TY - JOUR
T1 - Rearranging the domain order of a diabodybased IgG-like bispecific antibody enhances its antitumor activity and improves its degradation resistance and pharmacokinetics
AU - Asano, Ryutaro
AU - Shimomura, Ippei
AU - Konno, Shota
AU - Ito, Akiko
AU - Masakari, Yosuke
AU - Orimo, Ryota
AU - Taki, Shintaro
AU - Arai, Kyoko
AU - Ogata, Hiromi
AU - Okada, Mai
AU - Furumoto, Shozo
AU - Onitsuka, Masayoshi
AU - Omasa, Takeshi
AU - Hayashi, Hiroki
AU - Katayose, Yu
AU - Unno, Michiaki
AU - Kudo, Toshio
AU - Umetsu, Mitsuo
AU - Kumagai, Izumi
PY - 2014/9/1
Y1 - 2014/9/1
N2 - 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.
AB - 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.
KW - Antibody engineering
KW - Bispecific diabody
KW - CD3
KW - Cancer immunotherapy
KW - Effective domain order
KW - Epidermal growth factor receptor
KW - IgG-like bispecific antibody
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U2 - 10.4161/mabs.29445
DO - 10.4161/mabs.29445
M3 - Article
C2 - 25517309
AN - SCOPUS:84913588010
VL - 6
SP - 1243
EP - 1254
JO - mAbs
JF - mAbs
SN - 1942-0870
IS - 5
ER -