Requirement For CDK6 In MLL-Rearranged Acute Myeloid Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3782-3782
Author(s):  
Theresa Placke ◽  
Katrin Faber ◽  
Atsushi Nonami ◽  
Helmut R. Salih ◽  
Stephen M. Sykes ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Small Molecule ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Myeloid Differentiation ◽  
Cell Cycle Distribution ◽  
Hematopoietic Stem ◽  
Acute Myeloid

Abstract Chromosomal rearrangements involving the H3K4 methyltransferase MLL trigger aberrant gene expression programs in hematopoietic stem and progenitor cells and give rise to an aggressive subtype of acute myeloid leukemia (AML) that is associated with intermediate or poor survival. Insights into MLL fusion-mediated leukemogenesis have not yet translated into better therapies in the clinic, in part because mutant MLL is difficult to target directly and it is incompletely understood which of the genes affected by altered epigenetic regulation in MLL-rearranged AML are responsible for malignant transformation. To search for essential signaling pathways in MLL-rearranged AML that might serve as new therapeutic targets, we performed loss-of-function RNA interference (RNAi) screens in 5 AML cell lines (NOMO-1, THP-1, OCI-AML3, HL-60, U937) using a lentiviral short hairpin RNA (shRNA) library, and observed that the cell cycle regulator CDK6, but not its functional homolog CDK4, was preferentially required by MLL-AF9pos NOMO-1 and THP-1 cells. The enhanced CDK6 dependence of MLL-rearranged cells was confirmed in an expanded panel of AML cell lines (MLL-rearranged, n=6; MLL wildtype [WT], n=4) that also included cell lines harboring other MLL fusions (MLL-AF4 and MLL-AF6), and the RNAi-induced phenotype was countered by overexpression of an shRNA-resistant CDK6 cDNA. Stable knockdown of MLL-AF9 in MLL-AF9pos cell lines and overexpression of MLL-AF9 in WT MLL-expressing cell lines, normal human CD34pos cells, or Ba/F3 murine pro-B cells led to concordant changes in CDK6 mRNA and protein levels that resembled those of HOXA9, a known MLL-AF9 target, indicating that CDK6 is rendered essential via transcriptional activation by truncated MLL. Analysis of cell cycle distribution, apoptosis induction, and myeloid differentiation demonstrated that the differential growth-inhibitory effect of CDK6 suppression was mainly attributable to myeloid differentiation, as MLL-AF9pos cell lines upregulated CD11b expression and assumed a more mature, macrophage-like morphology upon CDK6 knockdown, effects not observed in WT MLL-expressing cells. Furthermore, the immature phenotype of NOMO-1 cells could be rescued by overexpression of an shRNA-resistant CDK6 cDNA. Consistent with the observations in AML cell lines, knockdown of Cdk6 also impaired the proliferation and in vitro clonogenic activity of primary murine bone marrow (BM) cells stably transduced with MLL-AF9, whereas cells expressing another leukemogenic fusion gene (MOZ-TIF2) and Ba/F3 cells were largely unaffected. We also expressed MLL-AF9 in unfractionated BM derived from Cdk6 knockout mice and observed that colony numbers were gradually reduced in cultures initiated with Cdk6+/- and Cdk6-/- BM compared to WT BM. Furthermore, most of the colonies obtained were small and displayed loose morphology in contrast to the large, dense, blast-like colonies seen in cultures initiated with transduced WT BM. We are currently investigating whether Cdk6 is also required for AML development and propagation in vivo using a murine BM transplantation model of MLL-AF9-induced leukemia. The context-dependent effects of lowering CDK6 expression could be recapitulated in cell lines and primary human AML specimens using palbociclib (also known as PD-0332991), a small-molecule inhibitor of CDK4 and CDK6 enzymatic activity that is in clinical development as an anticancer agent. We are currently devising strategies to combine this compound with cytotoxic chemotherapy as well as other targeted therapeutics, such as small-molecule bromodomain inhibitors, to maximize killing of MLL-rearranged AML cells. Together, our data (1) identify CDK6 as a critical and potentially “actionable” effector of MLL fusion proteins in leukemogenesis, (2) link the catalytic activity of CDK6 to arrested myeloid differentiation in MLL-rearranged AML, and (3) underscore that cell cycle regulators thought to normally act redundantly may have distinct functions in different genetic contexts. Disclosures: No relevant conflicts of interest to declare.

Small Molecule Inhibition of SIRT Activity: A Novel Therapeutic Approach for Acute Myeloid Leukemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 84-84
Author(s):  
Steffan T Nawrocki ◽  
Claudia M Espitia ◽  
Kevin R. Kelly ◽  
William G. Bornmann ◽  
Jennifer S Carew
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Small Molecule ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Clonogenic Survival ◽  
Altered Gene ◽  
Induced Apoptosis ◽  
Acute Myeloid ◽  
Novel Therapeutic

Abstract Abstract 84 New therapeutic strategies are urgently needed to improve clinical outcomes for patients with acute myeloid leukemia (AML), which is an extremely aggressive disease with very few long-term survivors. The sirtuin deacetylases (SIRTs) are critical regulators of genes that are essential for longevity, cell growth, tumor suppression, and apoptosis. Elevated SIRT expression has been reported in several types of cancer and may promote pathogenesis and drug resistance by increasing the lifespan and survival capacity of malignant cells. Our preliminary analysis of SIRT expression indicated that SIRT1 was consistently expressed at significantly higher levels in AML cell lines and primary AML blasts as compared with normal controls. In order to investigate the potential role of SIRT1 as a regulator of AML pathogenesis, we utilized shRNA to stably knockdown its expression in MV4-11 and KG-1 AML cells. Cells with targeted SIRT1 knockdown displayed an altered gene expression profile as compared with non-targeted controls. Moreover, antagonizing SIRT1 expression significantly impeded the progression of AML in a xenograft mouse model. A number of deacetylase inhibitors have been clinically evaluated for cancer therapy. However, disrupting SIRT function as an anticancer strategy remains to be rigorously investigated as none of these previously studied drugs significantly inhibit the activity of this class of NAD+-dependent deacteylases. Tenovin-6 is a novel small molecule SIRT inhibitor. We investigated the efficacy and pharmacodynamic effects of tenovin-6 in AML cell lines, primary blasts from patients with AML, and mouse models. Treatment with tenovin-6 induced apoptosis and dramatically diminished AML clonogenic survival. Tenovin-6 promoted a dose-dependent increase in the acetylated levels of the SIRT-regulated gene p53 in AML cells and triggered the induction of several p53 transcriptional targets including p21 and PUMA. Targeted knockdown of PUMA with shRNA significantly reduced the pro-apoptotic effects of tenovin-6, indicating that it is a critical mediator of its anti-leukemic activity. Notably, administration of tenovin-6 to mice implanted with AML cells was well-tolerated and led to a highly significant reduction in disease burden and increase in overall survival. Our collective findings demonstrate that SIRT1 is a promising novel therapeutic target in AML. Further investigation aimed to elucidate the safety, efficacy, and mechanism of action of tenovin-6 is warranted. Disclosures: No relevant conflicts of interest to declare.

Non-Canonical Autophagy during APL Therapy

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1621-1621
Author(s):  
Jing Jin ◽  
Magali Humbert ◽  
Mario P. Tschan
Keyword(s):  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Differentiation Markers ◽  
Hematopoietic Stem ◽  
Myeloid Development ◽  
Autophagy Pathway ◽  
Neutrophil Differentiation ◽  
Acute Myeloid

Abstract Autophagy is an intracellular degradation system that ensures a dynamic recycling of cytoplasmic contents. Autophagy is required for self-renewal and cell survival under stress caused by a variety of stimuli including starvation and chemotherapy. There is accumulating evidence for additional functions of autophagy during myeloid development and hematopoietic stem cell maintenance. In this study, we used primary acute myeloid leukemia (AML) samples and human APL/AML cell lines to investigate the autophagy pathway active in all-trans retinoic acid (ATRA) mediated neutrophil differentiation. By characterizing the autophagic pathway during neutrophil differentiation of APL cells in more detail, we identified a non-canonical autophagy pathway, which not necessarily requires a hierarchal involvement of all autophagy-related (ATG) proteins. In addition to previous findings, from us and others, showing that ATRA-induced autophagy in APL cells is Beclin-1 independent, we discovered that ATRA-induced autophagy during APL differentiation is dependent on only one ATG16L isoform. The ATG16L proteins ATG16L1 and L2 are part of the ubiquitin-like conjugation systems ATG12-ATG5-ATG16L1 and ATG8/LC3 that are essential for phagophore elongation and autophagosome maturation. ATG16L2 is an isoform of ATG16L1, which is dispensable for starvation-induced autophagy despite forming an ATG12-ATG5-ATG16L2 complex in COS-7 cells. By investigating ATG16 gene expression in acute myeloid leukemia (AML) blast cells, we found that ATG16L1 as well as L2 are significantly downregulated in primary AML patient samples. In addition, neutrophil differentiation of APL/AML cell lines and CD34+ myeloid progenitor cells resulted in a significant induction of ATG16L1 and ATG16L2 expression. Induction of ATG16L2 was clearly more prominent than that of ATG16L1. Importantly, knocking down ATG16L2 but not ATG16L1 significantly attenuated neutrophil differentiation of AML cells as evidenced by decreased expression of the differentiation markers CD11b, GCSFR and CEBPE. Moreover, inhibition of ATG16L2 but not ATG16L1 resulted in decreased autophagy induction upon ATRA-treatment. Conversely, silencing ATG16L1 but not ATG16L2 was able to inhibit canonical starvation but not ATRA-induced differentiation associated autophagy in APL cells. Our data reveal distinct functions of ATG16L1 and ATG16L2 in starvation and ATRA-induced autophagy. To investigate the transcriptional regulation of ATG16L2 during neutrophil differentiation, we screened the ATG16L2 promoter region for putative transcription factor binding sites. We identified PU.1 as a transcriptional regulator of ATG16L2 using chromatin immunoprecipitation, PU.1 knockdown APL cells and a PU.1 inducible AML cell line model. These findings are in line with our earlier findings that PU.1 activates transcription of the ATG genes WIPI1, ATG3, MAP1S and ATG4C during APL differentiation. Our data provide strong evidence for a particular, non-canonical subtype of autophagy operative during neutrophil differentiation of APL cells. ATG16L2, in contrast to ATG16L1 is essential for successful ATRA-induced neutrophil differentiation and autophagy. This is in sharp contrast to its lack of function during starvation-induced autophagy. Deciphering the particular autophagy pathway active during APL differentiation is a prerequisite to develop novel differentiation therapies that are based on autophagy modulation. Since our findings have been validated in non-APL cells, activation of autophagy might support neutrophil differentiation of AML cells in a more general way. Disclosures No relevant conflicts of interest to declare.

scholarly journals Pharmacologic Targeting of LIN28/Let-7 in Acute Myeloid Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4072-4072 ◽  
Author(s):  
Martina M Roos ◽  
Michelle Li ◽  
Pang Amara ◽  
John P Chute
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Small Molecule ◽  
Myeloid Leukemia ◽  
Driver Genes ◽  
Oncogenic Driver ◽  
Acute Myeloid

Abstract Acute myeloid leukemia (AML) is a genetically heterogeneous malignancy with high relapse rates and mortality due to the outgrowth of chemotherapy-resistant leukemic stem cells (LSCs). Thus, the development of novel therapeutic strategies capable of eradicating human AML represents a major area of unmet medical need. The RNA binding protein, LIN28, is a known driver of many cancer stem cells, AML included, wherein overexpression of LIN28 correlates with reduced patient survival. LIN28 blocks the function of the let-7 microRNA family, which exert tumor suppressive effects by repressing oncogenes and cell cycle regulators including MYC, RAS and CyclinD. Thus, LIN28 is an attractive mechanistic target for the purpose of inhibiting AML LSCs. Using a targeted high-throughput screen, we identified a class of small molecules which selectively block the LIN28/let-7 interaction (Roos et al., ACS Chem Biol, 2016). Preliminary studies demonstrate that a lead small molecule markedly impairs the proliferation and clonogenic capacity of human AML cell lines and primary patient AML samples. In vivo, systemic administration of a lead small molecule LIN28/let-7 inhibitor decreases leukemic tumor burden, reduces LSC numbers and significantly improves animal survival. Mechanistic studies revealed that targeted inhibition of the LIN28/let-7 axis restores let-7 microRNA levels in AML LSCs and subsequently inhibits a panoply of key oncogenic driver genes, including the NF-ĸB pathway, a hallmark for LSC proliferation. Furthermore, AML cell lines and primary patient cells treated with the LIN28/let-7 small molecule inhibitor showed a block at the G1/S phase interface and significantly decreased cell cycle progression. Consequently, LIN28/let-7 inhibition leads to LSC differentiation and ultimately leukemic cell death. In summary, we demonstrate for the first time the drugability of the LIN28/let-7 axis in vivo and reveal a novel pharmacological means to suppress a multitude of oncogenic driver genes in human AML. These results suggest that small molecule inhibition of LIN28/let-7 has high therapeutic potential as a new class of targeted therapies for AML. Disclosures No relevant conflicts of interest to declare.

scholarly journals CRNDE enhances the expression of MCM5 and proliferation in acute myeloid leukemia KG-1a cells by sponging miR-136-5p

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chen Liu ◽  
Liang Zhong ◽  
Chenlan Shen ◽  
Xuan Chu ◽  
Xu Luo ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Noncoding Rna ◽  
Colorectal Neoplasia ◽  
Luciferase Reporter ◽  
Cell Cycle Distribution ◽  
Cell Progression ◽  
Acute Myeloid

AbstractThe long-noncoding RNA colorectal neoplasia differentially expressed (CRNDE) gene has been considered to be crucial in tumor malignancy. Although CRNDE is highly expressed in acute myeloid leukemia (AML), its mechanism of action remains unknown. In this study, GEPIA and qRT-PCR were performed to confirm the expression of CRNDE in AML samples and cell lines, respectively. CRNDE shRNA vectors were transfected to explore the biological functions of CRNDE. The cell proliferation was assessed by the CCK8 assay, while apoptosis and cell cycle distribution were measured by flow cytometry and Western blotting. The results showed that CRNDE was overexpressed in both AML samples and cell lines. CRNDE silencing inhibited proliferation and increased apoptotic rate and cell cycle arrest of KG-1a cells. The luciferase reporter assay coupled with RIP assay revealed that CRNDE act as a ceRNA. Rescue assays demonstrated that the effects of CRNDE silencing could be reversed by miR-136-5p inhibitors. In conclusion, our results expound that the CRNDE/miR-136-5p/MCM5 axis modulates cell progression and provide a new regulatory network of CRNDE in KG-1a cells.

Antitumor Effect of Pomolic Acid in Acute Myeloid Leukemia Cells Involves Cell Death, Decreased Cell Growth and Topoisomerases Inhibition

2019 ◽  
Vol 18 (10) ◽  
pp. 1457-1468
Author(s):  
Michelle X.G. Pereira ◽  
Amanda S.O. Hammes ◽  
Flavia C. Vasconcelos ◽  
Aline R. Pozzo ◽  
Thaís H. Pereira ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Acute Myeloid Leukemia ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Natural Compound ◽  
Dna Topoisomerases ◽  
Human Dna ◽  
Pomolic Acid ◽  
Acute Myeloid

Background: Acute myeloid leukemia (AML) represents the largest number of annual deaths from hematologic malignancy. In the United States, it was estimated that 21.380 individuals would be diagnosed with AML and 49.5% of patients would die in 2017. Therefore, the search for novel compounds capable of increasing the overall survival rate to the treatment of AML cells is urgent. Objectives: To investigate the cytotoxicity effect of the natural compound pomolic acid (PA) and to explore the mechanism of action of PA in AML cell lines with different phenotypes. Methods: Three different AML cell lines, HL60, U937 and Kasumi-1 cells with different mechanisms of resistance were used to analyze the effect of PA on the cell cycle progression, on DNA intercalation and on human DNA topoisomerases (hTopo I and IIα) in vitro studies. Theoretical experiments of the inhibition of hTopo I and IIα were done to explore the binding modes of PA. Results: PA reduced cell viability, induced cell death, increased sub-G0/G1 accumulation and activated caspases pathway in all cell lines, altered the cell cycle distribution and inhibited the catalytic activity of both human DNA topoisomerases. Conclusion: Finally, this study showed that PA has powerful antitumor activity against AML cells, suggesting that this natural compound might be a potent antineoplastic agent to improve the treatment scheme of this neoplasm.

scholarly journals Loss of ISWI ATPase SMARCA5 (SNF2H) in Acute Myeloid Leukemia Cells Inhibits Proliferation and Chromatid Cohesion

2020 ◽  
Vol 21 (6) ◽  
pp. 2073
Author(s):  
Tomas Zikmund ◽  
Helena Paszekova ◽  
Juraj Kokavec ◽  
Paul Kerbs ◽  
Shefali Thakur ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Cell Cycle Progression ◽  
Mitotic Division ◽  
Cycle Progression ◽  
Hematopoietic Stem ◽  
Chromatid Cohesion ◽  
Acute Myeloid Leukemia Cells ◽  
Acute Myeloid

ISWI chromatin remodeling ATPase SMARCA5 (SNF2H) is a well-known factor for its role in regulation of DNA access via nucleosome sliding and assembly. SMARCA5 transcriptionally inhibits the myeloid master regulator PU.1. Upregulation of SMARCA5 was previously observed in CD34+ hematopoietic progenitors of acute myeloid leukemia (AML) patients. Since high levels of SMARCA5 are necessary for intensive cell proliferation and cell cycle progression of developing hematopoietic stem and progenitor cells in mice, we reasoned that removal of SMARCA5 enzymatic activity could affect the cycling or undifferentiated state of leukemic progenitor-like clones. Indeed, we observed that CRISPR/cas9-mediated SMARCA5 knockout in AML cell lines (S5KO) inhibited the cell cycle progression. We also observed that the SMARCA5 deletion induced karyorrhexis and nuclear budding as well as increased the ploidy, indicating its role in mitotic division of AML cells. The cytogenetic analysis of S5KO cells revealed the premature chromatid separation. We conclude that deleting SMARCA5 in AML blocks leukemic proliferation and chromatid cohesion.

RNAi-Based Functional Genomics Screening of the Human Kinome Identifies Major Growth Regulating Kinases in Acute Myeloid Leukemia Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2951-2951
Author(s):  
Raoul Tibes ◽  
Ashish Choudhary ◽  
Amanda Henrichs ◽  
Sadia Guled ◽  
Irma Monzon ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Acute Myeloid Leukemia ◽  
Gene Silencing ◽  
Functional Genomics ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Large Scale ◽  
Cell Cycle Checkpoint ◽  
Acute Myeloid

Abstract In order to improve treatment strategies for Acute Myeloid Leukemia (AML), we adapted a functional genomics approach using RNAi screening to identify molecular targets that are vital to the growth of AML. Herein we report the first large-scale kinome gene silencing screen in AML. A high throughput RNAi screen was developed for the efficient siRNA transfection of AML cell lines. Eight commercially available cationic lipid-based transfection reagents were tested for their ability to transfect several AML cell lines with siRNA. These extensive transfection optimization experiments identified two AML cells lines TF-1 and ML4 with up to 95–100 and 70–75% transfection efficiency respectively. Two independent replicate kinome screens were performed on both cell lines using a siRNA library targeting 572 kinase genes with 2 siRNA/gene. At 96 hours post transfection, cell proliferation was assessed and the B-score method was used to background correct and analyze the screening data. Several siRNA to specific kinases were identified that significantly inhibit cell proliferation of up to ~40–88%. Hits were defined at two thresholds: siRNA having a B-score of <−2 providing a statistically significance of p<0.05 (confidence of > 95%) and a cutoff B-score of <−1.5 providing greater than 87% confidence for each siRNA hit. Two different kinases (2 siRNA/gene/screen) were identified as major growth regulating kinases in TF1 cells with all 4 siRNA/gene having a B-score <−2. For two additional kinases, 3/4 siRNA for each gene had a Bscore <−2. Expanding the cutoff to a B-score <−1.5 three further kinases were targeted by at least 3/4 siRNA/gene. Similar analysis using the same criteria for ML4 cells identified one kinase targeted by 3/4 siRNA at a B-score <−2, seven kinases with 2/4 siRNA <−2 and two kinases with 3/4 siRNA/gene at a B-score of <−1.5. Common hits for both cell lines with at least 6/8 siRNA per gene from 4 screens performing at a B-score <−2 identified two kinases, one of them PLK1. Applying a B-score threshold of <−1.5, we identified five kinases for which at least 5/8 siRNA/gene from 4 screens met these criteria. Kinases/genes will be presented at the meeting.Confirmation of gene silencing and validation of growth response is currently underway for a subset of genes. Among the strongest hits are siRNA targeting PLK1, as well as siRNA targeting three other kinase-genes involved in regulating cell cycle progression and checkpoints and gene ontology (GO) analysis showed enrichment in cell cycle and cell cycle-checkpoint processes. Inhibitors against PLK1 and other kinase hits identified in the screen are in (pre)-clinical development and if confirmed, our experiments provide a strong rational to test these in AML. The application of RNAi based screening is useful in the identification of genes important in AML proliferation, which could serve as targets for therapeutic intervention and guide AML drug development. Furthermore, results from these types of functional genomics approaches hold promise to be rapidly translated into clinical application.

MicroRNA-29a Is An Oncogene in Acute Myeloid Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 683-683
Author(s):  
Christopher Y. Park ◽  
Yoon-Chi Han ◽  
Govind Bhagat ◽  
Jian-Bing Fan ◽  
Irving L Weissman ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Stem Cells ◽  
Acute Myeloid Leukemia ◽  
Hematopoietic Stem Cells ◽  
Mirna Expression ◽  
Myeloid Leukemia ◽  
Hematopoietic Stem ◽  
Acute Myeloid

Abstract microRNAs (miRNAs) are short, non-protein encoding RNAs that bind to the 3′UTR’s of target mRNAs and negatively regulate gene expression by facilitating mRNA degradation or translational inhibition. Aberrant miRNA expression is well-documented in both solid and hematopoietic malignancies, and a number of recent miRNA profiling studies have identified miRNAs associated with specific human acute myeloid leukemia (AML) cytogenetic groups as well as miRNAs that may prognosticate clinical outcomes in AML patients. Unfortunately, these studies do not directly address the functional role of miRNAs in AML. In fact, there is no direct functional evidence that miRNAs are required for AML development or maintenance. Herein, we report on our recent efforts to elucidate the role of miRNAs in AML stem cells. miRNA expression profiling of AML stem cells and their normal counterparts, hematopoietic stem cells (HSC) and committed progenitors, reveals that miR-29a is highly expressed in human hematopoietic stem cells (HSC) and human AML relative to normal committed progenitors. Ectopic expression of miR-29a in mouse HSC/progenitors is sufficient to induce a myeloproliferative disorder (MPD) that progresses to AML. During the MPD phase of the disease, miR-29a alters the composition of committed myeloid progenitors, significantly expedites cell cycle progression, and promotes proliferation of hematopoietic progenitors at the level of the multipotent progenitor (MPP). These changes are manifested pathologically by marked granulocytic and megakaryocytic hyperplasia with hepatosplenomegaly. Mice with miR-29a-induced MPD uniformly progress to an AML that contains a leukemia stem cell (LSC) population that can serially transplant disease with as few as 20 purified LSC. Gene expression analysis reveals multiple tumor suppressors and cell cycle regulators downregulated in miR-29a expressing cells compared to wild type. We have demonstrated that one of these genes, Hbp1, is a bona fide miR-29a target, but knockdown of Hbp1 in vivo does not recapitulate the miR-29a phenotype. These data indicate that additional genes are required for miR-29a’s leukemogenic activity. In summary, our data demonstrate that miR-29a regulates early events in normal hematopoiesis and promotes myeloid differentiation and expansion. Moreover, they establish that misexpression of a single miRNA is sufficient to drive leukemogenesis, suggesting that therapeutic targeting of miRNAs may be an effective means of treating myeloid leukemias.

MYB Overexpression Is Directly Involved in Acute Myeloid Leukemia Pathogenesis and Could Constitute a New Therapeutic Target for Patients with Aberrant Expression of This Gene.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2402-2402 ◽  
Author(s):  
Carmen Vicente ◽  
Ana Conchillo ◽  
Daphnie Pauwels ◽  
Iria Vazquez ◽  
Laura Garcia-Orti ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Complete Remission ◽  
Synergistic Effect ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Recurrent Event ◽  
Aberrant Expression ◽  
Acute Myeloid

Abstract Abstract 2402 Poster Board II-379 The MYB proto-oncogene encodes a nuclear transcription factor with an essential role in proliferation, lineage commitment, and differentiation of hematopoietic progenitor cells. Proper levels of MYB are known to be important during hematopoietic cell development, and the Myb gene is a frequent target of retroviral insertions in myeloid, B- and T-cell leukemias in the mouse. Overexpression of MYB in T-acute lymphoblastic leukemia (T-ALL) causes a differentiation block of the T cells, and it has been shown that NOTCH1 mutation and MYB duplication cooperate in the pathogenesis of T-ALL. Our aim was to study the role of MYB in the pathogenesis of acute myeloid leukemia (AML), and to investigate its potential as a target for therapy. We functionally characterized MYB in 15 AML cell lines. Twelve of the 15 cell lines tested had MYB overexpression. Knockdown of MYB by siRNA in these cell lines caused decreased cell viability and proliferation, and reduced the clonogenic capacity, that could be explained in some cell lines by changes on the stage of cell differentiation. These results show that MYB overexpression is involved in the pathogenesis of AML. Moreover, knockdown of MYB in combination with common AML treatments (Idarubicin, Cytarabine and Sorafenib) had a strong synergistic effect on proliferation and viability of cells, suggesting that MYB could be a new target for therapy in AML. These observations prompted us to quantify MYB expression in a cohort of 159 patients with AML at diagnosis. We detected MYB overexpression in 14.5% (23/159) patients, with a higher prevalence within the intermediate prognosis group (17/83, 20.5%), particularly in patients with normal karyotype (NK) (14/62, 22.6%). Interestingly, 33% of patients without FLT-3 ITD and NPM1 mutations had MYB overexpression. To study the prognosis impact of MYB overexpression in AML, we performed a survival analysis in a preliminary series of 100 AML patients at diagnosis. As expected, significant differences in OS according to age, complete remission and cytogenetic prognostic group were found (p<0.01). MYB overexpression had no significant impact in the OS; however, this genetic marker allowed distinguishing a group of patients with a worse outcome within the group that did not get complete remission after treatment. Recently it has been described that MYB duplication causes elevated MYB expression in T-ALL; we detected duplication of MYB in 2 of 13 AML cell lines and in 2 patients with MYB overexpression (2/23, 8.6%). In conclusion, these results show that aberrant expression of MYB is involved in the activation of pathways responsible for the increased proliferative and clonogenic capacity that is characteristic of AML, independently of other genetic aberrations. Moreover, we show that MYB overexpression is a recurrent event in AML, especially in the subgroup of patients with NK, and that MYB could cooperate with other mutations in the leukemic transformation, as described previously in T-ALL. The synergistic effect of combined treatments with MYB knockdown, suggest that MYB silencing could be a new target for therapy in patients with AML and MYB overexpression. Disclosures: No relevant conflicts of interest to declare.

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