Johan Aurelius

Immunotherapeutic strategies for relapse control in acute myeloid leukemia

Despite that the initial phases of chemotherapy induce disappearance of leukemic cells in many patients with acute myeloid leukemia (AML), the prevention of life-threatening relapses in the post-remission phase remains a significant clinical challenge. Allogeneic bone marrow transplantation, which is available for a minority of patients, efficiently prevents recurrences of leukemia by inducing immune-mediated elimination of leukemic cells, and over the past decades, numerous immunotherapeutic protocols have been developed aiming to mimic the graft-versus-leukemia reaction for the prevention of relapse. Here we review past and present strategies for relapse control with focus on overcoming leukemia-related immunosuppression in AML. We envisage future treatment protocols, in which systemic immune activators, such as vaccines, dendritic cell-based therapies, engineered variants of IL-2, or IL-15, are combined with agents that counter immunosuppression mediated by, e.g., the PD/PDL interaction, CTLA-4, CD200, reactive oxygen species, IDO expression, CXCR4, or the KIR/class I interaction, based on characteristics of the prevailing malignant clone. This combinatorial approach may pave the way for individualized immunotherapy in AML.

Remission maintenance in acute myeloid leukemia: impact of functional histamine H2 receptors expressed by leukemic cells

Post-consolidation immunotherapy with histamine dihydrochloride and interleukin-2 has been shown to improve leukemia-free survival in acute myeloid leukemia in a phase III trial. For this study, treatment efficacy was determined among 145 trial patients with morphological forms of acute myeloid leukemia as defined by the French-American-British classification. Leukemia-free survival was strongly improved in M4/M5 (myelomonocytic/monocytic) leukemia but not in M2 (myeloblastic) leukemia. We also analyzed histamine H2 receptor expression by leukemic cells recovered from 26 newly diagnosed patients. H2 receptors were typically absent from M2 cells but frequently expressed by M4/M5 cells. M4/M5 cells, but not M2 cells, produced reactive oxygen species that triggered apoptosis in adjacent natural killer cells. These events were significantly inhibited by histamine dihydrochloride. Our data demonstrate the presence of functional histamine H2 receptors on human AML cells and suggest that expression of these receptors by leukemic cells may impact on the effectiveness of histamine-based immunotherapy.

Post‐consolidation Immunotherapy with Histamine Dihydrochloride and Interleukin‐2 in AML

The initial chemotherapy in acute myeloid leukaemia (AML) comprises a first phase of induction and a second phase of consolidation. In the majority of patients, the induction treatment leads to complete remission (CR), defined as microscopic disappearance of leukaemic disease along with the return of normal haematopoiesis. However, despite the introduction of more efficacious consolidation regimens, a worryingly large proportion of AML patients in CR will subsequently experience relapses with poor prospects of long‐term survival. A relapse is assumed to be the result of expansion of residual leukaemic cells that have escaped the initial chemotherapy. The anti‐leukaemic functions of T cells and natural killer (NK) cells has formed the background to the use of interleukin‐2 (IL‐2), a T‐ and NK cell‐activating cytokine, with the aim to eliminate residual leukaemia and hence reduce the relapse rate in AML, but the clinical trials using IL‐2 monotherapy have yielded disappointment. A recent phase III study has demonstrated that post‐consolidation treatment with the combination of histamine dihydrochloride (HDC) and IL‐2 significantly prevents relapse in AML patients. Here we account for the preclinical background to the use of HDC/IL‐2 in AML along with a review of clinical results.

Monocytic AML cells inactivate antileukemic lymphocytes: role of NADPH oxidase/gp91(phox) expression and the PARP-1/PAR pathway of apoptosis.

Dysfunction of T cells and natural killer (NK) cells has been proposed to determine the course of disease in acute myeloid leukemia (AML), but only limited information is available on the mechanisms of lymphocyte inhibition. We aimed to evaluate to what extent human malignant AML cells use NADPH oxidase-derived reactive oxygen species (ROS) as an immune evasion strategy. We report that a subset of malignant myelomonocytic and monocytic AML cells (French-American-British [FAB] classes M4 and M5, respectively), recovered from blood or BM of untreated AML patients at diagnosis, expressed the NADPH oxidase component gp91(phox). Highly purified FAB M4/M5 AML cells produced large amounts of ROS on activation and triggered poly-[ADP-ribose] polymerase-1-dependent apoptosis in adjacent NK cells, CD4(+) T cells, and CD8(+) T cells. In contrast, immature (FAB class M1) and myeloblastic (FAB class M2) AML cells rarely expressed gp91(phox), did not produce ROS, and did not trigger NK or T-cell apoptosis. Microarray data from 207 AML patients confirmed a greater expression of gp91(phox) mRNA by FAB-M4/M5 AML cells than FAB-M1 cells (P < 10(-11)) or FAB-M2 cells (P < 10(-9)). Our data are suggestive of a novel mechanism by which monocytic AML cells evade cell-mediated immunity.

Immunotherapy with histamine dihydrochloride for the prevention of relapse in acute myeloid leukemia

Most patients with acute myeloid leukemia (AML) achieve complete remission (CR) after induction chemotherapy. Despite ensuing courses of consolidation chemotherapy, a large fraction of patients will experience relapses with poor prospects of long-term survival. Histamine dihydrochloride (HDC) in combination with the T-cell-derived cytokine IL-2 was recently approved within the EU as a remission maintenance immunotherapy in AML. HDC reduces myeloid cell-derived suppression of anti-leukemic lymphocytes, and aims to unravel a therapeutic benefit of IL-2 in AML by improving natural killer and T-cell activation. A randomized Phase III trial with 320 AML patients in CR demonstrated a significant reduction of relapse risk after immunotherapy with HDC plus low-dose IL-2 in the post-consolidation phase. HDC is the first approved therapeutic to target the state of immunosuppression in AML; further development in this area may comprise supplementary or alternative counter-suppressive agents with the aim to improve the efficacy of cancer immunotherapy.

Role of natural killer cell subsets and natural cytotoxicity receptors for the outcome of immunotherapy in acute myeloid leukemia.

In a phase IV trial, 84 patients (age 18–79) with acute myeloid leukemia (AML) in first complete remission (CR) received cycles of immunotherapy with histamine dihydrochloride (HDC) and low-dose human recombinant interleukin 2 (IL-2) for 18 months to prevent leukemic relapse. During cycles, the treatment resulted in expansion of CD56bright (CD3−/16−/56bright) and CD16+ (CD3−/16+/56+) natural killer (NK) cells in the blood along with increased NK cell expression of the natural cytotoxicity receptors (NCRs) NKp30 and NKp46. Multivariate analyses correcting for age and risk group demonstrated that high CD56bright NK cell counts and high expression of NKp30 or NKp46 on CD16+ NK cells independently predicted leukemia-free survival (LFS) and overall survival (OS). Our results suggest that the dynamics of NK cell subsets and their NCR expression may determine the efficiency of relapse-preventive immunotherapy in AML.

Redox Remodeling by Dendritic Cells Protects Antigen-Specific T Cells against Oxidative Stress

Microorganisms and microbial products induce the release of reactive oxygen species (ROS) from monocytes and other myeloid cells, which may trigger dysfunction and apoptosis of adjacent lymphocytes. Therefore, T cell-mediated immunity is likely to comprise mechanisms of T cell protection against ROS-inflicted toxicity. The present study aimed to clarify the dynamics of reduced sulfhydryl groups (thiols) in human T cells after presentation of viral and bacterial Ags by dendritic cells (DCs) or B cells. DCs, but not B cells, efficiently triggered intra- and extracellular thiol expression in T cells with corresponding Ag specificity. After interaction with DCs, the Ag-specific T cells acquired the capacity to neutralize exogenous oxygen radicals and resisted ROS-induced apoptosis. Our results imply that DCs provide Ag-specific T cells with antioxidative thiols during Ag presentation, which suggests a novel aspect of DC/T cell cross-talk of relevance to the maintenance of specific immunity in inflamed or infected tissue.

Role of the ERK Pathway for Oxidant-Induced Parthanatos in Human Lymphocytes

Reactive oxygen species (ROS) are formed by myeloid cells as a defense strategy against microorganisms. ROS however also trigger poly(ADP-ribose) polymerase 1- (PARP-1) dependent cell death (parthanatos) in adjacent lymphocytes, which has been forwarded as a mechanism of immune escape in several forms of cancer. The present study assessed the role of mitogen-activated protein kinases (MAPKs), in particular the extracellular signal-regulated kinase (ERK), in ROS-induced signal transduction leading to lymphocyte parthanatos. We report that inhibitors of ERK1/2 phosphorylation upheld natural killer (NK) cell-mediated cytotoxicity under conditions of oxidative stress and rescued NK cells and CD8+ T lymphocytes from cell death induced by ROS-producing monocytes. ERK1/2 phosphorylation inhibition also protected lymphocytes from cell death induced by exogenous hydrogen peroxide (H2O2) and from ROS generated by xanthine oxidase or glucose oxidase. Phosphorylation of ERK1/2 was observed in lymphocytes shortly after exposure to ROS. ROS-generating myeloid cells and exogenous H2O2 triggered PARP 1-dependent accumulation of poly ADP-ribose (PAR), which was prevented by ERK pathway inhibitors. ERK1/2 phosphorylation was induced by ROS independently of PARP-1. Our findings are suggestive of a role for ERK1/2 in ROS-induced lymphocyte parthanatos, and that the ERK axis may provide a therapeutic target for the protection of lymphocytes against oxidative stress.

Histamine Promotes the Development of Monocyte-Derived Dendritic Cells and Reduces Tumor Growth by Targeting the Myeloid NADPH Oxidase

The efficiency of immune-mediated clearance of cancer cells is hampered by immunosuppressive mediators in the malignant microenvironment, including NADPH oxidase–derived reactive oxygen species. We aimed at defining the effects of histamine, an inhibitor of the myeloid NADPH oxidase/NOX2, on the development of Ag-presenting dendritic cells (DCs) from myeloid precursors and the impact of these mechanisms for tumor growth. Histamine was found to promote the maturation of human DCs from monocytes by increasing the expression of HLA-DR and costimulatory molecules, which resulted in improved induction of Th cells with Th0 polarity. Experiments using wild-type and NOX2-deficient myelomonoblastic cells showed that histamine facilitated myeloid cell maturation only in cells capable of generating reactive oxygen species. Treatment of mice with histamine reduced the growth of murine EL-4 lymphomas in parallel with an increment of tumor-infiltrating DCs in NOX2-sufficient mice but not in NOX2-deficient (gp91phox−/−) mice. We propose that strategies to target the myeloid NADPH oxidase may facilitate the development of endogenous DCs in cancer.

TLR-Stimulated Neutrophils Instruct NK Cells To Trigger Dendritic Cell Maturation and Promote Adaptive T Cell Responses

Polymorphonuclear neutrophils (PMNs) are innate effector cells with pivotal roles in pathogen recognition, phagocytosis, and eradication. However, their role in the development of subsequent immune responses is incompletely understood. This study aimed to identify mechanisms of relevance to the cross talk between human neutrophils and NK cells and its potential role in promoting adaptive immunity. TLR-stimulated PMNs were found to release soluble mediators to attract and activate NK cells in vitro. PMN-conditioned NK cells displayed enhanced cytotoxicity and cytokine production, and responded vigorously to ensuing stimulation with exogenous and endogenous IL-12. The neutrophil-induced activation of NK cells was prevented by caspase-1 inhibitors and by natural antagonists to IL-1 and IL-18, suggesting a role for the NOD-like receptor family pyrin domain containing-3 inflammasome. In addition, PMN-conditioned NK cells triggered the maturation of monocyte-derived dendritic cells, which promoted T cell proliferation and IFN-γ production. These data imply that neutrophils attract NK cells to sites of infection to convert these cells into an active state, which drives adaptive immune responses via maturation of dendritic cells. Our results add to a growing body of evidence that suggests a sophisticated role for neutrophils in orchestrating the immune response to pathogens.