Probing the cellular Mechanisms of Transthyretin Cytotoxicity

甲状腺素运载蛋白细胞毒性的细胞机制探讨

• 批准号: 7980179
• 负责人: Natalia Reixach
• 金额: $ 38.93万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7980179
• 关键词: Abbreviations; Accounting; Affect; Affinity; Aging; Alzheimer' s Disease; Amyloid; Amyloid Fibrils; Amyloidosis; Animal Model; Antioxidants; Apoptotic; Biochemical; Cardiomyopathies; Cell Death; Cell Line; Cells; Cellular Assay; Cerebrospinal Fluid; Cessation of life; Data; Deposition; Disease; Dissociation; Evaluation; Event; Extracellular Matrix; Familial Amyloid Neuropathies; Family; Functional disorder; Generations; Glycosaminoglycans; Goals; HSF1; Hand; Human Pathology; Kinetics; Knowledge; Lead; Light; Mass Spectrum Analysis; Mediating; Modeling; Molecular; Mutate; Organ; Pathogenesis; Pathway interactions; Plants; Prealbumin; Predisposition; Process; Proteins; Proteomics; Reactive Oxygen Species; Receptor Cell; Resveratrol; Retinol Binding Proteins; Risk Factors; Role; Secondary Protein Structure; Specificity; System; Testing; Therapeutic Agents; Therapeutic Intervention; Thyroxine; Time; Tissues; Transgenic Model; Variant; Work; age related; analog; analytical ultracentrifugation; base; cell injury; conformer; cytotoxic; cytotoxicity; extracellular; heat-shock factor 1; in vivo; inhibitor/antagonist; insight; molecular marker; mutant; novel therapeutics; oxidation; polyphenol; prevent; protein misfolding; public health relevance; receptor; research study; response; small molecule; tissue culture; tool

DESCRIPTION (provided by applicant): The transthyretin (TTR) amyloidoses are fatal disorders of secondary protein structure characterized by extracellular deposition of aggregates, including amyloid fibrils, derived from the secreted transthyretin protein. Aging is a risk factor for both the senile systemic (normal sequence) and familial (mutated sequence) forms of these disorders. At this time, no pharmacologic treatment is available. Recent data suggest that oligomers generated early in the course of TTR misfolding may be critical in the in vivo pathogenesis. We have developed the cell and molecular biologic tools to precisely analyze the molecular mechanisms underlying cell damage. Such knowledge is critical to the long-term goal of finding new therapeutic strategies to treat these diseases. Our working hypotheses are 1) TTR cytotoxicity in tissue culture reflects the human pathology regarding the susceptibility of target tissues and the amyloidogenicity of different TTR variants. 2) There is a relationship between the age-related increase in protein oxidation and the onset of TTR amyloidoses. 3) The interaction between the appropiate TTR conformer and target cells results in the generation of reactive oxygen species which, in turn, trigger an apoptotic cascade leading to cell death. 4) There is a cell receptor mediating the cytotoxic response. 5) The sequence of events leading to cell death can be prevented with compounds structurally derived from the plant polyphenol resveratrol and other antioxidants. These hypotheses will be tested in our 3 specific aims. In aim 1 we will establish tissue culture systems that mimic the human pathology in terms of tissue specificity and TTR variant. With these models in hand we will precisely identify the responsible TTR cytotoxic species, and define the role of protein oxidation and of the extracellular matrix on TTR-induced cytotoxicity. In aim 2 we will characterize and identify the cell receptor involved in the cytotoxic response using biochemical, immunochemical and proteomic mass spectrometry. In aim 3 we will use the cell system to screen compounds that inhibit TTR-induced cytotoxicity to find substances that may prevent TTR amyloidosis in vivo. We will define the mechanism of inhibition of TTR- induced cytotoxicity by the active small molecules which might in turn, be applicable to other amyloidoses. The insights gained from these studies will provide lead compounds and potential new pathways for therapeutic intervention. PUBLIC HEALTH RELEVANCE: The transthyretin amyloidoses are a set of age-related, mostly untreatable, poorly understood and ultimately fatal diseases affecting millions in the US and many more worldwide. We will create relevant tissue culture models to define the cellular mechanism of transthyretin damage and to find new compounds that can be used to treat these disorders. The findings will open new avenues for therapeutic intervention such as blockade of the formation of transthyretin toxic species, or inhibition of the cellular pathways that lead to cell death.

描述（由申请人提供）：甲状腺转蛋白（TTR）淀粉样病变是二级蛋白结构的致命疾病，其特征是细胞外沉积聚集体，包括淀粉样原纤维，来源于分泌的甲状腺转蛋白。衰老是这些疾病的老年性系统性（正常序列）和家族性（突变序列）形式的一个危险因素。此时，没有药物治疗可用。最近的数据表明，在TTR错误折叠过程早期产生的低聚物可能在体内发病机制中起关键作用。我们已经开发了细胞和分子生物学工具来精确分析细胞损伤的分子机制。这些知识对于寻找治疗这些疾病的新治疗策略的长期目标至关重要。我们的工作假设是：1)TTR在组织培养中的细胞毒性反映了人类病理对靶组织的易感性和不同TTR变体的淀粉样变性。2) TTR淀粉样变的发病与年龄相关的蛋白氧化增加有关。3)适当的TTR构象与靶细胞之间的相互作用导致活性氧的产生，进而引发凋亡级联导致细胞死亡。4)有一种细胞受体介导细胞毒性反应。5)从植物多酚白藜芦醇和其他抗氧化剂结构上衍生的化合物可以预防导致细胞死亡的一系列事件。这些假设将在我们的三个具体目标中得到检验。在目标1中，我们将建立在组织特异性和TTR变异方面模仿人类病理的组织培养系统。有了这些模型，我们将精确地确定负责的TTR细胞毒性物种，并确定蛋白质氧化和细胞外基质在TTR诱导的细胞毒性中的作用。在目标2中，我们将使用生化，免疫化学和蛋白质组质谱法表征和鉴定参与细胞毒性反应的细胞受体。在目标3中，我们将使用细胞系统筛选抑制TTR诱导的细胞毒性的化合物，以发现可能在体内阻止TTR淀粉样变的物质。我们将明确活性小分子抑制TTR诱导的细胞毒性的机制，这可能反过来适用于其他淀粉样变性。从这些研究中获得的见解将为治疗干预提供先导化合物和潜在的新途径。