CHO 세포 배양에서 Hexose 혼합 비율 조절에 의한 Albumin-erythropoietin의 Sialylation 증대

초록

Sialylation is very important for in vivo half-life oftherapeutic proteins because sialic acids can reduce the ratesof clearance in hepatocyte. Many studies were developed forsupplementing various carbon sources which directly influcencedcellular metabolism and glycosylation. So it is necessaryto investigate interconversion of carbons sources in combinationbecause once they are absorbed into cells and synthesizedinto nucleotide sugars, they can be converted to differentkinds of sugars and nucleotide sugars. Albumin-erythropoietin(Alb-EPO) has three glycan sites with tetra-antennarystructure, so it can be added up to 14 sialic acids. Therefore,we demonstrated the effects of combination ratios of hexose(glucose:galactose:mannose) among the generally used carbonsources essential for cell growth on sialylation of a fusionprotein, Alb-EPO. As a result, there was no negative effect oncell growth except the condition of the highest concentration. The titer of Alb-EPO was increased depending on increasingconcentration of hexoses in order of mannose, glucose, andgalactose and reached up to 1.7-fold in comparison to the control. On the other hand, there was more than 1.5-fold increasein sialylation followed by changed nucleotide sugar pool atthe conditions supplemented with glucose and galactose athigh concentration. In conclusion, our strategy of controllingthe concentrations of available carbon sources offers an effectiveapproach for the improved production of highly sialylatedAlb-EPO without deleterious effect on growth.

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