MICA: Strategy for heart repair in Duchenne Muscular Dystrophy (DMD) using genetically engineered autologous Mesoangioblasts

MICA：利用基因工程自体中成血管细胞修复杜氏肌营养不良症 (DMD) 的心脏的策略

• 批准号: MR/X00466X/1
• 负责人: Francesco Galli
• 金额: $ 51.79万
• 依托单位: University of Manchester
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FX00466X%2F1
• 关键词: MICA; Strategy; heart; repair; Duchenne

The NHS official website indicates that around 70,000 people have Muscular Dystrophy (MD) or a related condition in the UK. Duchenne Muscular Dystrophy (DMD) is the most common and one of the most severe forms of MD. DMD is an inherited disease due to lack of expression of the dystrophin protein, causing a progressive waste of skeletal muscle which lead to progressive loss of ambulation, muscle weakness and muscle wasting in children. In the most severe cases, also heart muscle cells are affected, and this result in heart failure, the most common cause of premature death in DMD patients. All MDs still lack an effective therapy, even though administration of steroids (although with severe adverse effects), corrective surgery, assisted ventilation and drugs to support cardiac function have improved both the duration and the quality of life for patients.Many approaches based on either stem cells or gene therapy have entered clinical experimentation with the aim of repairing DMD heart, but none has reached significant clinical efficacy. The limited success of these therapies is due both to the difficulties of delivering the treatment to the heart and to the activation of the immune system of the patients towards donor cells or viral vectors. To overcome these limitations, I have developed a cell mediated exon skipping strategy, for skeletal muscle, currently being tested in a clinical trial in Manchester (EudrAct n.2019-001825-28), based upon transplantation of autologous Mesoangioblasts (Mabs), which are vessel-associated myogenic progenitors. Mabs are genetically corrected using a lentivector expressing a small nuclear RNA (snRNA) designed to induce exon-skipping on dystrophin exon 51 and to generate a short but functional version of dystrophin. The novelty of this approach is based on the ability of snRNA to diffuse along the regenerating muscle fibre and then correcting the resident dystrophic neighbouring nuclei inducing exon skipping. However, Mabs can be used only for the treatment of skeletal muscle as they cannot naturally differentiate into cardiac muscle: for this reason, the heart of the MD patients would remain untreated. In this project I will address this problem. It is well known that fibroblasts can be converted to cardiac muscle cells but, in comparison with Mabs, they cannot be delivered systematically, due to their inability to cross the vessel walls. This makes cardiac-converted fibroblasts good candidates to treat only localised lesions like myocardial infarct but not to treat progressive and widespread cardiomyopathies. My preliminary data show that Mabs can be converted to cardiomyocytes by transient over-expression of specific cardiac related genes in around 10-15 days. I therefore hypothesize that this time window will allow to deliver these Mabs-converted cardiomyocytes by cardiac catheterization in the whole heart. This project aims to evaluate the structural and functional amelioration of the heart in a mouse model of DMD after transplantation of Mabs-converted cardiomyocytes. Moreover, I will quantify the extent of the fusion of genetically corrected Mabs-converted cardiomyocytes with resident cardiomyocytes and subsequently the rate of correction, by exon-skipping, in the neighbouring nuclei quantifying the amount of dystrophin produced.The core element of this strategy is already in clinical experimentation for DMD skeletal muscle but not for the heart. This project will test the applicability of the cell mediated exon-skipping strategy to the heart. The succesful outcome of this project, demonstrating the efficacy of this strategy, will lead the way for a future clinical testing of this strategy in the heart of DMD patients and in heart diseases in general.

英国国家医疗服务体系官方网站显示，英国约有7万人患有肌肉萎缩症（MD）或相关疾病。杜氏肌营养不良症（DMD）是最常见和最严重的一种肌营养不良症。DMD是一种遗传性疾病，由于缺乏肌营养不良蛋白的表达，导致骨骼肌的进行性浪费，导致儿童逐渐丧失行走能力，肌肉无力和肌肉萎缩。在最严重的情况下，心肌细胞也会受到影响，导致心力衰竭，这是DMD患者过早死亡的最常见原因。所有MDs仍然缺乏有效的治疗方法，尽管使用类固醇（尽管有严重的不良反应）、矫正手术、辅助通气和支持心功能的药物已经改善了患者的生存时间和生活质量。许多基于干细胞或基因治疗的方法已进入临床实验阶段，目的是修复DMD心脏，但没有一种方法取得显著的临床疗效。这些疗法的有限成功是由于将治疗方法输送到心脏的困难，以及患者对供体细胞或病毒载体的免疫系统的激活。为了克服这些限制，我开发了一种细胞介导的外显子跳跃策略，用于骨骼肌，目前正在曼彻斯特的一项临床试验中进行测试（EudrAct .2019-001825-28），基于自体成血管细胞（mab）的移植，这是血管相关的肌源性祖细胞。单克隆抗体使用表达小核RNA （snRNA）的慢载体进行遗传校正，该慢载体设计用于诱导肌营养不良蛋白51外显子上的外显子跳跃，并产生短但功能的肌营养不良蛋白。这种方法的新颖性是基于snRNA沿着再生肌纤维扩散的能力，然后纠正引起外显子跳跃的营养不良的邻近细胞核。然而，单克隆抗体只能用于骨骼肌的治疗，因为它们不能自然分化为心肌，因此，MD患者的心脏将得不到治疗。在这个项目中，我将解决这个问题。众所周知，成纤维细胞可以转化为心肌细胞，但与单克隆抗体相比，它们不能系统地递送，因为它们不能穿过血管壁。这使得心脏转化成纤维细胞仅用于治疗局部病变如心肌梗死，而不用于治疗进展性和广泛性心肌病。我的初步数据显示，Mabs可以在10-15天左右通过短暂过表达特定的心脏相关基因转化为心肌细胞。因此，我假设这个时间窗口将允许通过心导管在整个心脏中输送这些mabs转化的心肌细胞。本项目旨在评估单克隆抗体转化心肌细胞移植后DMD小鼠模型心脏结构和功能的改善。此外，我将量化基因校正的单克隆抗体转化的心肌细胞与原有心肌细胞的融合程度，随后通过外显子跳变在邻近细胞核中校正的速率，从而量化产生的肌营养不良蛋白的数量。这一策略的核心要素已经在骨骼肌的临床试验中，但还没有在心脏。该项目将测试细胞介导的外显子跳跃策略在心脏中的适用性。该项目的成功结果证明了该策略的有效性，将为未来在DMD患者心脏和一般心脏病中对该策略进行临床试验铺平道路。