New disease models and therapeutic strategies for arrhythmogenic cardiomyopathy

Arrhythmogenic cardiomyopathy (ACM) is a rare genetic form of cardiomyopathy, manifesting with ventricular arrhythmias, syncope, and sudden cardiac death. Approximately half of ACM patients present pathogenic variations in one or more genes encoding proteins of the intercalated discs, junctional structures that provide mechanical and electro-metabolic coupling among cardiomyocytes. The disease is typically transmitted with an autosomal dominant pattern of inheritance; however, it is characterized by incomplete penetrance and variable expressivity, even within the same family, suggesting that different environmental factors might contribute to define the clinical phenotype. Although ACM has been first described almost thirty years ago, the pathogenic mechanisms leading to its development are still only partially known, therefore limiting the development of novel therapeutic options. Indeed, most of the management strategies aim at preventing arrhythmias and sudden cardiac death, while the two main ACM hallmarks, the progressive loss of cardiomyocytes and the substitution of myocardium with fibro-fatty tissue, remail untargeted. In this scenario, the main goal of this thesis project was to identify novel therapies to slow down fibro-fatty replacement specifically acting on cardiac fibro-adipogenic progenitor cells (cFAPs), which have been recognized as key players in the disease progression. We found these cells to be more abundant in hearts of mice belonging to an ACM murine model (Tg-hQ, previously developed by our research group) compared to wild-type animals. Moreover, we used cFAPs as in vitro system to test the effect of the pan-HDAC inhibitor Givinostat, selected as candidate drug because of its well reported anti-fibrotic action in the context of Duchenne Muscular Dystrophy. Here, Givinostat achieved promising results by reducing proliferation and fibro-fatty differentiation of cFAPs in vitro therefore being a promising drug for ACM treatment. The efficacy of the drug was subsequently evaluated in vivo on a limited number of Tg-hQ mice in which it was possible to observe a reduction in the expression of fibrotic genes upon the treatment. In parallel, the project aimed at characterizing a novel transgenic murine model (Tg-hG) presenting a cardiomyocyte-restricted overexpression of the human p.G100R desmoglein 2. These animals developed a series of ACM-related features over time, including cardiac inflammation, progressive fibrotic accumulation, and mild alterations in cardiac functionality at rest. Moreover, single-nucleus RNA sequencing helped to dissect the cellular composition of wild-type and transgenic hearts. Interactome analyses highlighted the involvement of fibroblast-secreted factors in three key ACM-related networks: extracellular matrix accumulation, inflammation of Wnt signaling, possibly opening to the identification of novel therapeutic targets.