Intermittent enzyme replacement therapy with recombinant human β-galactosidase prevents neuraminidase 1 deficiency

Amanda R Luu,Cara Wong,Vishal Agrawal,Nathan Wise,Britta Handyside,Melanie J Lo,Glenn Pacheco,Jessica B Felix,Alexander Giaramita,Alessandra D'Azzo,Jon Vincelette,Sherry Bullens,Stuart Bunting,Terri M Christianson,Charles M Hague,Jonathan H Lebowitz,Gouri Yogalingam

doi:10.1074/jbc.ra119.010794

Abstract

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 multienzyme 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 overexpression in GM1 gangliosidosis patient fibroblasts coincides with profound secondary NEU1 deficiency. A regimen of intermittent enzyme replacement therapy 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 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 intracerebroventricular enzyme replacement therapy 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.

Highlights

In humans autosomal recessive mutations in the galactosidase b 1 (GLB1) gene give rise to a deficiency of lysosomal b-galactosidase (b-Gal), a glycosidase that participates in the degradation of multiple galactose-containing substrates (1). b-Gal deficiency gives rise to the accumulation of three biochemically distinct classes of substrates in lysosomes: GM1 and GA1 gangliosides, the glycosaminoglycan, keratan sulfate, and oligosaccharides derived from glycoprotein metabolism (2)
We have previously demonstrated that research-grade rhbGal produced in Chinese hamster ovary (CHO) cells and purified in our laboratory exhibits cation-independent mannose-6-phosphate receptor (CI-MPR)–dependent cellular uptake in GM1 gangliosidosis patient fibroblasts (8)
Our results suggest that a chronic enzyme replacement therapy (ERT) dosing approach (Fig. 4), and a stable gene therapy approach (Fig. 6, A, C, and E) can augment b-Gal levels in GM1 gangliosidosis patient fibroblasts, both approaches have the potential to promote secondary neuraminidase 1 (NEU1) deficiency, when b-Gal levels are augmented for prolonged periods

Summary

Introduction

In humans autosomal recessive mutations in the GLB1 gene give rise to a deficiency of lysosomal b-galactosidase (b-Gal), a glycosidase that participates in the degradation of multiple galactose-containing substrates (1). b-Gal deficiency gives rise to the accumulation of three biochemically distinct classes of substrates in lysosomes: GM1 and GA1 gangliosides, the glycosaminoglycan, keratan sulfate, and oligosaccharides derived from glycoprotein metabolism (2). Following 24 h of cellular uptake, lysosome-delivered rhb-Gal–His[6] enzyme activity decays rapidly in PPCA-deficient galactosialidosis patient fibroblasts over a period of 40 days, with a t1/2 corresponding to 0.5 days (Fig. 2C).

Results

Conclusion