Title : Site-specific analysis of
N-linked oligosaccharides of recombinant lysosomal
arylsulfatase A produced in different cell lines
Abstract :
- Metachromatic leukodystrophy ( MLD ) is a lysosomal storage disease caused by a deficiency of the lysosomal enzyme arylsulfatase A ( ASA )
- Enzyme replacement therapy ( ERT ) is a therapeutic option for MLD and other lysosomal disorders
- This therapy depends on N-linked oligosaccharide-mediated delivery of intravenously injected recombinant enzyme to the lysosomes of patient cells
- Because of the importance of N-linked oligosaccharide side chains in ERT , we examined the com position of the three N-linked glycans of four different recombinant ASAs in a site-specific manner
- Depending on the culture conditions and the cell line expressing the enzyme , we detected a high variability of the high-mannose-type N-glycans which prevail at all glycosylation sites.
- Our data show that the com position of the glycans is largely determined by substantial trimming in the medium
- The susceptibility for trimming is different for the glycans at the three N-glycosylation sites
- Interestingly, which of the glycans is most susceptible to trimming also depends on production conditions
- CHO cells cultured under bioreactor conditions yielded recombinant ASA with the most preserved N-glycan structures, the highest mannose-6-phosphate content and the highest similarity to non-recombinant enzyme
- Notably, roughly one-third of the N-glycans released from the three glycosylation sites were fucosylated
- In the last years, numerous recombinant lysosomal enzymes were used for preclinical ERT trials
- Our data show that the oligosaccharide structures were very different in these trials making it difficult to draw common conclusions from the various investigations