157P Adoptive Cell Therapy with Cytokine-Induced Killer Cells Retargeted with Immunotools Against HER-2-expressing Breast Cancer

Cytokine-Induced Killer (CIK) cells are CD3+ CD56+ effector cells endowed with T and NK cells phenotypic and functional properties, easy to expand in clinically relevant numbers and able to exert an antibody-dependent cell-mediated cytotoxicity (ADCC) if combined with monoclonal antibodies (mAbs). In the present study, we evaluated the enhancement of CIK cell antigen-specific lytic activity in combination either with the bispecific antibody (bsAb) HER2xCD3, or with the engineered mAb Trastuzumab (TRS) V90Lec13. CIK cells were expanded from PBMCs of healthy donors by the addition of IFNγ, OKT3 and IL-2. The calcium flux of CIK cells and the immunotools binding proprieties was assessed by flow cytometry. The effector cell cytotoxicity and the immunotools dose-dependent activity were evaluated in a 4-hours Calcein-AM assay or in a 72-hours real-time cell analysis (XCELLigence) against HER-2-expressing breast cancer cell lines, together with the measured of the cytokines released in multiplex assay. The bsAb biodistribution was evaluated in vivo in NSG mice. HER2xCD3 bind exclusively to HER-2+ cell lines and to CIK cells eliciting a potent calcium efflux. The combination of CIK cells with both immunotools resulted in a significant improvement of the antigen-specific cytotoxic activity against breast cancer cell lines when compared to the combination of CIK cells with clinical mAb TRS. More important, the combination showed a remarkable cytotoxicity that completely restrains target cells growth even at a very low effector/target (E/T) ratio (0.1:1 E/T) and at very low immunotool concentrations. Moreover, cytokines released by CIK cells matched with a proinflammatory profile but are not correlated with the possible cytokines release syndrome (CRS). Interestingly, TRS-resistant tumor cell lines showed to be sensitive to HER2xCD3-redirected CIK cell lytic activity. Moreover, the bsAb efficiently accumulated in the tumor site, reaching the maximum concentration 8 hours after i.v. injection. Taken together, these results highlight the potentiality of using recombinant immunotools to improve the antigen-specific cytotoxic activity of CIK cells against HER-2-positive tumor cells.