Control of Recombinant Monoclonal Antibody Effector Functions by Fc N‐Glycan Remodeling in Vitro

Biotechnology Progress - Tập 21 Số 6 - Trang 1644-1652 - 2005
Jason Hodoniczky1,2, Yanbo Zheng1,2, David C. James2
1Contributed equally to this work.
2School of Engineering, University of Queensland, St. Lucia, QLD, Australia 4072

Tóm tắt

Abstract

N‐Glycans at Asn297 in the Fc domain of IgG molecules are required for Fc receptor‐mediated effector functions such as antibody‐dependent cell‐mediated cytotoxicity (ADCC) and complement‐dependent cytotoxicity (CDC). In this study we have specifically remodeled the Fc N‐glycans of intact recombinant IgG1 therapeutic monoclonal antibody (Mab) products, Rituxan and Herceptin, with a soluble recombinant rat β‐1,4‐N‐acetylglucosaminyltransferase III (rGnTIII) produced by baculovirus‐infected insect cells. N‐Glycan remodeling in vitro permitted a controlled and selective transfer of a bisecting β1,4‐linked GlcNAc to the core β‐linked mannose of degalactosylated Mab N‐glycans to yield Mabs varying in bisecting GlcNAc content from 31% to 85%. This was confirmed by analysis of N‐glycans by both normal phase HPLC and MALDI‐MS, the latter yielding the expected mass increase of 203.2 Da with no other oligosaccharide modifications evident. ADCC of remodeled Rituxan and Herceptin Mabs was determined using peripheral blood mononuclear cells as effectors and either CD20+ (SKW6.4 and SU‐DHL‐4) or Her2+ (SKBR‐3) target cells, respectively. A conserved 10‐fold increase in ADCC was observed for both remodeled therapeutic Mabs with high (>80%) bisecting GlcNAc content. In contrast, although the presence of a bisecting GlcNAc had minimal effect on CDC, degalactosylation of Rituxan reduced CDC by approximately half, relative to unmodified (variably galactosylated) control Mab. In summary, our data suggests that in vitro remodeling of therapeutic Mab Fc N‐glycans may be utilized to control the therapeutic efficacy of Mabs in vivo and to offer a more “humanized” glycoform profile for recombinant Mab products.

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Biotechnology Progress

Tập 21 Số 6

1644-1652

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