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Mol Ther. 2019 Jan 25. pii: S1525-0016(19)30005-X. doi: 10.1016/j.ymthe.2019.01.002. [Epub ahead of print]

Development of Novel Micro-dystrophins with Enhanced Functionality.

Author information

1
Molecular and Cellular Biology Program, University of Washington School of Medicine, Seattle, WA 98195, USA; Department of Neurology, University of Washington School of Medicine, Seattle, WA 98195, USA; Senator Paul D. Wellstone Muscular Dystrophy Specialized Research Center, Seattle, WA 98195, USA.
2
Department of Neurology, University of Washington School of Medicine, Seattle, WA 98195, USA; Senator Paul D. Wellstone Muscular Dystrophy Specialized Research Center, Seattle, WA 98195, USA.
3
Senator Paul D. Wellstone Muscular Dystrophy Specialized Research Center, Seattle, WA 98195, USA; Department of Biochemistry, University of Washington School of Medicine, Seattle, WA 98195, USA.
4
Molecular and Cellular Biology Program, University of Washington School of Medicine, Seattle, WA 98195, USA; Department of Neurology, University of Washington School of Medicine, Seattle, WA 98195, USA; Senator Paul D. Wellstone Muscular Dystrophy Specialized Research Center, Seattle, WA 98195, USA; Department of Biochemistry, University of Washington School of Medicine, Seattle, WA 98195, USA; Department of Medicine, University of Washington School of Medicine, Seattle, WA 98195, USA. Electronic address: jsc5@uw.edu.

Abstract

Gene therapies using adeno-associated viral (AAV) vectors have advanced into clinical trials for several diseases, including Duchenne muscular dystrophy (DMD). A limitation of AAV is the carrying capacity (∼5 kb) available for genes and regulatory cassettes (RCs). These size constraints are problematic for the 2.2-Mb dystrophin gene. We previously designed a variety of miniaturized micro-dystrophins (μDys) that displayed significant, albeit incomplete, function in striated muscles. To develop μDys proteins with improved performance, we explored structural modifications of the dystrophin central rod domain. Eight μDys variants were studied that carried unique combinations of between four and six of the 24 spectrin-like repeats present in the full-length protein, as well as various hinge domains. Expression of μDys was regulated by a strong but compact muscle-restricted RC (CK8e) or by the ubiquitously active cytomegalovirus (CMV) RC. Vectors were evaluated by intramuscular injection and systemic delivery to dystrophic mdx4cv mice, followed by analysis of skeletal muscle pathophysiology. Two μDys designs were identified that led to increased force generation compared with previous μDys while also localizing neuronal nitric oxide synthase to the sarcolemma. An AAV vector expressing the smaller of these (μDys5) from the CK8e RC is currently being evaluated in a DMD clinical trial.

KEYWORDS:

AAV; DMD; Duchenne muscular dystrophy; clinical trials; dystrophin; gene therapy

PMID:
30718090
DOI:
10.1016/j.ymthe.2019.01.002
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