Gene therapy is a promising approach to treating Duchenne Muscular Dystrophy (DMD). However, current methods typically require the addition of extra dystrophin genes to the genome or the lifelong re- administration of foreign genetic material that works transiently to restore dystrophin expression, both of which have significant safety and practical concerns. Furthermore, these strategies have been limited by an inability to deliver the large and complex dystrophin gene sequence. An appealing alternative to these gene replacement approaches is the targeted repair of the endogenous mutant dystrophin gene. This concept, known as genome editing, represents a potential cure to DMD without the need for permanent integration of or repeated exposure to foreign biological material. Furthermore, it corrects the problem at the source by correcting the mutation to the naturally occurring dystrophin gene. Genome editing has been made a reality for human gene therapy by the recent development of transformative technologies that use engineered enzymes to cut and paste DNA sequences at specific sites in the genome. In fact, genome editing is now in clinical trials for treating cancer and HIV. The most recently developed genome editing technology, known as CRISPR, is much more robust than previous technologies and has rapidly transformed all areas of biomedical research and biotechnology in less than two years. Several efforts are underway to use CRISPR to correct genetic diseases, and we have demonstrated that it is possible to restore dystrophin expression in muscle cells from DMD patients. However, for this to be viable for clinical translation, we must demonstrate successful genome editing in skeletal and cardiac muscle tissue in animal models of the disease. In this study, we will use adeno- associated virus to delivery CRISPR to skeletal and cardiac muscles of a mouse model of DMD and a mouse model carrying the human dystrophin gene. The overall objective of this research proposal is to develop methods to restore dystrophin expression via targeted genome editing in vivo. The central hypothesis is that nuclease-mediated gene correction will lead proper dystrophin expression and function in mouse models of DMD. This research plan is innovative because it capitalizes on the unfulfilled potential of the CRISPR genome editing technology to address the fundamental limitations of conventional gene therapies and the unmet need for a safe and effective permanent cure to DMD. Importantly, this approach is also broadly applicable to numerous genetic diseases in addition to DMD. Thus in addition to identifying a lead candidate nuclease and delivery method for treatment of DMD, this work will also lead to additional development and refinements of the CRISPR technology to broadly benefit patients affected by hereditary disorders. Finally, the development of technologies for in vivo genome editing in skeletal and cardiac muscle will be broadly useful for biotechnology and basic scientific research.
Gene therapy holds incredible potential for curing inherited diseases, including neuromuscular disorders. However, the potential of gene therapy has not been realized due to fundamental limitations of gene delivery technologies. This project uses an innovative approach to correct the disease-causing genetic mutations, rather than delivering extra genes, as a therapy for Duchenne Muscular Dystrophy.
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