Preparation and biological activities of anti-HER2 monoclonal antibodies with fully core-fucosylated homogeneous bi-antennary complex-type glycans.

Wataru Tsukimura,Akio Matsuda,Kaoru Takegawa,Kiyoshi Furukawa,Takashi Shirai,Masako Mori,Kenji Osumi,Masaki Kurogochi

doi:10.1080/09168451.2017.1394813

Wataru Tsukimura, Akio Matsuda + Show 6 more

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https://doi.org/10.1080/09168451.2017.1394813

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Abstract

Recently, the absence of a core-fucose residue in the N-glycan has been implicated to be important for enhancing antibody-dependent cellular cytotoxicity (ADCC) activity of immunoglobulin G monoclonal antibodies (mAbs). Here, we first prepared anti-HER2 mAbs having two core-fucosylated N-glycan chains with the single G2F, G1aF, G1bF, or G0F structure, together with those having two N-glycan chains with a single non-core-fucosylated corresponding structure for comparison, and determined their biological activities. Dissociation constants of mAbs with core-fucosylated N-glycans bound to recombinant Fcγ-receptor type IIIa variant were 10 times higher than those with the non-core-fucosylated N-glycans, regardless of core glycan structures. mAbs with the core-fucosylated N-glycans had markedly reduced ADCC activities, while those with the non-core-fucosylated N-glycans had high activities. These results indicate that the presence of a core-fucose residue in the N-glycan suppresses the binding to the Fc-receptor and the induction of ADCC of anti-HER2 mAbs.

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Since the monoclonal antibodies (mAbs) produced in CHO cells contain N-glycans with a mixture of structures in terms of core-fucosylation, β4-galactosylation, β-N-acetylglucosaminylation, and α-mannosylation,25) we first treated them with GST-tagged Endo S immobilized on beads and with Endo D, which, unlike Endo S, hydrolyzes oligomannose-type glycans present in a small quantity.15) The mAbs obtained contained two (±Fucα1→6) GlcNAcβ1→Asn groups
IgG contains two bi-antennary N-glycans attached to each Asn[297] of the heavy chain.28) These N-glycans are considered to be involved in maintaining the threedimensional structure of the Fc-portion of IgG.29) modification of the N-glycans by digestion with several kinds of exoglycosidases or treatment with synthetic inhibitors caused changes in Fc-mediated functions such as C1q-binding, Fc-receptor-binding, and antibody-dependent cellular cytotoxicity (ADCC) activity of IgG.30–33) IgG molecules having Nglycans without a core-fucose residue produced in fucosyltransferase 8-deficient CHO cells had higher ADCC activity than those produced in parent CHO cells,18) indicating that the presence or absence of a core-fucose residue attached to the N-glycan affects the ADCC activity of IgG

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Introduction

View supplementary material Submit your article to this journal View related articles Citing articles: 4 View citing articles. Wang and his group used a chemoenzymatic method for remodeling the N-glycans attached to rituximab produced in Chinese hamster ovary (CHO) cells.14) We used endoβ-N-acetylglucosaminidase S (Endo S) to remove Nglycans from recombinant anti-HER2 mAb produced in silkworm cocoon and to transfer N-glycans with A2, G2, G0, and M3 structures (Supplement 1) to produce a mAb having two N-glycans with homogeneous structures. By using these mAbs, it was shown that their structures influence their binding to a variant Fcγ receptor type IIIa (FcγRIIIa-V158) and their ADCC activities.15)

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