Enhancing Sarcolemmal Membrane Repair Using a Novel Protein Therapeutic Approach can Improve Membrane Integrity in Duchenne Muscular Dystrophy Cells and Mouse Models

Duchenne Muscular Dystrophy (DMD) is a fatal neuromuscular disease involving progressive cardiac and skeletal myocyte cell death and subsequent muscle degeneration. DMD and Becker muscular dystrophy (BMD) result from mutations in the dystrophin gene that compromise the structural integrity of the sarcolemma. While there has been important recent progress in the treatment of DMD, the currently available therapeutics have significant limitations. Thus, novel therapies that address the compromised sarcolemmal membrane integrity are an unmet medical need. Tripartite motif protein 72, or mitsugumin 53 (TRIM72/MG53), facilitates the sarcolemma repair response after disruption of the membrane. Loss of TRIM72/MG53 function in mice leads to a progressive myopathy. Exogenous delivery of recombinant human MG53 protein (rhMG53) can increase sarcolemmal membrane repair in many different cell types and ameliorate pathology in models of DMD and multiple limb girdle muscular dystrophies. Despite the promise of rhMG53 as a therapeutic invention the development of this protein to treat DMD has been complicated by findings showing that high levels of the protein correlate with metabolic dysfunction in animal models and some human patients. Deletion analysis of the major domains of the rhMG53 protein allowed us to design an improved version of rhMG53 called MyoTRIM. MyoTRIM contains amino acid replacements to eliminate metabolic effects and improve solubility while maintaining membrane repair effects. One set of modifications inactivate the E3 ligase activity of the protein, which revealed this function is dispensable for subcellular localization of the protein. TRIM72/MG53 is reported to mediate this effect by binding phosphatidylserine (PS) at membrane injury sites, so we demonstrate that MyoTRIM can bind PS-coated beads to illustrate that carboxy-terminal domains are required for effcient PS binding rather than the E3 ligase function. External application of MyoTRIM can increase membrane repair capacity in a variety of cell-based assays using DMD and BMD patient myoblasts and myotubes. We also establish that MyoTRIM can increase membrane repair and prevent eccentric contraction induced injury in a dystrophin deficient mouse model. Taken together, these results provide mechanistic insight into rhMG53 function and indicate that MyoTRIM can recapitulate the therapeutic effects on sarcolemmal membrane repair while eliminating E3 ligase activity and associated side effects. This work was supported by the Offce of the Assistant Secretary of Defense for Health Affairs through the Duchenne Muscular Dystrophy Research Program (DMDRP) under Translational Research Award HT9425-23-1-0940. The opinions, interpretations, conclusions, and recommendations are those of the author and are not necessarily endorsed by the Department of Defense. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.