Bispecific anti-CD3 x anti-HER2 antibody mediates T cell cytolytic activity to HER2-positive colorectal cancer in vitro and in vivo

Abstract

Targeting HER2 overexpressed breast cancer cells with anti‑HER2 monoclonal antibodies inhibits tumor growth. Here we investigated whether HER2 can serve as a target for T cell-mediated immunotherapy of human colorectal carcinoma. Specific cytolytic activity of activated T cells (ATCs) armed with anti‑CD3 x anti‑HER2 bispecific antibody (HER2Bi-Ab) against HER2+ tumor cells was evaluated by bioluminescent signal generated by luciferase reporter on tumor cells in vitro and in vivo. In contrast to unarmed ATCs, increased cytotoxic activity of HER2Bi-armed ATCs against HER2+ tumor cells was observed. Moreover, HER2Bi-armed ATCs expressed higher level of activation marker CD69 and secreted significantly higher levels of IFN-γ than the unarmed ATC counterpart. In addition, compared with anti‑HER2 mAb (Herceptin®) or unarmed ATC, HER2Bi-armed ATCs showed significant suppression against colorectal carcinoma cells. In colorectal tumor cell xenograft mice, infusion of HER2Bi-armed ATCs successfully inhibited the growth of Colo205-luc cells. The HER2Bi-armed ATCs with anti-tumor effects may provide a promising immunotherapy for colorectal carcinoma in the future.

Targeting bladder cancer using activated T�cells armed with bispecific antibodies

“…2B. Moreover, evaluation of the binding of Bi-Ab to CD3 + cells has been previously shown by a similar method (27)(28)(29).…”

“…CD3xanti-HER2 (HER2Bi-Ab) bispecific antibodies and arming of ATCs. Anti-EGFR (Erbitux ® ; Merck Serono, Darmstadt, Germany) or anti-HER2 (Herceptin ® ; Roche, Indianapolis, IN, uSA) was reacted with sulfo-SMCC, and anti-CD3 mAb (OKT3; eBioscience) was reacted with Traut's reagents using a previously described method (27)(28)(29). Cryopreserved ATCs were thawed, armed with EGFRBi-Ab or HER2Bi-Ab at a concentration of 50 ng/10 6 cells at room temperature for 30 min, and washed to eliminate unbound antibodies.…”

“…We next constructed two types of bispecific antibodies that recognized both CD3 on T cells and EGFR or HER2 on bladder cancer cells. Erbitux ® or Herceptin ® antibody was chemically hetero-conjugated to OKT3 and named EGFRBi-Ab or HER2Bi-Ab (27)(28)(29). The binding specificity of EGFRBi-Ab and HER2Bi-Ab was tested against EGFR and HER2, respectively.…”

Targeting immunotherapy for bladder cancer using anti‐CD3× B7‐H3 bispecific antibody

“…One is luciferase quantitative assay. [28][29][30][31][32] The other is collecting cell free supernatants, and the cytotoxicity was mostly measured with Lactate dehydrogenase activity kit (Sigma-Aldrich, St. Louis, MO, USA).…”

“…According to the data provided by the literature, 21,[28][29][30] 150 ng/10 6 cells was chosen as the concentration of B7-H3Bi-Ab, ATC mixed with OKT3 and anti-B7-H3 mAb were used as unarmed ATC controls. Both were interacted at 37°C for 18 hours.…”