Contributed by Lawrence Steinman, July 22, 2018 (sent for review May 22, 2018; reviewed by Emanuela Gussoni and Reinhard Hohlfeld)
Duchenne muscular dystrophy is a genetic disorder in which mutations in the dystrophin gene causes severe muscle wasting. A proposed treatment method involves systemic gene replacement therapy to introduce a functional dystrophin gene to the skeletal and cardiac muscles of the patient via rAAV6. A potentially serious problem may arise from immune recognition of dystrophin and the accompanying adeno-associated viral (AAV) vector. This unwanted immunity could interfere with successful long-term dystrophin expression by muscle cells. To suppress the unwanted immune response, a DNA vaccine was engineered to dampen immunity to both dystrophin and AAV6 capsid. Development of this engineered plasmid aims to improve gene replacement therapy and to overcome problems inherent with unwanted immunity to the encoded protein and to the delivery vector.
In gene therapy for Duchenne muscular dystrophy there are two potential immunological obstacles. An individual with Duchenne muscular dystrophy has a genetic mutation in dystrophin, and therefore the wild-type protein is “foreign,” and thus potentially immunogenic. The adeno-associated virus serotype-6 (AAV6) vector for delivery of dystrophin is a viral-derived vector with its own inherent immunogenicity. We have developed a technology where an engineered plasmid DNA is delivered to reduce autoimmunity. We have taken this approach into humans, tolerizing to myelin proteins in multiple sclerosis and to proinsulin in type 1 diabetes. Here, we extend this technology to a model of gene therapy to reduce the immunogenicity of the AAV vector and of the wild-type protein product that is missing in the genetic disease. Following gene therapy with systemic administration of recombinant AAV6-microdystrophin to mdx/mTRG2 mice, we demonstrated the development of antibodies targeting dystrophin and AAV6 capsid in control mice. Treatment with the engineered DNA construct encoding microdystrophin markedly reduced antibody responses to dystrophin and to AAV6. Muscle force in the treated mice was also improved compared with control mice. These data highlight the potential benefits of administration of an engineered DNA plasmid encoding the delivered protein to overcome critical barriers in gene therapy to achieve optimal functional gene expression.
↵1Present address: Department of Orthopedic Surgery, University of Pennsylvania, Philadelphia, PA 19104.
↵2Present address: Department of Cell and Developmental Biology, University of Pennsylvania, Philadelphia, PA 19104.
↵3Present address: Penn Institute of Regenerative Medicine, University of Pennsylvania, Philadelphia, PA 19104.
Author contributions: P.P.H., F.M., P.E.K., A.F., M.B., K.E.G.M., E.E.F., J.S.C., W.H.R., H.M.B., and L.S. designed research; P.P.H., L.J.L., F.M., P.E.K., A.F., M.B., K.E.G.M., E.E.F., J.S.C., and W.H.R. performed research; P.P.H., L.J.L., F.M., P.E.K., A.F., M.B., K.E.G.M., J.S.C., W.H.R., H.M.B., and L.S. analyzed data; and P.P.H., F.M., A.F., J.S.C., W.H.R., H.M.B., and L.S. wrote the paper.
Reviewers: E.G., Boston Children’s Hospital; and R.H., Institute of Clinical Neuroimmunology.
Conflict of interest statement: Stanford University has filed a provisional patent application on behalf of P.P.H. and L.S. claiming some of the concepts contemplated in this publication. L.S. and Reinhard Hohlfeld are coauthors on a 2017 Commentary.
See Commentary on page 9652.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1808648115/-/DCSupplemental.
Published under the PNAS license.
See related content: