Generation and characterization of a bispecific diabody targeting both EPH receptor A10 and CD3
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Authors: Kamada, Haruhiko; Taki, Shintaro; Nagano, Kazuya; Inoue, Masaki; Ando, Daisuke; Mukai, Yohei; Higashisaka, Kazuma; Yoshioka, Yasuo; Tsutsumi, Yasuo; Tsunoda, Shin-ichi
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. (C) 2014 Elsevier Inc. All rights reserved.
Expression profile of Eph receptors and ephrin ligands in healthy human B lymphocytes and chronic lymphocytic leukemia B-cells
LEUKEMIA RESEARCH
Authors: Alonso-C, Luis M.; Trinidad, Eva M.; de Garcillan, Beatriz; Ballesteros, Monica; Castellanos, Milagros; Cotillo, Ignacio; Munoz, Juan J.; Zapata, Agustin G.
Abstract
Increasing information relates some Eph receptors and their ligands, ephrins (EFN), with the immune system. Herein, we found that normal B-cells from peripheral blood (PB) and lymph nodes (LN) showed a differential expression of certain Eph/EFN members, some of them being modulated upon in vitro stimulation including EFNA1, EFNA4, EphB6 and EphA10. In contrast, PB CLL B-cells showed a more heterogeneous Eph/EFN profile than their normal PB B-cell counterparts, expressing Eph/EFN members frequently found within the LN and activated B-cells, specially EFNA4, EphB6 and EphA10. Two of them, EphB6 and EFNA4 were further related with the clinical course of CLL patients. EphB6 expression correlated with a high content of ZAP-70 mRNA and a poor prognosis. High serum levels of a soluble EFNA4 isoform positively correlated with increasing peripheral blood lymphocyte counts and lymphadenopathy. These findings suggest that Eph/EFN might be relevant in normal B-cell biology and could represent new potential prognostic markers and therapeutic targets for CLL. (c) 2008 Elsevier Ltd. All rights reserved.