Dendritic Cells (DCs) Can Be Successfully Generated From Leukemic Blasts in Individual Patients With AML or MDS

Abstract

Myeloid-leukemic cells (AML, MDS, CML) can be differentiated to leukemia-derived dendritic cell [DC (DCleu)] potentially presenting the whole leukemic antigen repertoire without knowledge of distinct leukemia antigens and are regarded as promising candidates for a vaccination strategy. We studied the capability of 6 serum-free DC culture methods, chosen according to different mechanisms, to induce DC differentiation in 137 cases of AML and 52 cases of MDS. DC-stimulating substances were cytokines ("standard-medium", "MCM-Mimic", "cytokine-method"), bacterial lysates ("Picibanil"), double-stranded RNA ["Poly (I:C)"] or a cytokine bypass method ("Ca-ionophore"). The quality/quantity of DC generated was estimated by flow cytometry studying (co) expressions of "DC"antigens, costimulatory, maturation, and blast-antigens. Comparing these methods on average 15% to 32% DC, depending on methods used, could be obtained from blast-containing mononuclear cells (MNC) in AML/MDS cases with a DC viability of more than 60%. In all, 39% to 64% of these DC were mature; 31% to 52% of leukemic blasts could be converted to DCleu and DCleu-proportions in the suspension were 2% to 70% (13%). Average results of all culture methods tested were comparable, however not every given case of AML could be differentiated to DC with 1 selected method. However performing a pre-analysis with 3 DC-generating methods (MCM-Mimic, Picibanil, Ca-ionophore) we could generate DC in any given case. Functional analyses provided proof, that DC primed T cells to antileukemia-directed cytotoxic cells, although an anti-leukemic reaction was not achieved in every case. In summary our data show that a successful, quantitative DC/DCleu generation is possible with the best of 3 previously tested methods in any given case. Reasons for different functional behaviors of DC-primed T cells must be evaluated to design a practicable DC-based vaccination strategy.

Cytokine Release Patterns in Mixed Lymphocyte Culture (MLC) of T-Cells with Dendritic Cells (DC) Generated from AML Blasts Contribute to Predict anti-Leukaemic T-Cell Reactions and Patients’ Response to Immunotherapy

“…DLIs can be applied therapeutically for treatment of relapse after SCT or prophylactically in order to stabilize remissions -indicating again the central role of T-cells (Roddie et al 2006;Barrett & Savani 2006;Schmid et al 2007). For an improvement of therapy of AML-/MDS-pts at relapse after SCT GM-CSF might be applied in the context of DLI in order to facilitate the generation of APC from leukaemic blasts and thereby to increase antileukaemic activities of donor T-cells against myeloid blasts, as already shown by others and us in ex vivo settings (Kufner et al 2005;Kremser et al 2010;Schmid et al 2011;Dreyßig et al 2011). We could already show that compared to 'MNC'-stimulation stimulation of T-cells with 'DC'/DC leu , as effective APCs presenting the complete leukaemic antigenic repertoire, gives rise to a better mediation of an antileukaemic T-cell response in vitro thereby potentially contributing to a better response to DLI after allogeneic SCT in vivo (Kufner et al 2005;Kremser et al 2010;Grabrucker et al 2010;Dreyßig et al 2011;Buhmann et al 2016).…”

“…Proportions of positive events in defined gates compared with the isotype controls were calculated using Cell Quest software (Becton Dickinson, BD, Heidelberg, Germany). Quantification and characterization of DC and DC leu were performed by Flow cytometry according to our gating strategy already described by former groups (Schmetzer et al 2007;Kremser et al 2010;Liepert et al 2010;Grabrucker et al 2010;Dreyßig et al 2011). Flow cytometric analyses were carried out on day 0 and 10 followed by a functional cytotoxic fluorolysis assay on day 10 as described below.…”

“…We and others could generate leukaemia derived DC (DC leu ) by transforming leukaemic cells from every given AML-patient in vitro directly to mature APC co-presenting 'DC'-typical antigens (DCA, e.g. CD40, CD86) using the best of three previously tested DC-generating methods (Choudhury et al 1999;Stripecke et al 2002;Kremser et al 2010;Dreyßig et al 2011;Schmetzer & Schmid 2012). Accordingly, we could show that autologous T-cells obtained from AML-pts or allogeneic donor T-cells can be stimulated by DC leu resulting in a very efficient lysis of naive blasts.…”

The characterization and role of leukemia cell-derived dendritic cells in immunotherapy for leukemic diseases

“…In all, 31% to 52% of leukemic blasts can be converted to leukemia-derived DCs and 39% to 64% of these DCs are mature (38,39). Lymphocytic leukemia cells can be also induced to differentiate into leukemic DCs with phenotypical and functional characterization of DCs (40).…”

“…Lymphocytic leukemia cells can be also induced to differentiate into leukemic DCs with phenotypical and functional characterization of DCs (40). At the current point in time, 2 to 5 cytokine combination profiles, consisting of GM-CSF, TNF, IL-4, CD40L, SCF, FL, and TGF-α, are mainly used to induce leukemic cells to differentiate (39,56) (Table 1). Methods of inducing leukemic cells to differentiate into DCs still need further improvement.…”