Histone deacetylase 4 (HDAC4) is a stress responsive factor that mediates multiple responses in skeletal muscle upon different stresses. We have clarified its protective role in maintaining skeletal muscle homeostasis after denervation, or in Amyotrophic Lateral Sclerosis, and in muscle regeneration upon injury. As a member of class IIa, HDAC4 shuttles from the nucleus to the cytoplasm in response to several stimuli. We have found HDAC4 to be upregulated prevalently in the cytoplasm of Duchenne Muscular Dystrophy (DMD) muscles in humans and mice. DMD is a genetic, progressive, incurable disorder, characterized by muscle degeneration and weakness. To delineate HDAC4 role in skeletal muscle in DMD, we generated and characterized mdx mice with a tissue-specific deletion of HDAC4 (mdx;KO). We demonstrated that deletion of HDAC4 in skeletal muscle worsens the pathological features of DMD, by enhancing muscle damage and reducing muscle regeneration, ultimately leading to a decrease in muscle performance. We proved that the mdx;KO phenotype is rescued by the cytoplasmic HDAC4 (HDAC4 L/A). Considering its protective role in mdx;KO muscles, we delivered HDAC4 L/A in mdx muscles via electroporation, ameliorating mdx muscle function and architecture. We propose to fully characterize the effects of cytoplasmic HDAC4 L/A in DMD in terms of muscle function, degeneration, regeneration, and lipid infiltration. Our results will provide innovative findings and suggestions for new pharmacological approaches (i.e. delivery of cytoplasmic HDAC4 or pharmacological extrusion of HDAC4 from the nucleus) for the treatment of DMD patients.
The potential progress towards improving the current therapeutic approaches for the treatment of DMD is the long-term aim of this proposal. To date there is no effective cure for DMD and more research must be done to further understand the molecular mechanisms underlying this pathology. The pan-HDAC inhibitor givinostat has been shown to be partially beneficial for dystrophic mice and patients. However, givinostat was not able to improve the dystrophic phenotype in adult mice or muscle performance in DMD patients. Moreover, as a pan-HDACi, givinostat nonspecifically blocks all HDAC members and long-term treatment with pan-HDACi has been associated with numerous side effects. Innovative approaches, in combination with pharmacological treatments targeting HDAC, could be much more efficient to counteract dystrophy; however, this will be possible only if the detailed mechanisms of action of different HDAC members are considered.
Our yet unpublished results clearly demonstrate a strikingly, novel, protective cytoplasmic role of HDAC4 in DMD. We believe that HDAC4 functions need to be fully characterized and preserved in DMD, rather than being inhibited by the use of pan-HDACi. Limiting the therapeutic approach to the use of Class I HDAC inhibitors may still in part influences the acetylation status of HDAC4 targets, since HDAC4 deacetylates via Class I HDACs. Thus, combinatorial approaches based on selective inhibition of some HDACs, while preserving or enhancing the activity of others, may be the solution towards a breakthrough improvement of DMD therapy for dystrophic patients.
We plan to deliver GFP-HDAC4 L/A in young mdx muscles to study how, muscle function, degeneration and regeneration and lipid infiltration are affected. This study will provide a proof-of-principle evidence that paves the way to alternative therapeutic approaches, which may include, for instance, the delivery of cytoplasmic HDAC4 cDNA or peptide via nanoparticles, and /or forced extrusion of HDAC4 from the nucleus by using drugs, such as PP2A inhibitors, for the treatment of DMD patients.