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영어

Glycosylation plays important roles m a therapeutic glycoprotein such as stability, efficiency, and pharmacokinetics. Especially, sialic acid expressed as an outer terminal unit on the glycoprotein is involved in half-life in serum. In particular, Neu5Gc, a non-human sialic acid, is often observed in therapeutic glycoproteins produced from mammalian cell lines. And exogenous Neu5Gc can be an immunogenic antigen in human cells. Therefore, the screening and quantitation of Neu5Gc in a therapeutic glycoprotein is highly required in the manufacturing and development processes. In this study, we developed an analytical method for selectively identifying and quantifying Neu5Gc with high sensitvity usmg mass spectrometry. The amount of Neu5Gc isolated from various mAb-based therapeutics such as infliximab and trastuzumab was correctly determined. Briefly, sialic acids were liberated from each therapeutic glycoprotein by chmical hydrolysis and further enriched using solid phases extraction with a PGC cartridge. The Neu5Gc was chromatographically separated on a PGC column, then analyzed by MRM-MS. The concentration of Neu5Gc from mAb-based therapeutics was determined at low nano mole levels with high reproducibility ( CV < 6%). Batch-to-batch variations of Neu5Gc from a therapeutic glycoprotein were quatitatively determined. This method will be a valuable platform for not only QbD processes of biotherapeutics but also QA/QC of products to assess impurites and immunogenecities