Conclusion
We found the glycosylation heterogeneity of hyperglycosylated
recombinant human interferon-β (rhIFN-β), R27T. The glycosylation site
occupancy showed approx. 94% double-glycosylated form R27T with 6%
single-glycosylated R27T. Microheterogeneity of R27T N-glycans had a
mixture of bi-, tri-, and tetra-antennary glycans, some with lactosamine
extensions, but neither outer arm fucose nor α-galactose was detected.
Sialic acid major variants, Neu5Ac and Neu5Gc were more abundant in R27T
than in Rebif. The major N-glycan, accounting for ~42%
of total N-glycans, had a di-sialylated, core-fucosylated biantennary
structure. These findings could assist the development of
second-generation rhIFN-β products, and help us better understand
glycosylation CQAs and their biological significance.