Deficiency of Antioxidative Paraoxonase 2 (Pon2) Leads to Increased Number of Phenotypic LT-HSCs and Disturbed Erythropoiesis

Background. Long-term hematopoietic stem cells (LT-HSCs) reside in bone marrow niches with tightly controlled reactive oxygen species (ROS) levels. ROS increase results into LT-HSC differentiation and stem cell exhaustion. Paraoxonase 2 (PON2) has been shown to be important for ROS control. Objectives. We investigate the effects of inactivation of the PON2 gene on hematopoietic cell differentiation and activity. Methods and Results. In young mice with inactivated Pon2 gene (Pon2-/-, -/- BM outcompeted WT BM at early time points. ROS levels were significantly increased in Pon2-/- whole BM, but not in Pon2-/- LT-HSCs. In more differentiated stages of hematopoiesis, Pon2 deficiency led to a misbalanced erythropoiesis both in physiologic and stress conditions. In older mice (>9 months), Pon2 depletion caused an increase in LT-HSCs as well as increased levels of granulocyte/macrophage progenitors (GMPs) and myeloid skewing, indicating a premature aging phenotype. No significant changes in ROS levels in old Pon2-/- LT- and short-term (ST-) HSCs were observed, but a significant reduction of spontaneous apoptotic cell death was measured. RNA-seq analysis in Pon2-/- LT-HSCs identified overrepresentation of genes involved in the C-X-C chemokine receptor type 4 (Cxcr4) signaling, suggesting compensatory mechanisms to overcome ROS-mediated accelerated aging in hematopoietic progenitor cells. Conclusions. In summary, our current data indicate that PON2 is involved in the regulation of HSC functions

Contrasting requirements during disease evolution identify EZH2 as a therapeutic target in AML

Epigenetic regulators, such as EZH2, are frequently mutated in cancer, and loss-of-function EZH2 mutations are common in myeloid malignancies. We have examined the importance of cellular context for Ezh2 loss during the evolution of acute myeloid leukemia (AML), where we observed stage-specific and diametrically opposite functions for Ezh2 at the early and late stages of disease. During disease maintenance, WT Ezh2 exerts an oncogenic function that may be therapeutically targeted. In contrast, Ezh2 acts as a tumor suppressor during AML induction. Transcriptional analysis explains this apparent paradox, demonstrating that loss of Ezh2 derepresses different expression programs during disease induction and maintenance. During disease induction, Ezh2 loss derepresses a subset of bivalent promoters that resolve toward gene activation, inducing a feto-oncogenic program that includes genes such as Plag1, whose overexpression phenocopies Ezh2 loss to accelerate AML induction in mouse models. Our data highlight the importance of cellular context and disease phase for the function of Ezh2 and its potential therapeutic implications.The Huntly laboratory is funded by CRUK (program C18680/ A25508), the European Research Council (grant 647685 COMAL), the Kay Kendall Leukaemia Fund, the Medical Research Council (MRC), Bloodwise, the Wellcome Trust, and the Cambridge National Institute of Health Research Biomedical Research Centre. F. Basheer is a recipient of a Wellcome Trust PhD for Clinicians award. P. Gallipoli is funded by the Wellcome Trust (109967/Z/15/Z). We acknowledge the Wellcome Trust/ MRC center grant (097922/Z/11/Z) and support from Wellcome Trust strategic award 100140. Research in the laboratory is also supported by core funding from the Wellcome Trust and MRC to the Wellcome-MRC Cambridge Stem Cell Institute. This research was supported by the Cambridge National Institute of Health Research Biomedical Research Centre Cell Phenotyping Hub

Frequent ZNF217 mutations lead to transcriptional deregulation of interferon signal transduction via altered chromatin accessibility in B cell lymphoma

Recent exome-wide studies discovered frequent somatic mutations in the epigenetic modifier ZNF217 in primary mediastinal B cell lymphoma (PMBCL) and related disorders. As functional consequences of ZNF217 alterations remain unknown, we comprehensively evaluated their impact in PMBCL. Targeted sequencing identified genetic lesions affecting ZNF217 in 33% of 157 PMBCL patients. Subsequent gene expression profiling (n = 120) revealed changes in cytokine and interferon signal transduction in ZNF217-aberrant PMBCL cases. In vitro, knockout of ZNF217 led to changes in chromatin accessibility interfering with binding motifs for crucial lymphoma-associated transcription factors. This led to disturbed expression of interferon-responsive and inflammation-associated genes, altered cell behavior, and aberrant differentiation. Mass spectrometry demonstrates that ZNF217 acts within a histone modifier complex containing LSD1, CoREST and HDAC and interferes with H3K4 methylation and H3K27 acetylation. Concluding, our data suggest non-catalytic activity of ZNF217, which directs histone modifier complex function and controls B cell differentiation-associated patterns of chromatin structure

NCAM1 (CD56) promotes leukemogenesis and confers drug resistance in AML

Neural cell adhesion molecule 1 (NCAM1; CD56) is expressed in up to 20% of acute myeloid leukemia (AML) patients. NCAM1 is widely used as a marker of minimal residual disease; however, the biological function of NCAM1 in AML remains elusive. In this study, we investigated the impact of NCAM1 expression on leukemogenesis, drug resistance, and its role as a biomarker to guide therapy. Beside t(8;21) leukemia, NCAM1 expression was found in most molecular AML subgroups at highly heterogeneous expression levels. Using complementary genetic strategies, we demonstrated an essential role of NCAM1 in the regulation of cell survival and stress resistance. Perturbation of NCAM1 induced cell death or differentiation and sensitized leukemic blasts toward genotoxic agents in vitro and in vivo. Furthermore, Ncam1 was highly expressed in leukemic progenitor cells in a murine leukemia model, and genetic depletion of prolonged disease latency and significantly reduced leukemia-initiating cells upon serial transplantation. To further analyze the mechanism of the NCAM1-associated phenotype, we performed phosphoproteomics and transcriptomics in different AML cell lines. NCAM1 expression strongly associated with constitutive activation of the MAPK-signaling pathway, regulation of apoptosis, or glycolysis. Pharmacological inhibition of MEK1/2 specifically inhibited proliferation and sensitized NCAM1 AML cells to chemotherapy. In summary, our data demonstrate that aberrant expression of NCAM1 is involved in the maintenance of leukemic stem cells and confers stress resistance, likely due to activation of the MAPK pathway. Targeting MEK1/2 sensitizes AML blasts to genotoxic agents, indicating a role for NCAM1 as a biomarker to guide AML treatment