ALS/FTD mutant C9orf72-induced genetic and nuclear pathology in iPS cell models

ALS/FTD 突变体 C9orf72 在 iPS 细胞模型中诱导遗传和核病理学

• 批准号: 8913279
• 负责人: Jeffrey D Rothstein
• 金额: $ 35.44万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8913279
• 关键词: 9p21; Address; Antisense Oligonucleotide Therapy; Antisense Oligonucleotides; Astrocytes; Autopsy; Binding; Biological Markers; C9ORF72; Cell Line; Cell Nucleus; Cell model; Cells; Chromosomes; Code; DNA-Binding Proteins; Data; Development; Disease; Disease model; Down-Regulation; Drug effect disorder; Event; Fibroblasts; Functional disorder; Future; Gene Expression; Genes; Genetic; Genetic study; Genomics; Human; Human Cell Line; Laboratories; Microarray Analysis; Modeling; Molecular; Motor Neurons; Mutation; Nerve Degeneration; Neuroglia; Neurons; Nuclear; Nuclear RNA; Oligodendroglia; Oligonucleotide Probes; Open Reading Frames; Pathologic; Pathology; Pathway interactions; Patients; Phenotype; Process; RNA; RNA Editing; RNA Splicing; RNA-Binding Proteins; Small Interfering RNA; Specificity; Technology; Tissue Sample; Tissues; Toxic effect; Untranslated RNA; Validation; Work; brain tissue; cell type; crosslinking and immunoprecipitation sequencing; design; drug discovery; experience; genetic profiling; human disease; human tissue; induced pluripotent stem cell; loss of function; molecular phenotype; mutant; novel; novel therapeutics; protein expression; public health relevance; success; therapeutic development; tool

DESCRIPTION (provided by applicant): The overall aim of this proposal is to study the pathology of the expanded hexanucleotide repeats in the C9orf72 gene using human iPS differentiated neurons and glia cells. The expanded GGGGCC hexanucleotide repeat in the non-coding region of the C9orf72 gene on chromosome 9p21 has been discovered as the cause of approximately 30-50% of familial and up to 10% of sporadic ALS cases as well as 12% of familial FTD cases, making this the most common known genetic cause of ALS/FTD to date. Expanded repeats are highly unstable and potentially yield toxic RNAs that accumulate in the nucleus and are hypothesized to cause cellular dysfunction via aberrant DNA and protein binding. Preliminary data from our laboratory confirm (GGGGCC)n nuclear RNA foci, aberrant gene expression and nuclear retention of RNA binding proteins (RBPs) in C9orf72 patient derived fibroblasts and iPS differentiated neurons. We therefore propose to elaborate on these early findings by generating an extensive genetic profile of ALS/FTD human iPS neurons and glia cells through the use of microarray and validate whether the iPS changes are relevant by comparing to human autopsy C9orf72 brain tissues. Furthermore, we will use C9orf72 iPS cell lines to investigate the RNA toxicity/pathology thru the identification of aberrant accumulation and binding of RNA binding proteins. Finally we will determine if we can abrogate C9orf72 genomic toxicity and pathology with antisense oligonucleotides already designed and validated in our laboratory. The likelihood of success will be greatly enhanced through a collaborative working relationship, the availability and experience of using iPS cells and human tissues. The extensive use of iPS cells to model disease, to cross correlate with human tissues and their use to validate ameliorative antisense therapy provides an important and possibly new direction for understanding disease pathophysiology and therapeutics development. !

描述(申请人提供)：这项建议的总体目标是利用人的iPS分化的神经元和神经胶质细胞来研究C9orf72基因中扩展的六核苷酸重复的病理学。染色体9p21上C9orf72基因非编码区扩增的GGGGCC六核苷酸重复序列被发现是大约30%-50%的家族性ALS病例和多达10%的散发性ALS病例以及12%的家族性FTD病例的原因，使其成为迄今已知的最常见的ALS/FTD遗传原因。扩展的重复序列是高度不稳定的，可能会产生有毒的RNA，积累在细胞核中，并被假设通过异常的DNA和蛋白质结合导致细胞功能障碍。我们实验室的初步数据证实，在C9orf72患者来源的成纤维细胞和iPS分化的神经元中，存在核RNA灶(GGGGCC)、RNA结合蛋白(RBPs)的异常基因表达和核保留。因此，我们建议通过使用微阵列生成ALS/FTD人类iPS神经元和神经胶质细胞的广泛遗传图谱来详细说明这些早期发现，并通过与人类尸检C9orf72脑组织进行比较来验证iPS变化是否相关。此外，我们将使用C9orf72 iPS细胞株，通过鉴定RNA结合蛋白的异常积累和结合来研究RNA的毒性/病理学。最后，我们将确定是否可以用我们实验室已经设计和验证的反义寡核苷酸来消除C9orf72的基因组毒性和病理学。通过合作工作关系、使用iPS细胞和人体组织的可用性和经验，成功的可能性将大大提高。IPS细胞被广泛用于模拟疾病、与人体组织相互关联以及用于验证改进性反义治疗，为理解疾病的病理生理学和治疗发展提供了一个重要的、可能的新方向。好了！