27. Therapeutic Efficacy of Intracranial and Intrathecal AAV2/9-PPT1 in Infantile Batten Disease
Authors: Charles Shyng, Hemanth R. Nelvagal, Josh T. Dearborn, Jonathan D. Cooper, Mark S. Sands
Journal: Molecular Therapy (2016)

Background The neuronal ceroid lipofuscinoses (NCLs) are a group of the most common pediatric neurodegenerative lysosomal storage disorders. Infantile NCL (INCL), caused by a deficiency in the lysosomal enzyme palmitoyl-protein thioesterase-1 (PPT1), is characterized clinically by progressive cognitive and motor decline, profound neurodegeneration and neuroinflammation, and accumulation of autofluorescent storage material (AFSM). Infantile NCL murine model recapitulates the human disease. AAV2/5-PPT1 intracranial (IC) delivery delayed the onset of INCL histopathological markers in the forebrain and cerebellum and improved preclinical outcome measures. However, overall disease progression was only partially corrected suggesting peripheral nervous system involvement. In collaboration with Dr. Jon Cooper (King's College, London), we discovered substantial progressive pathology in the spinal cord: neuronal loss and axon density, significant microgliosis and astrocytosis, and AFSM (Nelvagal H etal, manuscript in prep). These data suggest that the spinal cord could be an important therapeutic target. We hypothesize that IC and intrathecal (IT) gene therapy in combination will significantly improve the lifespan, preclinical outcome measures, and histopathological markers as compared to either therapy alone. Methods We generated five groups (n=10): PPT1-/-, wild type, and PPT1-/- injected with IC, IT, or the combination IC/IT AAV2/9-PPT1. For IC injections, 3-2µl bilateral intracranial injections were performed. For IT injections, one 15µl bolus injection into the lumbar subarachnoid space was performed. The AAV2/9-PPT1 virus was diluted to 1×1012 viral particles/ml. To date, we have collected 3, 5, and 7-month time points for all groups, and have generated a 9-month time point. Samples will be analyzed for PPT1 enzyme activity, AFSM, neuroinflammation and neurohistopathology, spinal cord pathology, and a histochemical stain for PPT1. Lifespan, behavior, and brain weight (gross measure of atrophy) will be analyzed. Significance was determined using a 2-way ANOVA test. Results PPT1-/- mice have a median lifespan of 35.8 weeks and rapid decline in rotarod performance beginning at 5 months. There is a progressive decline in PPT1-/- brain weight beginning at 3 months. IT AAV2/9-PPT1 mice have a median lifespan of 48.4 weeks and have a steady decline in rotarod performance beginning at 7 months. There is a progressive decline in the IT mice brain weight compared to wild-type, reaching significance at 7 months (p<. 001); however, it had significantly less atrophy than PPT1-/- brains until 7 months (p<0.05). IC AAV2/9-PPT1 mice have a median lifespan of 58.5 weeks and a rapid decline in rotarod performance beginning at 9 months. IC AAV2/9-PPT1 mice brain weight are not significantly different than wild-type. To date, at 66 weeks, all IC/IT AAV2/9-PPT1 mice are alive. There is a significant decline in IC/IT mice rotarod performance at 15 months. The IC/IT mice brain weight is not significantly different than wild-type. Data for the enzyme activity, neuroinflammatory markers, histopathology, and histochemical stain will be complete by April 2016. Conclusions To date, these data confirm that targeting the entire CNS will provide a significant step for INCL therapy. The combination therapy significantly increases the lifespan beyond that of an additive benefit. As expected, modifying the gene therapy vector from IC-AAV2/5 to IC-AAV2/9 significantly improved preclinical outcome measures. Lastly, the IT AAV2/9-PPT1 injections suggest that spinal cord disease plays an important role in INCL pathogenesis. These findings could form the basis for an effective therapeutic strategy that incorporates targeting multiple facets of INCL disease.