A tumor-selective cell surface localization of heat shock protein 70 (Hsp70), the major heat-inducible member of the Hsp70 group, correlates with an increased sensitivity to lysis mediated by human natural killer (NK) cells and, therefore, might be of clinical relevance. With the exception of mammary carcinomas, an Hsp70 plasma membrane expression was found on freshly isolated human biopsy material of colorectal, lung, neuronal, and pancreas carcinomas, liver metastases, and leukemic blasts of patients with acute myelogenous leukemia. Since normal tissues and bone marrow of healthy human individuals do not express Hsp70 on the cell surface, Hsp70 can be considered as a tumor-selective structure in vivo. Furthermore, we demonstrate that autologous, Hsp70-positive leukemic blasts can be killed by NK cells stimulated with low doses of interleukin 2 plus recombinant Hsp70 protein.

The success of hematopoietic stem cell transplantation (HSCT) lies with the ability of the engrafting immune system to remove residual leukemia cells via a graft-versus-leukemia effect (GvL), caused either spontaneously post-HSCT or via donor lymphocyte infusion. GvL effects can also be initiated by allogenic mismatched natural killer cells, antigen-specific T cells, and activated dendritic cells of leukemic origin. The history and further application of this GvL effect and the main mechanisms will be discussed and reviewed in this chapter.

The reciprocal interaction of tumor cells with the immune system is influenced by various members of the tumor necrosis factor (TNF)/TNF receptor (TNFR) family, and recently, glucocorticoid-induced TNFR-related protein (GITR) was shown to stimulate antitumor immunity in mice. However, GITR may mediate different effects in mice and men and impairs the reactivity of human natural killer (NK) cells. Here, we studied the role of GITR and its ligand (GITRL) in human acute myeloid leukemia (AML). Surface expression of GITRL was observed on AML cells in six of seven investigated cell lines, and 34 of 60 investigated AML patients whereas healthy CD34 + cells did not express GITRL. Furthermore, soluble GITRL (sGITRL) was detectable in AML patient sera in 18 of 55 investigated cases. While the presence of GITRL was not restricted to a specific AML subtype, surface expression was significantly associated with monocytic differentiation. Signaling via GITRL into patient AML cells induced the release of TNF and interleukin-10 (IL-10), and this was blocked by the inhibition of mitogen-activated protein kinases extracellular signal-regulated kinase 1/2. Furthermore, triggering GITR by surface-expressed and sGITRL impaired NK cell cytotoxicity and IFN-; production in cocultures with leukemia cells, and NK cell reactivity could be restored by blocking GITR and neutralization of sGITRL and IL-10. Thus, whereas a stimulatory role of the GITR-GITRL system in mouse antitumor immunity has been reported, our data show that in humans GITRL expression subverts NK cell immunosurveillance of AML. Our results provide useful information for therapeutic approaches in AML, which, like haploidentical stem cell transplantation, rely on a sufficient NK cell response.

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.

Expression of CD137 ligand (4-1BBL), a member of the TNF family of proteins, has been reported on several types of APCs, various carcinoma cells, and can be induced on activated T cells. In this study, we report that the soluble ligand was released constitutively at low levels from leukocytes and at higher levels following cellular activation. Release from cells was blocked by addition of a metalloproteinase inhibitor which concomitantly caused the accumulation of 4-1BBL on the cell surface. In addition, we show that a soluble form of 4-1BBL was present at high levels in the sera of some patients with various hematological diseases, but only at low levels in healthy donors. Soluble 4-1BBL was active in that it competed with recombinant 4-1BBL for binding to the 4-1BB receptor and was able to costimulate IL-2 and IFN-γ release from peripheral T cells. These results indicate that the release of soluble 4-1BBL from the cell surface is mediated by one or more sheddases and likely regulates 4-1BB-4-1BBL interactions between cells in vivo. Cleavage of 4-1BBL to an active soluble form would alter both proximal and distal cellular responses, including cell survival and costimulatory or inflammatory responses, that are mediated through the 4-1BB pathway. This, in turn, would likely alter disease progression or outcome.