Autosomal recessive mutations in the galactosidase beta 1 (GLB1) gene cause lysosomal βgalactosidase (Beta-Gal) deficiency, resulting in accumulation of galactose-containing substrates and onset of the progressive and fatal neurodegenerative lysosomal storage disease, GM1 gangliosidosis. Here, an enzyme replacement therapy (ERT) approach in fibroblasts from GM1 gangliosidosis patients with recombinant human Beta-Gal (rhBeta-Gal) produced in Chinese hamster ovary cells enabled direct and precise rhBeta-Gal delivery to acidified lysosomes. A single, low dose (3 nM) of rhBeta-Gal was sufficient for normalizing Beta-Gal activity and mediating substrate clearance for several weeks. We found that rhBeta-Gal uptake by the fibroblasts is dose dependent and saturable, and can be competitively inhibited by mannose-6-phophate, suggesting cation-independent, mannose-6phophate receptor-mediated endocytosis from the cell-surface. A single intracerebroventricular (ICV)-administered dose of rhBeta-Gal (100 micrograms) resulted in broad bilateral ____________________________ GM1 gangliosidosis exhibits an autosomal recessive mode of inheritance and arises from http://www.jbc.org/cgi/
Mutations in the galactosidase β 1 (GLB1) gene cause lysosomal β-galactosidase (β-Gal) deficiency and clinical onset of the neurodegenerative lysosomal storage disease, GM1 gangliosidosis. β-Gal and neuraminidase 1 (NEU1) form a multi-enzyme complex in lysosomes along with the molecular chaperone, protective protein cathepsin A (PPCA). NEU1 is deficient in the neurodegenerative lysosomal storage disease sialidosis, and its targeting to and stability in lysosomes strictly depend on PPCA. In contrast, β-Gal only partially depends on PPCA, prompting us to investigate the role that β-Gal plays in the multienzyme complex. Here, we demonstrate that β-Gal negatively regulates NEU1 levels in lysosomes by competitively displacing this labile sialidase from PPCA. Chronic cellular uptake of purified recombinant human β-Gal (rhβ-Gal) or chronic lentiviral-mediated GLB1 over-expression in GM1 gangliosidosis patient fibroblasts coincides with profound secondary NEU1 deficiency. A regimen of intermittent enzyme replacement therapy (ERT) dosing with rhβ-Gal, followed by enzyme withdrawal, is sufficient to augment β-Gal activity levels in GM1 gangliosidosis patient fibroblasts without promoting NEU1 deficiency. In the absence of β-Gal, NEU1 levels are elevated in the GM1 gangliosidosis mouse brain, which are restored to normal levels following weekly intracerebroventricular (ICV) dosing with rhβ-Gal. Collectively, our results highlight the need to carefully titrate the dose and dosing frequency of β-Gal augmentation therapy for GM1 gangliosidosis. They further suggest that intermittent ICV-ERT dosing with rhβ-Gal is a tunable approach that can safely augment β-Gal levels while maintaining NEU1 at physiological levels in the GM1 gangliosidosis brain.
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