The analysis of glycosylation is important when analyzing EPO and DPO. As requested in the ICH guidelines (Topic Q6B), the carbohydrate content (neutral sugars, amino sugars and sialic acids) should be determined. In addition, the structure of the carbohydrate chains, the oligosaccharide pattern (antennary profile), the glycosylation site(s) and occupancy should be analyzed.
EPO has three N-glycosylation sites at Asn24, Asn38 and Asn83 and one O-linked site at Ser126. DPO has an additional two N-linked glycosylation sites at Asn residues 30 and 86.
Glycan structures should be characterized and particular attention should be paid to their degree of sialylation, the glycans present at each of the sites and the degree of glycosylation at each site (glycosylation site occupancy).