Cytokine Induced Killer cells for cell therapy of acute myeloid leukemia: improvement of their immune activity by expression of CD33-specific chimeric receptors

Background. Cytokine induced killer cells are ex-vivo expanded cells with potent antitumoral activity. Cytokine induced killer cells infusion in acute myeloid leukemia patients relapsing after allogeneic hematopoietic stem cell transplant is well tolerated, but limited clinical responses were observed. To improve their effector functions against acute myeloid leukemia, we genetically modified cytokine induced killer cells with chimeric receptors specific for the CD33 myeloid antigen. Design and Methods. SFG-retroviral vectors coding for anti-CD33-zeta and anti-CD33-CD28-OX40-zeta chimeric receptors were used to transduce cytokine induced killer cells. Transduced cells were in vitro characterized for their ability to lyse leukemic targets (4-hour-51Chromium-release and 6-day co-cultures assays on human stromal mesenchymal cells), to proliferate (3H-thymidine-incorporation assay) and to secrete cytokines (Flow cytomix assay) after contact with acute myeloid leukemia cells. Their activity against normal CD34+ hematopoietic progenitors was evaluated by analyzing the colony forming unit capacity after co-incubation. Results. Cytokine induced killer cells were efficiently transduced with the anti-CD33 chimeric receptors, maintaining their native phenotype and functions and acquiring potent cytotoxicity (up to 80% lysis after 4-hour incubation) against different acute myeloid leukemia targets, as also confirmed in long-term killing experiments. Moreover, introduction of the anti-CD33 chimeric receptors was accompanied by a prominent CD33-specific proliferative activity, with a release of high levels of immunostimulatory cytokines. The presence of CD28-OX40 in chimeric receptor endodomain was associated with a significant amelioration of cytokine induced killer cells anti-leukemic activity. Importantly, even though anti-CD33 chimeric receptors-transduced cytokine induced killer cells showed toxicity against normal hematopoietic CD34+ progenitors, a residual clonogenic activity was preserved. Conclusions. Our results indicate that anti-CD33 chimeric receptors strongly enhance anti-leukemic cytokine induced killer functions, suggesting that cytokine induced killer cells transduced with these molecules might represent a promising optimized tool for acute myeloid leukemia immunotherapy