The absence of fucose but not the presence of galactose or bisecting N-acetylglucosamine of human IgG1 complex-type oligosaccharides shows the critical role of enhancing antibody-dependent cellular cytotoxicity.
An anti-human interleukin 5 receptor (hIL-5R) humanized immunoglobulin G1 (IgG1) and an anti-CD20 chimeric IgG1 produced by rat hybridoma YB2/0 cell lines showed more than 50-fold higher antibody-dependent cellular cytotoxicity (ADCC) using purified human peripheral blood mononuclear cells as effector than those produced by Chinese hamster ovary (CHO) cell lines. Monosaccharide composition and oligosaccharide profiling analysis showed that low fucose (Fuc) content of complex-type oligosaccharides was characteristic in YB2/0-produced IgG1s compared with high Fuc content of CHO-produced IgG1s. YB2/0-produced anti-hIL-5R IgG1 was subjected to Lens culinaris aggulutin affinity column and fractionated based on the contents of Fuc. The lower Fuc IgG1 had higher ADCC than the IgG1 before separation. In contrast, the content of bisecting GlcNAc of the IgG1 affected ADCC much less than that of Fuc. In addition, the correlation between Gal and ADCC was not observed. When the combined effect of Fuc and bisecting GlcNAc was examined in anti-CD20 IgG1, only a severalfold increase of ADCC was observed by the addition of GlcNAc to highly fucosylated IgG1. Quantitative PCR analysis indicated that YB2/0 cells had lower expression level of FUT8 mRNA, which codes alpha1,6-fucosyltransferase, than CHO cells. Overexpression of FUT8 mRNA in YB2/0 cells led to an increase of fucosylated oligosaccharides and decrease of ADCC of the IgG1. These results indicate that the lack of fucosylation of IgG1 has the most critical role in enhancement of ADCC, although several reports have suggested the importance of Gal or bisecting GlcNAc and provide important information to produce the effective therapeutic antibody.
Shinkawa
Toyohide
T
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., 3-6-6 Asahi-machi, Machida-shi, Tokyo 194-8533, Japan.
Nakamura
Kazuyasu
K
Yamane
Naoko
N
Shoji-Hosaka
Emi
E
Kanda
Yutaka
Y
Sakurada
Mikiko
M
Uchida
Kazuhisa
K
Anazawa
Hideharu
H
Satoh
Mitsuo
M
Yamasaki
Motoo
M
Hanai
Nobuo
N
Shitara
Kenya
K
D000117:Acetylglucosamine; D000818:Animals; D000920:Antibody-Dependent Cell Cytotoxicity; D001483:Base Sequence; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002460:Cell Line; D002846:Chromatography, Affinity; D006224:Cricetinae; D017931:DNA Primers; D005643:Fucose; D005690:Galactose; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D037102:Lectins; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D014162:Transfection
D017931:DNA Primers; D007074:Immunoglobulin G; D037102:Lectins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; D005643:Fucose; D000117:Acetylglucosamine; D005690:Galactose