Stimulators of Neuronal Transthyretin (TTR) Transcription as Alzheimer's Therapy

神经元运甲状腺素蛋白 (TTR) 转录刺激剂用于治疗阿尔茨海默病

• 批准号: 8331502
• 负责人: Joel N Buxbaum
• 金额: $ 28.43万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-15 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8331502
• 关键词: Abbreviations; Acids; Age; Alzheimer' s Disease; Amyloid; Amyloidosis; Antibodies; Appearance; Behavioral; Biological Assay; Brain; Cells; Cerebrospinal Fluid; Characteristics; Complex; Coupled; Defense Mechanisms; Dimethyl Sulfoxide; Enhancers; Enzyme-Linked Immunosorbent Assay; Enzymes; Florida; Genes; Genetic Transcription; Goals; Heat-Shock Response; Hepatic; Hippocampus (Brain); Histone Deacetylase; Histone Deacetylase Inhibitor; Human; In Vitro; Institution; Knock-out; Laboratories; Learning; Ligands; Link; Luciferases; Messenger RNA; Molecular Bank; Mus; Neuraxis; Neurodegenerative Disorders; Neurons; Pathologic; Peripheral; Phenotype; Physiological; Pilot Projects; Prealbumin; Production; Prophylactic treatment; Protein Precursors; Proteins; Proteomics; Reporter; Reporter Genes; Resistance; Serum; Sodium Dodecyl Sulfate; Staining method; Stains; Techniques; Time; Toxic effect; Transgenic Mice; Transgenic Model; Vorinostat; chromatin immunoprecipitation; cytotoxicity; disease phenotype; gene therapy; hepatocyte nuclear factor; high throughput screening; in vivo; mind control; mouse model; novel; promoter; small molecule; tissue culture; transcription factor

DESCRIPTION (provided by applicant): We have shown that over-expression of a wild type human transthyretin (TTR) gene can suppress the neuropathologic and behavioral phenotypes of a well-studied validated transgenic model of human Alzheimer's disease (AD). We have also shown that TTR interacts with A2 in vitro to inhibit oligomer and fibril formation and tissue culture cytotoxicity and in vivo with TTR-A2 complexes being co-immunoprecipitable from APP23 and some human AD brains with anti-sera to either protein. We have also noted that 70% of human cortical neurons from AD brains stain with an anti-TTR antibody (compared with 10% of age matched control brains) as do almost all the hippocampal and cortical neurons in the APP23 mouse model. TTR mRNA (by qtPCR) is 10 times higher in primary hippocampal neurons from APP23 mice than from controls and the ratio is even greater in hippocampal lysates. These findings coupled with the observation that the pathologic changes appear to be accelerated when the APP23 strain is crossed onto a mouse TTR knockout background suggest that TTR may be a normal physiologic neuronal defense mechanism in neurodegenerative disease of the AD type. In contrast the suppression of the features of AD by TTR over-expression may be viewed as pharmacologic. Our proposal is directed at finding small molecules that will enhance TTR transcription and protein production in cells of neuronal origin. To that end we have developed reporter constructs in which the human TTR promoter drives a Gaussia luciferase reporter gene which is readily detectable in a high throughput screening mode available at our institution. Pilot studies have shown that the assay is robust, reproducible and stable under a variety of conditions. We propose to use the assay to serially screen a group of available molecular libraries (Maybridge, LOPAC, MLPCN, Scripps Florida) to identify the 25 most potent, specific least toxic enhancers of TTR transcription in neuronally defined cells. We will characterize their mechanism of action using cell biologic, proteomic and microarray techniques established in our laboratory and ultimately assay their capacity to suppress the neuropathologic and behavioral manifestations displayed by the APP23 transgenic mouse model of AD.

描述（由申请人提供）：我们已经表明野生型人甲状腺素运载蛋白（TTR）基因的过表达可以抑制充分研究的验证的人阿尔茨海默病（AD）转基因模型的神经病理学和行为表型。我们还表明，TTR与A2在体外相互作用，以抑制寡聚体和原纤维的形成和组织培养细胞毒性，并在体内与TTR-A2复合物是从APP 23和一些人AD脑与抗血清免疫共沉淀的任一蛋白质。我们还注意到，来自AD脑的70%的人皮质神经元用抗TTR抗体染色（与年龄匹配的对照脑的10%相比），APP 23小鼠模型中几乎所有的海马和皮质神经元也是如此。TTR mRNA（通过qtPCR）在来自APP 23小鼠的原代海马神经元中比来自对照的高10倍，并且该比率在海马裂解物中甚至更高。这些发现加上当APP 23菌株与小鼠TTR敲除背景杂交时病理变化似乎加速的观察结果表明，TTR可能是AD型神经退行性疾病中的正常生理神经元防御机制。相反，TTR过表达对AD特征的抑制可被视为药理学。我们的建议是针对寻找小分子，将提高TTR的转录和蛋白质的生产在神经元起源的细胞。为此，我们已经开发了报告构建体，其中人TTR启动子驱动Gaussia荧光素酶报告基因，其在我们机构可用的高通量筛选模式中容易检测。初步研究表明，该测定法在各种条件下具有耐用性、重现性和稳定性。我们建议使用该方法连续筛选一组可用的分子库（Maybridge，LOPAC，MLPCN，Scripps佛罗里达），以确定25个最有效的，特定的毒性最小的TTR转录增强剂在神经元定义的细胞。我们将使用我们实验室建立的细胞生物学、蛋白质组学和微阵列技术来表征它们的作用机制，并最终测定它们抑制APP 23转基因AD小鼠模型所表现出的神经病理和行为表现的能力。