Immunotherapies, such as chimeric antigen receptor T cells, bispecific antibodies, and immune checkpoint inhibitors, have emerged as promising modalities in multiple hematologic malignancies. Despite ...the excitement surrounding immunotherapy, it is currently not possible to predict which patients will respond. Within solid tumors, the status of the immune microenvironment provides valuable insight regarding potential responses to immune therapies. Much less is known about the immune microenvironment within hematologic malignancies but the characteristics of this environment are likely to serve a similar predictive role. Acute myeloid leukemia (AML) is the most common hematologic malignancy in adults, and only 25% of patients are alive 5 years following their diagnosis. There is evidence that manipulation of the immune microenvironment by leukemia cells may play a role in promoting therapy resistance and disease relapse. In addition, it has long been documented that through modulation of the immune system following allogeneic bone marrow transplant, AML can be cured, even in patients with the highest risk disease. These concepts, along with the poor prognosis associated with this disease, have encouraged many groups to start exploring the utility of novel immune therapies in AML. While the implementation of these therapies into clinical trials for AML has been supported by preclinical rationale, many questions still exist surrounding their efficacy, tolerability, and the overall optimal approach. In this review, we discuss what is known about the immune microenvironment within AML with a specific focus on T cells and checkpoints, along with their implications for immune therapies.
Mutations in the IDH1 and IDH2 genes encoding isocitrate dehydrogenases are frequently found in human glioblastomas and cytogenetically normal acute myeloid leukaemias (AML). These alterations are ...gain-of-function mutations in that they drive the synthesis of the ‘oncometabolite’ R-2-hydroxyglutarate (2HG). It remains unclear how IDH1 and IDH2 mutations modify myeloid cell development and promote leukaemogenesis. Here we report the characterization of conditional knock-in (KI) mice in which the most common IDH1 mutation, IDH1(R132H), is inserted into the endogenous murine Idh1 locus and is expressed in all haematopoietic cells (Vav-KI mice) or specifically in cells of the myeloid lineage (LysM-KI mice). These mutants show increased numbers of early haematopoietic progenitors and develop splenomegaly and anaemia with extramedullary haematopoiesis, suggesting a dysfunctional bone marrow niche. Furthermore, LysM-KI cells have hypermethylated histones and changes to DNA methylation similar to those observed in human IDH1- or IDH2-mutant AML. To our knowledge, our study is the first to describe the generation and characterization of conditional IDH1(R132H)-KI mice, and also the first report to demonstrate the induction of a leukaemic DNA methylation signature in a mouse model. Our report thus sheds light on the mechanistic links between IDH1 mutation and human AML.
Mutations in the isocitrate dehydrogenase-1 gene (IDH1) are common drivers of acute myeloid leukemia (AML) but their mechanism is not fully understood. It is thought that IDH1 mutants act by ...inhibiting TET2 to alter DNA methylation, but there are significant unexplained clinical differences between IDH1- and TET2-mutant diseases. We have discovered that mice expressing endogenous mutant IDH1 have reduced numbers of hematopoietic stem cells (HSCs), in contrast to Tet2 knockout (TET2-KO) mice. Mutant IDH1 downregulates the DNA damage (DD) sensor ATM by altering histone methylation, leading to impaired DNA repair, increased sensitivity to DD, and reduced HSC self-renewal, independent of TET2. ATM expression is also decreased in human IDH1-mutated AML. These findings may have implications for treatment of IDH-mutant leukemia.
•Mutant IDH1 decreases hematopoietic stem cell (HSC) number and impairs self-renewal•Mutant IDH1 causes TET2-independent downregulation of ATM via methylation of H3K9•Mutant IDH1 causes accumulation of DNA damage and impairs DNA repair in HSCs•Mutant IDH1 increases HSC sensitivity to radiation and daunorubicin
Although strong evidence supports that IDH1 mutants act by inhibiting TET2 in hematological malignancies, there are clear clinical differences between mutations of these genes. Inoue et al. show that mutant IDH1 decreases ATM independent of TET2, leading to impaired DNA repair and reduced hematopoietic stem cells.
Nuclear factor interleukin-3 (Nfil3; also known as E4-binding protein 4) is a basic region leucine zipper transcription factor that has antiapoptotic activity in vitro under conditions of growth ...factor withdrawal. To study the role of Nfil3 in vivo, we generated gene-targeted Nfil3-deficient (Nfil3(-/-)) mice. Nfil3(-/-) mice were born at normal Mendelian frequency and were grossly normal and fertile. Although numbers of T cells, B cells, and natural killer (NK) T cells were normal in Nfil3(-/-) mice, a specific disruption in NK cell development resulted in severely reduced numbers of mature NK cells in the periphery. This defect was NK cell intrinsic in nature, leading to a failure to reject MHC class I-deficient cells in vivo and reductions in both interferon gamma production and cytolytic activity in vitro. Our results confirm the specific and essential requirement of Nfil3 for the development of cells of the NK lineage.
Recent studies have begun to define the role of micro-RNAs in regulating the immune response. Micro-RNA155 (mir-155) has been shown to play a role in germinal center formation, T cell inflammation, ...and regulatory T cell development. In this study, we evaluated the role of mir-155 in cytotoxic T cell function. We report in this study that mice lacking mir-155 have impaired CD8(+) T cell responses to infections with lymphocytic choriomeningitis virus and the intracellular bacteria Listeria monocytogenes. We show by a series of adoptive transfer studies that the impaired CD8(+) T cell response to L. monocytogenes is T cell intrinsic. In addition, we observed that CD8(+) T cells lacking mir-155 have impaired activation of the prosurvival Akt pathway after TCR cross-linking. These data suggest that mir-155 may be a good target for therapies aimed at modulating immune responses.
Mir‐155, a central modulator of T‐cell responses Lind, Evan F.; Ohashi, Pamela S.
European journal of immunology,
January 2014, 2014-Jan, 2014-01-00, 20140101, Volume:
44, Issue:
1
Journal Article
Peer reviewed
The activation of T cells is a tightly regulated process that has evolved to maximize protective immune responses to pathogens while minimizing damage to self‐tissues. A delicate balance of ...cell‐intrinsic, costimulatory, and transcriptional pathways as well as micro‐environmental cues such as local cytokines controls the magnitude and nature of T‐cell responses in vivo. The discovery of functional small noncoding RNAs called micro‐RNAs (miRNAs) has introduced new mechanisms that contribute to the regulation of protein translation and cellular responses to stimuli. miRNAs are short (approximately 22 bp) RNA species, which bind to mRNAs and suppress translation. Due to their short length and imperfect base pairing requirements, each miRNA has the potential to regulate various pathways through the translational inhibition of multiple mRNAs. The human and mouse genomes each encode hundreds of miRNAs, and studying the function of miRNAs has led to the realization that they play important roles in diverse biological processes from development and cancer to immunity. This review focuses on the function of mir‐155 in T cells and the impact of this miRNA on autoimmunity, tumor immunity, and pathogen‐induced immunity.
Development of the acquired immune response is dependent on the signaling of CD40 by its ligand, CD154. These molecules govern both the magnitude and quality of humoral- and cell-mediated immunity. A ...litany of studies have conclusively documented that blockade of this ligand-receptor pair can prevent, and also intervene in, the progression of antibody- and cell-mediated autoimmune diseases, and can instill long-lived allogeneic and xenogeneic graft tolerance. Many effector mechanisms of inflammation are abolished as a result of CD154 blockade, but we are now beginning to understand that CD154 blockade may, in some instances, engender long-lived, antigen-specific tolerance. In the context of transplantation tolerance, we present a hypothesis that alpha CD154 blockade is most effective at inducing long-lived allospecific tolerance if anergy and regulation can be elicited prior to the onslaught of inflammation that is induced by grafting (preemptive tolerance). This facet of alpha CD154-induced tolerance appears to co-opt the normal processes of peripheral tolerance induced by immature DCs and can be exploited to induce long-lived antigen-specific tolerance. The underlying science and the prospects for inducing long-lived antigen-specific tolerance in a model of allograft tolerance through CD154 blockade are presented and discussed.
To identify new therapeutic targets in acute myeloid leukemia (AML), we performed small-molecule and small-interfering RNA (siRNA) screens of primary AML patient samples. In 23% of samples, we found ...sensitivity to inhibition of colony-stimulating factor 1 (CSF1) receptor (CSF1R), a receptor tyrosine kinase responsible for survival, proliferation, and differentiation of myeloid-lineage cells. Sensitivity to CSF1R inhibitor GW-2580 was found preferentially in de novo and favorable-risk patients, and resistance to GW-2580 was associated with reduced overall survival. Using flow cytometry, we discovered that CSF1R is not expressed on the majority of leukemic blasts but instead on a subpopulation of supportive cells. Comparison of CSF1R-expressing cells in AML vs healthy donors by mass cytometry revealed expression of unique cell-surface markers. The quantity of CSF1R-expressing cells correlated with GW-2580 sensitivity. Exposure of primary AML patient samples to a panel of recombinant cytokines revealed that CSF1R inhibitor sensitivity correlated with a growth response to CSF1R ligand, CSF1, and other cytokines, including hepatocyte growth factor (HGF). The addition of CSF1 increased the secretion of HGF and other cytokines in conditioned media from AML patient samples, whereas adding GW-2580 reduced their secretion. In untreated cells, HGF levels correlated significantly with GW-2580 sensitivity. Finally, recombinant HGF and HS-5–conditioned media rescued cell viability after GW-2580 treatment in AML patient samples. Our results suggest that CSF1R-expressing cells support the bulk leukemia population through the secretion of HGF and other cytokines. This study identifies CSF1R as a novel therapeutic target of AML and provides a mechanism of paracrine cytokine/growth factor signaling in this disease.
•CSF1R inhibition reduces cell viability in >20% of AML patient samples and is expressed on a subpopulation of supportive cells.•CSF1R activation stimulates paracrine cytokine secretion (eg, HGF), suggesting that CSF1R is a novel target of AML support cells.
Display omitted
Oral azacitidine (oral-Aza) treatment results in longer median overall survival (OS) (24.7 vs. 14.8 months in placebo) in patients with acute myeloid leukemia (AML) in remission after intensive ...chemotherapy. The dosing schedule of oral-Aza (14 days/28-day cycle) allows for low exposure of Aza for an extended duration thereby facilitating a sustained therapeutic effect. However, the underlying mechanisms supporting the clinical impact of oral-Aza in maintenance therapy remain to be fully understood. In this preclinical work, we explore the mechanistic basis of oral-Aza/extended exposure to Aza through in vitro and in vivo modeling. In cell lines, extended exposure to Aza results in sustained DNMT1 loss, leading to durable hypomethylation, and gene expression changes. In mouse models, extended exposure to Aza, preferentially targets immature leukemic cells. In leukemic stem cell (LSC) models, the extended dose of Aza induces differentiation and depletes CD34+CD38- LSC. Mechanistically, LSC differentiation is driven in part by increased myeloperoxidase (MPO) expression. Inhibition of MPO activity either by using an MPO-specific inhibitor or blocking oxidative stress, a known mechanism of MPO, partly reverses the differentiation of LSC. Overall, our preclinical work reveals novel mechanistic insights into oral-Aza and its ability to target LSC.
Dendritic cells (DC) are mediators between innate and adaptive immune responses to pathogens and tumors. DC development is determined by signaling through the receptor tyrosine kinase Fms-like ...tyrosine kinase 3 (FLT3) in bone marrow myeloid progenitors. Recently the naming conventions for DC phenotypes have been updated to distinguish between "Conventional" DCs (cDCs) and plasmacytoid DCs (pDCs). Activating mutations of FLT3, including Internal Tandem Duplication (FLT3-ITD), are associated with poor prognosis for acute myeloid leukemia (AML) patients. Having a shared myeloid lineage it can be difficult to distinguish
DCs from AML tumor cells. To date, there is little information on the effects of FLT3-ITD in DC biology. To further elucidate this relationship we utilized CITE-seq technology in combination with flow cytometry and multiplex immunoassays to measure changes to DCs in human and mouse tissues. We examined the cDC phenotype and frequency in bone marrow aspirates from patients with AML to understand the changes to cDCs associated with FLT3-ITD. When compared to healthy donor (HD) we found that a subset of FLT3-ITD+ AML patient samples have overrepresented populations of cDCs and disrupted phenotypes. Using a mouse model of FLT3-ITD+ AML, we found that cDCs were increased in percentage and number compared to control wild-type (WT) mice. Single cell RNA-seq identified FLT3-ITD+ cDCs as skewed towards a cDC2 T-bet
phenotype, previously shown to promote Th17 T cells. We assessed the phenotypes of CD4
T cells in the AML mice and found significant enrichment of both Treg and Th17 CD4
T cells in the bone marrow and spleen compartments.
stimulation of CD4
T cells also showed increased Th17 phenotype in AML mice. Moreover, co-culture of AML mouse-derived DCs and naïve OT-II cells preferentially skewed T cells into a Th17 phenotype. Together, our data suggests that FLT3-ITD+ leukemia-associated cDCs polarize CD4
T cells into Th17 subsets, a population that has been shown to be negatively associated with survival in solid tumor contexts. This illustrates the complex tumor microenvironment of AML and highlights the need for further investigation into the effects of FLT3-ITD mutations on DC phenotypes and their downstream effects on Th polarization.
PDF
