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

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.

Duchenne Muscular Dystrophy (DMD) is an inherited severe muscle-wasting disease due to lack of expression of the dystrophin protein. The main cause of death in DMD patients is heart failure caused by the wasting of cardiac muscle that compromises heart functions. Currently, the only recognized therapy for dystrophic patients is represented by steroid administration. However, these can only delay the progression of DMD, and there are multiple side effects. Many novel therapeutical approaches have been proposed in the last few years, but none have yet showed a clinical efficacy. I have recently developed a new approach involving cell-mediated exon skipping and intra-arterial delivery of Mesoangioblasts (Mabs). Mabs are isolated from dystrophic patients and transduced with a snRNA inducing the skipping of a specific exon of dystrophin generating a shorter but functional dystrophin protein. The snRNA assembles in the cytoplasm with nuclear proteins and then diffuses along the muscle fibres correcting the resident neighbouring nuclei and amplifying the therapeutical effect. However, as Mabs do not spontaneously differentiate into cardiomyocytes, the approach is used for skeletal muscle but not the heart.
The aim of my project is to overcome this problem by developing a strategy to treat the heart of DMD patients using genetically corrected Mabs-derived cardiomyocyte. Preliminary data show that Mabs can be converted into functional cardiomyocytes through transient overexpression of three specific cardiac-related genes. Systemic delivery genetically corrected and differentiated cardiac-derived Mabs will potentially treat all the diseased myocardium and fusing with resident cardiomyocytes will correct them and amplify the therapeutical effect, similar to skeletal muscles, though to a minor extent. The combination of these two strategies will lead to a cure for DMD and has the potential to become a strategy for many other dilated cardiomyopathies.