Evaluation of the role of RNA toxicity in SCA2 pathogenesis using genome editing in patient iPSCs

使用患者 iPSC 基因组编辑评估 RNA 毒性在 SCA2 发病机制中的作用

基本信息

批准号：
9803833
负责人：
Lisa M Ellerby
金额：
$ 50.58万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-07-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9803833
关键词：
Alleles Antibodies CAG repeat Cell model Cells Clustered Regularly Interspaced Short Palindromic Repeats Complex Data Development Disease Epitopes Evaluation Exposure to Fibroblasts Fractionation Future Generations Genes Goals Human Herpesvirus 4 Huntington Disease In Situ Lead Length MJD1 protein Machado-Joseph Disease Metabolism Microsatellite Repeats Modeling Mutation Neurodegenerative Disorders Neurons Neurotoxins Nuclear RNA Pathogenesis Pathology Pathway interactions Patients Phenotype Play Property Proteins Protocols documentation RNA RNA Processing RNA Splicing RNA-Binding Proteins Ribosomal RNA Role SCA2 protein Shapes Structure Techniques Testing Therapeutic Toxic effect Transcript Translating Type 2 Spinocerebellar Ataxia base experimental study genome editing homologous recombination induced pluripotent stem cell mutant neurotoxic neurotoxicity novel polyglutamine protein expression therapy development tool vector

项目摘要

Spinocerebellar ataxia type 2 (SCA2) is a devastating neurodegenerative disease caused by a CAG repeat expansion in the gene ataxin-2 (ATXN2). The CAG repeat is translated into a polyglutamine (polyQ) tract in the mutant ATXN2 protein that has neurotoxic properties. Current therapeutic efforts are focused at suppressing the expression of the mutant ATXN2 protein, or targeting downstream pathways of neurotoxicity. We and others have shown that mutant CAG/CTG repeat-containing transcripts also contribute to pathogeneses of several repeat diseases, including Huntington's disease (HD), Huntington's disease-like 2 (HDL2) and spinocerebellar ataxia type 3 and 8 (SCA3 and SCA8). Therefore, in addition to targeting the neurotoxicity of mutant proteins, a successful therapy may require targeting the mutant transcripts and/or pathways downstream of the mutant transcripts. Our preliminary evidence supports the idea that in SCA2, in addition to toxic ATXN2 protein, both sense ATXN2 RNA and a transcript antisense to ATXN2 (ATXN2-AS) containing an expanded CUG repeat, contribute to SCA2 pathogenesis. We therefore propose to use genome editing approach to modify normal and SCA2 iPSC line into novel, isogenic iPSC lines that model either protein or RNA-induced mechanisms of SCA2 pathogenesis. These lines will be used to (1) further test the novel hypothesis of SCA2 by which mutant ATXN2/ATXN2-AS transcripts disrupt RNA processing in SCA2, including RNA export, rRNA metabolism and splicing, and (2) determine the relative contribution of RNA neurotoxicity to SCA2 pathogenesis. The results obtained from this study will not only facilitate our understanding of the increasingly complex pathogenesis of SCA2, but also that of other CAG/CTG repeat diseases in which both bi-directionally encoded transcripts and protein contribute to pathology. The results will help guide future development of SCA2 therapy, as well as therapies of other microsatellite repeat expansion diseases.

脊椎小脑共济失调2型（SCA2）是由CAG重复引起的毁灭性神经退行性疾病基因共生2（ATXN2）的扩展。 CAG重复被翻译成多谷氨酰胺（Polyq）。具有神经毒性特性的突变ATXN2蛋白。当前的治疗努力集中在抑制突变ATXN2蛋白的表达，或针对神经毒性下游途径的表达。我们和其他人已经表明，突变的CAG/CTG重复转录本也有助于几个病原体重复疾病，包括亨廷顿氏病（HD），亨廷顿疾病样2（HDL2）和脊椎队共济失调3和8（SCA3和SCA8）。因此，除了靶向突变蛋白的神经毒性外，成功的治疗可能需要针对突变体下游的突变转录本和/或途径成绩单。我们的初步证据支持这样的观念：在SCA2中，除了有毒ATXN2蛋白外，两者都 sense atxn2 rnA和对atxn2（atxn2-as）的转录反义，其中包含扩展的cug重复，有助于SCA2发病机理。因此，我们建议使用基因组编辑方法来修改正常和 SCA2 IPSC线融入新型的等生IPSC线，该线模拟蛋白质或RNA诱导的SCA2机理发病。这些线将用于（1）进一步测试SCA2的新假设 ATXN2/ATXN2-AS转录本破坏SCA2中的RNA处理，包括RNA导出，RRNA代谢和剪接，（2）确定RNA神经毒性对SCA2发病机理的相对贡献。结果从这项研究获得的不仅会促进我们对日益复杂的发病机理的理解 SCA2，以及其他CAG/CTG重复疾病的SCA2，其中双向编码的转录本和蛋白质有助于病理学。结果将有助于指导SCA2疗法的未来发展，以及其他微卫星重复扩张疾病的疗法。