Interferon Gamma Secretion of Adaptive and Innate Immune Cells as a Parameter to Describe Leukaemia-Derived Dendritic-Cell-Mediated Immune Responses in Acute Myeloid Leukaemia in vitro

Abstract

<b><i>Introduction:</i></b> Myeloid leukaemic blasts can be converted into leukaemia-derived dendritic cells (DC_leu), characterised by the simultaneous expression of dendritic- and leukaemia-associated antigens, which have the competence to prime and enhance (leukaemia-specific) immune responses with the whole leukaemic antigen repertoire. To display and further specify dendritic cell (DC)- and DC_leu-mediated immune responses, we analysed the interferon gamma (IFNy) secretion of innate and adaptive immune cells. <b><i>Methods:</i></b> DC/DC_leu were generated from leukaemic whole blood (WB) with (blast)modulatory Kit-I (granulocyte-macrophage colony-stimulating factor [GM-CSF] + Picibanil [OK-432]) and Kit-M (GM-CSF + prostaglandin E1) and were used to stimulate T cell-enriched immunoreactive cells. Initiated anti-leukaemic cytotoxicity was investigated with a cytotoxicity fluorolysis assay. Initiated IFNy secretion of T, NK, CIK, and iNKT cells was investigated with a cytokine secretion assay (CSA). IFNy positivity was additionally evaluated with an intracellular cytokine assay (ICA). Recent activation of leukaemia-specific cells was verified through addition of leukaemia-associated antigens (LAA; WT-1 and Prame) <b><i>Results:</i></b> We found Kit-I and Kit-M competent to generate mature DC and DC_leu from leukaemic WB without induction of blast proliferation. Stimulation of immunoreactive cells with DC/DC_leu regularly resulted in an increased anti-leukaemic cytotoxicity and increased IFNy secretion of T, NK, and CIK cells, pointing to the significant role of DC/DC_leu in leukaemia-specific alongside anti-leukaemic reactions. Interestingly, an addition of LAA did not further increase IFNy secretion, suggesting an efficient activation of leukaemia-specific cells. Here, both the CSA and ICA yielded comparable frequencies of IFNy-positive cells. Remarkably, the anti-leukaemic cytotoxicity positively correlated with the IFNy secretion in T^CD3+, T^CD4+, T^CD8+, and NK^CD56+ cells. <b><i>Conclusion:</i></b> Ultimately, the IFNy secretion of innate and adaptive immune cells appeared to be a suitable parameter to assess and monitor the efficacy of in vitro and potentially in vivo acute myeloid leukaemia immunotherapy. The CSA in this regard proved to be a convenient and reproducible technique to detect and phenotypically characterise IFNy-secreting cells. In respect to our studies on DC-based immunomodulation, we were able to display the potential of DC/DC_leu to induce or improve leukaemia-specific and anti-leukaemic activity.