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LSD1 inhibition by tranylcypromine derivatives interferes with GFI1-mediated repression of PU.1 target genes and induces differentiation in AML

ABSTRACT: Lsd1KO and ATRA treatment in Hoxa9/Meis1- and MN1-transformed myeloid progenitor cells LSD1 has emerged as a promising epigenetic target in the treatment of acute myeloid leukemia (AML). Inhibition of LSD1 has been shown to induce differentiation and facilitate the responsiveness of AML cells to all-trans retinoic acid. We used two murine AML models based on retroviral overexpression of Hoxa9/Meis1 (H9M) or MN1 to study the effect of Lsd1 knockout in AML. The conditional knockout of Lsd1 resulted in differentiation with both granulocytic and monocytic features in H9M-induced AML cells and enhanced their responsiveness towards ATRA treatment, which was not seen in MN1-driven AML. It also extended the survival of mice with H9M-driven AML. To identify molecular changes associated with the loss of Lsd1, we performed whole transcriptome analysis by RNA sequencing of H9M and MN1 Lsd1 fl/fl (Ctrl) and Lsd1cKO cells with or without ATRA treatment. The experiment was done in triplicates. The conditional knockout led to a broad increase in the expression of multiple genes regulated by the important myeloid transcription factors GFI1 and PU.1. These include the transcription factors GFI1B and IRF8, which may be relevant for the observed differentiation response.

ORGANISM(S): Mus musculus

PROVIDER: GSE110178 | GEO | 2019/01/08

REPOSITORIES: GEO

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Project description:Myeloid differentiation is blocked in acute myeloid leukemia (AML), but the molecular mechanisms are not well characterized. MN1 is overexpressed in some AML patients and confers resistance to all-trans retinoic acid (ATRA)-induced differentiation. To understand the role of MN1 as a transcriptional regulator in myeloid differentiation, we fused transcriptional activation (VP16) or repression (M33) domains with MN1 and characterized these cells in vivo. Transcriptional activation of MN1 target genes induced myeloproliferative disease with long latency and differentiation potential to mature neutrophils. A large proportion of differentially expressed genes between leukemic MN1 and differentiation-permissive MN1VP16 cells belonged to the immune response pathway like Irf8 and Ccl9. As MN1 is a co-factor of MEIS1 and RARA, we compared chromatin occupancy between MN1, MEIS1 and RARA. Immune response genes that were upregulated in MN1VP16 cells were co-targeted by MN1 and MEIS1, but not RARA, suggesting that myeloid differentiation is blocked through transcriptional repression of shared target genes of MN1 and MEIS1. Constitutive expression of Irf8 or its target gene Ccl9 identified these genes as potent inhibitors of MN1-induced leukemia. Our data show that MN1 prevents activation of the immune response pathway, and suggest that restoration of Irf8 signalling as a novel therapeutic target in AML. C57BL/6J bone marrow cells were harvested from mice treated for 4 days with 150 mg 5-fluorouracil/kg and stimulated for 48 hours in DMEM supplemented with 15% FBS, 10 ng/mL hIL6, 6ng/mL mIL3, and 20ng/mL mSCF. The cells were infected with MN1 or MN1VP16 retroviral constructs by cocultivation with irradiated E86 viral producer cells in the presence of 5μg/mL protamine sulfate (Sigma) for 48 hours and then transplanted into lethally irradiated syngeneic recipient mice. 4 weeks after transplantation MN1, MN1VP16 GFP+ cells and Gr1+/CD11b+ bone marrow cells were FACS-sorted and analyzed by Affymetrix GeneChip Mouse 430 2.0 (43.000 probes) microarrays

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LSD1 inhibition by tranylcypromine derivatives interferes with GFI1-mediated repression of PU.1 target genes and induces differentiation in AML

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