Modulation of Jagged1/Pitpna in DMD as a means of therapy

DMD 中 Jagged1/Pitpna 的调节作为治疗手段

• 批准号: 10447111
• 负责人: LOUIS M KUNKEL
• 金额: $ 38.55万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10447111
• 关键词: Affect; Bioinformatics; Blood flow; Canis familiaris; Cardiopulmonary; Cell Line; Cell model; Cessation of life; Clinical Research; Complement; Control Animal; DNA Sequence Alteration; Data; Disease; Disease Progression; Down-Regulation; Duchenne muscular dystrophy; Dystrophin; Evaluation; Exhibits; Failure; Fibrosis; Gene Expression Profiling; Generations; Genes; Genetic; Genetic study; Goals; Grant; Human; In Vitro; Injections; Investigation; Life; Longevity; Messenger RNA; Modeling; Mus; Muscle; Muscle Cells; Muscle function; Muscular Atrophy; Muscular Dystrophies; Myoblasts; Outcome; PTEN gene; Pathology; Pathway interactions; Patient-Focused Outcomes; Patients; Performance; Pharmacology; Phenotype; Phosphatidylinositol Transfer Protein; Predisposition; Proteins; Proto-Oncogene Proteins c-akt; Reading Frames; Replacement Therapy; Research; Role; Sarcolemma; Signal Pathway; Signal Transduction; Skeletal Muscle; Swimming; Symptoms; Terminator Codon; Testing; Therapeutic; Transgenic Organisms; Translations; Viral; Zebrafish; adeno-associated viral vector; base; comparative; disease-causing mutation; exon skipping; experimental study; follow-up; heart function; improved; in vivo; inhibitor; inhibitor therapy; knock-down; mini-dystrophin; mouse model; muscle degeneration; muscular structure; overexpression; premature; prevent; restoration; targeted treatment; therapeutic target; treatment strategy

PROJECT SUMMARY/ABSTRACT Duchenne Muscular Dystrophy (DMD) is a degenerative muscle wasting disease caused by mutations in the dystrophin gene. The absence of dystrophin protein at the muscle sarcolemma results in increased muscle susceptibility to contraction-induced damage as well as dysregulation of secondary signaling pathways, many of which remain poorly understood. Despite advances in treatment strategies under investigation aimed at restoration of dystrophin expression including viral delivery of mini-dystrophin, read-through of translation stop codons, and exon skipping to restore the reading frame, there is no cure for DMD, and the identification of therapies that improve pathology independent of dystrophin would be of significant value to patients. In this vain, genetic modifiers of DMD are emerging as potential therapeutic targets. Our lab recently identified Jagged1 and Pitpna as genetic modifiers of DMD pathology in a Golden Retriever Muscular Dystrophy (GRMD) dog colony in which two exceptional “escaper” dogs exhibited a drastically milder phenotype than typical GRMD dogs despite being dystrophin-deficient. Normally, GRMD dogs show a severe phenotype similar to human DMD including early progressive muscle degeneration, fibrosis, and premature death often within the first 2 years of life due to cardiopulmonary failure. Gene expression analyses of the escaper dogs compared to severely affected GRMD dogs and control animals revealed that Jagged1 overexpression (OE) and decreased Pitpna expression were hallmarks of the mild phenotype, which including maintained ambulation and normal lifespan. In subsequent genetic studies of dystrophin-deficient zebrafish, we demonstrated that modulation of Jagged1 and Pitpna prevents manifestation of the dystrophic muscle phenotype, increases long-term survival, and improves swim performance. In primary myoblasts derived from normal and DMD patients, we also showed that Jagged1 overexpression and Pitpna inhibition impact AKT/PTEN signaling and improve myoblast fusion. Given this positive data, we will now extend our research of Jagged1 and Pitpna modulation into the well-characterized mdx5cv mouse model of DMD. We will also investigate genetic and pharmacological inhibition of PDE10A, which we have shown to elicit decreased Pitpna expression and improve dystrophic pathology in dystrophin-deficient zebrafish. The long-term goal in this project is to assess and validate the therapeutic potential of the genetic modifiers Jagged1 and Pitpna to ameliorate DMD pathology, and to identify a viable pharmacologic modulator to advance into clinical studies. To accomplish this, we will implement experiments in accordance to the following three specific aims: 1) Determine the effect of reduced Pitpna expression on dystrophic pathology in mdx5cv mice, 2) Characterize the functional role of Jagged1 overexpression in mdx5cv mice, and 3) Assess the therapeutic potential of PDE10A inhibition on dystrophic pathology in primary human muscle cells, zebrafish, and mouse models of DMD.

项目概要/摘要 杜氏肌营养不良症 (DMD) 是一种由基因突变引起的退行性肌肉萎缩疾病 肌营养不良蛋白基因。肌肉肌膜处缺乏抗肌营养不良蛋白导致肌肉增加 对收缩引起的损伤以及次级信号通路失调的敏感性，许多 其中仍然知之甚少。尽管正在研究的治疗策略取得了进展，旨在 恢复肌营养不良蛋白表达，包括病毒递送微型肌营养不良蛋白、通读翻译终止点 密码子和外显子跳跃来恢复阅读框架，DMD 无法治愈，并且识别 独立于肌营养不良蛋白改善病理学的疗法对患者具有重要价值。在这个 然而，DMD 的基因修饰剂正在成为潜在的治疗靶点。我们的实验室最近发现 Jagged1 和 Pitpna 作为金毛寻回犬肌营养不良症 DMD 病理学的遗传修饰剂 （GRMD）狗群，其中两只特殊的“逃亡者”狗表现出比其他狗温和得多的表型 典型的 GRMD 狗，尽管缺乏肌营养不良蛋白。通常，GRMD 狗表现出严重的表型 与人类 DMD 相似，包括早期进行性肌肉退化、纤维化和过早死亡 出生后 2 年内因心肺衰竭。逃亡犬的基因表达分析 与严重受影响的 GRMD 狗和对照动物相比，发现 Jagged1 过度表达 (OE) Pitpna 表达减少是轻度表型的标志，其中包括维持 行走能力和正常寿命。在随后的抗肌营养不良蛋白缺陷斑马鱼的遗传学研究中，我们 证明 Jagged1 和 Pitpna 的调节可预防营养不良性肌肉的表现 表型，增加长期存活率，并提高游泳表现。在原代成肌细胞中 正常人和 DMD 患者，我们还表明 Jagged1 过表达和 Pitpna 抑制的影响 AKT/PTEN 信号传导并改善成肌细胞融合。鉴于这些积极的数据，我们现在将扩展我们的研究 Jagged1 和 Pitpna 调制到特征良好的 mdx5cv DMD 小鼠模型中。我们还将 研究 PDE10A 的遗传和药理学抑制，我们已证明 PDE10A 可以引起 Pitpna 降低 表达并改善肌营养不良蛋白缺陷斑马鱼的营养不良病理学。这方面的长期目标 该项目旨在评估和验证基因修饰剂 Jagged1 和 Pitpna 的治疗潜力 改善 DMD 病理学，并确定可行的药理调节剂以推进临床研究。 为此，我们将根据以下三个具体目标进行实验：1） 确定 Pitpna 表达减少对 mdx5cv 小鼠营养不良病理学的影响，2) 表征 Jagged1 过表达在 mdx5cv 小鼠中的功能作用，以及 3) 评估 PDE10A 的治疗潜力 抑制原代人类肌肉细胞、斑马鱼和 DMD 小鼠模型的营养不良病理。