Expression of a passenger miR-9* predicts favorable outcome in adults with acute myeloid leukemia less than 60 years of age.

In double-stranded miRNA/miRNA* duplexes, one of the strands represents an active miRNA, whereas another, known as a passenger strand (miRNA*), is typically degraded. MiR-9* is not detectable in normal myeloid cells. Here we show that miR-9* is expressed in 59% of acute myeloid leukemia (AML) cases and we investigate its clinical impact in 567 adults with de novo AML (age⩽60 years). AML cases with detectable miR-9* included a lower percentage of cases with favorable risk (P<0.001) as compared with those with no detectable miR-9*. High levels of miR-9* expression independently predicted for higher complete remission (odds ratio=1.28, P=0.013) and better event-free survival (EFS) (hazard ratio (HR)=0.86, P=0.001), relapse-free survival (RFS) (HR=0.84, P=0.008) and overall survival (OS) (HR=0.86, P=0.002). Among the subgroup of adverse risk patients, high miR-9* expressers had strikingly longer median survival than low miR-9* expressers (EFS: 16 vs 5 months, P=0.020; RFS: 12 vs 4, P=0.060; OS: 23 vs 8, P=0.021). Comparative transcriptome analysis suggests that miR-9* regulates genes involved in leukemogenesis, for example, MN1 and MLLT3. This is the first report showing that an miRNA* has prognostic value in AML.

Aberrant expression of miR-9/9* in myeloid progenitors inhibits neutrophil differentiation by post-transcriptional regulation of ERG.

Aberrant post-transcriptional regulation by microRNAs (miRNAs) has been shown to be involved in the pathogenesis of acute myeloid leukemia (AML). In a previous study, we performed a large functional screen using a retroviral barcoded miRNA expression library. Here, we report that overexpression of miR-9/9* in myeloid 32D cell line (32D-miR-9/9*) had profound impact on granulocyte colony-stimulating factor-induced differentiation. Further in vitro studies showed that enforced expression of miR-9/9* blocked normal neutrophil development in 32D and in primary murine lineage-negative bone marrow cells. We examined the expression of miR-9/9* in a cohort of 647 primary human AMLs. In most cases, miR-9 and miR-9* were significantly upregulated and their expression levels varied according to AML subtype, with the highest expression in MLL-related leukemias harboring 11q23 abnormalities and the lowest expression in AML cases with t(8;21) and biallelic mutations in CEBPA. Gene expression profiling of AMLs with high expression of miR-9/9* and 32D-miR-9/9* identified ETS-related gene (Erg) as the only common potential target. Upregulation of ERG in 32D cells rescued miR-9/9*-induced block in neutrophil differentiation. Taken together, this study demonstrates that miR-9/9* are aberrantly expressed in most of AML cases and interfere with normal neutrophil differentiation by downregulation of ERG.

Novel role of WD40 and SOCS box protein-2 in steady-state distribution of granulocyte colony-stimulating factor receptor and G-CSF-controlled proliferation and differentiation signaling.

Signals induced by granulocyte colony-stimulating factor (G-CSF), the major cytokine involved in neutrophil development, are tightly controlled by ligand-induced receptor internalization. Truncated G-CSF receptors (G-CSF-Rs) that fail to internalize show sustained proliferation and defective differentiation signaling. Steady-state forward routing also determines cell surface levels of cytokine receptors, but mechanisms controlling this are poorly understood. Here, we show that WD40 and suppressor of cytokine signaling (SOCS) box protein-2 (Wsb-2), an SOCS box-containing WD40 protein with currently unknown function, binds to the COOH-terminal region of G-CSF-R. Removal of this region did not affect internalization, yet resulted in increased membrane expression of G-CSF-R and enhanced proliferation signaling at the expense of differentiation induction. Conversely, Wsb-2 binding to the G-CSF-R reduced its cell surface expression and inhibited proliferation signaling. These effects depended on the SOCS box involved in ubiquitylation and on cytosolic lysines of G-CSF-R and imply a major role for ubiquitylation through the G-CSF-R C-terminus in forward routing of the receptor. Importantly, the Wsb-2 gene is commonly disrupted by virus integrations in mouse leukemia. We conclude that control of forward routing of G-CSF-R is essential for a balanced response of myeloid progenitors to G-CSF and suggest that disturbance of this balance may contribute to myeloid leukemia.

Granulocyte colony-stimulating factor and its receptor in normal hematopoietic cell development and myeloid disease.

Hematopoiesis, the process of blood cell formation, is orchestrated by cytokines and growth factors that stimulate the expansion of different progenitor cell subsets and regulate their survival and differentiation into mature blood cells. Granulocyte colony-stimulating factor (G-CSF) is the major hematopoietic growth factor involved in the control of neutrophil development. G-CSF is now applied on a routine basis in the clinic for treatment of congenital and acquired neutropenias. G-CSF activates a receptor of the hematopoietin receptor superfamily, the G-CSF receptor (G-CSF-R), which subsequently triggers multiple signaling mechanisms. Here we review how these mechanisms contribute to the specific responses of hematopoietic cells to G-CSF and how perturbations in the function of the G-CSF-R are implicated in various types of myeloid disease.