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成本方面具有重大的社会和经济影响。路易 身体痴呆（LBD）是痴呆症最常见原因之一，包括帕金森氏病 痴呆症（PDD）和痴呆症具有Lewy身体（DLB）。大约30％的阿尔茨海默氏病（AD）患者 与单独的AD相比，LBD还会遭受LBD的快速和严重认知下降。 LBD是关联的 大脑中称为α-突触核蛋白（α-syn）的蛋白质异常沉积。大量验尸研究 Braak等。证明α-Syn病理以PDD的刻板印象传播，电动机的发作 症状是在黑质（SN）中丧失多巴胺能神经元的丧失，最终约有80％的患者 在皮质中伴有α-Syn病理学的PDD。简而言之，致病α-Syn的传播充当主要 LBD中认知障碍（CI）的驱动力。最近的研究支持致病性α-syn行为的观念 以类似于表现出不同生化和病理表型的菌株特异性prions的方式。甚至 重组α-Syn聚集体（一个菌株）可以在循环聚集后转换为另一个菌株，这两个可以转换为 菌株暴露于用蛋白酶K消化后的不同神经毒性和免疫印迹模式。注射时 脑内MSA（多个系统萎缩）脑匀浆明显促进α-Syn病理 与PD匀浆相比，扩散表明MSA和PD具有不同的α-Syn菌株。拉紧- 在神经系统疾病的迹象，疾病发作的时间，形态学的形态中观察到了具体差异 大脑α-syn沉积物和诱导聚集体的构象特性。而且，不同的应变 靶向不同的细胞种群和大脑内的细胞类型，概括了选择性靶向 观察到α-核核病。为了研究α-Syn菌株在PDD进程中的作用，我们收集了 临床表征良好的患者的脑脊液（CSF）样品纵向遵循。在 双盲方式，我们扩大了由患者CSF（包含α-Syn种子）模板的α-Syn聚集体， 使用公认的应变扩增技术-PMCA（蛋白质错误折叠环状扩增）。我们 表征了这些α-SYN聚集体来自具有良好神经毒性，生化和 生物物理测定。但是，鉴于错误折叠的α-Syn聚集体暴露了异质应变特性 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，它是 建立一种指纹方法来定义LBD患者的这些α-SYN菌株所必需的。幸运的是，很小 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曲线 成千上万的小分子到某些蛋白质靶标。通过比较和 将这些亲和力问题作为指纹收缩可以帮助我们识别和区分α-Syn菌株。我们有 使用我们已建立的大环化合物“ Rapafucin”库生成了3维SMM，并进行了筛选 针对体外衍生的不同α-Syn菌株的SMM。我们发现这些菌株暴露了不同 3D-SMM（指纹标志）中的结合模式。我们建议将3D-SMM应用于化学 指纹识别平台以识别和区分患者的α-Syn菌株并开发相关疗法 战略。