Acute myeloid leukemia cells fail to activate native human dendritic cells: a potential mechanism of immune evasion

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Natural killer (NK) cells and T cells are considered be important in the immunosurveillance of acute myeloid leukemia (AML).1 NK cells as potent effectors of innate immunity possess efficient immunomodulatory and cytotoxic properties, which essentially contribute to the elimination of AML cells. CD8+ cytotoxic T cells (CTLs) and CD4+ T cells represent important effectors in adaptive antitumor immunity. CD8+ CTLs are able to recognize and destroy AML cells. CD4+ T cells enhance the capacity of dendritic cells (DCs) to induce CD8+ CTLs by the interaction between CD40 on DCs and CD40 ligand on activated CD4+ T cells. In addition, they provide help for the maintenance and expansion of CD8+ CTLs by secreting cytokines such as interleukin (IL)-2 and are able to eradicate tumor cells directly. Owing to their various antitumor effects, NK cells and T cells emerged as promising candidates for immunotherapeutic strategies in AML therapy.1 However, recent studies revealed that human AML cells can efficiently evade the control of these potent immune effector cells. Thus, it has been shown that AML cells downregulate the entire mismatched human leukocyte antigen haplotype to avoid powerful antileukemia effects mediated by donor T cells after haploidentical stem cell transplantation.2 In addition, it has been shown that cytokine production and cytotoxic activity of NK cells can be markedly impaired by AML cells. This effect is mediated by the interaction between the glucocorticoid-induced tumor necrosis factor (TNF) receptor-related protein on NK cells and its AML cell membrane-associated or -soluble ligand.3 More recently, Baessler et al.4 reported that the interaction of CD137 ligand on AML cells and CD137 on NK cells can also efficiently inhibit NK-cell-mediated antitumor effects.

Whereas these reports offer potential explanations how AML cells can evade the control of NK cells and T cells, little is known about the impact of primary AML cells on native human DCs, which have a major role in the activation of innate and adaptive antitumor responses. Recently, we defined 6-sulfo LacNAc (slan)DCs (formerly termed M-DC8+ DCs) as a major subpopulation of proinflammatory myeloid human blood DCs, which mainly differ from other blood DC subsets by their selective phenotype (6-sulfo LacNAc+, CD1c, CD11c+, CD14, CD16+, CD45RA+, C5aR+).5, 6 Functional data revealed that slanDCs can mediate tumor-directed cytotoxicity and efficiently activate tumor-reactive NK cells and CD8+ CTLs.7 To get novel insights into the impact of primary human AML cells on native human DCs, we investigated the capacity of AML cells to modulate the secretion of TNF-α and IL-12 by slanDCs. In previous studies, we have shown that activated slanDCs produce large amounts of TNF-α, which essentially contributes to their tumor-directed cytotoxicity.5, 7 In addition, we have shown that activated slanDCs are principal producers of IL-12, which favors the differentiation of naive CD4+ T cells into Th1 cells and is important in NK cell activation.6, 8

Blood samples were obtained from healthy donors with informed consent. Immunomagnetic isolation of slanDCs (purity: >90%) from peripheral blood mononuclear cells was performed as previously described.6 Primary AML cells were obtained from the bone marrow of patients who had been included into two consecutive multicenter trials (SHG96 and DSIL2003) after informed consent and approval by the institutional review board. The AML samples used for the experiments were classified according to French–American–British subtypes and percentages of myeloblasts: AML1 (M1, 95%), AML2 (M1, 93,5%), AML3 (M1, 93,5%), AML4 (M1, 92%), AML5 (M1, 92,5%), AML6 (M1, 91,5%), AML7 (M1, 90,5%), AML8 (M5a, 92%), AML9 (M5a, 91,5%), AML10 (M5a, 92,5%), AML11 (M5a, 95%). To evaluate whether AML cells influence the cytokine expression of unstimulated slanDCs, freshly isolated DCs (1 × 105 per well) were co-cultivated with AML cells (1 × 104 per well) in round-bottomed 96-well plates. After 24 h, supernatants were collected and the concentration of TNF-α and IL-12p70 was determined by enzyme-linked immunosorbent assay. As shown in Figures 1a and b, AML cells were not able to induce the secretion of TNF-α or IL-12 in slanDCs.

Figure 1

Impact of primary AML cells on cytokine secretion of unstimulated slanDCs. Freshly isolated slanDCs were cultured in the presence or absence of AML cells derived from 10 patients for 24 h. Subsequently, supernatants were collected and the concentration of (a) TNF-α and (b) IL-12p70 was determined by ELISA. Columns represent means ± s.e. of results obtained from three healthy donors. For each donor the mean of triplicate determination was used.

In further experiments, the impact of AML cells on activated slanDCs was investigated. Therefore, freshly isolated slanDCs were plated in round-bottomed 96-well plates at 1 × 105 per well and incubated with AML cells (1 × 104 per well). After 6 h, 1 μg/ml lipopolysaccharide was added for additional 18 h to stimulate cytokine release by slanDCs. Supernatants were collected and the concentration of TNF-α and IL-12p70 was analyzed. As depicted in Table 1, 8 of 10 primary AML cell samples did not alter the secretion of TNF-α and IL-12 by activated slanDCs. Interestingly, two AML cell samples significantly impair the capability of activated slanDCs to produce these cytokines. The evaluated AML cell samples did not secrete TNF-α and IL-12 under these conditions as determined by flow cytometry (data not shown).

Table 1 Impact of primary AML cells on cytokine secretion of stimulated slanDCs

In summary, we showed that none of the 10 evaluated AML cell samples induces the secretion of the cytotoxic effector molecule TNF-α or the immunomodulatory cytokine IL-12 in unstimulated slanDCs. The failure to trigger slanDC activation efficiently can be explained by an insufficient expression of ‘danger’ molecules by AML cells and/or the production of inhibitory molecules. As a consequence, the capacity of slanDCs to mediate tumor-directed cytotoxicity7 and to activate tumor-reactive NK cells8 and T cells6 may be impaired. In addition, we found that two AML cell samples significantly inhibit TNF-α and IL-12 secretion by activated slanDCs. Besides the potential of primary AML cells to evade the control of NK cells and T cells,2, 3, 4 their failure to trigger unstimulated native human DCs as well as their capability to inhibit the cytokine production of activated DCs may represent additional immune escape mechanisms. These various mechanisms may be important in the development and relapse of AML despite immunosurveillance. In addition, they may also have implications for the design of immunotherapeutic strategies for AML.


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The technical assistance of Bärbel Löbel, Karin Günther, Jule Kriegel and Anne Seltmann (all from the Institute of Immunology) is greatly appreciated. We also thank Silke Soucek and Michael Kramer for providing the characteristics of the AML cells. This work was supported by grants from the Deutsche Forschungsgemeinschaft to MB and GE (SFB655, project B2 and B7) and from the Medical Faculty, Technical University of Dresden to RW.

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Correspondence to M Schmitz.

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