| Facioscapulohumeral muscular dystrophy is the most common inherited muscular dystrophy though no treatment exists. The lack of therapeutic development for FSHD is directly linked to insufficient understanding of how the disease is caused. The goals of the studies presented here were to gain a better understanding of the pathogenic mechanisms of FSHD and to develop targeted translational therapies to treat the disease.;FSHD is associated with D4Z4 repeat contraction on human chromosome 4q35, which does not result in complete loss or mutation of any gene. Consequently, the major obstacle to discerning the underlying pathogenic mechanism is to identify the cause. Although no gene was conclusively linked to FSHD development, evidence supported a role for the D4Z4-encoded DUX4 gene. In Chapter 3, our objective was to test the in vivo myopathic potential of DUX4. We delivered DUX4 to zebrafish and mouse muscle by transposon-mediated transgenesis and adeno-associated viral vectors, respectively. We found over-expression of DUX4 caused abnormalities associated with muscular dystrophy in both animal models. This toxicity required DNA binding, since a DUX4 DNA binding domain mutant produced no abnormalities. We also found the toxic effects of DUX4 were p53-dependent. This study demonstrated the myopathic potential of DUX4 in animal muscle and provided a p53-dependent mechanism for DUX4-induced toxicity. Considering previous studies showed DUX4 was elevated in FSHD patient muscles, our data support the hypothesis that DUX4 over-expression contributes to FSHD development.;With DUX4 as a potential target, gene silencing approaches could provide treatment for FSHD. With as many as 29 different gene mutations responsible for other dominant myopathies, gene silencing approaches could have a broad impact. Feasible mechanisms to silence dominant disease genes have lagged behind gene replacement strategies, but with the discovery of RNA interference (RNAi) and its subsequent development into a promising new gene silencing tool, the landscape has changed. In Chapter 4, our objective was to demonstrate proof-of-principle for RNAi therapy of a dominant myopathy in vivo. We tested the potential of AAV-delivered therapeutic microRNAs, targeting the human Facioscapulohumeral muscular dystrophy (FSHD) Region Gene 1 (FRG1), to correct myopathic features in mice expressing toxic levels of human FRG1 (FRG1-high mice). We found that FRG1 gene silencing improved muscle mass, strength, and histopathological abnormalities associated with muscular dystrophy in FRG1-high mice, thereby demonstrating therapeutic promise for treatment of FSHD and other dominantly inherited myopathies using RNAi.;Next we applied this therapeutic strategy to FSHD by targeting DUX4. Several recent studies support an FSHD pathogenesis model involving over-expression of the myopathic DUX4 gene making it the most promising therapeutic target. In Chapter 5, we tested a pre-clinical RNAi-based DUX4 gene silencing approach as a prospective treatment for FSHD. We found that AAV vector-delivered therapeutic microRNAs corrected DUX4-associated myopathy in mouse muscle. These results provide proof-of-principle for RNAi therapy of FSHD through DUX4 inhibition. Together these studies have helped define the main pathogenic insult in FSHD and laid out a plausible, targeted therapy to treat the disease. |
