Tuning a MAb glycan profile in cell culture: Supplementing N-acetylglucosamine to favour G0 glycans without compromising productivity and cell growth

• Published in: Journal of biotechnology 2015-11, Vol.214, p.105-112
• Main Authors: Blondeel, Eric J.M., Braasch, Katrin, McGill, Thomas, Chang, David, Engel, Christina, Spearman, Maureen, Butler, Michael, Aucoin, Marc G.
• Format: Article
• Language: English
• Subjects: Acetylation; Acetylglucosamine - metabolism; Animals; Antibodies, Monoclonal - analysis; Antibodies, Monoclonal - chemistry; Antibodies, Monoclonal - metabolism; Biotechnology; CHO Cells; Complexity; Cricetinae; Cricetulus; Culture Media - chemistry; Culture Media - metabolism; Glucosamine - metabolism; Glucosamine GlcN; Glucose - metabolism; Glycan; Glycosylation; Monoclonal antibodies; N-acetylglucosamine GlcNAc; Nucleotide sugars; Nucleotides - metabolism; Precursors; Protein quality; Proteins; Tuning
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2015.09.014

•Supplementing glucosamine (GlcN) causes mAb glycans to favour G0 types.•Glucosamine severely restricts cell growth in mammalian cells.•N-acetylglucosamine (GlcNAc) produces same effect to glycans as GlcN.•GlcNAc does not restrict cell growth.•Acetylation of GlcN to make GlcNAc restricts cell growth in mammalian cells. Glycosylation is a critical quality attribute of many therapeutic proteins, particularly monoclonal antibodies (MAbs). Nucleotide-sugar precursors supplemented to growth medium to affect the substrate supply chain of glycosylation has yielded promising but varied results for affecting glycosylation. Glucosamine (GlcN), a precursor for N-acetylglucosamine (GlcNAc), is a major component of mammalian glycans. The supplementation of GlcN to CHO cells stably-expressing a chimeric heavy-chain monoclonal antibody, EG2-hFc, reduces the complexity of glycans to favour G0 glycoforms, while also negatively impacting cell growth. Although several researchers have examined the supplementation of glucosamine, no clear explanation of its impact on cell growth has been forthcoming. In this work, the glucosamine metabolism is examined. We identified the acetylation of GlcN to produce GlcNAc to be the most likely cause for the negative impact on growth due to the depletion of intracellular acetyl-CoA pools in the cytosol. By supplementing GlcNAc in lieu of GlcN to CHO cells producing EG2-hFc, we achieve the same shift in glycan complexity with marginal impacts on the cell growth and protein production.