Interleukin 4 induces apoptosis of acute myeloid leukemia cells in a Stat6-dependent manner.

Cytokines provide signals that regulate immature normal and acute myeloid leukemia (AML) cells in the bone marrow microenvironment. We here identify interleukin 4 (IL4) as a selective inhibitor of AML cell growth and survival in a cytokine screen using fluorescently labeled AML cells. RNA-sequencing of the AML cells revealed an IL4-induced upregulation of Stat6 target genes and enrichment of apoptosis-related gene expression signatures. Consistent with these findings, we found that IL4 stimulation of AML cells induced Stat6 phosphorylation and that disruption of Stat6 using CRISPR/Cas9-genetic engineering rendered cells partially resistant to IL4-induced apoptosis. To evaluate whether IL4 inhibits AML cells in vivo, we expressed IL4 ectopically in AML cells transplanted into mice and also injected IL4 into leukemic mice; both strategies resulted in the suppression of the leukemia cell burden and increased survival. Notably, IL4 exposure caused reduced growth and survival of primary AML CD34+CD38- patient cells from several genetic subtypes of AML, whereas normal stem and progenitor cells were less affected. The IL4-induced apoptosis of AML cells was linked to Caspase-3 activation. Our results demonstrate that IL4 selectively induces apoptosis of AML cells in a Stat6-dependent manner-findings that may translate into new therapeutic opportunities in AML.

Modeling chronic myeloid leukemia in immunodeficient mice reveals expansion of aberrant mast cells and accumulation of pre-B cells.

Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm that, if not treated, will progress into blast crisis (BC) of either myeloid or B lymphoid phenotype. The BCR-ABL1 fusion gene, encoding a constitutively active tyrosine kinase, is thought to be sufficient to cause chronic phase (CP) CML, whereas additional genetic lesions are needed for progression into CML BC. To generate a humanized CML model, we retrovirally expressed BCR-ABL1 in the cord blood CD34(+) cells and transplanted these into NOD-SCID (non-obese diabetic/severe-combined immunodeficient) interleukin-2-receptor γ-deficient mice. In primary mice, BCR-ABL1 expression induced an inflammatory-like state in the bone marrow and spleen, and mast cells were the only myeloid lineage specifically expanded by BCR-ABL1. Upon secondary transplantation, the pronounced inflammatory phenotype was lost and mainly human mast cells and macrophages were found in the bone marrow. Moreover, a striking block at the pre-B-cell stage was observed in primary mice, resulting in an accumulation of pre-B cells. A similar block in B-cell differentiation could be confirmed in primary cells from CML patients. Hence, this humanized mouse model of CML reveals previously unexplored features of CP CML and should be useful for further studies to understand the disease pathogenesis of CML.

Succinate independently stimulates full platelet activation via cAMP and phosphoinositide 3-kinase-ß signaling.

BACKGROUND: The citric cycle intermediate succinate has recently been identified as a ligand for the G-protein-coupled receptor (GPCR) SUCNR1. We have previously found that this receptor is one of the most highly expressed GPCRs in human platelets. OBJECTIVE: The aim of this study was to investigate the role of SUCNR1 in platelet aggregation and to explore the signaling pathways of this receptor in platelets. METHODS AND RESULTS: Using real-time-PCR, we demonstrated that SUCNR1 is expressed in human platelets at a level corresponding to that of the P2Y(1) receptor. Light transmission aggregation experiments showed dose-dependent aggregation induced by succinate, reaching a maximum response at 0.5 mM. The effect of succinate on platelet aggregation was confirmed with flow cytometry, showing increased surface expression of activated glycoprotein IIb-IIIa and P-selectin. Intracellular SUCNR1 signaling was found to result in decreased cAMP levels, Akt phosphorylation mediated by phosphoinositide 3-kinase-ß activation, and receptor desensitization. Furthermore, succinate-induced platelet aggregation was demonstrated to depend on Src, generation of thromboxane A(2), and ATP release. Platelet SUCNR1 is subject to desensitization through both homologous and heterologous mechanisms. In addition, the P2Y(12) receptor inhibitor ticagrelor completely prevented platelet aggregation induced by succinate. CONCLUSIONS: Our experiments show that succinate induces full aggregation of human platelets via SUCNR1. Succinate-induced platelet aggregation depends on thromboxane A(2) generation, ATP release, and P2Y(12) activation.