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Chinese hamster ovary cells can produce galactose-α-1,3-galactose antigens on proteins

Nature Biotechnology volume 28, pages 1153–1156 (2010)Cite this article

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An Addendum to this article was published on 06 May 2011

This article has been updated

To the Editor:

Chinese hamster ovary (CHO) cells are widely used for the manufacture of biotherapeutics, in part because of their ability to produce proteins with desirable properties, including 'human-like' glycosylation profiles. For biotherapeutics production, control of glycosylation is critical because it has a profound effect on protein function, including half-life and efficacy. Additionally, specific glycan structures may adversely affect their safety profile. For example, the terminal galactose-α-1,3-galactose (α-Gal) antigen can react with circulating anti α-Gal antibodies present in most individuals1. It is now understood that murine cell lines, such as SP2 or NSO, typical manufacturing cell lines for biotherapeutics, contain the necessary biosynthetic machinery to produce proteins containing α-Gal epitopes2,3,4. Furthermore, the majority of adverse clinical events associated with an induced IgE-mediated anaphylaxis response in patients treated with the commercial antibody Erbitux (cetuximab) manufactured in a murine myeloma cell line have been attributed to the presence of the α-Gal moiety4. Even so, it is generally accepted that CHO cells lack the biosynthetic machinery to synthesize glycoproteins with α-Gal antigens5. Contrary to this assumption, we report here the identification of the CHO ortholog of N-acetyllactosaminide 3-α-galactosyltransferase-1, which is responsible for the synthesis of the α-Gal epitope. We find that the enzyme product of this CHO gene is active and that glycosylated protein products produced in CHO contain the signature α-Gal antigen because of the action of this enzyme. Furthermore, characterizing the commercial therapeutic protein abatacept (Orencia) manufactured in CHO cell lines, we also identified the presence of α-Gal. Finally, we find that the presence of the α-Gal epitope likely arises during clonal selection because different subclonal populations from the same parental cell line differ in their expression of this gene. Although the specific levels of α-Gal required to trigger anaphylaxis reactions are not known and are likely product specific, the fact that humans contain high levels of circulating anti-α-Gal antibodies suggests that minimizing (or at least controlling) the levels of these epitopes during biotherapeutics development may be beneficial to patients. Furthermore, the approaches described here to monitor α-Gal levels may prove useful in industry for the surveillance and control of α-Gal levels during protein manufacture.

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Figure 1: Structural complex of CHO Ggta1 exons 8–9 gene product with UDP-2F-Gal donor and LacNAc acceptor substrate.
Figure 2: MS/MS fragmentation analysis of N-glycan species observed in abatacept.

Change history

  • 12 April 2011

    There is an addendum to this correspondence. Please see the PDF for details.

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Acknowledgements

The Linkage analysis study was supported in part by the National Institutes of Health National Center for Research Resources (NIH/NCRR)-funded grant entitled 'Integrated Technology Resource for Biomedical Glycomics' (grant no. P41 RR018502-01) to the Complex Carbohydrate Research Center. We would like to thank E. Arevalo, S. Wudyka, T. Carbeau and S. Prabhakar for valuable technical assistance and J. Robblee for critical review of the manuscript.

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  1. Momenta Pharmaceuticals, Cambridge, Massachusetts, USA

    Carlos J Bosques, Brian E Collins, James W Meador III, Hetal Sarvaiya, Jennifer L Murphy, Guy DelloRusso, Dorota A Bulik, I-Hsuan Hsu, Nathaniel Washburn, Sandra F Sipsey, James R Myette & Ganesh Venkataraman

  2. Harvard-MIT Division of Health Sciences and Technology, Department of Biological Engineering, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Rahul Raman, Zachary Shriver & Ram Sasisekharan

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Correspondence to Ganesh Venkataraman.

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Competing interests

Momenta Pharmaceuticals is a biotechnology company specializing in the detailed characterization and engineering of complex drugs including glycoprotein therapeutics. Momenta currently does not have marketed protein products but has products in development (http://www.momentapharma.com/). Contributing authors from Momenta (all except R.R., Z.S. & R.S.) hold stocks and stock options in the company.

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Supplementary Figs. 1–5, Supplementary Table 1 and Supplementary Methods (PDF 799 kb)

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Bosques, C., Collins, B., Meador, J. et al. Chinese hamster ovary cells can produce galactose-α-1,3-galactose antigens on proteins. Nat Biotechnol 28, 1153–1156 (2010). https://doi.org/10.1038/nbt1110-1153

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