Combined activity of the redox-modulating compound setanaxib (GKT137831) with cytotoxic agents in the killing of acute myeloid leukemia cells

Acute myeloid leukemia (AML) cells harbor elevated levels of reactive oxygen species (ROS), which promote cell proliferation and cause oxidative stress. Therefore, the inhibition of ROS formation or elevation beyond a toxic level have been considered as therapeutic strategies. ROS elevation has recently been linked to enhanced NADPH oxidase 4 (NOX4) activity. Therefore, the compound Setanaxib (GKT137831), a clinically advanced ROS-modulating substance, which has initially been identified as a NOX1/4 inhibitor, was tested for its inhibitory activity on AML cells. Setanaxib showed antiproliferative activity as single compound, and strongly enhanced the cytotoxic action of anthracyclines such as daunorubicin in vitro. Setanaxib attenuated disease in a mouse model of FLT3-ITD driven myeloproliferation in vivo. Setanaxib did not significantly inhibit FLT3-ITD signaling, including FLT3 autophosphorylation, activation of STAT5, AKT, or extracellular signal regulated kinase 1 and 2 (ERK1/2). Surprisingly, the effects of Setanaxib on cell proliferation appeared to be independent of the presence of NOX4 and were not associated with ROS quenching. Instead, Setanaxib caused elevation of ROS levels in the AML cells and importantly, enhanced anthracycline-induced ROS formation, which may contribute to the combined effects. Further assessment of Setanaxib as potential enhancer of cytotoxic AML therapy appears warranted

A JAK of all trades: how global phosphoproteomics reveal the Achilles heel of MPNs

While Janus-kinase (JAK)-inhibitors effectively reduce the inflammatory phenotype of myeloproliferative neoplasms (MPN), they do not affect disease burden or presence of the mutated clone to a major extent. Here, we show how Janus-kinase 2 (JAK2)-mutated cells persist through maintenance of the mitogen-activated protein kinase Interacting Serine/Threonine Kinase 1 (MKNK1) – Extracellular Signal-regulated Kinase (ERK)-axis by hijacking the splicing machinery through post-translational modifications

Immunoproteasome function maintains oncogenic gene expression in KMT2A-complex driven leukemia

Abstract Pharmacologic targeting of chromatin-associated protein complexes has shown significant responses in KMT2A-rearranged (KMT2A-r) acute myeloid leukemia (AML) but resistance frequently develops to single agents. This points to a need for therapeutic combinations that target multiple mechanisms. To enhance our understanding of functional dependencies in KMT2A-r AML, we have used a proteomic approach to identify the catalytic immunoproteasome subunit PSMB8 as a specific vulnerability. Genetic and pharmacologic inactivation of PSMB8 results in impaired proliferation of murine and human leukemic cells while normal hematopoietic cells remain unaffected. Disruption of immunoproteasome function drives an increase in transcription factor BASP1 which in turn represses KMT2A-fusion protein target genes. Pharmacologic targeting of PSMB8 improves efficacy of Menin-inhibitors, synergistically reduces leukemia in human xenografts and shows preserved activity against Menin-inhibitor resistance mutations. This identifies and validates a cell-intrinsic mechanism whereby selective disruption of proteostasis results in altered transcription factor abundance and repression of oncogene-specific transcriptional networks. These data demonstrate that the immunoproteasome is a relevant therapeutic target in AML and that targeting the immunoproteasome in combination with Menin-inhibition could be a novel approach for treatment of KMT2A-r AML

JAK2-V617F promotes venous thrombosis through β1/β2 integrin activation.

JAK2-V617F-positive chronic myeloproliferative neoplasia (CMN) commonly displays dysfunction of integrins and adhesion molecules expressed on platelets, erythrocytes, and leukocytes. However, the mechanism by which the 2 major leukocyte integrin chains, β1 and β2, may contribute to CMN pathophysiology remained unclear. β1 (α4β1; VLA-4) and β2 (αLβ2; LFA-1) integrins are essential regulators for attachment of leukocytes to endothelial cells. We here showed enhanced adhesion of granulocytes from mice with JAK2-V617F knockin (JAK2+/VF mice) to vascular cell adhesion molecule 1- (VCAM1-) and intercellular adhesion molecule 1-coated (ICAM1-coated) surfaces. Soluble VCAM1 and ICAM1 ligand binding assays revealed increased affinity of β1 and β2 integrins for their respective ligands. For β1 integrins, this correlated with a structural change from the low- to the high-affinity conformation induced by JAK2-V617F. JAK2-V617F triggered constitutive activation of the integrin inside-out signaling molecule Rap1, resulting in translocation toward the cell membrane. Employing a venous thrombosis model, we demonstrated that neutralizing anti-VLA-4 and anti-β2 integrin antibodies suppress pathologic thrombosis as observed in JAK2+/VF mice. In addition, aberrant homing of JAK2+/VF leukocytes to the spleen was inhibited by neutralizing anti-β2 antibodies and by pharmacologic inhibition of Rap1. Thus, our findings identified cross-talk between JAK2-V617F and integrin activation promoting pathologic thrombosis and abnormal trafficking of leukocytes to the spleen