ALS therapies and genomics for mutant TDP-43 and TLS/FUS

描述（申请人提供）：本申请涉及两个广泛的挑战领域（15）转化科学，包括特定的挑战主题15-NS-104（早期治疗开发）和15-NS-103（“概念验证”的演示）和广泛的挑战领域（08）基因组学，特定挑战主题03-NS-101（跨疾病研究，以确定低发病率和遗传复杂性高发病率疾病的共同机制）。肌萎缩侧索硬化症（ALS）是一种成人发病的神经退行性疾病，其中上下运动神经元的过早丧失会导致致命的瘫痪，典型的病程为一到五年。虽然在过去的15年里，大多数了解ALS发病机制的努力都集中在由普遍表达的超氧化物歧化酶突变引起的疾病上，（占所有ALS病例的约2%），在过去的14个月中，通过发现两种惊人相似的DNA/RNA结合蛋白的突变，TAR DNA结合蛋白（TDP-43）和融合肉瘤（FUS/TLS）也是ALS的主要原因。在正常情况下，这两种蛋白质都与RNA转录和选择性RNA剪接有关（FUS/TLS是剪接体的组成部分）。这提出了一种挑衅性的可能性，即RNA代谢的改变，特别是选择性RNA剪接的错误，在ALS发病机制中起着至关重要的作用。随着进一步认识到含有TDP-43的细胞质内含物存在于绝大多数ALS散发性患者和包括额颞痴呆（FTD）在内的几种其他神经退行性疾病中，TDP-43中的错误可能是理解疾病机制的核心，不仅是ALS，还有其他神经退行性疾病。在此，我们提出1）将联合收割机转基因和基因靶向方法结合起来，开发TDP-43和FUS/TLS介导的ALS的新小鼠模型，以及2）利用新开发的与高通量测序相关的方法来确定TDP-43和FUS/TLS在RNA代谢调节中的作用，作为了解其改变如何成为发病机制的基础并鉴定治疗开发的新靶点的手段。 公共卫生相关性：在过去的14个月里，在理解致命的运动神经元疾病肌萎缩性侧索硬化症中出了什么问题方面的范式转变已经开始，因为发现一对RNA结合蛋白（TDP-43和FUS/TLS）的突变是该疾病的主要原因。通过构建和分析这些遗传性ALS的遗传模拟小鼠，将确定这些突变如何导致运动神经元过早死亡。新的、非常高通量的DNA/RNA测序方法将用于鉴定其表达因突变而改变的基因，从而鉴定新的靶点和方法，以设计成功的ALS治疗方法。

DESCRIPTION (provided by applicant): This application addresses two Broad Challenge areas (15) Translational Science, including specific Challenge Topics 15-NS-104 (Early-stage therapy development) and 15-NS-103 (Demonstration of "proof-of-concept") and Broad Challenge area (08) Genomics, specific Challenge Topic 03-NS-101 (Cross-disease research to identify mechanisms common to Mendelian disorders of low incidence and genetically complex, high incidence disorders). Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder in which premature loss of upper and lower motor neurons leads to fatal paralysis with a typical disease course of one to five years. While most efforts to understand ALS pathogenesis over the last 15 years have focused on disease caused by mutation in the ubiquitously expressed superoxide dismutase (representing ~2% of all instances of ALS), during the past 14 months, a paradigm shift in understanding ALS has been initiated by discovery that mutations in two strikingly similar DNA/RNA binding proteins, TAR DNA-binding protein (TDP-43) and Fused in sarcoma (FUS/TLS), are also primary causes of ALS. In the normal context, both proteins have been linked to RNA transcription and alternative RNA splicing (FUS/TLS is an integral component of the spliceosome). This has raised the provocative possibility that alterations of RNA metabolism, especially errors of alternative RNA splicing, play a crucial role in ALS pathogenesis. With the further realization that TDP-43-containing cytoplasmic inclusions are found in the vast majority of ALS sporadic patients and in several other neurodegenerative diseases including Frontal Temporal Dementia (FTD), errors in TDP-43 may be central to understanding disease mechanism not just in ALS by also other neurodegenerative disorders. Here, we propose 1) to combine transgenic and gene- targeting approaches to develop new mouse models for TDP-43 and FUS/TLS mediated ALS and 2) to utilize newly developed methodologies linked to high-throughput sequencing to determine the role of TDP-43 and FUS/TLS in the regulation of RNA metabolism as a means to understand how their alterations underlie pathogenesis and to identify new targets for therapy development. PUBLIC HEALTH RELEVANCE: Over the last 14 months, a paradigm shift in understanding what goes wrong in the fatal motor neuron disease Amyotrophic Lateral Sclerosis has been initiated by the discoveries that mutations in a pair of RNA binding proteins (TDP-43 and FUS/TLS) are primary causes of the disease. Through construction and analysis of mice that are genetic mimics of these instances of inherited ALS, how these mutations cause premature death of the motor neurons will be identified. New, very high through put DNA/RNA sequencing methods will be used to identify genes whose expression is altered by the mutations, thereby identifying new targets and approaches for devising a successful therapy for ALS.