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Blockade of miR-150 Maturation by MLL-Fusion/MYC/LIN-28 Is Required for MLL-Associated Leukemia

Expression of microRNAs (miRNAs) is under stringent regulation at both transcriptional and posttranscriptional levels. Disturbance at either level could cause dysregulation of miRNAs. Here, we show that MLL fusion proteins negatively regulate production of miR-150, an miRNA widely repressed in acute leukemia, by blocking miR-150 precursors from being processed to mature miRNAs through MYC/LIN28 functional axis. Forced expression of miR-150 dramatically inhibited leukemic cell growth and delayed MLL-fusion-mediated leukemogenesis, likely through targeting FLT3 and MYB and thereby interfering with the HOXA9/MEIS1/FLT3/MYB signaling network, which in turn caused downregulation of MYC/LIN28. Collectively, we revealed a MLL-fusion/MYC/LIN28⊣miR-150⊣FLT3/MYB/HOXA9/MEIS1 signaling circuit underlying the pathogenesis of leukemia, where miR-150 functions as a pivotal gatekeeper and its repression is required for leukemogenesis.

Co-inhibition of NF-κB and JNK is synergistic in TNF-expressing human AML

TNF signaling inactivation sensitizes AML cells to NF-kB inhibition but protects healthy hematopoietic stem progenitor cells from this treatment.

p27kip1 maintains a subset of leukemia stem cells in the quiescent state in murine MLL‐leukemia

MLL (mixed‐lineage leukemia)‐fusion genes induce the development of leukemia through deregulation of normal MLL target genes, such as HOXA9 and MEIS1. Both HOXA9 and MEIS1 are required for MLL‐fusion gene‐induced leukemogenesis. Co‐expression of HOXA9 and MEIS1 induces acute myeloid leukemia (AML) similar to that seen in mice in which MLL‐fusion genes are over‐expressed. p27kip1 (p27 hereafter), a negative regulator of the cell cycle, has also been defined as an MLL target, the expression of which is up‐regulated in MLL leukemic cells (LCs). To investigate whether p27 plays a role in the pathogenesis of MLL‐leukemia, we examined the effects of p27 deletion (p27−/−) on MLL‐AF9 (MA9)‐induced murine AML development. HOXA9/MEIS1 (H/M)‐induced, p27 wild‐type (p27+/+) and p27−/− AML were studied in parallel as controls. We found that LCs from both MA9‐AML and H/M‐AML can be separated into three fractions, a CD117‐CD11bhi differentiated fraction as well as CD117+CD11bhi and CD117+CD11blo, two less differentiated fractions. The CD117+CD11blo fraction, comprising only 1–3% of total LCs, expresses higher levels of early hematopoietic progenitor markers but lower levels of mature myeloid cell markers compared to other populations of LCs. p27 is expressed and is required for maintaining the quiescent and drug‐resistant states of the CD117+CD11blo fraction of MA9‐LCs but not of H/M‐LCs. p27 deletion significantly compromises the leukemogenic capacity of CD117+CD11blo MA9‐LCs by reducing the frequency of leukemic stem cells (LSCs) but does not do so in H/M‐LCs. In addition, we found that p27 is highly expressed and required for cell cycle arrest in the CD117‐CD11bhi fraction in both types of LCs. Furthermore, we found that c‐Myc expression is required for maintaining LCs in an undifferentiated state independently of proliferation. We concluded that p27 represses the proliferation of LCs, which is specifically required for maintaining the quiescent and drug‐resistant states of a small subset of MA9‐LSCs in collaboration with the differentiation blockage function of c‐Myc.

Sensitizing acute myeloid leukemia cells to induced differentiation by inhibiting the RIP1/RIP3 pathway

Tumor necrosis factor-α (TNF-α)-induced RIP1/RIP3 (receptor-interacting protein kinase 1/receptor-interacting protein kinase 3)-mediated necroptosis has been proposed as an alternative strategy for treating apoptosis-resistant leukemia. However, we found that most acute myeloid leukemia (AML) cells, especially M4 and M5 subtypes, produce TNF and show basal level activation of RIP1/RIP3/MLKL signaling, yet do not undergo necroptosis. TNF, through RIP1/RIP3 signaling, prevents degradation of SOCS1, a key negative regulator of interferon-γ (IFN-γ) signaling. Using both pharmacologic and genetic assays, we show here that inactivation of RIP1/RIP3 resulted in reduction of SOCS1 protein levels and partial differentiation of AML cells. AML cells with inactivated RIP1/RIP3 signaling show increased sensitivity to IFN-γ-induced differentiation. RIP1/RIP3 inactivation combined with IFN-γ treatment significantly attenuated the clonogenic capacity of both primary AML cells and AML cell lines. This combination treatment also compromised the leukemogenic ability of murine AML cells in vivo. Our studies suggest that inhibition of RIP1/RIP3-mediated necroptotic signaling might be a novel strategy for the treatment of AML when combined with other differentiation inducers.