Abstract
A cornerstone of autologous cell therapy for Duchenne muscular dystrophy is the engineering of suitable cells to express dystrophin in a stable fashion upon differentiation to muscle fibers. Most viral transduction methods are typically restricted to the expression of truncated recombinant dystrophin derivatives and by the risk of insertional mutagenesis, while non-viral vectors often suffer from inefficient transfer, expression and/or silencing. Here we addressed such limitations by using plasmid vectors containing nuclear matrix attachment regions (MAR). Using in vitro transfection and intra muscular transplantation in nude and immunosuppressed mdx mice, we show that clones of mesoangioblast skeletal muscle progenitors can be generated to mediate stable expression from MAR-containing vectors, while maintaining their ability to differentiate in vitro and in vivo and to express dystrophin after transplantation in dystrophic mouse muscles. We conclude that the incorporation of MARs into plasmid vectors may improve non-viral plasmid-based cell therapy feasibility.
Highlights
Duchenne muscular dystrophy is an X-linked progressive muscular wasting disease that affects skeletal and cardiac muscles, and for which there is currently no cure
Gene therapy approaches aim to engineer vectors that efficiently transduce myofibers with a dystrophin expression cassette, whereas cell therapy approaches aim to deliver the dystrophin transgene to the myofiber by stem/progenitor cells, while preferably replenishing the satellite cell pool with genetically corrected or complemented autologous cells
A panel of human and animal matrix attachment regions (MAR) elements was tested for their effect on the establishment of stable cell clones and for transgene expression level and stability using murine mesoangioblasts (Table 1)
Summary
Duchenne muscular dystrophy is an X-linked progressive muscular wasting disease that affects skeletal and cardiac muscles, and for which there is currently no cure. It is caused by mutations in the dystrophin gene, resulting in the lack or reduction of the protein. This deficit leads to a disruption of the dystroglycan complex and destabilization of the sarcolemma, resulting in progressive muscle wasting [1]. Promising experimental approaches that aim to restore the dystrophin complex at the sarcolemmal membrane include i) exon skipping by pharmacological strategies, ii) systemic gene therapy and iii) cell therapy [2,3]. We assessed a novel approach to express full-length dystrophin in a cell therapy setting
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