Prestimulation of monocytes by the cytokines GM-CSF or IL-2 enhances the antibody dependent cellular cytotoxicity of monoclonal antibody 17-1A

Previously, we have shown that the cytokines IFN-alpha, IFN-gamma and IL-2 significantly enhance the antibody dependent cellular cytotoxicity (ADCC) exerted by the monoclonal antibody (mAb) 17-1A which recognizes the tumor associated antigen EpCAM. ADCC was assessed by a new flow cytometric cytotoxicity assay using the PKH2 labeled colorectal tumor cell line HT29 as target cells and peripheral blood mononuclear cells as effecters. Monocytes are assumed to be one of the major effectors for ADCC. However, isolated monocytes have a rather low ADCC capacity while addition of CD4(+) lymphocytes optimizes ADCC. Since such an interaction between immune cells may act through cytokines we investigated whether a seven-day-prestimulation of monocytes by the cytokines M-CSF, CM-CSF, IFN-gamma, IFN-alpha and IL-2 enhances ADCC. Thereafter, we added for three days IL-2 and IFN-alpha with or without the mAb 17-1A for terminal activation of monocytes. Interestingly, GM-CSF which was ineffective in terminal activation, significantly enhanced ADCC of monocytes when it was used for prestimulation. Similar results were obtained with IL-2. IFN-gamma and M-CSF were also active but less than GM-CSF. Astonishingly, IFN-gamma and IFN-alpha prestimulation of monocytes suppressed the enhancement of ADCC exerted by CM-CSF and IL-2, respectively. Our experiments suggest that the timing of cytokine application is critical for the induction of optimal ADCC. Subcutaneous pretreatment with CM-CSF or IL-2 followed by the combination of IL- 2/IFN-alpha/17-1A should be evaluated in a phase I clinical trial in patients with colorectal cancer.