title: Novel Vector Design and Hexosaminidase Variant Enabling Self-Complementary Adeno-Associated Virus for the Treatment of Tay-Sachs Disease.
abstract: GM2 gangliosidosis is a family of three genetic neurodegenerative disorders caused by the accumulation of GM2 ganglioside (GM2) in neuronal tissue. Two of these are due to the deficiency of the heterodimeric (α-β), "A" isoenzyme of lysosomal β-hexosaminidase (HexA). Mutations in the α-subunit (encoded by HEXA) lead to Tay-Sachs disease (TSD), whereas mutations in the β-subunit (encoded by HEXB) lead to Sandhoff disease (SD). The third form results from a deficiency of the GM2 activator protein (GM2AP), a substrate-specific cofactor for HexA. In their infantile, acute forms, these diseases rapidly progress with mental and psychomotor deterioration resulting in death by approximately 4 years of age. After gene transfer that overexpresses one of the deficient subunits, the amount of HexA heterodimer formed would empirically be limited by the availability of the other endogenous Hex subunit. The present study used a new variant of the human HexA α-subunit, μ, incorporating critical sequences from the β-subunit that produce a stable homodimer (HexM) and promote functional interactions with the GM2AP- GM2 complex. We report the design of a compact adeno-associated viral (AAV) genome using a synthetic promoter-intron combination to allow self-complementary (sc) packaging of the HEXM gene. Also, a previously published capsid mutant, AAV9.47, was used to deliver the gene to brain and spinal cord while having restricted biodistribution to the liver. The novel capsid and cassette design combination was characterized in vivo in TSD mice for its ability to efficiently transduce cells in the central nervous system when delivered intravenously in both adult and neonatal mice. This study demonstrates that the modified HexM is capable of degrading long-standing GM2 storage in mice, and it further demonstrates the potential of this novel scAAV vector design to facilitate widespread distribution of the HEXM gene or potentially other similar-sized genes to the nervous system.
authors.affiliation: 1 Gene Therapy Center, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina.
authors.forename: Subha
authors.initials: S
authors.lastname: Karumuthil-Melethil
authors.affiliation: 1 Gene Therapy Center, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina.
authors.forename: Sahana
authors.initials: S
authors.lastname: Nagabhushan Kalburgi
authors.affiliation: 2 Medical Genetics/Departments of Pediatrics, Queen's University , Kingston, Ontario, Canada .
authors.forename: Patrick
authors.initials: P
authors.lastname: Thompson
authors.affiliation: 3 Genetics and Genome Biology, SickKids, Toronto, Ontario, Canada .
authors.forename: Michael
authors.initials: M
authors.lastname: Tropak
authors.affiliation: 4 New Hope Research Foundation , North Oaks, Minnesota.
authors.forename: Michael D
authors.initials: MD
authors.lastname: Kaytor
authors.affiliation: 4 New Hope Research Foundation , North Oaks, Minnesota.
authors.forename: John G
authors.initials: JG
authors.lastname: Keimel
authors.affiliation: 5 Department of Microbiology, University of Manitoba , Winnipeg, Manitoba, Canada .
authors.forename: Brian L
authors.initials: BL
authors.lastname: Mark
authors.affiliation: 3 Genetics and Genome Biology, SickKids, Toronto, Ontario, Canada .|6 Department of Laboratory Medicine and Pathology, University of Toronto, Toronto, Ontario, Canada .
authors.forename: Don
authors.initials: D
authors.lastname: Mahuran
authors.affiliation: 2 Medical Genetics/Departments of Pediatrics, Queen's University , Kingston, Ontario, Canada .
authors.forename: Jagdeep S
authors.initials: JS
authors.lastname: Walia
authors.affiliation: 1 Gene Therapy Center, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina.|7 Department of Ophthalmology, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina.
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Gray
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D000229:Dependovirus / Q000235:genetics*; D004195:Disease Models, Animal; D005260:Female; D005678:G(M2) Ganglioside / Q000378:metabolism; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000008:administration & dosage*; D006596:Hexosaminidases / Q000235:genetics*; D008099:Liver / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation / Q000235:genetics*; D013661:Tay-Sachs Disease / Q000235:genetics / Q000628:therapy*
chemical_list: D005678:G(M2) Ganglioside; D006596:Hexosaminidases