权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Developing resources to alleviate muscle atrophy in FSHD by genome engineering

通过基因组工程开发资源缓解 FSHD 肌肉萎缩

基本信息

批准号：
8532067
负责人：
Brian P. Chadwick
金额：
$ 17.16万
依托单位：
FLORIDA STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-01 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8532067
关键词：
10q 4q35 Affect Alleles Atrophic Cell Transplantation Cells Chromatin Chromosomes Contracts Custom D4Z4 DNA DNA Sequence DNA-Binding Proteins Data Degenerative Disorder Development Disease Disease model Distal Duchenne muscular dystrophy Engineering Enzymes Excision Face Facioscapulohumeral Muscular Dystrophy Foundations General Population Generations Genetic Genetic Transcription Genome Genome engineering Goals Haplotypes Health Heterochromatin Homeobox Human Human Chromosomes Individual Inherited Life Live Birth Methods Molecular Models Muscle Muscular Atrophy Muscular Dystrophies Mutation Myotonic Dystrophy Open Reading Frames Patients Poly A Polyadenylation Reporting Resources Sequence Homologs Sequence-Specific DNA Binding Protein Shoulder Signal Transduction Site Skeletal Muscle Stem cells Symptoms Technology Testing Therapeutic Intervention Transcript Translations Upper arm Variant Zinc Fingers base cell type design effective therapy gain of function induced pluripotent stem cell innovation molecular modeling muscle degeneration novel nuclease repaired telomere transcription factor

项目摘要

DESCRIPTION (provided by applicant): Facioscapulohumeral muscular dystrophy (FSHD) is the third most common inherited form of muscular dystrophy after Duchenne and Myotonic dystrophy, affecting 1 in 20,000 live births. FSHD is primarily characterized by progressive weakness and atrophy of skeletal muscle of the face, shoulders and upper arms that typically manifest in a patients second or third decade of life. Currently there is no cure or effective treatment for this disease. FSHD is an autosomal dominant disorder and in almost all cases the genetic basis for disease involves a contraction in the size of the macrosatellite repeat D4Z4 in the subtelomeric region of chromosome 4q. D4Z4 is composed of a tandem array of a 3.3kb sequence with between 10-150 repeat units defining a single allele in healthy individuals. Intriguingly, whereas reduction in the size of D4Z4 to fewer than 10 repeat units is associated with FSHD, contraction alone is not sufficient to cause disease. Indeed, D4Z4 alleles comparable in size to those observed in FSHD patients have been detected in unaffected individuals, whereas in others complete loss of the macrosatellite has been reported in the absence of FSHD symptoms. Instead, FSHD is exclusively associated with contraction of D4Z4 on specific variants of the subtelomeric region of human chromosome 4q. Recent studies have demonstrated that permissive chromosomes share a canonical polyadenylation signal that upon a contracted allele stabilizes transcripts originating from the most distal D4Z4 repeat unit resulting in a toxic gain of function in muscle. The objective of this proposal is to generate the resources necessary to nullify pathogenic D4Z4 alleles. Recent technological advances have made possible genome engineering in human cells through custom-built zinc finger nucleases (ZFN) and TAL effector nucleases (TALEN). Both approaches involve generation of sequence specific DNA binding proteins tethered to nuclease enzymes that induce double strand breaks at a desired sequence. Introduction of ZFNs and TALENs into cells along with a modified homologous sequence to the target site results in the introduction of desired changes into the endogenous locus as the supplied template is used to repair the damage. In Aim 1, we will develop ZFNs and TALENs directed to sequences proximal to the D4Z4 array. These will be used in combination with a repair template that introduces a telomere seeding sequence to remove the macrosatellite, resulting in a truncated chromosome lacking D4Z4 similar to that observed in unaffected individuals. In Aim 2, we will develop ZFNs and TALENs designed to sequences immediately distal to D4Z4 to convert the polyadenylation signal to a sequence found on non-permissive chromosomes. Generation of such resources will permit exploration of a novel therapy for individuals afflicted with this debilitating progressive disorder.

描述（由申请人提供）：面肩肱型肌营养不良症（FSHD）是继杜氏肌营养不良症和强直性肌营养不良症之后第三种最常见的遗传性肌营养不良症，每20，000例活产婴儿中就有1例受影响。FSHD的主要特征在于面部、肩部和上臂的骨骼肌的进行性虚弱和萎缩，其通常在患者生命的第二或第三个十年中显现。目前还没有治愈或有效治疗这种疾病。FSHD是一种常染色体显性遗传疾病，几乎所有病例的遗传基础都涉及染色体4 q亚端粒区大卫星重复序列D4 Z4的大小收缩。D4 Z4由3.3kb序列的串联阵列组成，具有在健康个体中定义单个等位基因的10-150个重复单元。有趣的是，尽管D4 Z4的大小减少到少于10个重复单位与FSHD相关，但单独收缩不足以引起疾病。事实上，在未受影响的个体中检测到与FSHD患者中观察到的大小相当的D4 Z4等位基因，而在其他人中，在没有FSHD症状的情况下报告了宏卫星的完全丢失。相反，FSHD仅与D4 Z4在人类染色体4 q的亚端粒区域的特定变体上的收缩相关。最近的研究表明，允许染色体共享一个典型的多聚腺苷酸化信号，其在收缩的等位基因稳定来自最远端D4 Z4重复单元的转录本，导致肌肉中的毒性功能获得。该提案的目的是产生使致病性D4 Z4等位基因无效所需的资源。最近的技术进步使得通过定制的锌指核酸酶（ZFN）和TAL效应物核酸酶（TALEN）在人类细胞中进行基因组工程成为可能。这两种方法都涉及产生与核酸酶连接的序列特异性DNA结合蛋白，所述核酸酶在所需序列处诱导双链断裂。将ZFN和TALEN与修饰的靶位点同源序列一起沿着引入细胞导致将所需的变化引入内源基因座，因为所提供的模板用于修复损伤。在目标1中，我们将开发针对D4 Z4阵列近端序列的ZFN和TALEN。这些将与修复模板结合使用，该修复模板引入端粒播种序列以去除大卫星，从而产生与未受影响的个体中观察到的类似的缺少D4 Z4的截短染色体。在目标2中，我们将开发ZFN和TALEN，其被设计为紧邻D4 Z4远端的序列，以将聚腺苷酸化信号转化为在非允许染色体上发现的序列。这些资源的产生将允许探索一种新的治疗方法，用于患有这种使人衰弱的进行性疾病的个体。