Project Summary NeuroGT, Inc is a start-up company founded by Dr. Haiyan Fu, an associate professor in the Gene Therapy Center at University of North Carolina at Chapel Hill, with the mission of develop and commercialize effective gene therapy products to treat rare neurogenetic diseases in humans. The goal of this project is to develop an effective gene therapy product targeting the root cause for treating Mucopolysaccharidosis (MPS) I is a fatal lysosomal storage disease (LSD) caused by autosomal recessive defects in α-L-iduronidase (IDUA). Severe form of MPS I (MPS IH, Hurler syndrome) represents the majority of known cases, with premature deaths usually before age 10 years, predominantly due to neurological deterioration and cardiorespiratory failure. No effective treatment is available for neurological indications of MPS IH. Because of the global diffuse neuropathy and the blood brain barrier (BBB), MPS IH is not amenable to either recombinant enzyme replacement therapy or bone marrow transplantation, which are the standard of care for treating somatic symptoms of MPS I. Gene replacement therapy targeting the root cause has been demonstrated to be an ideal strategy for treating monogenic diseases. Numerous studies have demonstrated successful in IV or intrathecal (IT) delivery of trans-BBB-neurotropic AAV9 for treating neurogenetic diseases. Importantly, the efficacy and safety profiles of IV and IT rAAV9 delivery have been demonstrated to be highly reproducible across different neurogenetic diseases, including LSDs. For optimal therapeutic potential, we have developed a novel self-complementary (sc) AAV vector, scAAV- mCMV-∆hIDUAop, to deliver a miniaturized human IDUA cDNA with codon-optimization. When tested in vitro in human MPS IH cells, this scAAV-∆hIDUAop vector construct was shown to express functional IDUA protein at a level 8-fold higher, than the single-stranded (ss) rAAV-hIDUAop vector. Importantly, the transduction with scAAV--∆hIDUAop vector also lead to 11-fold increase in the secretion of rIDUA, which can enter non- transduce MPS I cells and clear GAG storage there. Our preliminary data strongly support the potential of further development towards clinical application in humans. In this proposed project, for clinical relevance, we will test this new scAAV-∆hIDUAop vector in MPS I mouse model using AAV9, via IV, intrathecal (IT) and IV+IT delivery, to assess the therapeutic potential and determine the optimal regimen for treating MPS I. The proposed project will allow us to generate rigorous preclinical efficacy and safety data, to support the subsequent clinical development and commercialization.