Characterization of Interleukin-15-Transpresenting Dendritic Cells for Clinical Use.

Personalized dendritic cell- (DC-) based vaccination has proven to be safe and effective as second-line therapy against various cancer types. In terms of overall survival, there is still room for improvement of DC-based therapies, including the development of more immunostimulatory DC vaccines. In this context, we redesigned our currently clinically used DC vaccine generation protocol to enable transpresentation of interleukin- (IL-) 15 to IL-15Rßγ-expressing cells aiming at boosting the antitumor immune response. In this study, we demonstrate that upon electroporation with both IL-15 and IL-15Rα-encoding messenger RNA, mature DC become highly positive for surface IL-15, without influencing the expression of prototypic mature DC markers and with preservation of their cytokine-producing capacity and their migratory profile. Functionally, we show that IL-15-transpresenting DC are equal if not better inducers of T-cell proliferation and are superior in tumor antigen-specific T-cell activation compared with DC without IL-15 conditioning. In view of the clinical use of DC vaccines, we evidence with a time- and cost-effective manner that clinical grade DC can be safely engineered to transpresent IL-15, hereby gaining the ability to transfer the immune-stimulating IL-15 signal towards antitumor immune effector cells.

Medical costs of treatment and survival of patients with acute myeloid leukemia in Belgium.

The advent of new cell-based immunotherapies for leukemia offers treatment possibilities for certain leukemia subgroups. The wider acceptability of these new technologies in clinical practice will depend on its impact on survival and costs. Due to the small patient groups who have received it, these aspects have remained understudied. This non-randomized single-center study evaluated medical costs and survival for acute myeloid leukemia between 2005 and 2010 in 50 patients: patients treated with induction and consolidation chemotherapy (ICT) alone; patients treated with ICT plus allogeneic hematopoietic stem cell transplantation (HCT), which is the current preferred post-remission therapy in patients with intermediate- and poor-risk AML with few co-morbidities, and patients treated with ICT plus immunotherapy using autologous dendritic cells (DC) engineered to express the Wilms' tumor protein (WT1). Total costs including post- consolidation costs on medical care at the hematology ward and outpatient clinic, pharmaceutical prescriptions, intensive care ward, laboratory tests and medical imaging were analyzed. Survival was markedly better in HCT and DC. HCT and DC were more costly than ICT. The median total costs for HCT and DC were similar. These results need to be confirmed to enable more thorough cost-effectiveness analyses, based on observations from multicenter, randomized clinical trials and preferably using quality-adjusted life-years as an outcome measure.

Natural killer cell immune escape in acute myeloid leukemia.

As central players of the innate immune system, natural killer (NK) cells can exert direct and indirect anti-tumor effects via their cytotoxic and immune regulatory capacities, pivotal in the induction of an effective adaptive anti-tumor immune response. Hence, NK cells are considered to be important in the immune surveillance of cancer. In acute myeloid leukemia (AML) patients, however, significantly impaired NK cell functions can facilitate escape from immune surveillance and affect patient outcome. Here, we review various NK cell defects and AML evasion mechanisms to escape from NK cell-mediated immune surveillance and we discuss NK cell-related parameters as prediction factors of AML patient outcome. On the basis of these observations, novel immunotherapeutic strategies capitalizing on the potentiation of NK cell functions have emerged in AML immunotherapy, as discussed in this review. Increased knowledge on AML escape routes from NK cell immune surveillance will further aid in the design of novel NK cell-based immunotherapy approaches for the treatment of AML.

Interferon-α in acute myeloid leukemia: an old drug revisited.

Interferon-α (IFN-α), a type I IFN, is a well-known antitumoral agent. The investigation of its clinical properties in acute myeloid leukemia (AML) has been prompted by its pleiotropic antiproliferative and immune effects. So far, integration of IFN-α in the therapeutic arsenal against AML has been modest in view of the divergent results of clinical trials. Recent insights into the key pharmacokinetic determinants of the clinical efficacy of IFN along with advances in its pharmaceutical formulation, have sparked renewed interest in its use. This paper reviews the possible applicability of IFN-α in the treatment of AML and provides a rational basis to re-explore its efficacy in clinical trials.

Quantification of IFN-gamma produced by human purified NK cells following tumor cell stimulation: comparison of three IFN-gamma assays.

Interferon (IFN)-gamma released by natural killer (NK) cells has become a subject of major interest, given its importance in bridging the innate and adaptive immune system. Interestingly, reports concerning tumor cell stimulation of NK cells show divergent data on which stimuli induce IFN-gamma production. Here, the question remains whether tumor cell recognition is sufficient to trigger IFN-gamma or whether a second signal is required such as type I IFN. While IFN-gamma detection methods are abundantly used with peripheral blood mononuclear cells or purified T cell fractions as responder populations, only limited data is available about comparison of these assays with purified NK cells. In this study, we assessed the relationship between stimulation of human purified resting peripheral blood NK cells with one (tumor cell or IFN-alpha) and two (tumor cell+IFN-alpha) signals by measuring IFN-gamma using three different assays. We performed the enzyme-linked immunosorbent assay (ELISA), the enzyme-linked immunospot (ELISPOT) assay and intracellular cytokine staining (ICS) assay in parallel per donor and determined whether there was a correlation between these assays. Our results show that two-signal stimulation of human resting NK cells induces significantly more IFN-gamma as compared to one-signal stimulation, readily picked up by all assays. Moreover, statistical analysis points towards a positive correlation between these assays for IFN-gamma produced following two-signal stimulation. Importantly, we show that tumor cell stimulation alone is enough to trigger secretion of IFN-gamma, but this finding was only evidenced by ELISPOT. These results reveal that the choice of IFN-gamma detection method can markedly influence the outcome regarding induction of NK cell IFN-gamma by tumor cells.

Immunosuppression induced by immature dendritic cells is mediated by TGF-beta/IL-10 double-positive CD4+ regulatory T cells.

Dendritic cells (DC) have important functions in T cell immunity and T cell tolerance. Previously, it was believed that T cell unresponsiveness induced by immature DC (iDC) is caused by the absence of inflammatory signals in steady-state in vivo conditions and by the low expression levels of costimulatory molecules on iDC. However, a growing body of evidence now indicates that iDC can also actively maintain peripheral T cell tolerance by the induction and/or stimulation of regulatory T cell populations. In this study, we investigated the in vitro T cell stimulatory capacity of iDC and mature DC (mDC) and found that both DC types induced a significant increase in the number of transforming growth factor (TGF)-beta and interleukin (IL)-10 double-positive CD4(+) T cells within 1 week of autologous DC/T cell co-cultures. In iDC/T cell cultures, where antigen-specific T cell priming was significantly reduced as compared to mDC/T cell cultures, we demonstrated that the tolerogenic effect of iDC was mediated by soluble TGF-beta and IL-10 secreted by CD4(+)CD25(-)FOXP3(-) T cells. In addition, the suppressive capacity of CD4(+) T cells conditioned by iDC was transferable to already primed antigen-specific CD8(+) T cell cultures. In contrast, addition of CD4(+) T cells conditioned by mDC to primed antigen-specific CD8(+) T cells resulted in enhanced CD8(+) T cell responses, notwithstanding the presence of TGF-beta(+)/IL-10(+) T cells in the transferred fraction. In summary, we hypothesize that DC have an active role in inducing immunosuppressive cytokine-secreting regulatory T cells. We show that iDC-conditioned CD4(+) T cells are globally immunosuppressive, while mDC induce globally immunostimulatory CD4(+) T cells. Furthermore, TGF-beta(+)/IL-10(+) T cells are expanded by DC independent of their maturation status, but their suppressive function is dependent on immaturity of DC.

Proinflammatory response of human leukemic cells to dsRNA transfection linked to activation of dendritic cells.

Leukemic cells exert immunosuppressive effects that interfere with dendritic cell (DC) function and hamper effective antileukemic immune responses. Here, we sought to enhance the immunogenicity of leukemic cells by loading them with the double-stranded (ds) RNA Toll-like receptor 3 (TLR3) ligand polyriboinosinic polyribocytidylic acid (poly(I:C)), mimicking viral infection of the tumor cells. Given the responsiveness of DC to TLR ligands, we hypothesized that the uptake of poly(I:C)-loaded leukemic cells by immature DC (iDC) would lead to DC activation. Primary acute myeloid leukemia (AML) cells and AML cell lines markedly responded to poly(I:C) electroporation by apoptosis, upregulation of TLR3 expression, enhanced expression of major histocompatibility complex (MHC) and costimulatory molecules and by production of type I interferons (IFN). Upon phagocytosis of poly(I:C)-electroporated AML cells, DC maturation and activation were induced as judged by an increased expression of MHC and costimulatory molecules, production of proinflammatory cytokines and an increase of T helper 1 (T(H)1)-polarizing capacity. These immune effects were suboptimal when AML cells were passively pulsed with poly(I:C), indicating the superiority of poly(I:C) transfection over pulsing. Our results demonstrate that poly(I:C) electroporation is a promising strategy to increase the immunogenicity of AML cells and to convert iDC into activated mature DC following the phagocytosis of AML cells.