Myeloblastin is an Myb target gene: mechanisms of regulation in myeloid leukemia cells growth-arrested by retinoic acid

Blood ◽  
2001 ◽  
Vol 97 (8) ◽  
pp. 2449-2456 ◽  
Author(s):  
Pierre G. Lutz ◽  
Anne Houzel-Charavel ◽  
Christel Moog-Lutz ◽  
Yvon E. Cayre
Keyword(s):  
Retinoic Acid ◽  
Myeloid Leukemia ◽  
Target Genes ◽  
Growth Arrest ◽  
Constitutive Expression ◽  
Myeloid Cell ◽  
Leukemia Cells ◽  
Down Regulation ◽  
Novel Target ◽  
Stimulating Factor

Abstract A pivotal role has been assigned to Myb in the control of myeloid cell growth. Although Myb is a target of retinoic acid, little is known about the mechanisms by which it may contribute to induced growth arrest in leukemia cells. Indeed, few Myb target genes are known to be linked to proliferation. Myeloblastin is involved in the control of proliferation in myeloid leukemia cells. It is expressed early during hematopoiesis and is a granulocyte colony-stimulating factor–responsive gene. Myeloblastin can confer factor-independent growth to hematopoietic cells, an early step in leukemia transformation. The myeloblastin promoter contains PU.1, C/EBP, and Myb binding sites, each of which are critical for constitutive expression in myeloid cells. Inhibition of myeloblastin expression in leukemia cells growth-arrested by retinoic acid is demonstrated to depend on Myb down-regulation. Myb is shown to induce myeloblastin expression and abolish its down-regulation by retinoic acid. Altogether, the data offer a clue as to how a myeloid-specific transcriptional machinery can be accessible to regulation by retinoic acid and point to myeloblastin as a novel target of Myb. This link between Myb and myeloblastin suggests a previously nonidentified Myb pathway through which growth arrest is induced by retinoic acid in myeloid leukemia cells.

Olfactomedin 4 Mediates All-Trans Retinoic Acid Induced Growth Inhibition, Differentiation and Apoptosis in Myeloid Leukemia Cells and Inhibits 4E-BP1 Phosphorylation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2395-2395
Author(s):  
Wenli Liu ◽  
Hyun W Lee ◽  
Griffin P Rodgers
Keyword(s):  
Retinoic Acid ◽  
Chronic Myeloid Leukemia ◽  
Growth Inhibition ◽  
Myeloid Leukemia ◽  
Leukemia Cells ◽  
Over Expression ◽  
Retinoic Acids ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Novel Target

Abstract Abstract 2395 Poster Board II-372 Introduction: All-trans retinoic acid (ATRA) has been shown to induce cellular differentiation and growth inhibition of acute promyelocytic leukemia cells. Clinical application of ATRA has proved extremely successful in inducing clinical remission in most acute promyelocytic leukemia patients. Although the mechanisms of retinoid-dependent gene transcription regulation are well understood, the target genes that mediate retinoid-induced biological responses still remain to be defined. Olfactomedin 4 (OLFM4, also called hGC-1 and GW112) is a member of olfactomedin-related glycoprotein family. It is constitutively expressed in myeloid cells and gastrointestinal tract. It has been recently reported that OLFM4 expression is up-regulated in stomach and colon cancer patients. The purpose of this study is to examine its expression in myeloid leukemia patients, its regulation by ATRA and potential biological functions in myeloid leukemia. Results: 1) In this study, we found that OLFM4 expression was up-regulated in the peripheral leukocytes of chronic myeloid leukemia patients (91%, 22 cases) and acute myeloid leukemia patients (30%, 10 cases). OLFM4 expression in accelerated phase of chronic myeloid leukemia patients was significantly higher than that in chronic phase. 2) We identified that OLFM4 is a novel target gene of retinoic acids in myeloid leukemia cells. Treatment of HL-60 cells with ATRA and 9-cis-RA induced OLFM4 expression. The expression level of OLFM4 is correlated with the myeloid cell differentiation stage. Deletion analysis led to the identification of a positive retinoic acid response element (DR5) and a negative response element (DR1) within OLFM4 promoter. Furthermore, electrophoretic mobility-shift assays and transfection study in COS-7 cells demonstrated that RARα/RXRα binds to the DR5 site and mediates ATRA induced transactivation of OLFM4 promoter. 3) We showed that OLFM4 over-expression in HL-60 cells lead to growth inhibition, differentiation and apoptosis and potentates ATRA mediated these effects. Conversely, silencing of endogenous OLFM4 by lentiviral shRNA against OLFM4 in AML-193 cells reduces ATRA induced growth inhibition, differentiation and apoptosis. 4) We further investigated the molecular mechanism that OLFM4 is involved in leukemia cell growth and differentiation. We found that over-expression of OLFM4 in HL60 cells inhibited ATRA induced phosphorylation of translation repressor 4E-BP1. This inhibitory effect was further confirmed in 293T cells, in which over-expression of OLFM4 inhibits 4E-BP1 phosphorylation at all four phosphorylation sites (Thr37/46, Thr70 and Ser65). Application of lentiviral shRNA against OLFM4 in AML-193 increased phosphorylation of 4E-BP1 compared with control shRNA. Over-expression of OLFM4 in HL60 cells does not affect phosphorylation of Akt, p70S6 kinase, GSK3β and ERK1/2 . Conclusion: 1) OLFM4 expression is up-regulated in chronic and acute myeloid leukemia patients and is correlated with the stage of chronic myeloid leukemia. 2) OLFM4 is a novel target gene of retinoic acids. RARα/RXRα binds to the DR5 site of OLFM4 promoter and mediates the ATRA induced transactivation of OLFM4. 3) OLFM4 mediates ATRA induced growth inhibition, differentiation and apoptosis of myeloid leukemia cells. 4) OLFM4 acts an inhibitor of 4E-BP1 phosphorylation down stream of Akt and mTOR, suggesting OLFM4 may inhibit protein synthesis that is observed with the differentiation of myeloid leukemia cells. Our results suggest that OLFM4 up-regulation in the leukocytes of leukemia patients might have a feedback effect to restrain cell growth, and induce differentiation and apoptosis rather than act as an initiator of leukemiagenesis. As OLFM4 is a secreted glycoprotein, it could represent a promising therapeutic agent in the treatment of myeloid leukemia patients. The effect of purified OLFM4 on leukemia cells is currently under investigation. Disclosures: No relevant conflicts of interest to declare.

Upregulation of hGC-1 Expression by Hypomethylation, Retinoic Acid and Interferons in Human Myeloid Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4486-4486
Author(s):  
Wenli Liu ◽  
Griffin P Rodgers
Keyword(s):  
Retinoic Acid ◽  
Myeloid Leukemia ◽  
Target Gene ◽  
Leukemia Cells ◽  
Luciferase Reporter ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Response Element ◽  
Gene Assay ◽  
Stimulating Factor

Abstract hGC-1 (Granulocyte colony stimulating factor induced gene-1, also called GW112 and olfactomedin 4) was first identified in human myeloid precursor cells induced by granulocyte colony stimulating factor (G-CSF). It is a myeloid lineage and differentiation stage specific gene. Its expression, regulation and biological function, specifically in myeloid cells, are still poorly understood. In this study, we analyzed the hGC-1 gene expression in leukemia patients and further investigated the mechanism of hGC-1 gene regulation in leukemia cells. We found that hGC-1 was overexpressed in myeloid leukemia patients compared with normal individuals in peripheral blood leukocytes (p<0.01) and its expression in accelerated phase of chronic myeloid leukemia (CML) patients was significantly higher than that in chronic phase (p<0.01) using a dot blot and quantitative RT-PCR analysis. Hypomethylation of CpG sites in the promoter of hGC-1 gene were observed in CML patients by pyrosequence and 5-aza-2′-deoxycytidine induced hGC-1 expression in myeloid leukemia cells, suggesting that promoter CpG methylation status affects the expression of hGC-1 gene. All-trans-retinoic acid (ATRA) and interferons (IFNs) are active anti-leukemia agents. ATRA and IFNs have shown synergistic interactions in various experimental conditions and represent a potentially useful therapeutic combination in the treatment of various types of leukemia. However, the target genes and molecular basis of these interactions still needs to be further elucidated. Here, we identified that hGC-1 was a target gene of RA in myeloblastic leukemia cells. Treatment with ATRA induced hGC-1 expression in HL-60 cells and enhanced hGC-1 expression in AML-193 and GDM-1 cells. Deletion analysis led to the identification of a positive retinoic acid response element (DR5) and a negative response element (DR1) within hGC-1 promoter. Furthermore, electrophoretic mobility-shift assays demonstrated that RARa/RXRa binds to the DR5 site. Transfection study in COS-7 cells revealed RARa/RXRa mediated the RA induced transactivation of hGC-1 promoter. We also found that hGC-1 was an early responsive gene of IFN a and b in myeloid leukemia cells (HL-60, AML-193 and GDM-1). An effective interferon-stimulated response element (ISRE) was identified in the promoter of hGC-1 gene by examining the deletion mutants in luciferase reporter gene assay. Combined application of ATRA and IFNa and IFNb enhanced hGC-1 expression synergistically. Taken together, hGC-1 is identified as a novel target gene of methylation modification, RA and IFNs in myeloid leukemia cells. Our results suggest that hGC-1 is a potential marker for CML stage and may play a role in retinoic acid and interferon induced biological effects in leukemia cells.

scholarly journals Olfactomedin 4 is a novel target gene of retinoic acids and 5-aza-2′-deoxycytidine involved in human myeloid leukemia cell growth, differentiation, and apoptosis

Blood ◽  
2010 ◽  
Vol 116 (23) ◽  
pp. 4938-4947 ◽  
Author(s):  
Wenli Liu ◽  
Hyun Woo Lee ◽  
Yueqin Liu ◽  
Ruihong Wang ◽  
Griffin P. Rodgers
Keyword(s):  
Acute Myeloid Leukemia ◽  
Retinoic Acid ◽  
Growth Inhibition ◽  
Myeloid Leukemia ◽  
Target Gene ◽  
Down Regulation ◽  
Receptor Alpha ◽  
Retinoic Acids ◽  
Novel Target ◽  
Acute Myeloid

Abstract Clinical application of retinoic acids (RAs) and demethylation agents has proven to be effective in treating certain myeloid leukemia patients. However, the target genes that mediate these antileukemia activities are still poorly understood. In this study, we identified olfactomedin 4 (OLFM4), a myeloid-lineage–specific gene from the olfactomedin family, as a novel target gene for RAs and the demethylation agent, 5-aza-2′-deoxycytidine. We demonstrated that the retinoic acid receptor alpha/retinoic X receptor alpha heterodimer binds to a retinoic acid response-element (DR5) site in the OLFM4 promoter and mediates all-trans-retinoic acid (ATRA)–induced transactivation of the OLFM4 gene. OLFM4 overexpression in HL-60 cells led to growth inhibition, differentiation, and apoptosis, and potentiated ATRA induction of these effects. Conversely, down-regulation of endogenous OLFM4 in acute myeloid leukemia-193 cells compromised ATRA-induced growth inhibition, differentiation, and apoptosis. Overexpression of OLFM4 in HL-60 cells inhibited constitutive and ATRA-induced phosphorylation of the eukaryote initiation factor 4E-binding protein 1 (4E-BP1), whereas down-regulation of OLFM4 protein in acute myeloid leukemia-193 cells increased 4E-BP1 phosphorylation, suggesting that OLFM4 is a potent upstream inhibitor of 4E-BP1 phosphorylation/deactivation. Thus, our study demonstrates that OLFM4 plays an important role in myeloid leukemia cellular functions and induction of OLFM4-mediated effects may contribute to the therapeutic value of ATRA.

AZD1152, a novel and selective aurora B kinase inhibitor, induces growth arrest, apoptosis, and sensitization for tubulin depolymerizing agent or topoisomerase II inhibitor in human acute leukemia cells in vitro and in vivo

Blood ◽  
2007 ◽  
Vol 110 (6) ◽  
pp. 2034-2040 ◽  
Author(s):  
Jing Yang ◽  
Takayuki Ikezoe ◽  
Chie Nishioka ◽  
Taizo Tasaka ◽  
Ayuko Taniguchi ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Topoisomerase Ii ◽  
Growth Arrest ◽  
Leukemia Cells ◽  
Aurora B ◽  
Thymidine Uptake ◽  
Aurora B Kinase ◽  
Aurora Kinases ◽  
Topoisomerase Ii Inhibitor

Abstract Aurora kinases play an important role in chromosome alignment, segregation, and cytokinesis during mitosis. We have recently shown that hematopoietic malignant cells including those from acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) aberrantly expressed Aurora A and B kinases, and ZM447439, a potent inhibitor of Aurora kinases, effectively induced growth arrest and apoptosis of a variety of leukemia cells. The present study explored the effect of AZD1152, a highly selective inhibitor of Aurora B kinase, on various types of human leukemia cells. AZD1152 inhibited the proliferation of AML lines (HL-60, NB4, MOLM13), ALL line (PALL-2), biphenotypic leukemia (MV4-11), acute eosinophilic leukemia (EOL-1), and the blast crisis of chronic myeloid leukemia K562 cells with an IC50 ranging from 3 nM to 40 nM, as measured by thymidine uptake on day 2 of culture. These cells had 4N/8N DNA content followed by apoptosis, as measured by cell-cycle analysis and annexin V staining, respectively. Of note, AZD1152 synergistically enhanced the antiproliferative activity of vincristine, a tubulin depolymerizing agent, and daunorubicin, a topoisomerase II inhibitor, against the MOLM13 and PALL-2 cells in vitro. Furthermore, AZD1152 potentiated the action of vincristine and daunorubicin in a MOLM13 murine xenograft model. Taken together, AZD1152 is a promising new agent for treatment of individuals with leukemia. The combined administration of AZD1152 and conventional chemotherapeutic agent to patients with leukemia warrants further investigation.

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