title: Bicistronic lentiviral vector corrects beta-hexosaminidase deficiency in transduced and cross-corrected human Sandhoff fibroblasts.
abstract: Sandhoff disease is an autosomal recessive neurodegenerative disease characterized by a GM2 ganglioside intralysosomal accumulation. It is due to mutations in the beta-hexosaminidases beta-chain gene, resulting in a beta-hexosaminidases A (alphabeta) and B (betabeta) deficiency. Mono and bicistronic lentiviral vectors containing the HEXA or/and HEXB cDNAs were constructed and tested on human Sandhoff fibroblasts. The bicistronic SIV.ASB vector enabled a massive restoration of beta-hexosaminidases activity on synthetic substrates and a 20% correction on the GM2 natural substrate. Metabolic labeling experiments showed a large reduction of ganglioside accumulation in SIV.ASB transduced cells, demonstrating a correct recombinant enzyme targeting to the lysosomes. Moreover, enzymes secreted by transduced Sandhoff fibroblasts were endocytosed in deficient cells via the mannose 6-phosphate pathway, allowing GM2 metabolism restoration in cross-corrected cells. Therefore, our bicistronic lentivector supplying both alpha- and beta-subunits of beta-hexosaminidases may provide a potential therapeutic tool for the treatment of Sandhoff disease.
authors.affiliation: Laboratoire de Génétique, Institut Cochin (Université René Descartes Paris 5, INSERM U567, CNRS UMR 8104), 24 rue du faubourg St-Jacques, 75014 Paris, France.
authors.forename: Audrey
authors.initials: A
authors.lastname: Arfi
authors.forename: Christophe
authors.initials: C
authors.lastname: Bourgoin
authors.forename: Luisa
authors.initials: L
authors.lastname: Basso
authors.forename: Carla
authors.initials: C
authors.lastname: Emiliani
authors.forename: Brunella
authors.initials: B
authors.lastname: Tancini
authors.forename: Vanna
authors.initials: V
authors.lastname: Chigorno
authors.forename: Yu-Teh
authors.initials: YT
authors.lastname: Li
authors.forename: Aldo
authors.initials: A
authors.lastname: Orlacchio
authors.forename: Livia
authors.initials: L
authors.lastname: Poenaru
authors.forename: Sandro
authors.initials: S
authors.lastname: Sonnino
authors.forename: Catherine
authors.initials: C
authors.lastname: Caillaud
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D015536:Down-Regulation / Q000502:physiology; D004705:Endocytosis / Q000502:physiology; D005347:Fibroblasts / Q000201:enzymology / Q000821:virology; D005678:G(M2) Ganglioside / Q000096:biosynthesis; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D054818:Hexosaminidase A; D054819:Hexosaminidase B; D006801:Humans; D007527:Isoenzymes / Q000172:deficiency / Q000235:genetics; D016086:Lentivirus / Q000235:genetics; D008247:Lysosomes / Q000201:enzymology / Q000235:genetics; D008360:Mannosephosphates / Q000378:metabolism; D012497:Sandhoff Disease / Q000201:enzymology / Q000235:genetics / Q000628:therapy*; D014161:Transduction, Genetic / Q000379:methods*; D001619:beta-N-Acetylhexosaminidases / Q000172:deficiency* / Q000235:genetics*
chemical_list: D007527:Isoenzymes; D008360:Mannosephosphates; D005678:G(M2) Ganglioside; C027693:mannose-6-phosphate; D054818:Hexosaminidase A; D054819:Hexosaminidase B; D001619:beta-N-Acetylhexosaminidases