249. Minimal Dose of AAV8/DC190-ASM Necessary for Therapeutic Efficacy in a Mouse Model of Niemann-Pick Disease

Abstract

Niemann-Pick disease type B (NPD) is a congenital defect caused by a deficiency of the lysosomal hydrolase, acid sphingomyelinase (ASM). This panethnic disorder results in the pathological accumulation of sphingomyelin (SPM) in the macrophages of lung, spleen, liver and bone marrow. While a broad continuum of clinical manifestations occurs in NPD patients, significant morbidity results as a consequence of progressively worsening pulmonary involvement. Previously, we reported that systemic administration of 3x1011 drp of a recombinant adeno-associated virus vector (AAV8) encoding human ASM under the transcriptional control of a liver-restricted promoter (DC190) into acid sphingomyelinase knockout mice (ASMKO) resulted in expression and secretion of therapeutic levels of the enzyme, as measured by SPM clearance in visceral tissues. Furthermore, substrate depletion in lung was accompanied by significant reduction in the levels of proinflammatory cytokines and complete reversal of lung pathology. The goal of the present study was to define the minimal dose of AAV8/DC190-ASM necessary for therapeutic efficacy in the ASMKO mice. As expected, administration of lower doses (3X1010 to 1X1011 ) of AAV8/DC190-ASM resulted in expression and secretion of correspondingly lower levels of ASM. Encouragingly, while uptake of ASM was found to be dose-dependent, all doses were capable of substantially reducing SPM levels in the affected tissues, including the lung, by day 60 post-treatment. To assess the kinetics of clearance, ASMKO were administered various doses of AAV8/DC190-ASM and groups were sacrificed at day 14 or day 28 post-administration. Maximal tissue levels of ASM were achieved as early as 14 days post-injection. At the highest dose (3X1011 drp), SPM levels in the liver, spleen, lung and bone marrow were largely normalized by day 14. Importantly, as an extension of our previous observations, we found that decreased SPM levels in the lung coincided with a reduction in the levels of pro-inflammatory mediators (e.g. MIP-1|[alpha]|) in the bronchoalveolar lavages of ASMKO mice. It was also noted that liver-restricted expression of human ASM promoted immune tolerance to the expressed enzyme in the ASMKO mice at all the doses tested. Hence, AAV8-mediated liver-restricted expression of ASM was capable of correcting the visceral storage disease and abrogating the chronic pulmonary inflammatory manifestations shown associated with NPD. Therapeutic efficacy could be attained with a 10-fold lower dose of vector (3X1010 drp/mouse) than previously studied and near-complete correction was observed within two weeks post-administration. Based on these findings, it is anticipated that yet lower doses of AAV8/DC190-ASM may be therapeutically efficacious in the ASMKO mice.