Rearranging the domain order of a diabodybased IgG-like bispecific antibody enhances its antitumor activity and improves its degradation resistance and pharmacokinetics

Ryutaro Asano, Ippei Shimomura, Shota Konno, Akiko Ito, Yosuke Masakari, Ryota Orimo, Shintaro Taki, Kyoko Arai, Hiromi Ogata, Mai Okada, Shozo Furumoto, Masayoshi Onitsuka, Takeshi Omasa, Hiroki Hayashi, Yu Katayose, Michiaki Unno, Toshio Kudo, Mitsuo Umetsu, Izumi Kumagai

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

One approach to creating more beneficial therapeutic antibodies is to develop bispecific antibodies (bsAbs), particularly IgG-like formats with tetravalency, which may provide several advantages such as multivalent binding to each target antigen. Although the effects of configuration and antibody-fragment type on the function of IgG-like bsAbs have been studied, there have been only a few detailed studies of the influence of the variable fragment domain order. Here, we prepared four types of hEx3-scDb-Fc, IgG-like bsAbs, built from a single-chain hEx3-Db (humanized bispecific diabody [bsDb] that targets epidermal growth factor receptor and CD3), to investigate the infl uence of domain order and fusion manner on the function of a bsDb with an Fc fusion format. Higher cytotoxicities were observed with hEx3-scDb-Fcs with a variable light domain (VL)-variable heavy domain (VH) order (hEx3-scDb-Fc-LHs) compared with a VH-VL order, indicating that differences in the Fc fusion manner do not affect bsDb activity. In addition, fl ow cytometry suggested that the higher cytotoxicities of hEx3-scDb-Fc-LH may be attributable to structural superiority in cross-linking. Interestingly, enhanced degradation resistance and prolonged in vivo half-life were also observed with hEx3-scDb-Fc-LH. hEx3-scDb-Fc-LH and its IgG2 variant exhibited intense in vivo antitumor effects, suggesting that Fc-mediated effector functions are dispensable for effective anti-tumor activities, which may cause fewer side effects. Our results show that merely rearranging the domain order of IgG-like bsAbs can enhance not only their antitumor activity, but also their degradation resistance and in vivo half-life, and that hEx3-scDb-Fc-LHs are potent candidates for next-generation therapeutic antibodies.

Original languageEnglish
Pages (from-to)1243-1254
Number of pages12
JournalmAbs
Volume6
Issue number5
DOIs
Publication statusPublished - 2014 Sep 1

Keywords

  • Antibody engineering
  • Bispecific diabody
  • CD3
  • Cancer immunotherapy
  • Effective domain order
  • Epidermal growth factor receptor
  • IgG-like bispecific antibody

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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