The goal of this project is to demonstrate the efficacy of Sleeping Beauty (SB) transposons as vectors for non-viral gene delivery to treat mucopolysaccharidosis (MPS) diseases, specifically Type I, a-iduronidase (IDUA). The project has two sub-goals: 1) Complete experiments that reveal the efficacy of delivery of SB transposons to the mouse to treat a//of the effects of IDUA-deficiency;in particular restoration of IDUA activity in the brain (Aim 1). 2) Reduce potential cytotoxic effects of SB transposon integrants by directing transposition to safe harbors in the mouse genome (Aims 2 - 4).
Aim 1 will complete our assessment ofthe SB system as a vector for gene delivery in the mouse. The next step for development ofthe SB system for humans is to maximize the safety of using transposons that randomly integrate into genomes by directing their integration to sites that are thought to be safe. The principal hvpotheses of this proposal are: 1) Transposons can be delivered directly into the liver and brain for uptake and expression ofthe gene(s) they carry. 2) Certain features of SB Transposons can be exploited to direct their integration into specific sequences in genomes that are considered """"""""safe"""""""" and thereby avoid some ofthe issues associated with viruses used as gene delivery vectors.
The Specific Aims of the proposal are to 1) determine the efficacy of SB transposon delivery to the brain in MPS I mice, 2) develop a 'universal'targeting system to direct SB transposons into selected regions of a DNA molecule, 3) evaluate the ability of a 'universal'targeting system to direct SB transposons to selected regions ofthe mouse genome in vitro, and 4) evaluate the ability of a 'universal'targeting system to direct SB transposons to selected regions ofthe genome in the liver of a living mouse. IMPACT: There are two major impacts of our proposed studies. 1) We will demonstrate that SB transposons can be effectively used to achieve long-term expression in the brain as well as in other tissues and cell types without using viral vectors. 2) By targeting integration of SB transposons to specific sites in the mouse genome, the concerns of insertional mutagenesis will be reduced considerably.

Public Health Relevance

Lysosomal storage disorders are a rare group of inherited diseases caused by genetic deficiency in which patients suffer from skeletal abnormalities, heart and breathing problems, mental retardation and death. It is envisioned in this grant application that one way to treat these diseases would be to restore the missing gene in patients'central nen/ous system (in the brain) to prevent neurodegeneration. Targeting of transposon vectors to specific sites will increase the safety of use of this vector in humans. PROJECT/PERFORIVIANCE SiTE(S) (If additional space is needed, use Project/

National Institute of Health (NIH)
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
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University of Minnesota Twin Cities
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Ou, Li; Przybilla, Michael J; Whitley, Chester B (2018) Metabolomics profiling reveals profound metabolic impairments in mice and patients with Sandhoff disease. Mol Genet Metab :
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