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.

项目概要 就直接医疗和社会护理成本而言，痴呆症具有重大的社会和经济影响。 身体痴呆（LBD）是痴呆最常见的原因之一，包括帕金森病 大约 30% 的阿尔茨海默病 (AD) 患者患有痴呆 (PDD) 和路易体痴呆 (DLB)。 患有 LBD 的人也会比单独的 AD 更快速、更严重地导致认知能力下降。 大量尸检研究表明，大脑中存在一种名为 α-突触核蛋白 (α-syn) 的蛋白质异常沉积。 Braak 等人表明，PDD 中 α-syn 病理学以典型的方式传播，并且运动的发生 症状随着黑质 (SN) 多巴胺能神经元的丧失而出现，约 80% 的患者最终 皮质中 α-syn 病理学进展为 PDD。 简而言之，致病性 α-syn 的传播是主要因素。 最近的研究支持致病性 α-syn 可能表现的观点。 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。 重组α-syn聚集体（一种菌株）在循环聚集后可以转化为另一种菌株，并且这两种 菌株在用蛋白酶 K 消化后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）脑匀浆显着促进 α-syn 病理学 与 PD 匀浆相比，MSA 和 PD 具有不同的 α-syn 菌株。 在神经系统疾病的体征、发病时间、神经系统形态等方面观察到了具体差异。 大脑α-syn沉积物和诱导聚集体的构象特性此外，不同菌株。 针对大脑内不同的细胞群和细胞类型，概括了选择性靶向 为了研究 α-突触核蛋白病中观察到的 α-突触核蛋白菌株在 PDD 进展中的作用，我们收集了 对来自临床特征良好的患者的脑脊液（CSF）样本进行了纵向跟踪。 双盲方式，我们扩增了以患者脑脊液（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）。 表征了这些 α-syn 聚集体来自具有明确的神经毒性、生化和 然而，考虑到错误折叠的 α-syn 聚集体表现出异质应变特性。 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， 幸运的是，有必要建立一种指纹识别方法来定义来自 LBD 患者的这些 α-syn 菌株。 分子微阵列（SMM）筛选提供了一个平台来结合不同面板的亲和力特征 通过比较和分析这些“大数据”，将数以万计的小分子转化为某些蛋白质目标。 将这些亲和探针收缩为指纹可以帮助我们识别和区分 α-syn 菌株。 使用我们建立的大环化合物“rapafucin”库生成3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表现。 3D-SMM 中的结合模式（指纹签名）我们建议将 3D-SMM 作为化学物质应用。 指纹识别平台，用于识别和区分患者的 α-syn 菌株并开发相关疗法 战略。