Protein Supplementation Therapy to Increase Membrane Repair to Treat Duchenne Muscular Dystrophy

Abstract Our previous studies show synthesized recombinant human MG53 (rhMG53) protein binds at membrane damage sites to enhance membrane repair in cultured cells and dystrophic mouse models when applied outside the cell. Based on our studies showing that rhMG53 therapy ameliorates disease pathology in dystrophin-null mice, we now seek to develop a new therapy to enhance repair and restore muscle function that is compromised in Duchenne muscular dystrophy (DMD). In this proposed project, we will show new data where we optimized the original rhMG53 construct to engineer a new version of rhMG53 (that we now call MyoTRIM) to eliminate the E3 ligase activity of the protein that caused off-target effects on metabolism. We found MyoTRIM could increase membrane repair in myoblasts from DMD patients, illustrating the translational potential of this therapeutic. Thus, we propose in this Translational Research Award (TRA) proposal to expand on our existing preclinical translational data in support an appropriate therapeutic development path for MyoTRIM to treat DMD. We hypothesize that MyoTRIM can be effectively manufactured and that it will treat a DMD mouse model without toxic side effects. Optimization of protein production from CHO cell cultures will be conducted in the Weisleder Lab. These findings will be transferred to Cellthon, a CMO focused on therapeutic protein production in CHO cells. Randomized cohorts of D2-mdx mice will be treated with MyoTRIM generated in either CHO cells or E. coli over 28 weeks and changes in the dystrophy phenotype will be determined through various measurements of skeletal and cardiac muscle structure and function. Toxicity studies will be conducted at Charles River Laboratories, which has a long history of conducting such studies. We will also monitor potential immunogenicity of MyoTRIM in these toxicity studies. The proposed project will have an impact on multiple FY22 Translational Research Award focus areas by: 1) improving the function and quality of life, and extending the lifespan, of all individuals with DMD of all ages and harboring any mutation type, 2) providing a translational study using animal models to test a novel interventions targeting both cardiac and skeletal muscle, and 3) expanding existing preclinical translational data in support of a specific therapeutic development path. Because MyoTRIM targets a fundamental membrane repair process unrelated to the primary DMD mutations, it should be effective against all forms of muscular dystrophy that result from unstable muscle membranes. MyoTRIM is fundamentally innovative because it does not depend on direct targeting of specific mutations. The efficacy pf MyoTRIM is also not dependent on the presence of antibodies against AAV or the age of the DMD patient, thus this approach can be effective in all individuals with DMD. Additionally, rhMG53 is effective in both skeletal and heart muscle, thus MyoTRIM may be useful to treat both skeletal and cardiac muscle defects involved with DMD.