427. AAV2- and AAV5-Mediated CNS Delivery of Human CLN2 Reduces Lysosomal Storage in a Mouse Model of Late Infantile Neuronal Ceroid Lipofuscinosis

Late infantile neuronal ceroid lipofuscinosis (LINCL) is a neurometabolic disorder caused by mutations in CLN2, which encodes lysosomal tripeptidyl peptidase (TPP-1). Lack of TPP-1 results in intracellular accumulation of autofluorescent storage material within the CNS, neurodegeneration and death typically in childhood. In this study, we injected clinical-grade AAV vectors containing the human CLN2 cDNA into the LINCL mouse brain to determine therapeutic efficacy. A total of 3.6 |[times]| 109 genome copies of AAV2CUhCLN2 (n=5) or AAV5CUhCLN2 (n=6) were stereotaxically injected into the motor cortex, thalamus, and cerebellum of both hemispheres at 6 weeks of age. At 13 weeks post-injection (19 weeks of age), mice were sacrificed and brains hemispheres were separated from each other. The left hemisphere was analyzed for TPP-1 enzyme activity, and the right hemisphere was processed for immunohistochemistry and detection of autoflourescent storage material. Brains treated with AAV2CUhCLN2 contained TPP-1 activity at each of three injection tracts equivalent to that of levels in the brains of heterozygote which are phenotypically normal. In contrast, AAV5CUhCLN2-injected brains had approximately two fold wild-type levels of enzyme activity within these tracts. Immunohistochemistry and confocal microscopy of the right hemispheres showed the presence of TPP-1 protein in the lysosomes and neuronal processes of cells at the injection sites. Compared to control animals (AAV2CUNull, sham, uninjected), there was a marked reduction of autofluorescent storage material with AAV2CUCLN2 and AAV5CUCLN2 in the injected regions which include motor cortex, thalamus, cerebellum, as well as other regions such as the striatum and the hippocampus. There was no statistical difference in the amount of autofluorescent reduction mediated by the two serotype vectors despite the higher levels of enzyme activity with AAV5CUCLN2. Behavioral testing on the rotorod showed improvement of cerebellar motor function at 8 and 10 weeks post-injection with both viral vectors. This study demonstrates that the LINCL brain is responsive to TPP-1 replacement with different AAV serotype vectors, and that heterozygote levels of enzyme activity is sufficient to reverse lysosomal pathology and partially restore behavioral function.