PROJECT SUMMARY In this Direct-to-Phase II SBIR, BioStrategies LC proposes to advance the development of a novel AAV- based gene therapy for mucopolysaccharidosis type IIIA (MPS IIIA, also known as Sanfilippo A syndrome). In contrast to other gene therapies in development for MPS IIIA, BioStrategies’ approach is designed to achieve a systemic therapeutic effect, including delivery of a functional replacement enzyme across the blood brain barrier (BBB) without intrathecal or intraventricular administration. Further development and commercialization of BioStrategies’ gene therapy has the potential to deliver the first and only clinically effective treatment for MPS IIIA, potentially rescuing patients from the debilitating degenerative effects and early mortality associated with this genetic disorder. It would also serve as strong proof-of-principle for a new approach to gene therapy in a broad array of other genetic disorders that require the delivery of functional enzymes to the central nervous system in addition to the periphery. In preliminary studies, BioStrategies developed a novel approach that would use AAV-based gene therapy to deliver genes to the liver or other organs outside of the CNS to produce functional SGSH combined with non-toxic carbohydrate-binding subunit B of ricin (RTB), a lectin that has strong affinity for a broad array of glycoproteins and glycolipids on mammalian cell surfaces. This RTB fusion protein is designed to infiltrate multiple tissues, including crossing the BBB to achieve therapeutic levels in the brain. In animal models of MPS IIIA, intravenous administration of this SGSH:RTB fusion protein reliably and robustly achieved broad distribution, including across the BBB, restoring heparan sulfate to wild-type levels in the brain and peripheral tissues. The team then demonstrated that AAV-based gene therapy targeting the liver achieved continuous production of SGSH:RTB, with broad distribution and reversal of the MPS IIIA phenotype in brain and peripheral tissues. In this Direct-to-Phase II, BioStrategies now proposes to assess the utility of an array of AAV vectors to optimize SGSH:RTB production and distribution in vivo followed by robust, fully-powered efficacy testing of the best performing AAV construct in an MPS IIIA model. Aim 1. Optimize SGSH:RTB product design, transfection efficacy, and safety. Milestone: Select a lead candidate based on a decision matrix that prioritizes the highest efficacy in the CNS with the lowest dose, best safety profile, and commercialization potential (full details on decision considerations provided in the Research Strategy). Aim 2. Evaluate preclinical efficacy of AAV-based SGSH:RTB gene therapy in MPS IIIA mice. Milestones: Demonstrate that an intravenously-delivered AAV-based SGSH:RTB gene therapy 1) establishes continuous production of SGSH:RTB, 2) achieves therapeutic-levels in brain and other organs/tissues of interest, 3) prevents progression of the MPS IIIA phenotype (i.e.,...