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In this issue of PIH, we asked four researchers to write about basic research on the molecular mechanisms of the development of myeloid malignancies, in particular two epigenetic regulation and two space- and time-dependent factors. Regarding epigenomic regulation, Dr. Yang reviewed ASXL1 , a polycomb modifier gene that is often mutated in myeloid malignancies, but also in clonal hematopoiesis in healthy elderly people, and Dr. Vu reviewed RNA modifications, which are critical for development and tissue homeostasis, and are now recognized as an important driver for cancer development. Regarding spatiotemporal factors, Dr. Inoue reviewed the role of extracellular vesicles in leukemic stem cell niches. As some cancers develop preferentially in infancy or old age, Dr. Osato discussed the time-specific development of leukemia involving the RUNX1-ETO mutation, which is often found in leukemia in adolescents and young adults. Recent studies on hematopoietic development have shown that hematopoietic stem cells do not generate multipotent progenitor cells, but that these cells develop in parallel. We hope that reconsideration of the definition of leukemic stem cells and their origin will help us understand the regulatory mechanisms of these cells, but also enable us to develop future therapies by targeting factors that regulate the leukemic stem cell and the niche.

High Mobility Group AT-hook 2 (HMGA2) is a chromatin modifier and its overexpression has been found in patients with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Level of Hmga2 expression is fine-tuned by Lin28b-Let-7 axis and Polycomb Repressive Complex 2, in which deletion of Ezh2 leads to activation of Hmga2 expression in hematopoietic stem cells. To elucidate the mechanisms by which the overexpression of HMGA2 helps transformation of stem cells harboring a driver mutation of TET2 , we generated an Hmga2- expressing Tet2 -deficient mouse model showing the progressive phenotypes of MDS and AML. The overexpression of Hmga2 remodeled the transcriptional program of Tet2-deficient stem and progenitor cells, leading to the impaired differentiation of myeloid cells. Furthermore, Hmga2 was bound to a proximal region of Igf2bp2 oncogene, and activated its transcription, leading to enhancing self-renewal of Tet2 -deficient stem cells that was suppressed by inhibition of the DNA binding of Hmga2. These combinatory effects on the transcriptional program and cellular function were not redundant to those in Tet2 -deficient cells. The present results elucidate that Hmga2 targets key oncogenic pathways during the transformation and highlight the Hmga2-Igf2bp2 axis as a potential target for therapeutic intervention.

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is an aggressive subtype of acute leukemia, the cell of origin of which is considered to be precursors of plasmacytoid dendritic cells (pDCs). Since translocation (6;8)(p21;q24) is a recurrent anomaly for BPDCN, we demonstrate that a pDC-specific super-enhancer of RUNX2 is associated with the MYC promoter due to t(6;8). RUNX2 ensures the expression of pDC-signature genes in leukemic cells, but also confers survival and proliferative properties in BPDCN cells. Furthermore, the pDC-specific RUNX2 super-enhancer is hijacked to activate MYC in addition to RUNX2 expression, thereby promoting the proliferation of BPDCN. We also demonstrate that the transduction of MYC and RUNX2 is sufficient to initiate the transformation of BPDCN in mice lacking Tet2 and Tp53 , providing a model that accurately recapitulates the aggressive human disease and gives an insight into the molecular mechanisms underlying the pathogenesis of BPDCN. Translocation of (6;8)(p21;q24), a recurrent abnormality of blastic plasmacytoid dendritic cell neoplasm, involves adjacent MYC and RUNX2 regions. Here, the authors show that the RUNX2 super-enhancer is hijacked to activate MYC in addition to RUNX2 expression, promoting the development of this cancer.

High mobility group AT-hook 2 (Hmga2) is a chromatin modifier protein that plays a critical role in fetal development and leukemia propagation by binding to chromatin and DNA via its AT-hook domains. However, the molecular mechanisms by which Hmga2 activates the expression of target genes to drive the self-renewal of hematopoietic stem cells (HSCs) remain unclear. We generated Rosa26 locus Hmga2 conditional knock-in mice and found that overexpression of Hmga2 promoted self-renewal of normal HSCs, but maintained their fitness in bone marrow, and consequently was not sufficient to initiate malignancy. This result is consistent with previous findings showing that Hmga2 is a proto-oncogene. We also assessed the cellular functions of Hmga2 mutants lacking functional domains and demonstrated that the C-terminus acidic domain of Hmga2 and the domain’s linker region were critical for activating genes involved in stem cell signatures, such as the Igf2bp2 gene, to drive proliferation of HSCs. In contrast, overexpression of Hmga1, a member of the Hmga family with a different linker region, did not drive proliferation of HSCs. Our results reveal a critical role for the acidic domain of Hmga2 and the domain’s linker region in modulating the transcription and self-renewal functions of HSCs.

EZH1 and EZH2 are enzymatic components of polycomb repressive complex (PRC) 2, which catalyzes histone H3K27 tri-methylation (H3K27me3) to repress the transcription of PRC2 target genes. We previously reported that the hematopoietic cell-specific Ezh2 deletion ( Ezh2 Δ/Δ ) induced a myelodysplastic syndrome (MDS)-like disease in mice. We herein demonstrated that severe PRC2 insufficiency induced by the deletion of one allele Ezh1 in Ezh2 -deficient mice ( Ezh1 +/− Ezh2 Δ/Δ ) caused advanced dyserythropoiesis accompanied by a differentiation block and enhanced apoptosis in erythroblasts. p53, which is activated by impaired ribosome biogenesis in del(5q) MDS, was specifically activated in erythroblasts, but not in hematopoietic stem or progenitor cells in Ezh1 +/− Ezh2 Δ/Δ mice. Cdkn2a , a major PRC2 target encoding p19 Arf , which activates p53 by inhibiting MDM2 E3 ubiquitin ligase, was de-repressed in Ezh1 +/− Ezh2 Δ/Δ erythroblasts. The deletion of Cdkn2a as well as p53 rescued dyserythropoiesis in Ezh1 +/− Ezh2 Δ/Δ mice, indicating that PRC2 insufficiency caused p53-dependent dyserythropoiesis via the de-repression of Cdkn2a . Since PRC2 insufficiency is often involved in the pathogenesis of MDS, the present results suggest that p53-dependent dyserythropoiesis manifests in MDS in the setting of PRC2 insufficiency.

Loss-of-function mutations of EZH2 , a catalytic component of polycomb repressive complex 2 (PRC2), are observed in ~\\n10% of patients with myelodysplastic syndrome (MDS), but are rare in acute myeloid leukaemia (AML). Recent studies have shown that EZH2 mutations are often associated with RUNX1 mutations in MDS patients, although its pathological function remains to be addressed. Here we establish an MDS mouse model by transducing a RUNX1S291fs mutant into hematopoietic stem cells and subsequently deleting Ezh2 . Ezh2 loss significantly promotes RUNX1S291fs-induced MDS. Despite their compromised proliferative capacity of RUNX1S291fs/Ezh2 -null MDS cells, MDS bone marrow impairs normal hematopoietic cells via selectively activating inflammatory cytokine responses, thereby allowing propagation of MDS clones. In contrast, loss of Ezh2 prevents the transformation of AML via PRC1-mediated repression of Hoxa9 . These findings provide a comprehensive picture of how Ezh2 loss collaborates with RUNX1 mutants in the pathogenesis of MDS in both cell autonomous and non-autonomous manners. Mutations in the EZH2 gene are found in myelodysplastic syndrome (MDS) and are often accompanied by mutations in RUNX1 . Here, the authors develop a mouse model of MDS and show that EZH2 loss enhances the RUNX1-mediated MDS pathology.

The negative regulator of p53, MDM2, is frequently overexpressed in acute myeloid leukemia (AML) that retains wild-type TP53 alleles. Targeting of p53-MDM2 interaction to reactivate p53 function is therefore an attractive therapeutic approach for AML. Here we show that an orally active inhibitor of p53-MDM2 interaction, DS-5272, causes dramatic tumor regressions of MLL-AF9-driven AML in vivo with a tolerable toxicity. However, the antileukemia effect of DS-5272 is markedly attenuated in immunodeficient mice, indicating the critical impact of systemic immune responses that drive p53-mediated leukemia suppression. In relation to this, DS-5272 triggers immune-inflammatory responses in MLL-AF9 cells including upregulation of Hif1α and PD-L1, and inhibition of the Hif1α-PD-L1 axis sensitizes AML cells to p53 activation. We also found that NK cells are important mediators of antileukemia immunity. Our study showed the potent activity of a p53-activating drug against AML, which is further augmented by antitumor immunity. MDM2 is frequently overexpressed in acute myeloid leukaemia leading to p53 inactivation. Here, the authors are demonstrating that an inhibitor of p53-MDM2 interaction, DS-5272, induce in vivo tumour regression through immune response regulation.

Cholangiocarcinoma (CCA) is the second most common type of hepatic cancer. In east and southeast Asia, intrahepatic CCA is caused predominantly by infection of Opisthorchis viverrini and Clonorchis sinensis, two species of parasitic liver flukes. In this review, we present molecular evidence that liver fluke-associated CCAs have enhanced features of epithelial–mesenchymal transition (EMT) in bile duct epithelial cells (cholangiocytes) and that some of those features are associated with mis-regulation at the epigenetic level. We hypothesize that both direct and indirect mechanisms underlie parasitic infection-induced EMT in CCA.

Polycomb proteins function in the maintenance of gene silencing via post-translational modifications of histones and chromatin compaction. Genetic and biochemical studies have revealed that the repressive function of Polycomb repressive complexes (PRCs) in transcription is counteracted by the activating function of Trithorax-group complexes; this balance fine-tunes the expression of genes critical for development and tissue homeostasis. The function of PRCs is frequently dysregulated in various cancer cells due to altered expression or recurrent somatic mutations in PRC genes. The tumor suppressive functions of EZH2-containing PRC2 and a PRC2-related protein ASXL1 have been investigated extensively in the pathogenesis of hematological malignancies, including myeloproliferative neoplasms (MPN). BCOR, a component of non-canonical PRC1, suppresses various hematological malignancies including MPN. In this review, we focus on recent findings on the role of PRCs in the pathogenesis of MPN and the therapeutic impact of targeting the pathological functions of PRCs in MPN.

Epigenetic regulation is required not only for development, but also for tissue homeostasis, which is maintained via the self-renewal and differentiation of somatic stem cells. Accumulating evidence suggests that epigenetic regulators play critical roles in the maintenance of both self-renewing hematopoietic stem cells and leukemic stem cells. Recent genome-wide comprehensive analyses have identified mutations in epigenetic regulator genes, including genes whose products modify DNA and histones in hematological malignancies. Among these epigenetic regulators, repressive histone modifications by Polycomb-group complexes have been most fully characterized in hematopoietic stem cells, and are recognized as general regulators of stem cells. Hematopoietic stem cells are controlled by both cell-intrinsic and -extrinsic regulators, including transcription factors, signal transduction pathways, and niche factors. However, there is little insight into the mechanism of how epigenetic regulators act in concert with these factors to ensure blood homeostasis. In this review, we highlight recent findings in epigenetic regulation of hematopoiesis with emphasis on the role of Polycomb-group proteins and DNA-methylation modulators in hematopoietic stem cells and their progeny.