A MIR-486/DOCK3 SIGNALING AXIS MODULATES DYSTROPHIN-DEFICIENT PATHOLOGY

MIR-486/DOCK3 信号轴调节肌营养不良蛋白缺乏的病理学

• 批准号: 10404518
• 负责人: MATTHEW Scott ALEXANDER
• 金额: $ 32.51万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-17 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10404518
• 关键词: Affect; Age; Apoptosis; Arrhythmia; Base Pairing; Base Sequence; Binding; Biological Markers; Biopsy; Birth; Cardiac; Cell Death; Cell Line; Cell fusion; Cells; Cessation of life; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Data; Development; Disease; Disease Progression; Drug Compounding; Duchenne muscular dystrophy; Dystrophin; Evaluation; Exercise; Gene Expression; Genes; Genetic; Histology; Human; Impairment; Intramuscular; Knock-out; Knockout Mice; Laboratories; Link; Mediating; Membrane; Membrane Proteins; Messenger RNA; Methods; MicroRNAs; Molecular; Mus; Muscle; Muscle Cells; Muscle function; Muscular Atrophy; Myocardium; Myopathy; Outcome; PTEN gene; Pathogenesis; Pathologic; Pathology; Pathway interactions; Patients; Performance; Phenotype; Play; Protein Family; Proteins; Proto-Oncogene Proteins c-akt; RHOA gene; Regulation; Respiratory Failure; Role; Seeds; Serum; Severities; Severity of illness; Signal Pathway; Signal Transduction; Site; Skeletal Muscle; Stress; Symptoms; Testing; Therapeutic; Time; Viral; adeno-associated viral vector; boys; combinatorial; crosslinking and immunoprecipitation sequencing; emerging adult; engineered exosomes; exon skipping; exosome; experience; heart function; human disease; human model; improved; in vivo; loss of function mutation; male; mdx mouse; member; miRNA expression profiling; mouse model; notch protein; novel; novel strategies; novel therapeutics; overexpression; predictive marker; respiratory; skeletal; skeletal muscle wasting

PROJECT SUMMARY Duchenne muscular dystrophy (DMD) is an X-linked disorder that affects 1:5000 live male births in the US each year. DMD is caused by loss-of-function mutations in the DYSTROPHIN (DMD) gene that results in severe muscle wasting, loss of ambulation by age 12, cardiac arrhythmia, and death in early adulthood due to respiratory failure. While there are several novel strategies (such as exon-skipping compounds to create partially-functional Dystrophin protein) in clinical trials, none of them deal with the secondary signaling pathways and subsequent muscle pathogenesis that occurs in DMD. There is growing evidence that genetic modifiers of DMD have a significant impact on the outcomes and severity of DMD-related pathologies in patients. These DMD genetic modifiers have the potential to serve as novel therapeutic entry points for the treatment of diseases. Previously my laboratory identified miR-486 as a muscle-enriched microRNA that is decreased in expression in DMD patient skeletal muscle biopsies. In this proposal, we will test the hypothesis that miR-486 and its target (DOCK3) play a significant role in the regulation of skeletal muscle. We have strong data that demonstrates that miR-486 expression levels decrease and an mRNA target (DOCK3) increases corresponding with the degree of dystrophic pathology. In Aim 1, we will characterize the functional role of miR-486 on skeletal and heart muscles via characterizing muscle performance and cardiac remodeling in miR-486 knockout (KO) mice on normal and mdx5cv backgrounds. Aim 2 focuses on identifying the consequences of manipulating miR-486 and Dock3 expression on key downstream signaling pathways including PTEN/AKT and Rac1/RhoA using a novel Dock3 conditional muscle knockout mouse. Additionally, we will identify novel miR-486 target genes using Ago2-sequencing (CLIP-seq) strategy to identify in vivo skeletal muscle targets of miR-486. Lastly, Aim 3 focuses on using transient miR-486 overexpression strategies (Adeno-associated viral vectors and engineered exosomes) delivered intramuscularly and systemically to improve dystrophic symptoms at key developmental time points in mdx5cv mice. Our combinatorial use of DMD patient muscle cell lines, miR-486 KO and Dock3 conditional muscle KO mice, and experience evaluating genetic and drug compounds in mouse models of human diseases will allow us to rigorously evaluate the use of a microRNA (miR-486) overexpression as a novel therapy for the treatment of DMD.

项目概要 杜氏肌营养不良症 (DMD) 是一种 X 连锁疾病，影响美国每 1:5000 名活产男婴 年。 DMD 是由肌营养不良蛋白 (DMD) 基因功能丧失突变引起的，导致严重的 肌肉萎缩、12岁时丧失行走能力、心律失常以及成年早期因呼吸系统疾病死亡 失败。虽然有几种新颖的策略（例如外显子跳跃化合物来创建部分功能的 肌营养不良蛋白）在临床试验中，没有一个涉及次级信号通路和后续信号通路 DMD 中发生的肌肉发病机制。越来越多的证据表明 DMD 的基因修饰剂具有 对患者 DMD 相关病理的结果和严重程度产生重大影响。这些DMD遗传 修饰剂有可能成为疾病治疗的新治疗切入点。之前 我的实验室鉴定出 miR-486 是一种富含肌肉的 microRNA，在 DMD 患者中表达下降 骨骼肌活检。 在本提案中，我们将检验以下假设：miR-486 及其靶点 (DOCK3) 在 骨骼肌的调节。我们有强有力的数据表明 miR-486 表达水平 随着营养不良病理程度的增加，mRNA 靶标 (DOCK3) 会增加。在 目标 1，我们将通过表征来表征 miR-486 对骨骼肌和心肌的功能作用 正常和 mdx5cv 背景下 miR-486 敲除 (KO) 小鼠的肌肉性能和心脏重塑。 目标 2 侧重于确定操纵 miR-486 和 Dock3 表达对关键 使用新型 Dock3 条件肌肉的下游信号通路，包括 PTEN/AKT 和 Rac1/RhoA 淘汰赛小鼠。此外，我们将使用 Ago2 测序 (CLIP-seq) 鉴定新的 miR-486 靶基因 鉴定 miR-486 体内骨骼肌靶标的策略。最后，目标 3 重点关注使用瞬态 miR-486 肌内递送的过表达策略（腺相关病毒载体和工程外泌体） 并系统地改善 mdx5cv 小鼠关键发育时间点的营养不良症状。我们的 DMD患者肌肉细胞系、miR-486 KO和Dock3条件肌肉KO小鼠的组合使用， 在人类疾病小鼠模型中评估遗传和药物化合物的经验将允许 我们严格评估 microRNA (miR-486) 过度表达作为一种新疗法的用途 DMD 的治疗。

项目成果

Mechanics of dystrophin deficient skeletal muscles in very young mice and effects of age.

幼年小鼠肌营养不良蛋白缺陷骨骼肌的机制和年龄的影响。

• DOI: 10.1152/ajpcell.00155.2019
• 发表时间: 2021
• 期刊: American journal of physiology. Cell physiology
• 作者: Lopez,MichaelA;Bontiff,Sherina;Adeyeye,Mary;Shaibani,AzizI;Alexander,MatthewS;Wynd,Shari;Boriek,AladinM
• 通讯作者: Boriek,AladinM

The Interplay of Mitophagy and Inflammation in Duchenne Muscular Dystrophy.

• DOI: 10.3390/life11070648
• 发表时间: 2021-07-04
• 期刊: Life (Basel, Switzerland)
• 作者: Reid AL;Alexander MS
• 通讯作者: Alexander MS

DOCKopathies: A systematic review of the clinical pathologies associated with human DOCK pathogenic variants.

• DOI: 10.1002/humu.24398
• 发表时间: 2022-09
• 期刊: HUMAN MUTATION
• 影响因子: 3.9
• 作者: Samani, Adrienne;English, Katherine G.;Lopez, Michael A.;Birch, Camille L.;Brown, Donna M.;Kaur, Gurpreet;Worthey, Elizabeth A.;Alexander, Matthew S.
• 通讯作者: Alexander, Matthew S.

Extracellular serine and glycine are required for mouse and human skeletal muscle stem and progenitor cell function.

• DOI: 10.1016/j.molmet.2020.101106
• 发表时间: 2021-01
• 期刊: Molecular metabolism
• 影响因子: 8.1
• 作者: Gheller BJ;Blum JE;Lim EW;Handzlik MK;Hannah Fong EH;Ko AC;Khanna S;Gheller ME;Bender EL;Alexander MS;Stover PJ;Field MS;Cosgrove BD;Metallo CM;Thalacker-Mercer AE
• 通讯作者: Thalacker-Mercer AE

DOCK3-Associated Neurodevelopmental Disorder-Clinical Features and Molecular Basis.

• DOI: 10.3390/genes14101940
• 发表时间: 2023-10-14
• 期刊: GENES
• 影响因子: 3.5
• 作者: Alexander, Matthew S.;Velinov, Milen
• 通讯作者: Velinov, Milen