title: Generation and characterization of a bispecific diabody targeting both EPH receptor A10 and CD3.
abstract: The EPH receptor A10 (EphA10) is up-regulated in breast cancer but is not normally expressed in healthy tissue, thus it has been suggested that EphA10 may be a useful target for cancer therapy. This study reports a diabody, an antibody derivative binding two different target molecules, EphA10 expressed in tumor cells and CD3 expressed in T cells, which showed T cell dependent-cytotoxicity. The diabody, which has His-tagged and FLAG-tagged chains, was expressed in Escherichia coli and purified in both heterodimer (Db-1) and homodimer (Db-2) formulations by liquid chromatography. Flow cytometry analysis using EphA10-expressing cells showed that binding activity of heterodimers was stronger than that of homodimers. Addition of diabodies to PBMC cultures resulted in T-cell mediated redirected lysis, and the bioactivity was consistent with the stronger binding activity of heterodimeric diabody formulations. Our results indicate that diabodies recognizing both EphA10 and CD3 could have a range of potential applications in cancer therapy, such as breast cancers that express the EPH receptor A10, especially triple negative breast cancer.
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; The Center for Advanced Medical Engineering and Informatics, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Kamada
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; Laboratory of Biomedical Innovation, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Shintaro
authors.initials: S
authors.lastname: Taki
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan.
authors.forename: Kazuya
authors.initials: K
authors.lastname: Nagano
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Inoue
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; Laboratory of Biomedical Innovation, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Ando
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; Laboratory of Innovative Antibody Engineering and Design (iAED), Center for Drug Innovation and Screening, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Mukai
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; Laboratory of Toxicology and Safety Science, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Kazuma
authors.initials: K
authors.lastname: Higashisaka
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; The Center for Advanced Medical Engineering and Informatics, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan; Laboratory of Toxicology and Safety Science, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Yoshioka
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; The Center for Advanced Medical Engineering and Informatics, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan; Laboratory of Innovative Antibody Engineering and Design (iAED), Center for Drug Innovation and Screening, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; Laboratory of Toxicology and Safety Science, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Tsutsumi
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan; The Center for Advanced Medical Engineering and Informatics, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan; Laboratory of Biomedical Innovation, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan; Laboratory of Innovative Antibody Engineering and Design (iAED), Center for Drug Innovation and Screening, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka 567-0085, Japan. Electronic address: tsunoda@nibio.go.jp.
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Tsunoda
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis* / Q000276:immunology*; D017252:CD3 Complex / Q000276:immunology*; D045744:Cell Line, Tumor; D003602:Cytotoxicity, Immunologic; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D051379:Mice; D011485:Protein Binding; D036081:Receptors, Eph Family / Q000276:immunology*; D014162:Transfection
chemical_list: D018033:Antibodies, Bispecific; D017252:CD3 Complex; C568447:EPHA10 protein, human; D036081:Receptors, Eph Family
keywords: Bispecific antibody; CD3; Diabody; EPHA10; Redirected T cell response