Bicistronic lentiviral vector corrects beta-hexosaminidase deficiency in transduced and cross-corrected human Sandhoff fibroblasts.

Sandhoff disease is an autosomal recessive neurodegenerative disease characterized by a GM2 ganglioside intralysosomal accumulation. It is due to mutations in the β-hexosaminidases β-chain gene, resulting in a β-hexosaminidases A (αβ) and β (ββ) 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 B-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 α- and β-subunits of β-hexosaminidases may provide a potential therapeutic tool for the treatment of Sandhoff disease.