiple system atrophy) brain homogenates remarkably promote α-syn pathology spreading compared to PD homogenates, suggesting that MSA and PD have different strains of α-syn. Strain- specific difference were observed in the signs of neurological illness, time to disease onset, morphology of cerebral α-syn deposits and the conformation properties of the induced aggregates. Moreover, different strains targeted distinct cellular populations and cell types within the brain, recapitulating the selective targeting observed among α-synucleinopathies. To investigate the role of α-syn strains in PDD progression, we collected cerebrospinal fluid (CSF) samples from the clinically well-characterized patients followed longitudinally. In a double-blinded manner, we amplified α-syn aggregates templated by CSF (containing α-syn seeds) of patients, using a well-established strain amplification technique--PMCA (protein misfolding cyclic amplification). We characterized these α-syn aggregates from patients with well-established neurotoxicity, biochemical, and biophysical assays. However, given that misfolded α-syn aggregates exhibit heterogeneous strain properties and dynamic conversion, particularly in the complicated interplay with environmental, genetic, aging factors, it is necessary to build a fingerprinting method to define these α-syn strains from LBD patients. Fortunately, small molecule microarray (SMM) screening has provided a platform to combine the affinity profile of a diverse panel of tens of thousands of small molecules to certain protein targets. Analysis of this “big data” by comparing and contracting these affinity probes as fingerprints can help us identify and differentiate α-syn strains. We have generated a 3-dimensional SMM using our established macrocyclic compound “rapafucin” library, and screened the SMM against in-vitro-derived distinct α‐syn strains. We have found that these strains exhibited different binding patterns in the 3D-SMM (fingerprinting signatures). We propose to apply the 3D-SMM as a chemical fingerprinting platform to identify and differentiate α‐syn strains from patients and develop related therapeutic strategy.

项目摘要 痴呆症在直接医疗和社会护理费用方面具有重大的社会和经济影响。路易 身体痴呆（LBD）是痴呆症的最常见原因之一，包括帕金森病， 痴呆（PDD）和路易体痴呆（DLB）。大约30%的阿尔茨海默病（AD）患者 也患有LBD，导致比单独的AD更快和更严重的认知下降。LBD相关 大脑中有一种叫做α-突触核蛋白（α-syn）的蛋白质异常沉积。大量尸检研究， Braak等人表明，α-syn病理学在PDD中以一种刻板的方式传播，并且运动神经元的发病与PDD的发病有关。 症状发生时，黑质（SN）中的多巴胺能神经元丢失，约80%的患者最终 进展为皮质中存在α-syn病理的PDD。简而言之，致病性α-syn的传播是一个主要的 LBD中认知障碍（CI）的驱动因素。最近的研究支持致病性α-syn可能表现为 以类似于菌株特异性朊病毒的方式表现出不同的生化和病理表型。甚至 重组α-syn聚集体（一种菌株）可以在循环聚集后转化为另一种菌株，并且这两种重组α-syn聚集体（一种菌株）可以在循环聚集后转化为另一种菌株。 用蛋白酶K消化后，菌株表现出不同的神经毒性和免疫印迹模式。注射时 在脑内，MSA（多系统萎缩）脑匀浆显著促进α-syn病理 与PD匀浆相比，MSA和PD具有不同的α-syn菌株。应变- 在神经系统疾病的体征、疾病发作时间、 脑α-syn沉积和诱导聚集体的构象性质。此外，不同的菌株 靶向脑内不同的细胞群体和细胞类型，概括了选择性靶向 在α-突触核蛋白病中观察到。为了研究α-syn菌株在PDD进展中的作用，我们收集了 对临床特征良好的患者的脑脊液（CSF）样本进行纵向随访。中 双盲方式，我们扩增了以患者CSF（含有α-syn种子）为模板的α-syn聚集体， 使用成熟的菌株扩增技术--PMCA（蛋白质错误折叠循环扩增）。我们 表征这些α-syn聚集体的患者具有明确的神经毒性，生化， 生物物理测定。然而，由于错误折叠的α-syn聚集体表现出不均匀的应变性质， 和动态转换，特别是在与环境，遗传，衰老因素的复杂相互作用中， 有必要建立指纹图谱方法来确定LBD患者的这些α-syn菌株。幸运的是，小 分子微阵列（SMM）筛选提供了一个平台，用于联合收割机组合不同面板的亲和特性 成千上万的小分子对特定的蛋白质目标。通过对比分析这一“大数据”， 将这些亲和探针作为指纹图谱，可以帮助我们鉴定和区分α-syn菌株。我们有 使用我们建立的大环化合物“rapafucin”文库产生三维SMM，并筛选 SMM针对体外衍生的不同α-syn菌株。我们发现这些菌株表现出不同的 3D-SMM中的绑定模式（指纹签名）。我们建议将3D-SMM用作化学品 指纹识别平台，用于识别和区分患者的α-syn菌株，并开发相关治疗药物 战略