ABR, a Novel Inducer Of Transcription Factor C/EBPα, Is Necessary For Myeloid Differentiation and a Prognostic Factor In Acute Myeloid Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3814-3814
Author(s):  
Carolina Yaeko Namasu ◽  
Dennis Gerloff ◽  
Alexander Arthur Wurm ◽  
Daniela Braeuer-Hartmann ◽  
Jens-Uwe Hartmann ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Transcription Factor ◽  
Prognostic Factor ◽  
Myeloid Leukemia ◽  
Myeloid Differentiation ◽  
U937 Cells ◽  
Acute Myeloid

Abstract ABR (Active BCR-related) is the only protein in humans and mice closely homologous to BCR. BCR acts as a tumor suppressor in different cancers, such as chronic myeloid leukemia and meningiomas. A putative anti-oncogenic role of ABR has been shown in tumors of the central nervous system, such as medulloblastoma and astrocytomas, in which deletion of ABR was found. However, the role of ABR in hematopoiesis or leukemia remains unclear. We hypothesized that ABR might be important for myelopoiesis via increasing the expression of C/EBPα, a transcription factor known to be pivotal for myeloid differentiation and functionally impaired in acute myeloid leukemia (AML). In fact, we found that ABR expression is dramatically down-regulated (Figure 1, p=0.01) in bone marrow from AML patients (pts; n=63) compared to bone marrow (BM) mononuclear cells from healthy donors (n=3). In agreement with this finding, Abr is significantly increased during M-CSF and G-CSF-stimulated differentiation of primary wild type mouse BM cells (p<0.05). Additionally, we observe that ABR is necessary for monopoiesis induced by PMA (phorbol 12-myristate 13-acetate), since ABR knockdown in leukemic U937 cells results in a significant reduction of about 50% in the number of CD11b+ cells 48h after PMA treatment (p<0.05). Enforced ABR expression induces C/EBPα and its targets M-CSFR, G-CSFR and microRNA (miR)-223 in U937 cells (p<0.01). Moreover, we prove that ABR knockdown prevents induction of CEBPA, M-CSFR and G-CSFR during PMA-mediated differentiation (p<0.05). ABR overexpression blocks cell-cycle progression and down-regulates the known C/EBPα inhibitor E2F1 (p<0.01) in U937 cells, indicating the functional role of ABR as tumor suppressor. Those data suggest that ABR might induce CEBPA expression via inhibition of cell cycle activator E2F1. Finally, we are the first to identify ABR as a good prognostic factor in AML: patients with high ABR expression (median cut) survive significantly longer after allogeneic hematopoietic stem cell transplantation (Figure 2, p=0.04, log-rank test). Furthermore, high ABR expression associates with a low percentage of blasts in the peripheral blood (p=0.006) and high levels of antileukemic miR-181a (p<0.001). In conclusion, these data indicate that ABR, a novel inducer of C/EBPα, is necessary for myelopoiesis and a prognostic factor in AML. Raising ABR levels might be a goal for future therapeutics in AML. Disclosures: No relevant conflicts of interest to declare.

C/EBPα and MiR-182 Generate a Negative Feedback Loop Which Is Dysregulated in Acute Myeloid Leukemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 776-776
Author(s):  
Alexander Arthur Wurm ◽  
Dennis Gerloff ◽  
Daniela Braeuer-Hartmann ◽  
Christiane Katzerke ◽  
Jens-Uwe Hartmann ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Feedback Loop ◽  
Mononuclear Cells ◽  
Myeloid Differentiation ◽  
Negative Feedback Loop ◽  
U937 Cells ◽  
Leukemia Development ◽  
Acute Myeloid

Abstract The transcription factor CCAAT enhancer binding protein alpha (C/EBPα) is a master regulator of granulopoiesis and is silenced in approximately 50% of all acute myeloid leukemia (AML) cases. There are several mechanisms known how C/EBPα is inactivated in AML, including promoter hypermethylation, posttranslational modifications and mutations in the ORF of the CEBPA gene. MicroRNAs, a class of small non-coding RNAs, were identified as important regulators of normal hematopoiesis and leukemia development. We have already shown that microRNAs, such as miR-223, miR-34a and miR-30c, are essential elements in C/EBPα triggered granulocytic differentiation. But to our knowledge nothing is known about inactivation of C/EBPα by microRNAs in acute myeloid leukemia. In this study, we identified a novel network between C/EBPα and miR-182. In a next generation sequencing approach based on inducible K562-C/EBPα-ER cell line, we found miR-182 strongly downregulated by wildtype C/EBPα. We could further demonstrate an inverse correlation between C/EBPα protein amount and miR-182 expression level in several in vitro systems, including leukemic cell lines and G-CSF treated primary human CD34+progenitor cells. Additionally, C/EBPα and miR-182 showed reciprocal expression in sorted murine bone marrow subpopulations in vivo. To discover the mechanism how miR-182 is blocked by C/EBPα, we analyzed the minimal promoter region of miR-182 and performed chromatin immunoprecipitation (ChIP). Here, we could demonstrate a strong binding of C/EBPα to the miR-182 promoter, particularly to a conserved E2F binding site. Because E2F is a well known inhibitor of C/EBPα function, we tested whether E2F also effects miR-182 expression. An overexpression of E2F1 in U937 cells leads to an elevated miR-182 expression level. In addition, we measured the expression of miR-182 in bone marrow from AML patients regarding to their CEBPA mutation status. We could show that only patients with mutations in the C-terminal region of C/EBPα showed elevated miR-182 expression, while patients with N-terminal CEBPA mutations revealed no abnormal miR-182 expression compared to healthy donors or AML patients with no CEBPA mutation. The C-terminal domain of C/EBPα is necessary for E2F inhibition. These findings illustrate the importance of C/EBPα-E2F interaction during miR-182 regulation. Next, we found a highly conserved binding site of miR-182 in the 3’UTR of CEBPA itself, suggesting a possible negative feedback loop. To test this, we performed overexpression of miR-182 in U937 cells, umbilical cord blood mononuclear cells (UCB-MNCs) and primary blasts from AML patients. Here, we observed a strong reduction of C/EBPα protein after miR-182 overexpression in all cell types. Furthermore, we could demonstrate a direct binding of miR-182 to the 3’UTR of CEBPA via luciferase activity assay. Finally, we were interested in the functional impact of miR-182 in myeloid differentiation and leukemia development. We showed that enforced expression of miR-182 in U937 cells reduced the percentage of Mac-1 positive myeloid cells after treatment with all-trans retinoid acid (ATRA). Additionally, lentiviral overexpression of miR-182 induces a block of differentiation and hyperproliferation in G-CSF treated 32D cells and an enhanced replating capacity of primary mouse bone marrow mononuclear cells. Taken together, we identified miR-182 as novel oncogenic microRNA that directly blocks C/EBPα during myeloid differentiation and leukemia development. Thus, our data display a potential new strategy for therapeutics in C/EBPα dysregulated AML. Disclosures No relevant conflicts of interest to declare.

The Role of TWIST1 Gene Expression and Hypoxic Microenvironment in Acute Myeloid Leukemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3839-3839
Author(s):  
Emilia Carolina Malafaia ◽  
A. Mario Marcondes ◽  
Ekapun Karoopongse ◽  
Daniele Serehi ◽  
Maria de Lourdes L. F. Chauffaille ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Leukemia Cells ◽  
Twist1 Expression ◽  
Disease Biology ◽  
Acute Myeloid

Abstract TWIST1, a basic helix-loop-helix (bHLH) transcription factor, plays a critical role in mesodermal development and organogenesis. Overexpressed TWIST1 has been thoroughly related to epithelial-mesenchymal transition (EMT) in solid tumors (QIN Q et al., 2012) and has been described as an emerging risk factor in hematological neoplasms (MERINDOL et al., 2014). . Many questions remain to be addressed concerning to the role of TWIST1 in acute myeloid leukemia (AML). The understanding of TWIST1 in leukemia cells and its interaction with microenvironment can offer new insights in regards to disease biology and therapeutic targets for patients with AML. Objectives: 1) to evaluate the role of stroma contact and hypoxia in TWIST1 expression in myeloid cell lines. 2) To evaluate the functional impact of overexpressing TWIST1 on KG1a and PL21 cells. 3) To evaluate TWIST1 expression in primary cells of AML patients. Methods: In order to mimic bone marrow microenvironment, myeloid cells were co-cultured with mesenchymal HS5 cell line and PO2 1% was established with Smart -Trak¨ 2 (Sierra Instruments, Inc.) equipment. Quantitative mRNA was determined using TaqMan¨ Universal Master Mix (Applied Biosystems, Foster City, CA) and 3-step standard cycling conditions with sequence-specific primer TWIST1 normalized to the expression of β-actin. KG1a and PL21 cells were transduced with lentivirus vector carrying e-GFP ("enhanced green fluorescence protein") for stable expression of TWIST1. Transduced cells were sorted by FITC fluorochrome and then verified through western blot analysis with TWIST1 antibody. For quantification of apoptosis, cells were labeled with PE-conjugated antibody using annexin V-phycoerythrin and propidium iodide (BD Biosciences, USA). DAPI (4',6- diamidino-2-phenylindole dihydrochloride) was used to stain DNA and determine cell cycle information . Apoptosis and cell cycle were analyzed by FACS -Becton Dickinson Canto II (BD Biosciences). Statistical analysis was assessed with unpaired t test. Results: Hypoxia induced TWIST1 mRNA expression in OCIAML3, PL21, KG1a and ML1 cell lines (fold-increased 46.3, 29.8, 12.9 and 2.3 respectively). Cells expressing endogenous TWIST1 protein (OCIAML3 and ML1) showed resistance to apoptosis in a hypoxic microenvironment (normoxia versus hypoxia: OCI/AML3, 22.6 % vs 11.7% and ML1, 29.8% vs. 7.5%) in contrast, cells not expressing endogenous TWIST1 protein (KG1a and PL21) went to apoptosis in the same conditions. Thus, overexpressing TWIST1 in KG1a and PL21 induced apoptosis protection in hypoxia (KG1a unmodified vs. modified: 17.6 ± 6.3 vs. 2.8 ± 6.3, p=0.04; PL21 unmodified vs. modified: 26.9 ± 10.9 vs. 3.2 ± 0.6, p=0.04) (fig 1). We found increased TWIST1 mRNA levels in bone marrow samples of 23 AML patients (3.88 ± 1.59) compared with 5 healthy controls (0.54 ±0.25) (p= 0.02) (fig 2). Patients in the highest tertile of TWIST1 expression did not show differences in percentage of blasts in bone marrow and complete remission after treatment compared with patients in low and middle tertile. Conclusion: Our data suggest TWIST1 gene expression protects acute myeloid leukemia cells from apoptosis in a hypoxic microenvironment. Moreover, our results showed increased expression of TWIST1 in AML patients. Thus, TWIST1 is a potential gene involved in leukemogenesis and should be further explored to understand disease biology and potential therapeutic targets. Disclosures No relevant conflicts of interest to declare.

scholarly journals ABR, a novel inducer of transcription factor C/EBPα, contributes to myeloid differentiation and is a favorable prognostic factor in acute myeloid leukemia

Oncotarget ◽  
2017 ◽  
Vol 8 (61) ◽  
pp. 103626-103639 ◽  
Author(s):  
Carolina Yaeko Namasu ◽  
Christiane Katzerke ◽  
Daniela Bräuer-Hartmann ◽  
Alexander Arthur Wurm ◽  
Dennis Gerloff ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Transcription Factor ◽  
Prognostic Factor ◽  
Myeloid Leukemia ◽  
Myeloid Differentiation ◽  
Favorable Prognostic Factor ◽  
Factor C ◽  
Acute Myeloid

scholarly journals Leukemia stem cell-bone marrow microenvironment interplay in acute myeloid leukemia development

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Yiyi Yao ◽  
Fenglin Li ◽  
Jiansong Huang ◽  
Jie Jin ◽  
Huafeng Wang
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Chemotherapy Resistance ◽  
Bone Marrow Microenvironment ◽  
Cytotoxic Effects ◽  
High Relapse Rate ◽  
Underlying Mechanisms ◽  
Acute Myeloid

AbstractDespite the advances in intensive chemotherapy regimens and targeted therapies, overall survival (OS) of acute myeloid leukemia (AML) remains unfavorable due to inevitable chemotherapy resistance and high relapse rate, which mainly caused by the persistence existence of leukemia stem cells (LSCs). Bone marrow microenvironment (BMM), the home of hematopoiesis, has been considered to play a crucial role in both hematopoiesis and leukemogenesis. When interrupted by the AML cells, a malignant BMM formed and thus provided a refuge for LSCs and protecting them from the cytotoxic effects of chemotherapy. In this review, we summarized the alterations in the bidirectional interplay between hematopoietic cells and BMM in the normal/AML hematopoietic environment, and pointed out the key role of these alterations in pathogenesis and chemotherapy resistance of AML. Finally, we focused on the current potential BMM-targeted strategies together with future prospects and challenges. Accordingly, while further research is necessary to elucidate the underlying mechanisms behind LSC–BMM interaction, targeting the interaction is perceived as a potential therapeutic strategy to eradicate LSCs and ultimately improve the outcome of AML.

scholarly journals High HSPA8 expression predicts adverse outcomes of acute myeloid leukemia

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Jun Li ◽  
Zheng Ge
Keyword(s):  
Overall Survival ◽  
Acute Myeloid Leukemia ◽  
Prognostic Factor ◽  
Myeloid Leukemia ◽  
Cell Function ◽  
Adverse Outcomes ◽  
High Expression ◽  
Micro Rnas ◽  
Acute Myeloid

Abstract Background Acute myeloid leukemia (AML) remains one of the most common hematological malignancies, posing a serious challenge to human health. HSPA8 is a chaperone protein that facilitates proper protein folding. It contributes to various activities of cell function and also is associated with various types of cancers. To date, the role of HSPA8 in AML is still undetermined. Methods In this study, public datasets available from the TCGA (Cancer Genome Atlas) and GEO (Gene Expression Omnibus) were mined to discover the association between the expression of HSPA8 and clinical phenotypes of CN-AML. A series of bioinformatics analysis methods, including functional annotation and miRNA-mRNA regulation network analysis, were employed to investigate the role of HSPA8 in CN-AML. Results HSPA8 was highly expressed in the AML patients compared to the healthy controls. The high HSPA8 expression had lower overall survival (OS) rate than those with low HSPA8 expression. High expression of HSPA8 was also an independent prognostic factor for overall survival (OS) of CN-AML patients by multivariate analysis. The differential expressed genes (DEGs) associated with HSPA8 high expression were identified, and they were enriched PI3k-Akt signaling, cAMP signaling, calcium signaling pathway. HSPA8 high expression was also positively associated with micro-RNAs (hsa-mir-1269a, hsa-mir-508-3p, hsa-mir-203a), the micro-RNAs targeted genes (VSTM4, RHOB, HOBX7) and key known oncogenes (KLF5, RAN, and IDH1), and negatively associated with tumor suppressors (KLF12, PRKG1, TRPS1, NOTCH1, RORA). Conclusions Our research revealed HSPA8 as a novel potential prognostic factor to predict the survival of CN-AML patients. Our data also revealed the possible carcinogenic mechanism and the complicated microRNA-mRNA network associated with the HSPA8 high expression in AML.

scholarly journals Gpr44 Mediates the Selenium-Dependent Anti-Leukemic Effect in Acute Myeloid Leukemia

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1835-1835
Author(s):  
Fenghua Qian ◽  
Fenghua Qian ◽  
Diwakar Tukaramrao ◽  
Jiayan Zhou ◽  
Nicole Palmiero ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Murine Model ◽  
Myeloid Leukemia ◽  
Hematopoietic Stem ◽  
Cell Counts ◽  
Pparγ Agonist ◽  
Acute Myeloid

Abstract Objectives The relapse of acute myeloid leukemia (AML) remains a significant concern due to persistent leukemia stem cells (LSCs) that are not targeted by existing therapies. LSCs show sensitivity to endogenous cyclopentenone prostaglandin J (CyPG) metabolites that are increased by dietary trace element selenium (Se), which is significantly decreased in AML patients. We investigated the anti-leukemic effect of Se supplementation in AML via mechanisms involving the activation of the membrane-bound G-protein coupled receptor 44 (Gpr44) and the intracellular receptor, peroxisome proliferator-activated receptor gamma (PPARγ), by endogenous CyPGs. Methods A murine model of AML generated by transplantation of hematopoietic stem cells (HSCs- WT or Gpr44−/−) expressing human MLL-AF9 fusion oncoprotein, in the following experiments: To investigate the effect of Se supplementation on the outcome of AML, donor mice were maintained on either Se-adequate (Se-A; 0.08–0.1 ppm Se) or Se-supplemented (Se-S; 0.4 ppm Se) diets. Complete cell counts in peripheral blood were analyzed by hemavet. LSCs in bone marrow and spleen were analyzed by flow cytometry. To determine the role of Gpr44 activation in AML, mice were treated with Gpr44 agonists, CyPGs. LSCs in bone marrow and spleen were analyzed. Mice transplanted with Gpr44−/- AML cells were compared with mice transplanted with wild type AML cells and the progression of the disease was followed as above. To determine the role of PPARγ activation in AML, PPARγ agonist (Rosiglitazone, 6 mg/kg, i.p, 14 d) and antagonist (GW9662, 1 mg/kg, i.p. once every other day, 7 injections) were applied to Se-S mice transplanted with Gpr44−/- AML cells and disease progression was followed. Results Se supplementation at supraphysiological levels alleviated the disease via the elimination of LSCs in a murine model of AML. CyPGs induced by Se supplementation mediate the apoptosis in LSCs via the activation of Gpr44 and PPARγ. Conclusions Endogenous CyPGs produced upon supplementation with Se at supraphysiological levels improved the outcome of AML by targeting LSCs to apoptosis via the activation of two receptors, Gpr44 and PPARg. Funding Sources NIH DK 07,7152; CA 175,576; CA 162,665. Office of Dietary Supplements, USDA Hatch funds PEN04605, Accession # 1,010,021 (KSP, RFP).

scholarly journals Silencing long non-coding RNA XIST suppresses drug resistance in acute myeloid leukemia through down-regulation of MYC by elevating microRNA-29a expression

2020 ◽  
Vol 26 (1) ◽  
Author(s):  
Chong Wang ◽  
Lingling Li ◽  
Mengya Li ◽  
Weiqiong Wang ◽  
Yanfang Liu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Drug Resistance ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Bone Marrow Cells ◽  
Cellular Viability ◽  
Down Regulation ◽  
Acute Myeloid

Abstract Background Long non-coding RNAs (lncRNAs) are biomarkers participating in multiple disease development including acute myeloid leukemia (AML). Here, we investigated molecular mechanism of X Inactive-Specific Transcript (XIST) in regulating cellular viability, apoptosis and drug resistance in AML. Methods XIST, miR-29a and myelocytomatosis oncogene (MYC) expression in AML bone marrow cells collected from 62 patients was evaluated by RT-qPCR and Western blot analysis. Besides, the relationship among XIST, miR-29a and MYC was analyzed by dual luciferase reporter assay, RIP, and RNA pull down assays. AML KG-1 cells were treated with anti-tumor drug Adriamycin. The role of XIST/miR-29a/MYC in cellular viability, apoptosis and drug resistance in AML was accessed via gain- and loss-of-function approaches. At last, we evaluated role of XIST/miR-29a/MYC on tumorigenesis in vivo. Results XIST and MYC were up-regulated, and miR-29a was down-regulated in AML bone marrow cells. Silencing XIST inhibited cellular activity and drug resistance but promoted cellular apoptosis of KG-1 cells by down-regulating MYC. XIST inhibited miR-29a expression to up-regulate MYC. Moreover, silencing XIST inhibited tumorigenesis of AML cells in vivo. Conclusions Overall, down-regulation of XIST decreased MYC expression through releasing the inhibition on miR-29a, thereby reducing drug resistance, inhibiting viability and promoting apoptosis of AML cells.

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