DMD, an X-linked recessive disorder, is the most frequent hereditary myopathy affecting approximately 1 in 3500 boys of all races. It is caused by mutations in the gene for the myocyte structural protein dystrophin. The loss of functional dystrophin in striated muscles causes extreme myocyte fragility and damage. The skeletal muscles degenerate continuously becoming fibrotic with adipose infiltrates. Clinical symptoms first manifest in boys between 18 months and 3 years of age. Muscle wasting occurs as the disease progresses and the patients become nonambulatory between 9 to 12 years of age. Mortality results from continuous loss of skeletal muscle causing spinal deformation, breathing difficulties, and cardiomyopathies. The disease is always fatal, often in the second decade and invariably in the third decade of life. There are no cures for DMD, and the current therapeutics regimens involve steroidal anti-inflamatories to limit immune responses to muscle fiber damage and only palliative treatments. Recombinant AAV may be used to restore dystrophin expression to skeletal muscles, either using a protein replacement approach, e.g. vectors that express micro-dystrophin, or an exon-skipping approach. For either application, producing sufficient quantities of rAAV to attain meaningful endpoints in the clinically relevant canine model of DMD has been the major obstacle for clinical trials. Most of the rAAV vector we produce contain a non-structural gene expressing a modified U7 RNA that interferes with processing the primary dystrophin transcript. Using specific anti-sense elements, our collaborators determined that specific exons may be omitted from the mature mRNA. This exon-skipping approach has been validated in murine DMD models and limited trials in canine DMD models. Based on these encouraging results, studies are underway using rAAV that express reporter proteins to establish the pharmacological parameters for treating DMD. From these results, the bio-distribution, routes of administration, vector toxicity are determined. Finally, using the data obtained with reporter constructs, doses that produce phenotypic improvements in large animals DMD models can then be established.

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Project End
Budget Start
Budget End
Support Year
16
Fiscal Year
2010
Total Cost
$1,695,186
Indirect Cost
Name
National Heart, Lung, and Blood Institute
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Type
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Barbash, I M; Cecchini, S; Faranesh, A Z et al. (2013) MRI roadmap-guided transendocardial delivery of exon-skipping recombinant adeno-associated virus restores dystrophin expression in a canine model of Duchenne muscular dystrophy. Gene Ther 20:274-82
Li, Lina; Dimitriadis, Emilios K; Yang, Yu et al. (2013) Production and characterization of novel recombinant adeno-associated virus replicative-form genomes: a eukaryotic source of DNA for gene transfer. PLoS One 8:e69879
Airenne, Kari J; Hu, Yu-Chen; Kost, Thomas A et al. (2013) Baculovirus: an insect-derived vector for diverse gene transfer applications. Mol Ther 21:739-49
Denard, Jerome; Beley, Cyriaque; Kotin, Robert et al. (2012) Human galectin 3 binding protein interacts with recombinant adeno-associated virus type 6. J Virol 86:6620-31
Handel, Eva-Maria; Gellhaus, Katharina; Khan, Kafaitullah et al. (2012) Versatile and efficient genome editing in human cells by combining zinc-finger nucleases with adeno-associated viral vectors. Hum Gene Ther 23:321-9
Bish, Lawrence T; Sleeper, Meg M; Forbes, Sean C et al. (2012) Long-term restoration of cardiac dystrophin expression in golden retriever muscular dystrophy following rAAV6-mediated exon skipping. Mol Ther 20:580-9
Cecchini, Sylvain; Virag, Tamas; Kotin, Robert M (2011) Reproducible high yields of recombinant adeno-associated virus produced using invertebrate cells in 0.02- to 200-liter cultures. Hum Gene Ther 22:1021-30
Kotin, Robert M (2011) Large-scale recombinant adeno-associated virus production. Hum Mol Genet 20:R2-6
Virag, Tamas; Cecchini, Sylvain; Kotin, Robert M (2009) Producing recombinant adeno-associated virus in foster cells: overcoming production limitations using a baculovirus-insect cell expression strategy. Hum Gene Ther 20:807-17
Cecchini, Sylvain; Negrete, Alejandro; Virag, Tamas et al. (2009) Evidence of prior exposure to human bocavirus as determined by a retrospective serological study of 404 serum samples from adults in the United States. Clin Vaccine Immunol 16:597-604

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