Non-fucosylated Therapeutic Antibodies: The Next Generation of Therapeutic Antibodies

  • Mitsuo Satoh
  • Shigeru Iida
  • Naoko Yamane-Ohnuki
  • Katsuhiro Mori
  • Yutaka Kanda
  • Reiko Kuni-Kamochi
  • Ryosuke Nakano
  • Harue Imai-Nishiya
  • Akira Okazaki
  • Toyohide Shinkawa
  • Akihito Natsume
  • Rinpei Niwa
  • Kenya Shitara
Conference paper
Part of the Animal Cell Technology: Basic & Applied Aspects book series (ANICELLTECH, volume 15)


Therapeutic antibody IgG1 has two N-linked oligosaccharide chains bound to the Fc region. The oligosaccharides are of the complex biantennary type, composed of a trimannosyl core structure with the presence or absence of core fucose, bisecting N-acetylglucosamine (GlcNAc), galactose, and terminal sialic acid, which gives rise to structural heterogeneity. Both human serum IgG and therapeutic antibodies are well known to be heavily fucosulated. Recently, ­antibody-dependent cellular cytotoxicity (ADCC), a lytic attack on antibody-targeted cells, has been found to be one of the critical effector functions responsible for the clinical efficacy of therapeutic antibodies such as anti-CD20 IgG1 rituximab (RituxanR) and anti-Her2/neu IgG1 trastuzumab (HerceptinR). ADCC is triggered upon the binding of lymphocyte receptors (FcγRs) to the antibody Fc region. The activity is dependent on the amount of fucose attached to the innermost GlcNAc of N-linked Fc oligosaccharide via an α-1,6-linkage, and is dramatically enhanced by a reduction in fucose. Non-fucosylated therapeutic antibodies show more potent efficacy than their fucosylated counterparts both in vitro and in vivo [7, 8, 9, 10, 11, 12, 13, 14], and are not likely to be immunogenic because their carbohydrate structures are a normal component of natural human serum IgG. Thus, the application of non-fucosylated antibodies is expected to be a powerful and elegant approach to the design of the next generation therapeutic antibodies with improved efficacy. In this review, we discuss the importance of the oligosaccharides attached to the Fc region of therapeutic antibodies, especially regarding the inhibitory effect of fucosylated therapeutic antibodies on the efficacy of non-fucosylated counterparts in one medical agent. The impact of completely non-fucosylated therapeutic antibodies on therapeutic fields will be also discussed.


Therapeutic antibody N-linked Fc oligosaccharide core-fucosylation α-1,6-fucosyltransferase (FUT8) knockout Chinese hamster ovary (CHO) ADCC FcγRIIIa binding human plasma IgG 


  1. 1.
    T. W. Rademacher, R. B. Parekh, and R. A. Dwek, Glycobiology, Annu. Rev. Biochem. 57, 785–838 (1988).CrossRefPubMedGoogle Scholar
  2. 2.
    T. Mizuochi, T. Taniguchi, A. Shimizu, and A. Kobata, Structural and numerical variations of the carbohydrate moiety of immunoglobulin G, J. Immunol. 129, 2016–2020 (1982).PubMedGoogle Scholar
  3. 3.
    H. Harada, M. Kamei, Y. Tokumoto, S. Yui, F. Koyama, N. Kochibe, T. Endo, and A. Kobata, Systematic fractionation of oligosaccharides of human immunoglobulin G by serial affinity chromatography on immobilized lectin columns, Anal. Biochem. 164, 374–381 (1987).CrossRefPubMedGoogle Scholar
  4. 4.
    R. Jefferis, Glycosylation of human IgG antibodies: Relevance to therapeutic applications, BioPharm. 14, 19–26 (2002).Google Scholar
  5. 5.
    J. S. de Bono and E. K. Rowinsky, The ErbB receptor family: A therapeutic target for cancer, Trends Mol. Med. 8(Suppl 4), S19–S26 (2002).CrossRefPubMedGoogle Scholar
  6. 6.
    A. Forero and A. F. Lobuglio, History of antibody therapy for non-Hodgkin’s lymphoma, Semin. Oncol. 30, 1–5 (2003).CrossRefPubMedGoogle Scholar
  7. 7.
    A. J. Grillo-Lopez, Rituximab (Rituxan/Mab Thera): The first decade (1993–2003), Expert Rev. Anticancer Ther. 3, 767–769 (2003).CrossRefPubMedGoogle Scholar
  8. 8.
    C. L. Vogel and S. X. Franco, Clinical experience with trastuzumab (Herceptin), Breast J. 9, 452–462 (2003).CrossRefPubMedGoogle Scholar
  9. 9.
    G. Cartron, L. Dacheux, G. Salles, P. Solal-Celigny, P. Bardos, P. Colombat, and H. Watier, Therapeutic activity of humanized anti-CD20 monoclonal antibody and polymorphism in IgG Fc receptor Fc gamma RIIIa gene, Blood 99, 754–758 (2002).CrossRefPubMedGoogle Scholar
  10. 10.
    S. Dall’Ozzo, S. Tartas, G. Paintaud, G. Cartron, P. Colombat, P. Bardos, H. Watier, and G. Thibault, Rituximab-dependent cytotoxicity by natural killer cells: Influence of FCGR3A polymorphism on the concentration-effect relationship, Cancer Res. 64, 4664–4669 (2004).CrossRefPubMedGoogle Scholar
  11. 11.
    J. H. Anolik, D. Campbell, R. E. Felgar, F. Young, I. Sanz, J. Rosenblatt, and R. J. Looney, The relationship of FcγRIIIa genotype to degree of B cell depletion by rituximab in the treatment of systemic lupus erythematosus, Arthritis Rheum. 48, 455–459 (2003).CrossRefPubMedGoogle Scholar
  12. 12.
    W. K. Weng, and R. Levy, Two immunoglobulin G fragment C receptor polymorphisms independently predict response to rituximab in patients with follicular lymphoma, J. Clin. Oncol. 21, 3940–3947 (2003).CrossRefPubMedGoogle Scholar
  13. 13.
    R. Gennari, S. Menard, F. Fagnoni, L. Ponchio, M. Scelsi, E. Tagliabue, F. Castiglioni, L. Villani, C. Magalotti, N. Gibelli, B. Oliviero, B. Ballardini, G. D. Prada, A. Zambelli, and A. Costa, Pilot study of the mechanism of action of preoperative trastuzumab in patients with primary operable breast tumors overexpressing HER2, Clin. Cancer Res. 10, 5650–5655 (2004).CrossRefPubMedGoogle Scholar
  14. 14.
    J. M. Reishert, C. J. Rosensweig, L. B. Faden, and M. C. Dewitz, Monoclonal antibody successes in the clinic, Nat. Biotechnol. 23, 1073–1078 (2005).CrossRefGoogle Scholar
  15. 15.
    M. Baker, Upping the ante on antibodies, Nat. Biotechnol. 23, 1065–1072 (2005).CrossRefPubMedGoogle Scholar
  16. 16.
    J. Baselga and J. Albanell, Mechanism of action of anti-HER2 monoclonal antibodies, Ann. Oncol. 12(Suppl 1), S35–S41 (2001).CrossRefPubMedGoogle Scholar
  17. 17.
    N. L. Berinstein, A. J. Grillo-Lopez, C. A. White, I. Bence-Bruckler, D. Maloney, M. Czuczman, D. Green, J. Rosenberg, P. McLaughlin, and D. Shen, Association of serum rituximab (IDEC-C2B8) concentration and anti-tumor response in the treatment of recurrent low-grade or follicular non-Hodgkin’s lymphoma, Ann. Oncol. 9, 995–1001 (1998).CrossRefPubMedGoogle Scholar
  18. 18.
    M. M. Goldenberg, Trastuzumab, a recombinant DNA-derived humanized monoclonal antibody, a novel agent for the treatment of metastatic breast cancer, Clin. Ther. 21, 309–318 (1999).CrossRefPubMedGoogle Scholar
  19. 19.
    R. L. Shields, A. K. Namenuk, K. Hong, Y. G. Meng, J. Rae, J. Briggs, D. Xie, J. Lai, A. Stadlen, B. Li, J. A. Fox, and L. G. Presta, Resolution mapping of the binding site on human IgG1 for FcγRI, FcγRII, FcγRIII, and FcRn and design of IgG1 variants with improved binding to the FcγR, J. Biol. Chem. 276, 6591–6604 (2001).CrossRefPubMedGoogle Scholar
  20. 20.
    G. A. Lazar, W. Dang, S. Karki, O. Vafa, J. S. Peng, L. Hyun, C. Chan, H. S. Chung, A. Eivazi, S. C. Yoder, J. Vielmetter, D. F. Carmichael, R. J. Hayes, and B. I. Dahiyat, Engineered antibody Fc variants with enhanced effector function, Proc. Natl. Acad. Sci. USA 103, 4005–4010 (2006).CrossRefPubMedGoogle Scholar
  21. 21.
    H. H. van Ojika, L. Bevaart, C. E. Dahle, A. Bakker, M. J. Jansen, M. J. van Vugt, J. G. van de Winkel, and G. J. Weiner, CpG-A and B oligodeoxynucleotides enhance the efficacy of antibody therapy by activating different effector cell population, Cancer Res. 63, 5595–5600 (2003).Google Scholar
  22. 22.
    B. Jahrsdorfer, and G. J. Weiner, Immunostimulatory CpG oligodeoxynucleotides and antibody therapy of cancer, Semin. Oncol. 30, 476–482 (2003).CrossRefPubMedGoogle Scholar
  23. 23.
    J. W. Friedberg, D. Neuberg, J. G. Gribben, D. C. Fisher, C. Canning, M. Koval, C. M. Poor, L. M. Green, J. Daley, R. Soiffer, J. Ritz, and A. S. Freedman, Combination immunotherapy with rituximab and interleukin 2 in patients with relapsed or refractory follicular non-­Hodgkin’s lymphoma, Br. J. Haematol. 117, 828–834 (2002).CrossRefPubMedGoogle Scholar
  24. 24.
    B. Stockmeyer, D. Elsasser, M. Dechant, R. Repp, M. Gramatzki, M. J. Glennie, J. G. van de Winkel, and T. Valerius, Mechanisms of G-CSF- or GM-CSF- stimulated tumor cell killing by Fc receptor-directed bispecific antibodies, J. Immunol. Methods 248, 103–111 (2001).CrossRefPubMedGoogle Scholar
  25. 25.
    R. L. Shields, J. Lai, R. Keck, L. Y. O’Connell, K. Hong, Y. G. Meng, S. H. Weikert, and L. Prest, Lack of fucose on human IgG1 N-linked oligosaccharide improves binding to human FcγRIII and antibody-dependent cellular toxicity, J. Biol. Chem. 277, 26733–26740 (2002).CrossRefPubMedGoogle Scholar
  26. 26.
    T. Shinkawa, K. Nakamura, N. Yamane, E. Shoji-Hosaka, Y. Kanda, M. Sakurada, K. Uchida, H. Anazawa, M. Satoh, M. Yamasaki, N. Hanai, and K. Shitara, 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, J. Biol. Chem. 278, 3466–3473 (2003).CrossRefPubMedGoogle Scholar
  27. 27.
    N. Yamane-Ohnuki, S. Kinoshita, M. Inoue-Urakubo, M. Kusunoki, S. Iida, R. Nakano, M. Wakitani, R. Niwa, M. Sakurada, K. Uchida, K. Shitara, and M. Satoh, Establishment of FUT8 knockout Chinese hamster ovary cells: An ideal host cell line for producing completely defucosylated antibodies with enhanced antibody-dependent cellular cytotoxicity, Biotechnol. Bioeng. 87, 614–622 (2004).CrossRefPubMedGoogle Scholar
  28. 28.
    K. Mori, R. Kuni-Kamochi, N. Yamane-Ohnuki, M. Wakitani, K. Yamano, H. Imai, Y. Kanda, R. Niwa, S. Iida, K. Uchida, K. Shitara, and M. Satoh, Engineering Chinese hamster ovary cells to maximize effector function of produced antibodies using FUT8 siRNA, Biotechnol. Bioeng. 88, 901–908 (2004).CrossRefPubMedGoogle Scholar
  29. 29.
    A. Okazaki, E. Shoji-Hosaka, K. Nakamura, M. Wakitani, K. Uchida, S. Kakita, K. Tsumoto, I. Kumagai, and Shitara, K, Fucose depletion from human IgG1 oligosaccharide enhances binding enthalpy and association rate between IgG1 and FcγRIIIa, J. Mol. Biol. 336, 1239–1249 (2004).CrossRefPubMedGoogle Scholar
  30. 30.
    Y. Kanda, N. Yamane-Ohnuki, N. Sakai, K. Yamano, R. Nakano, M. Inoue, H. Misaka, S. Iida, M. Wakitani, Y. Konno, K. Yano, K. Shitara, S. Hosoi, and M. Satoh, Comparison of cell lines for stable production of fucose-negative antibodies with enhanced ADCC, Biotechnol. Bioeng. 94, 680–688 (2006).CrossRefPubMedGoogle Scholar
  31. 31.
    R. Niwa, E. Shoji-Hosaka, M. Sakurada, T. Shinkawa, K. Uchida, K. Matsushima, R. Ueda, N. Hanai, and K. Shitara, Defucosylated anti-CC chemokine receptor 4 IgG1 with enhanced antibody-dependent cellular cytotoxicity shows potent therapeutic activity to T cell leukemia and lymphoma, Cancer Res. 64, 2127–2133 (2004).CrossRefPubMedGoogle Scholar
  32. 32.
    R. Niwa, S. Hatanaka, E. Shoji-Hosaka, M. Sakurada, Y. Kobayashi, A. Uehara, H. Yokoi, K. Nakamura, and K. Shitara, Enhancement of the antibody-dependent cellular cytotoxicity of low-fucose IgG1 is independent of FcγRIIIa functional polymorphism, Clin. Cancer Res. 10, 6248–6255 (2004).CrossRefPubMedGoogle Scholar
  33. 33.
    R. Niwa, M. Sakurada, Y. Kobayashi, A. Uehara, K. Matsushima, R. Ueda, K. Nakamura, and K. Shitara, Enhanced natural killer cell binding and activation by low-fucose IgG1 antibody results in potent antibody-dependent cellular cytotoxicity induction at lower antigen density, Clin. Cancer Res. 11, 2327–2336 (2005).CrossRefPubMedGoogle Scholar
  34. 34.
    R. Niwa, A. Natsume, A. Uehara, M. Wakitani, S. Iida, K. Uchida, M. Satoh, and K. Shitara, IgG subclass-independent improvement of antibody-dependent cellular cytotoxicity by fucose removal from Asn297-linked oligosaccharides, J. Immunol. Methods 306, 151–160 (2005).CrossRefPubMedGoogle Scholar
  35. 35.
    A. Natsume, M. Wakitani, N. Yamane-Ohnuki, E. Shoji-Hosaka, R. Niwa, K. Uchida, M. Satoh, and K. Shitara, Fucose removal from complex-type oligosaccharide enhances the antibody-dependent cellular cytotoxicity of single-gene-encoded antibody comprising a ­single-chain antibody linked the antibody constant region, J. Immunol. Methods 306, 93–103 (2005).CrossRefPubMedGoogle Scholar
  36. 36.
    R. Jefferis, Glycosylation of recombinant antibody therapeutics, Biotechnol. Prog. 21, 11–16 (2005).CrossRefPubMedGoogle Scholar
  37. 37.
    S. Iida, H. Misaka, M. Inoue, M. Shibata, R. Nakano, N. Yamane-Ohnuki, M. Wakitani, K. Yano, K. Shitara, and M. Satoh, Non fucosylated therapeutic IgG1 antibody can evade the inhibitory effect of serum immunoglobulin G on antibody-dependent cellular cytotoxicity through its high binding to FcγIIIa, Clin. Cancer Res. 12, 2879–2887 (2006).CrossRefPubMedGoogle Scholar
  38. 38.
    M. Satoh, S. Iida, and K. Shitara, Non-fucosylated therapeutic antibodies as next-generation therapeutic antibodies, Expert Opin. Biol. Ther. in press.Google Scholar
  39. 39.
    M. X. Sliwkowski, J. A. Lofgren, G. D. Lewis, T. E. Hotaling, B. M. Fendly, and J. A. Fox, Nonclinical studies addressing the mechanism of action of trastuzumab (Herceptin), Semin. Oncol. 26, 60–70 (1999).PubMedGoogle Scholar
  40. 40.
    G. D. Lewis, L. Figari, B. Fendly, W. L. Wong, P. Carter, C. Gorman, and H. M. Shepard, Differential responses of human tumor cell lines to anti-p185HER2 monoclonal antibodies, Cancer Immunol. Immunother. 37, 255–263 (1993).CrossRefPubMedGoogle Scholar
  41. 41.
    Y. Vugmeyster, and K. Howell, Rituximab-mediated depletion of cynomolgus monkey B cells in vitro in different matrices: Possible inhibitory effect of IgG, Int. Immunopharmacol. 4, 1117–1124 (2004).CrossRefPubMedGoogle Scholar
  42. 42.
    S. Preithner, S. Elm, S. Lippold, M. Locher, A. Wolf, A. J. da Silva, P. A. Baeuerle, and N. S. Prang, High concentrations of therapeutic IgG1 antibodies are needed to compensate for inhibition of antibody-dependent cellular cytotoxicity by excess endogenous immnuno­globulin G, Mol. Immunol. 43, 1183–1193 (2006).CrossRefPubMedGoogle Scholar
  43. 43.
    S. Kamoda, C. Nomura, M. Kinoshita, S. Nishiura, R. Ishikawa, K. Kakehi, N. Kawasaki, and T. Hayakawa, Profiling analysis of oligosaccharides in antibody pharmaceuticals by capillary electrophoresis, J. Chromatogr. A 1050, 211–216 (2004).PubMedGoogle Scholar
  44. 44.
    M. A. Schenerman, J. N. Hope, C. Kletke, J. K. Singh, R. Kimura, E. I. Tsao, and G. Folena-Wasserman, Comparability testing of a humanized monoclonal antibody (SynagisR) to support cell line stability, process validation, and scale-up for manufacturing, Biologicals 27, 203–215 (1999).CrossRefPubMedGoogle Scholar
  45. 45.
    N. Uozumi, S. Yanagidani, E. Miyoshi, Y. Ihara, T. Sakuma, C. X. Gao, T. Teshima, S. Fujii, T. Shiba, and N. Taniguchi, Purification and cDNA cloning of porcine brain GDP-l-Fuc:N-acetyl-beta-d-glucosaminide alpha1,6-fucosyltransferase, J. Biol. Chem. 271, 27810–27817 (1996).CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Mitsuo Satoh
    • 1
  • Shigeru Iida
    • 1
  • Naoko Yamane-Ohnuki
    • 1
  • Katsuhiro Mori
    • 1
  • Yutaka Kanda
    • 1
  • Reiko Kuni-Kamochi
    • 1
  • Ryosuke Nakano
    • 1
  • Harue Imai-Nishiya
    • 1
  • Akira Okazaki
    • 1
  • Toyohide Shinkawa
    • 1
  • Akihito Natsume
    • 1
  • Rinpei Niwa
  • Kenya Shitara
    • 1
  1. 1.Tokyo Research laboratoriesKyowa Hakko Kogyo Co., Ltd.TokyoJapan

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