Significance: Niemann-Pick Type C1 disease (NPC1) is a devastating inherited neurodegenerative lysosomal storage disease caused by mutations in the NPC1 gene, which encodes for an intracellular membrane cholesterol transporter. Currently, there are few treatment options for NPC1. Protein replacement therapy is not possible, because the NPC1 protein is an insoluble membrane transporter. Gene therapy of either human NPC1, or a long standing mouse model of NPC1, has not been attempted, owing to the large size of the NPC1 mRNA, which exceeds the capacity of common gene therapy viral vectors. In contrast, large size genes can be encapsulated in nonviral delivery systems such as Trojan horse liposomes (THLs). THLs are manufactured with a receptor-specific monoclonal antibody (MAb), which acts as a molecular Trojan horse to ferry the THL from blood into the nuclear compartment of brain cells. In prior work, THL-mediated delivery of plasmid DNA has produced therapeutic effects in mouse models of neural disease including, Parkinson's disease, brain cancer, or lysosomal storage disease. Hypothesis: The hypothesis tested in the present work is that the NPC1 gene can be effectively replaced in brain, and in peripheral organs, with regular intravenous (IV) administration of THLs encapsulating plasmid DNA encoding the NPC1 gene. This hypothesis is supported by prior work in the NPC1 mouse model wherein embryos were transfected with the wild type NPC1 gene and these transgenic mice were cross-bred with the NPC1 mouse. Replacement of the NPC1 gene in brain effectively cured the disease in the NPC1 mouse. Preliminary Data: Prior work with a lysosomal storage disease mouse model has shown it is possible to achieve replacement of the wild type gene in mouse brain with IV administration of THLs carrying the expression plasmid DNA encoding the lysosomal enzyme gene. The THLs are targeted with a MAb specific for the mouse transferrin receptor (TfR). Prior work has also shown that repeat, chronic IV administration of THLs causes no toxicity or immune reactions.
Specific Aims : First, THLs will be manufactured, and these THLs are targeted with the MAb specific for the mouse TfR, and encapsulate an expression plasmid DNA encoding for the NPC1 cDNA. The expression plasmid DNA will be engineered with methods used previously for therapeutic plasmid DNA. Second, the potency of the THLs will be assessed in cell culture using human NPC1 fibroblasts. Third, a colony of NPC1 mice will be generated for this project producing at least 36 homozygote mice. The mice will undergo treatment for 12 weeks with weekly IV administration of THLs carrying the NPC1 DNA, beginning at the age of 6 weeks. Treatment efficacy will be assessed with survival, body and organ weights, brain histology, peripheral organ histology, as well as brain biochemical parameters of NPC1 gene expression. This research can be translated to human NPC1, because prior work as shown it is possible to deliver transgenes to virtually all cells in the brain of the adult non-human primate with IV administration of THLs.
Niemann-Pick disease Type C1 (NPC1) is a devastating inherited neurodegenerative condition caused by mutations in the NPC1 gene, which encodes an intracellular membrane transporter for cholesterol and other lipids. Current gene therapy with viral vectors is not under development owing to the large size of the NPC1 gene. The present project develops a new technology for delivery of large size plasmid DNA, called Trojan Horse Liposomes, for treatment of the brain in the NPC1 mouse. If successful, this research can be translated to the development of new plasmid DNA based therapeutics for human NPC1.