Aav-based gene therapy approaches for the treatment of canavan disease

Abstract

Background: The enzyme Aspartoacylase (ASPA) is normally expressed in oligodendrocytes, the myelin-forming cells in the central nervous system (CNS). ASPA gene mutations cause Canavan Disease (CD), a devastating neurological disorder characterized by psychomotor retardation, and spongiform degeneration of central white matter in affected children. The lack of ASPA leads to the enrichment in its substrate N-acetyl aspartate (NAA) which is a biomarker of CD. With no available treatment and a pathology restricted to the CNS CD has been trialled by gene therapy. However, gene replacement approaches using neurotropic recombinant adeno-associated viral (rAAV) vectors have proved unsuccessful. It was shown recently that promoter specificity targets AAV-mediated transgene expression to glia in normal adult rodents but this approach has not been exploited in a disease model. Purpose: Here we aimed at preclinical efficacy in a CD mouse model using AAV vectors with glial tropism. Methods: An AAV2 expression cassette including the myelin basic protein (MBP) promoter to drive expression of ASPA or greenfluorescent protein (GFP) was packaged as rh39 or AAV1/2. Vectors were delivered bilaterally to striatum, thalamus and cerebellum of presymptomatic ASPA-lacZ knock-in mutants or normal littermates at postnatal day (P) 10. Animals were analysed at 4e9 months of age. Viral spread and specificity of transgene expression was determined by immunohistochemistry.Nissl stained sections were used for semi-quantitation of brain vacuolisation and NAA levels were measured by 1H-NMR. Results: Pilot experiments revealed that direct vector injection at P10 yielded robust oligodendroglial specificity and vector spread regardless of the serotype. Ventricle size, vacuolisation, astrocytosis, and NAA concentration did not fully reach wild-type levels but were significantly reduced in ASPA-deficient animals that received AAV-ASPA compared with controls. Conclusion: Timed AAV-mediated ASPA expression in oligoendrocytes using myelin gene promoters may ameliorate CNS dysfunction in CD.