Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells

Wook Dong Kim; Miwako Tokunaga; Hiroyuki Ozaki; Takuya Ishibashi; Kohsuke Honda; Hiroyuki Kajiura; Kazuhito Fujiyama; Ryutaro Asano; Izumi Kumagai; Takeshi Omasa; Hisao Ohtake

doi:10.1007/s00253-009-2152-z

Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells

Wook Dong Kim, Miwako Tokunaga, Hiroyuki Ozaki, Takuya Ishibashi, Kohsuke Honda, Hiroyuki Kajiura, Kazuhito Fujiyama, Ryutaro Asano, Izumi Kumagai, Takeshi Omasa, Hisao Ohtake

ENG - Biomolecular Engineering

Research output: Contribution to journal › Article › peer-review

18 Citations (Scopus)

Abstract

The glycosylation pattern of a humanized anti-EGFR×anti-CD3 bispecific single-chain diabody with an Fc portion (hEx3-scDb-Fc) produced by recombinant Chinese hamster ovary cells was evaluated and compared with those of a recombinant humanized anti-IL-8 antibody (IgG1) and human serum IgG. N-Linked oligosaccharide structures were estimated by two-dimensional high-performance liquid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. No sialylation was observed with purified hEx3-scDb-Fc and the anti-IL-8 antibody. From the analysis of neutral oligosaccharides, approximately more than 90% of the N-linked oligosaccharides of hEx3-scDb-Fc and the anti-IL-8 antibody were alpha-1,6-fucosylated. The galactosylated biantennary oligosaccharides comprise over 40% of the total N-linked oligosaccharides in both hEx3-scDb-Fc and the anti-IL-8 antibody. The fully galactosylated biantennary oligosaccharides from hEx3-scDb-Fc and the anti-IL-8 antibody accounted for only 10% of the N-linked; however, more than 20% of the N-linked oligosaccharides were fully galactosylated biantennary oligosaccharides in human serum IgG. The glycosylation pattern of hEx3-scDb-Fc was quite similar to that of the anti-IL-8 antibody.

Original language	English
Pages (from-to)	535-542
Number of pages	8
Journal	Applied Microbiology and Biotechnology
Volume	85
Issue number	3
DOIs	https://doi.org/10.1007/s00253-009-2152-z
Publication status	Published - 2010 Jan

Keywords

Bispecific IgG-like antibody
Bispecific diabody
Chinese hamster ovary cells
Glycosylation
Immunoglobulin G

ASJC Scopus subject areas

Biotechnology
Applied Microbiology and Biotechnology

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Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells. / Kim, Wook Dong; Tokunaga, Miwako; Ozaki, Hiroyuki; Ishibashi, Takuya; Honda, Kohsuke; Kajiura, Hiroyuki; Fujiyama, Kazuhito; Asano, Ryutaro; Kumagai, Izumi; Omasa, Takeshi; Ohtake, Hisao.

In: Applied Microbiology and Biotechnology, Vol. 85, No. 3, 01.2010, p. 535-542.

Research output: Contribution to journal › Article › peer-review

Kim, Wook Dong ; Tokunaga, Miwako ; Ozaki, Hiroyuki ; Ishibashi, Takuya ; Honda, Kohsuke ; Kajiura, Hiroyuki ; Fujiyama, Kazuhito ; Asano, Ryutaro ; Kumagai, Izumi ; Omasa, Takeshi ; Ohtake, Hisao. / Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells. In: Applied Microbiology and Biotechnology. 2010 ; Vol. 85, No. 3. pp. 535-542.

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title = "Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells",

abstract = "The glycosylation pattern of a humanized anti-EGFR×anti-CD3 bispecific single-chain diabody with an Fc portion (hEx3-scDb-Fc) produced by recombinant Chinese hamster ovary cells was evaluated and compared with those of a recombinant humanized anti-IL-8 antibody (IgG1) and human serum IgG. N-Linked oligosaccharide structures were estimated by two-dimensional high-performance liquid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. No sialylation was observed with purified hEx3-scDb-Fc and the anti-IL-8 antibody. From the analysis of neutral oligosaccharides, approximately more than 90% of the N-linked oligosaccharides of hEx3-scDb-Fc and the anti-IL-8 antibody were alpha-1,6-fucosylated. The galactosylated biantennary oligosaccharides comprise over 40% of the total N-linked oligosaccharides in both hEx3-scDb-Fc and the anti-IL-8 antibody. The fully galactosylated biantennary oligosaccharides from hEx3-scDb-Fc and the anti-IL-8 antibody accounted for only 10% of the N-linked; however, more than 20% of the N-linked oligosaccharides were fully galactosylated biantennary oligosaccharides in human serum IgG. The glycosylation pattern of hEx3-scDb-Fc was quite similar to that of the anti-IL-8 antibody.",

keywords = "Bispecific IgG-like antibody, Bispecific diabody, Chinese hamster ovary cells, Glycosylation, Immunoglobulin G",

author = "Kim, {Wook Dong} and Miwako Tokunaga and Hiroyuki Ozaki and Takuya Ishibashi and Kohsuke Honda and Hiroyuki Kajiura and Kazuhito Fujiyama and Ryutaro Asano and Izumi Kumagai and Takeshi Omasa and Hisao Ohtake",

note = "Funding Information: Acknowledgments The present work is partially supported by grants from the NEDO of Japan, the Program for the Promotion of Fundamental Studies in Health Sciences of the NIBIO, and the Grant-in-Aid for Scientific Research of the JSPS. Copyright: Copyright 2010 Elsevier B.V., All rights reserved.",

year = "2010",

month = jan,

doi = "10.1007/s00253-009-2152-z",

language = "English",

volume = "85",

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T1 - Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells

AU - Kim, Wook Dong

AU - Tokunaga, Miwako

AU - Ozaki, Hiroyuki

AU - Ishibashi, Takuya

AU - Honda, Kohsuke

AU - Kajiura, Hiroyuki

AU - Fujiyama, Kazuhito

AU - Asano, Ryutaro

AU - Kumagai, Izumi

AU - Omasa, Takeshi

AU - Ohtake, Hisao

N1 - Funding Information: Acknowledgments The present work is partially supported by grants from the NEDO of Japan, the Program for the Promotion of Fundamental Studies in Health Sciences of the NIBIO, and the Grant-in-Aid for Scientific Research of the JSPS. Copyright: Copyright 2010 Elsevier B.V., All rights reserved.

PY - 2010/1

Y1 - 2010/1

N2 - The glycosylation pattern of a humanized anti-EGFR×anti-CD3 bispecific single-chain diabody with an Fc portion (hEx3-scDb-Fc) produced by recombinant Chinese hamster ovary cells was evaluated and compared with those of a recombinant humanized anti-IL-8 antibody (IgG1) and human serum IgG. N-Linked oligosaccharide structures were estimated by two-dimensional high-performance liquid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. No sialylation was observed with purified hEx3-scDb-Fc and the anti-IL-8 antibody. From the analysis of neutral oligosaccharides, approximately more than 90% of the N-linked oligosaccharides of hEx3-scDb-Fc and the anti-IL-8 antibody were alpha-1,6-fucosylated. The galactosylated biantennary oligosaccharides comprise over 40% of the total N-linked oligosaccharides in both hEx3-scDb-Fc and the anti-IL-8 antibody. The fully galactosylated biantennary oligosaccharides from hEx3-scDb-Fc and the anti-IL-8 antibody accounted for only 10% of the N-linked; however, more than 20% of the N-linked oligosaccharides were fully galactosylated biantennary oligosaccharides in human serum IgG. The glycosylation pattern of hEx3-scDb-Fc was quite similar to that of the anti-IL-8 antibody.

AB - The glycosylation pattern of a humanized anti-EGFR×anti-CD3 bispecific single-chain diabody with an Fc portion (hEx3-scDb-Fc) produced by recombinant Chinese hamster ovary cells was evaluated and compared with those of a recombinant humanized anti-IL-8 antibody (IgG1) and human serum IgG. N-Linked oligosaccharide structures were estimated by two-dimensional high-performance liquid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. No sialylation was observed with purified hEx3-scDb-Fc and the anti-IL-8 antibody. From the analysis of neutral oligosaccharides, approximately more than 90% of the N-linked oligosaccharides of hEx3-scDb-Fc and the anti-IL-8 antibody were alpha-1,6-fucosylated. The galactosylated biantennary oligosaccharides comprise over 40% of the total N-linked oligosaccharides in both hEx3-scDb-Fc and the anti-IL-8 antibody. The fully galactosylated biantennary oligosaccharides from hEx3-scDb-Fc and the anti-IL-8 antibody accounted for only 10% of the N-linked; however, more than 20% of the N-linked oligosaccharides were fully galactosylated biantennary oligosaccharides in human serum IgG. The glycosylation pattern of hEx3-scDb-Fc was quite similar to that of the anti-IL-8 antibody.

KW - Bispecific IgG-like antibody

KW - Bispecific diabody

KW - Chinese hamster ovary cells

KW - Glycosylation

KW - Immunoglobulin G

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