Lysosomal storage disorders (LSDs) are a group of inherited metabolic diseases characterized by a dysfunction in lysosomes, with cumulative frequency of 1 in 7000 live births (although individually rare). Over 2/3 of LSD patients present an involvement of the central nerve system (CNS) with a broad spectrum of severity (nLSD), which makes LSDs the most common cause of pediatric neurodegenerative disease. Allogeneic hematopoietic stem cell transplantation (HSCT) or enzyme replacement therapy (ERT) by periodical injection of recombinant enzyme are main treatment options for nLSD. However, they are largely unsuccessful in reversing neurological complications due to the poor penetration of the enzymes across the blood-brain-barrier (BBB) to the CNS, a major obstacle in treating nLSD. The cation-independent mannose-6-phosphate receptor (M6PR, also called IGF2R) plays a critical role in lysosomal enzyme trafficking and intercellular transfer of most lysosomal enzymes, which is essential for metabolic cross-correction in treating LSDs. Developmental decline of M6PR on the BBB during early postnatal period in mouse and human has been documented, which is attributable to the lack of CNS enzyme delivery. Using a dual luciferase reporter system with site-mutagenesis, we have recently discovered that microRNA-143 (miR143) modulate M6PR protein levels on BBB-forming brain capillary endothelial cells (BrMV) by targeting to 3' untranslated region of M6PR mRNA. Using a mouse model of Hurler syndrome (severe mucopolysaccharidosis type I, MPS I), which is caused by the deficiency of ?-L-iduronidase (IDUA), we further demonstrated functional rescue of M6PR-mediated IDUA transfer in the brain of double- knockout (MPS/miR-143KO) mice with long-term CNS therapeutic benefits, as well as in human vascular endothelial cells by down-regulation of miR-143 with miR-143-sponge sequences. The data provide strong scientific premise for the development of a novel CNS-targeted approach that would be applicable in treating many neurologic LSDs involving M6PR pathway, or in delivering brain therapeutics that can adapting M6PR- mediated transcytosis pathway. In this proposal, we aim to develop a novel adeno-associated viral vector (AAV)- based translatable platform to ?restore? M6PR pathway on mature BBB for advanced delivery of therapeutic enzymes into the CNS with 3 aims, including developing optimal artificial miR143 inhibitor (143in) and expression cassette(s) for efficient and targeted reduction of miR143 on BrMV (aim 1), examination of biodistribution and ?off-target? expression and effects in mice with AAV-143in delivery (aim 2), as well as preclinical evaluation of BBB-targeted AAV/miR143in in correcting CNS abnormalities in MPS I mice by enzyme therapy derived from genetically modified erythroid/ megakaryocytic lineages (aim 3). The impact of the study is driven by the unmet medical need for efficient treatment of inherited nLSDs AND the major limitation of drug-delivery across the BBB.
It has been a major unmet medical need for the development of efficient transcytosis systems across the blood- brain barrier with broad distribution of the therapeutic macromolecules to many brain cell types in treating a wide variety of neurological diseases. The goal of this project is to develop novel therapeutic strategies that coalesce in vivo epigenetic alteration with microRNA manipulation, targeted gene transfer and transgene expression for advanced treatment of a neurologic lysosomal storage disease (nLSD), that have general and significant applicability for the treatment of neuronopathic diseases from other rare nLSDs to major public health concerns such as Parkinson and Alzheimer diseases.
