Kamada Haruhiko, Taki Shintaro, Nagano Kazuya, Inoue Masaki, Ando Daisuke, Mukai Yohei, Higashisaka Kazuma, Yoshioka Yasuo, Tsutsumi Yasuo, Tsunoda Shin-ichi
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 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.
Biochem Biophys Res Commun. 2015 Jan 24;456(4):908-12. doi: 10.1016/j.bbrc.2014.12.030. Epub 2014 Dec 17.
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.
EPH受体A10（EphA10）在乳腺癌中上调，但在健康组织中通常不表达，因此有人认为EphA10可能是癌症治疗的一个有用靶点。本研究报道了一种双抗体，它是一种能结合两种不同靶分子的抗体衍生物，即肿瘤细胞中表达的EphA10和T细胞中表达的CD3，该双抗体表现出T细胞依赖性细胞毒性。这种带有组氨酸标签和FLAG标签链的双抗体在大肠杆菌中表达，并通过液相色谱法以异二聚体（Db-1）和同二聚体（Db-2）形式进行纯化。使用表达EphA10的细胞进行的流式细胞术分析表明，异二聚体的结合活性强于同二聚体。将双抗体添加到外周血单核细胞培养物中会导致T细胞介导的重定向裂解，并且其生物活性与异二聚体双抗体制剂更强的结合活性一致。我们的结果表明，识别EphA10和CD3的双抗体在癌症治疗中可能有一系列潜在应用，例如表达EPH受体A10的乳腺癌，尤其是三阴性乳腺癌。
