RESUMO
AML is a genetically heterogeneous disease and understanding how different co-occurring mutations cooperate to drive leukemogenesis will be crucial for improving diagnostic and therapeutic options for patients. MIR142 mutations have been recurrently detected in IDH-mutated AML samples. Here, we have used a mouse model to investigate the interaction between these two mutations and demonstrate a striking synergy between Mir142 loss-of-function and IDH2R140Q, with only recipients of double mutant cells succumbing to leukemia. Transcriptomic analysis of the non-leukemic single and leukemic double mutant progenitors, isolated from these mice, suggested a novel mechanism of cooperation whereby Mir142 loss-of-function counteracts aberrant silencing of Hoxa cluster genes by IDH2R140Q. Our analysis suggests that IDH2R140Q is an incoherent oncogene, with both positive and negative impacts on leukemogenesis, which requires the action of cooperating mutations to alleviate repression of Hoxa genes in order to advance to leukemia. This model, therefore, provides a compelling rationale for understanding how different mutations cooperate to drive leukemogenesis and the context-dependent effects of oncogenic mutations.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Proteínas de Homeodomínio/metabolismo , Isocitrato Desidrogenase/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , MicroRNAs/metabolismo , Animais , Carcinogênese/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Feminino , Regulação Leucêmica da Expressão Gênica/genética , Genótipo , Proteínas de Homeodomínio/genética , Humanos , Isocitrato Desidrogenase/genética , Leucemia Mieloide Aguda/genética , Masculino , Camundongos , MicroRNAs/genética , Mutação/genéticaRESUMO
Acute myeloid leukaemia (AML) is the most common adult acute leukaemia with the lowest survival rate. It is characterised by a build-up of immature myeloid cells anchored in the protective niche of the bone marrow (BM) microenvironment. The CXCL12/CXCR4 axis is central to the pathogenesis of AML as it has fundamental control over AML cell adhesion into the protective BM niche, adaptation to the hypoxic environment, cellular migration and survival. High levels of CXCR4 expression are associated with poor relapse-free and overall survival. The CXCR4 ligand, CXCL12 (SDF-1), is expressed by multiple cells types in the BM, facilitating the adhesion and survival of the malignant clone. Blocking the CXCL12/CXCR4 axis is an attractive therapeutic strategy providing a 'multi-hit' therapy that both prevents essential survival signals and releases the AML cells from the BM into the circulation. Once out of the protective niche of the BM they would be more susceptible to destruction by conventional chemotherapeutic drugs. In this review, we disentangle the diverse roles of the CXCL12/CXCR4 axis in AML. We then describe multiple CXCR4 inhibitors, including small molecules, peptides, or monoclonal antibodies, which have been developed to date and their progress in pre-clinical and clinical trials. Finally, the review leads us to the conclusion that there is a need for further investigation into the development of a 'multi-hit' therapy that targets several signalling pathways related to AML cell adhesion and maintenance in the BM.
Assuntos
Quimiocina CXCL12/fisiologia , Leucemia Mieloide Aguda/metabolismo , Proteínas de Neoplasias/fisiologia , Receptores CXCR4/fisiologia , Transdução de Sinais/fisiologia , Animais , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/sangue , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Benzilaminas , Medula Óssea/patologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/fisiologia , Hipóxia Celular , Movimento Celular/fisiologia , Micropartículas Derivadas de Células , Ensaios Clínicos como Assunto , Ciclamos , Avaliação Pré-Clínica de Medicamentos , Resistencia a Medicamentos Antineoplásicos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/fisiologia , Compostos Heterocíclicos/farmacologia , Compostos Heterocíclicos/uso terapêutico , Humanos , Leucemia Mieloide Aguda/sangue , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Terapia de Alvo Molecular , Proteínas de Neoplasias/antagonistas & inibidores , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Peptídeos/uso terapêutico , Peptídeos Cíclicos/uso terapêutico , Piridinas/uso terapêutico , Receptores CXCR4/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Nicho de Células-Tronco , Células Estromais/metabolismo , Células Estromais/patologia , Microambiente TumoralRESUMO
Our ability to manage acute myeloid leukemia (AML) is limited by our incomplete understanding of the epigenetic disruption central to leukemogenesis, including improper histone methylation. Here we examine 16 histone H3 genes in 434 primary AML samples and identify Q69H, A26P, R2Q, R8H and K27M/I mutations (1.6%), with higher incidence in secondary AML (9%). These mutations occur in pre-leukemic hematopoietic stem cells (HSCs) and exist in the major leukemic clones in patients. They increase the frequency of functional HSCs, alter differentiation, and amplify leukemic aggressiveness. These effects are dependent on the specific mutation. H3K27 mutation increases the expression of genes involved in erythrocyte and myeloid differentiation with altered H3K27 tri-methylation and K27 acetylation. The functional impact of histone mutations is independent of RUNX1 mutation, although they at times co-occur. This study establishes that H3 mutations are drivers of human pre-cancerous stem cell expansion and important early events in leukemogenesis.
Assuntos
Epigenômica , Regulação Leucêmica da Expressão Gênica/fisiologia , Histonas/metabolismo , Leucemia Mieloide Aguda/metabolismo , Animais , Animais Geneticamente Modificados , Antineoplásicos/farmacologia , Sequência de Bases , Células da Medula Óssea , Diferenciação Celular , Transformação Celular Neoplásica , DNA/genética , Drosophila melanogaster/genética , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Camundongos , Mutação , Neoplasias ExperimentaisRESUMO
Endothelin receptor type A (EDNRA) is known as a mediator of cell proliferation and survival. Aberrant regulation of EDNRA has been shown to play a role in tumor growth and metastasis. Using a global gene expression screen, we found that expression of Ednra was upregulated in murine leukemia inducing cells co-expressing Hoxa9 and Meis1 compared to cells only expressing Hoxa9. The aim of this study was to explore the role of Ednra in leukemogenesis further. In a murine bone marrow transplantation model, mice transplanted with cells overexpressing Ednra and Hoxa9 succumbed to leukemia significantly earlier than mice transplanted with cells overexpressing Hoxa9 only. Furthermore, overexpression of Ednra led to increased proliferation and resistance to apoptosis of bone marrow cells in vitro. We could also show that Meis1 binds to the Ednra promoter region, suggesting a regulatory role for Meis1 in Ednra expression. Taken together, our results suggest a role for Ednra in Hoxa9/Meis1-driven leukemogenesis.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Proteínas de Homeodomínio/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteína Meis1/metabolismo , Receptor de Endotelina A/metabolismo , Animais , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Knowledge of programmed death ligand 1 (PD-L1) expression and its regulation in B-cell lymphoma cells is limited. Investigating mechanisms that control PD-L1 expression in B-cell lymphoma cells might identify biomarkers that predict the efficacy of immunotherapy with anti-programmed death-1/PD-L1 antibodies. In addition, identification of mechanisms that regulate PD-L1 may identify molecules that can be targeted to improve the clinical efficacy of immune checkpoint inhibitors. In this study, we used proteomic approaches and patient-derived B-cell lymphoma cell lines to investigate mechanisms that regulate PD-L1 expression. We found that PD-L1 expression, particularly in nongerminal center B cell-derived diffuse large B-cell lymphoma (DLBCL), is controlled and regulated by several interactive signaling pathways, including the B-cell receptor (BCR) and JAK2/STAT3 signaling pathways. We found that that BCR-mediated NFATc1 activation upregulates IL-10 chemokine expression in PD-L1+ B-cell lymphoma cells. Released IL-10 activates the JAK2/STAT3 pathway, leading to STAT3-induced PD-L1 expression. IL-10 antagonist antibody abrogates IL-10/STAT3 signaling and PD-L1 protein expression. We also found that BCR pathway inhibition by BTK inhibitors (ibrutinib, acalabrutinib, and BGB-3111) blocks NFATc1 and STAT3 activation, thereby inhibiting IL-10 and PD-L1 expression. Finally, we validated the PD-L1 signaling network in 2 primary DLBCL cohorts consisting of 428 and 350 cases and showed significant correlations among IL-10, STAT3, and PD-L1. Thus, our findings reveal a complex signaling network regulating PD-L1 expression in B-cell lymphoma cells and suggest that PD-L1 expression can be modulated by small molecule inhibitors to potentiate immunotherapies.
Assuntos
Antígeno B7-H1/biossíntese , Regulação Leucêmica da Expressão Gênica/fisiologia , Linfoma Difuso de Grandes Células B/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Transdução de Sinais/fisiologia , Humanos , Interleucina-10/metabolismo , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Fatores de Transcrição NFATC/metabolismo , Fator de Transcrição STAT3/metabolismoRESUMO
Leukemic stem cells (LSCs) are an emerging target of curative anti-leukemia therapy. In acute lymphoblastic leukemia (ALL), LSCs frequently express CD34 and often lack CD38. However, little is known about markers and targets expressed in ALL LSCs. We have examined marker- and target expression profiles in CD34+/CD38- LSCs in patients with Ph+ ALL (nâ¯=â¯22) and Ph- ALL (nâ¯=â¯27) by multi-color flow cytometry and qPCR. ALL LSCs expressed CD19 (B4), CD44 (Pgp-1), CD123 (IL-3RA), and CD184 (CXCR4) in all patients tested. Moreover, in various subgroups of patients, LSCs also displayed CD20 (MS4A1) (10/41â¯=â¯24%), CD22 (12/20â¯=â¯60%), CD33 (Siglec-3) (20/48â¯=â¯42%), CD52 (CAMPATH-1) (17/40â¯=â¯43%), IL-1RAP (13/29â¯=â¯45%), and/or CD135 (FLT3) (4/20â¯=â¯20%). CD25 (IL-2RA) and CD26 (DPPIV) were expressed on LSCs in Ph+ ALL exhibiting BCR/ABL1p210, whereas in Ph+ ALL with BCR/ABL1p190, LSCs variably expressed CD25 but did not express CD26. In Ph- ALL, CD34+/CD38- LSCs expressed IL-1RAP in 6/18 patients (33%), but did not express CD25 or CD26. Normal stem cells stained negative for CD25, CD26 and IL-1RAP, and expressed only low amounts of CD52. In xenotransplantation experiments, CD34+/CD38- and CD34+/CD38+ cells engrafted NSG mice after 12-20 weeks, and targeting with antibodies against CD33 and CD52 resulted in reduced engraftment. Together, LSCs in Ph+ and Ph- ALL display unique marker- and target expression profiles. In Ph+ ALL with BCR/ABL1p210, the LSC-phenotype closely resembles the marker-profile of CD34+/CD38- LSCs in chronic myeloid leukemia, confirming the close biologic relationship of these neoplasms. Targeting of LSCs with specific antibodies or related immunotherapies may facilitate LSC eradication in ALL.
Assuntos
ADP-Ribosil Ciclase 1/metabolismo , Antígenos CD34/metabolismo , Leucemia Mieloide Aguda/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco/metabolismo , Animais , Biomarcadores Tumorais/metabolismo , Linhagem Celular , Feminino , Regulação Leucêmica da Expressão Gênica/fisiologia , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Camundongos , Camundongos Endogâmicos NODRESUMO
The treatment of acute leukemia is still challenging due in part to the development of resistance and relapse. This chemotherapeutics resistance is established by clonal selection of resistant variants of the cancer cells. Recently, a horizontal transfer of chemo-resistance among cancer cells via extracellular vesicles (EVs) has been suggested. The aim of this research was to investigate the role of EVs in chemo-resistance in acute myeloid leukemia. For this purpose, the sensitive strain of the promyelocytic leukemia HL60 cell line was studied along with its multi-resistant strain, HL60/AR that overexpresses the multidrug resistance protein 1 (MRP-1). A chemo-resistance transfer between the two strains was established by treating HL60 cells with EVs generated by HL60/AR. This study reveals that EVs from HL60/AR can interact with HL60 cells and transfer at least partially, their chemo-resistance. EVs-treated cells begin to express MRP-1 probably due to a direct transfer of MRP-1 and nucleic acids transported by EVs. In this context, two microRNAs were highlighted for their high differential expression in EVs related to sensitive or chemo-resistant cells: miR-19b and miR-20a. Because circulating microRNAs are found in all biological fluids, these results bring out their potential clinical use as chemo-resistance biomarkers in acute myeloid leukemia.
Assuntos
Resistencia a Medicamentos Antineoplásicos/fisiologia , Vesículas Extracelulares/metabolismo , Leucemia Mieloide Aguda/patologia , MicroRNAs/metabolismo , Resistência a Múltiplos Medicamentos/fisiologia , Regulação Leucêmica da Expressão Gênica/fisiologia , Células HL-60 , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/biossínteseRESUMO
Chromosomal translocations are one of the hallmarks of acute myeloid leukemia (AML), often leading to gene fusions and expression of an oncofusion protein. Over recent years it has become clear that most of the AML associated oncofusion proteins molecularly adopt distinct mechanisms for inducing leukemogenesis. Still these unique molecular properties of the chimeric proteins converge and give rise to a common pathogenic molecular mechanism. In the present study we compared genome-wide DNA binding and transcriptome data associated with AML1-ETO, CBFB-MYH11 and PML-RARA oncofusion protein expression to identify unique and common features. Our analyses revealed targeting of oncofusion binding sites to RUNX1 and ETS-factor occupied genomic regions. In addition, it revealed a highly comparable global histone acetylation pattern, similar expression of common target genes and related enrichment of several biological pathways critical for maintenance of AML, suggesting oncofusion proteins deregulate common gene programs despite their distinct binding signatures and mechanisms of action.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Histonas/metabolismo , Leucemia Mieloide Aguda/genética , Proteínas de Fusão Oncogênica/genética , Acetilação , Carcinogênese/genética , Carcinogênese/metabolismo , Imunoprecipitação da Cromatina , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Proteínas de Fusão Oncogênica/metabolismo , Proteína Proto-Oncogênica c-ets-1/genética , Proteína 1 Parceira de Translocação de RUNX1 , TranscriptomaRESUMO
Mutations that activate RAS proto-oncogenes and their effectors are common in acute myeloid leukemia (AML); however, efforts to therapeutically target Ras or its effectors have been unsuccessful, and have been hampered by an incomplete understanding of which effectors are required for AML proliferation and survival. We investigated the role of Ras effector pathways in AML using murine and human AML models. Whereas genetic disruption of NRAS(V12) expression in an NRAS(V12) and Mll-AF9-driven murine AML induced apoptosis of leukemic cells, inhibition of phosphatidylinositol-3-kinase (PI3K) and/or mitogen-activated protein kinase (MAPK) signaling did not reproduce this effect. Conversely, genetic disruption of RALB signaling induced AML cell death and phenocopied the effects of suppressing oncogenic Ras directly - uncovering a novel role for RALB signaling in AML survival. Knockdown of RALB led to decreased phosphorylation of TBK1 and reduced BCL2 expression, providing mechanistic insight into RALB survival signaling in AML. Notably, we found that patient-derived AML blasts have higher levels of RALB-TBK1 signaling compared to normal blood leukocytes, supporting a pathophysiologic role for RALB signaling for AML patients. Overall, our work provides new insight into the specific roles of Ras effector pathways in AML and has identified RALB signaling as a key survival pathway.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Mieloide Aguda/metabolismo , Proteínas ral de Ligação ao GTP/metabolismo , Proteínas ras/metabolismo , Animais , Xenoenxertos , Humanos , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos SCID , Transdução de Sinais/fisiologia , Células Tumorais CultivadasAssuntos
DNA (Citosina-5-)-Metiltransferases/genética , Regulação Leucêmica da Expressão Gênica/fisiologia , Genes Homeobox/genética , Leucemia Mieloide Aguda/genética , Proteína de Leucina Linfoide-Mieloide/genética , Complexo Repressor Polycomb 1/genética , Animais , Linhagem Celular Tumoral , DNA Metiltransferase 3A , Humanos , Camundongos , MutaçãoRESUMO
The RNA binding proteins RBM binding motif protein 38 (RBM38) and DEAD END 1 (DND1) selectively stabilize mRNAs by attenuating RNAse activity or protecting them from micro(mi)RNA-mediated cleavage. Furthermore, both proteins can efficiently stabilize the mRNA of the cell cycle inhibitor p21(CIP1). Since acute myeloid leukemia (AML) differentiation requires cell cycle arrest and RBM38 as well as DND1 have antiproliferative functions, we hypothesized that decreased RBM38 and DND1 expression may contribute to the differentiation block seen in this disease. We first quantified RBM38 and DND1 mRNA expression in clinical AML patient samples and CD34(+) progenitor cells and mature granulocytes from healthy donors. We found significantly lower RBM38 and DND1 mRNA levels in AML blasts and CD34(+) progenitor cells as compared to mature neutrophils from healthy donors. Furthermore, the lowest expression of both RBM38 and DND1 mRNA correlated with t(8;21). In addition, neutrophil differentiation of CD34(+) cells in vitro with G-CSF (granulocyte colony stimulating factor) resulted in a significant increase of RBM38 and DND1 mRNA levels. Similarly, neutrophil differentiation of NB4 acute promyelocytic leukemia (APL) cells was associated with a significant induction of RBM38 and DND1 expression. To address the function of RBM38 and DND1 in neutrophil differentiation, we generated two independent NB4RBM38 as well as DND1 knockdown cell lines. Inhibition of both RBM38 and DND1 mRNA significantly attenuated NB4 differentiation and resulted in decreased p21(CIP1) mRNA expression. Our results clearly indicate that expression of the RNA binding proteins RBM38 and DND1 is repressed in primary AML patients, that neutrophil differentiation is dependent on increased expression of both proteins, and that these proteins have a critical role in regulating p21(CIP1) expression during APL differentiation.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Mieloide Aguda/patologia , Leucemia Promielocítica Aguda/patologia , Proteínas de Neoplasias/biossíntese , Neutrófilos/patologia , Proteínas de Ligação a RNA/biossíntese , Adolescente , Adulto , Idoso , Western Blotting , Diferenciação Celular , Feminino , Técnicas de Silenciamento de Genes , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Adulto JovemRESUMO
A reciprocal translocation of the ABL1 gene to the BCR gene results in the expression of the oncogenic BCR-ABL1 fusion protein, which characterizes human chronic myeloid leukemia (CML), a myeloproliferative disorder considered invariably fatal until the introduction of the imatinib family of tyrosine kinase inhibitors (TKI). Nonetheless, insensitivity of CML stem cells to TKI treatment and intrinsic or acquired resistance are still frequent causes for disease persistence and blastic phase progression experienced in patients after initial successful therapies. Here, we investigated a possible role for the MAPK15/ERK8 kinase in BCR-ABL1-dependent autophagy, a key process for oncogene-induced leukemogenesis. In this context, we showed the ability of MAPK15 to physically recruit the oncogene to autophagic vesicles, confirming our hypothesis of a biologically relevant role for this MAP kinase in signal transduction by this oncogene. Indeed, by modeling BCR-ABL1 signaling in HeLa cells and taking advantage of a physiologically relevant model for human CML, i.e. K562 cells, we demonstrated that BCR-ABL1-induced autophagy is mediated by MAPK15 through its ability to interact with LC3-family proteins, in a LIR-dependent manner. Interestingly, we were also able to interfere with BCR-ABL1-induced autophagy by a pharmacological approach aimed at inhibiting MAPK15, opening the possibility of acting on this kinase to affect autophagy and diseases depending on this cellular function. Indeed, to support the feasibility of this approach, we demonstrated that depletion of endogenous MAPK15 expression inhibited BCR-ABL1-dependent cell proliferation, in vitro, and tumor formation, in vivo, therefore providing a novel "druggable" link between BCR-ABL1 and human CML.
Assuntos
Autofagia , Carcinogênese/metabolismo , Proliferação de Células/fisiologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas de Fusão bcr-abl/metabolismo , Oncogenes/fisiologia , Apoptose/fisiologia , Autofagia/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/fisiologia , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologiaRESUMO
A growing body of evidence suggests that the resistance of CLL cells to apoptosis is partly mediated through the interactions between leukemia cells and adjacent stromal cells residing in the lymphatic tissue or bone marrow microenvironment. Mcl-1, an anti-apoptotic protein that is associated with failure to treatment is up-regulated in CLL lymphocytes after interaction with microenvironment. However, the regulation of its expression in context to microenvironment is unclear. We evaluated and compared changes in Mcl-1 in CLL B-cells in suspension culture and when co-cultured on stromal cells. The blockade of apoptosis in co-cultured CLL cells is associated with diminution in caspase-3 and PARP cleavage and is not dependent on cytogenetic profile or prognostic factors of the disease. Stroma-derived resistance to apoptosis is associated with a cascade of transcriptional events such as increase in levels of total RNA Pol II and its phosphorylation at Ser2 and Ser5, increase in the rate of global RNA synthesis, and amplification of Mcl-1 transcript levels. The latter is associated with increase in Mcl-1 protein level without an impact on the levels of Bcl-2 and Bcl-xL. Post-translational modifications of protein kinases show increased phosphorylation of Akt at Ser473, Erk at Thr202/Tyr204 and Gsk-3ß at Ser9 and augmentation of total Mcl-1 accumulation along with phosphorylation at Ser159/Thr163 sites. Collectively, stroma-induced apoptosis resistance is mediated through signaling proteins that regulate transcriptional and translational expression and post-translational modification of Mcl-1 in CLL cells in context to bone marrow stromal microenvironment.
Assuntos
Células da Medula Óssea/metabolismo , Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Linfocítica Crônica de Células B/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Adulto , Idoso , Proliferação de Células , Sobrevivência Celular , Feminino , Citometria de Fluxo , Humanos , Leucemia Linfocítica Crônica de Células B/sangue , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-bcl-2/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Células Estromais/metabolismo , Células Tumorais CultivadasRESUMO
Suppressor with morphogenetic effect on genitalia family member (SMG1) belongs to a family of phosphoinositide 3-kinase-related kinases and is the main kinase involved in nonsense-mediated mRNA decay. Recently, SMG1 was suggested as a novel potential tumor suppressor gene, particularly in hypoxic tumors. To investigate the function of SMG1 in acute myeloid leukemia (AML), we performed methylation-specific polymerase chain reaction and found that SMG1 was hypermethylated in the promoter region. SMG1 hypermethylation was found in 66% (33/50) of AML samples compared with none (0/14) of the normal controls. SMG1 mRNA was down-regulated in AML patients with hypermethylation status whereas it was readily expressed in patients without methylation. Moreover, treatment of AML cells with demethylating agent 5-aza-2'-deoxycytidine (decitabine) inhibited AML cell growth and induced apoptosis by reversing SMG1 methylation status and restoring SMG1 expression. On the other hand, knockdown of SMG1 by RNA interference inhibited apoptosis. We also found that mTOR expression level was negatively correlated to SMG1 expression in AML patients which indicated that SMG1 and mTOR maybe act antagonistically to regulate AML cell growth. In conclusion, our results indicate that SMG1 acts as a potential tumor suppressor with epigenetic regulation in AML.
Assuntos
Metilação de DNA , DNA de Neoplasias/genética , Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Mieloide Aguda/metabolismo , Proteínas de Neoplasias/fisiologia , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Medula Óssea/metabolismo , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Decitabina , Regulação para Baixo , Feminino , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/genética , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mielomonocítica Aguda/genética , Leucemia Mielomonocítica Aguda/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Fosfatidilinositol 3-Quinases/genética , Regiões Promotoras Genéticas , Proteínas Serina-Treonina Quinases , Interferência de RNA , RNA Mensageiro/biossíntese , RNA Neoplásico/biossíntese , Serina-Treonina Quinases TOR/fisiologia , Proteínas Supressoras de Tumor/genética , Adulto JovemRESUMO
Tyrosine kinase inhibitors such as imatinib can effectively target the BCR-ABL oncoprotein in a majority of patients with chronic myeloid leukemia (CML). Unfortunately, some patients are resistant primarily to imatinib and others develop drug resistance, prompting interest in the discovery of new drug targets. Although much of this resistance can be explained by the presence of mutations within the tyrosine kinase domain of BCR-ABL, such mutations are not universally identified. Interferon regulatory factor-8 (IRF-8) is a transcription factor that is essential for myelopoiesis. Depressed IRF-8 levels are observed in a majority of CML patients and Irf-8(-/-) mice exhibit a CML-like disease. The underlying mechanisms of IRF-8 loss in CML are unknown. We hypothesized that BCR-ABL suppresses transcription of IRF-8 through STAT5, a proximal BCR-ABL target. Treatment of primary cells from newly diagnosed CML patients in chronic phase as well as BCR-ABL(+) cell lines with imatinib increased IRF-8 transcription. Furthermore, IRF-8 expression in cell line models was necessary for imatinib-induced antitumor responses. We have demonstrated that IRF-8 is a direct target of STAT5 and that silencing of STAT5 induced IRF-8 expression. Conversely, activating STAT5 suppressed IRF-8 transcription. Finally, we showed that STAT5 blockade using a recently discovered antagonist increased IRF-8 expression in patient samples. These data reveal a previously unrecognized BCR-ABL-STAT5-IRF-8 network, which widens the repertoire of potentially new anti-CML targets.
Assuntos
Fatores Reguladores de Interferon/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Adulto , Idoso , Animais , Antineoplásicos/farmacologia , Sequência de Bases , Benzamidas/farmacologia , Feminino , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/fisiologia , Técnicas de Silenciamento de Genes , Genes Supressores de Tumor/fisiologia , Humanos , Mesilato de Imatinib , Fatores Reguladores de Interferon/metabolismo , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Piperazinas/farmacologia , Regiões Promotoras Genéticas/fisiologia , Pirimidinas/farmacologia , Fator de Transcrição STAT5/genética , Transcrição Gênica/efeitos dos fármacos , Transcrição Gênica/fisiologia , Proteínas Supressoras de Tumor/genéticaRESUMO
The ETS transcription factor ERG plays a central role in definitive hematopoiesis, and its overexpression in acute myeloid leukemia (AML) is associated with a stem cell signature and poor prognosis. Yet how ERG causes leukemia is unclear. Here we show that pan-hematopoietic ERG expression induces an early progenitor myeloid leukemia in transgenic mice. Integrated genome-scale analysis of gene expression and ERG binding profiles revealed that ERG activates a transcriptional program similar to human AML stem/progenitor cells and to human AML with high ERG expression. This transcriptional program was associated with activation of RAS that was required for leukemia cells growth in vitro and in vivo. We further show that ERG induces expression of the Pim1 kinase oncogene through a novel hematopoietic enhancer validated in transgenic mice and human CD34(+) normal and leukemic cells. Pim1 inhibition disrupts growth and induces apoptosis of ERG-expressing leukemic cells. The importance of the ERG/PIM1 axis is further underscored by the poorer prognosis of AML highly expressing ERG and PIM1. Thus, integrative genomic analysis demonstrates that ERG causes myeloid progenitor leukemia characterized by an induction of leukemia stem cell transcriptional programs. Pim1 and the RAS pathway are potential therapeutic targets of these high-risk leukemias.
Assuntos
Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Mieloide Aguda/genética , Proteínas Proto-Oncogênicas c-pim-1/metabolismo , Transativadores/genética , Fatores de Transcrição/metabolismo , Animais , Antineoplásicos , Elementos Facilitadores Genéticos/genética , Genômica , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Células Progenitoras Mieloides/fisiologia , Transplante de Neoplasias , Transcrição Gênica/fisiologia , Regulador Transcricional ERGRESUMO
It has been previously shown that acute myeloid leukemia (AML) patients with higher levels of GATA1 expression have poorer outcomes. Furthermore, pediatric Down syndrome (DS) patients with acute megakaryocytic leukemia (AMKL), whose blast cells almost universally harbor somatic mutations in exon 2 of the transcription factor gene GATA1, demonstrate increased overall survival relative to non-DS pediatric patients, suggesting a potential role for GATA1 in chemotherapy response. In this study, we confirmed that amongst non-DS patients, GATA1 transcripts were significantly higher in AMKL blasts compared to blasts from other AML subgroups. Further, GATA1 transcript levels significantly correlated with transcript levels for the anti-apoptotic protein Bcl-xL in our patient cohort. ShRNA knockdown of GATA1 in the megakaryocytic cell line Meg-01 resulted in significantly increased cytarabine (ara-C) and daunorubicin anti-proliferative sensitivities and decreased Bcl-xL transcript and protein levels. Chromatin immunoprecipitation (ChIP) and reporter gene assays demonstrated that the Bcl-x gene (which transcribes the Bcl-xL transcripts) is a bona fide GATA1 target gene in AMKL cells. Treatment of the Meg-01 cells with the histone deacetylase inhibitor valproic acid resulted in down-regulation of both GATA1 and Bcl-xL and significantly enhanced ara-C sensitivity. Furthermore, additional GATA1 target genes were identified by oligonucleotide microarray and ChIP-on-Chip analyses. Our findings demonstrate a role for GATA1 in chemotherapy resistance in non-DS AMKL cells, and identified additional GATA1 target genes for future studies.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Fator de Transcrição GATA1/genética , Leucemia Megacarioblástica Aguda/genética , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Células Cultivadas , Criança , Pré-Escolar , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/fisiologia , Humanos , Leucemia Megacarioblástica Aguda/tratamento farmacológico , Análise em Microsséries , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Ácido Valproico/farmacologiaRESUMO
Chronic myelomonocytic leukemia (CMML) shares with other myeloid diseases a number of somatic gene mutations. These mutations can now be integrated within the framework of evolution theory to address the mechanisms of the disease. Several evidences indicate that the disease emerges in adult hematopoietic stem cells (HSC) through the age-dependent accumulation of DNA damage, leading stochastically to a driver mutation that confers a competitive advantage to the cell. A mutation in TET2 gene could be one of these driver mutations provoking the emergence of clonality. After a long latency, secondary lesions, such as mutations in the SRSF2 gene, contribute to progression to full-blown malignancy, with abnormal differentiation. Additional mutations accumulate and branching arising mostly through mitotic recombination generates clonal heterogeneity. Modifications in the microenvironment probably affect this clonal dynamics, whereas epigenetic alterations, such as hypermethylation of the TIF1γ gene promoter, may generate phenotypic diversification of otherwise clonal populations. The preserved although deregulated myeloid differentiation that characterizes CMML, with granulomonocyte expansion and various cytopenias, may depend on early clonal dominance in the hematopietic cell hierarchy. Progression to acute myeloid leukemia observed in 25-30% of the patients may arise from the massive expansion of a clone with novel genetic lesions, providing a high fitness to previously minor subclones when in chronic phase of the disease. This review discusses the various models of disease emergence and progression and how this recent knowledge could drive rational therapeutic strategies.
Assuntos
Evolução Clonal/fisiologia , Epigênese Genética/fisiologia , Regulação Leucêmica da Expressão Gênica/fisiologia , Leucemia Mielomonocítica Crônica/genética , Leucemia Mielomonocítica Crônica/patologia , Humanos , Leucemia Mielomonocítica Crônica/fisiopatologia , Mutação/fisiologiaRESUMO
TCL1 oncogene is overexpressed in aggressive form of human chronic lymphocytic leukemia (CLL) and its dysregulation in mouse B cells causes a CD5-positive leukemia similar to the aggressive form of human CLLs. To identify oncogenes that cooperate with Tcl1, we performed genetic screen in Eµ-TCL1 mice using Sleeping Beauty transposon-mediated mutagenesis. Analysis of transposon common insertion sites identified 7 genes activated by transposon insertions. Overexpression of these genes in mouse CLL was confirmed by real time reverse transcription-polymerase chain reaction. Interestingly, the main known function of 4 of 7 genes (Nfkb1, Tab2, Map3K14, and Nfkbid) is participation in or activation of the nuclear factor-kB (NF-kB) pathway. In addition, activation of the NF-kB is 1 of main functions of Akt2, also identified in the screen. These findings demonstrate cooperation of Tcl1 and the NF-kB pathway in the pathogenesis of aggressive CLL. Identification cooperating cancer genes will result in the development of combinatorial therapies to treat CLL.
Assuntos
Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/metabolismo , Subunidade p50 de NF-kappa B/genética , Subunidade p50 de NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Modelos Animais de Doenças , Regulação Leucêmica da Expressão Gênica/fisiologia , Testes Genéticos/métodos , Estimativa de Kaplan-Meier , Leucemia Linfocítica Crônica de Células B/mortalidade , Camundongos , Camundongos Transgênicos , Mutagênese Insercional/métodos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/fisiologia , Transposases/genética , Quinase Induzida por NF-kappaBRESUMO
Previously, we have shown that an AP-1 family member, FRA-2, is constitutively expressed in adult T-cell leukemia/lymphoma (ATL) and, together with JUND, upregulates CCR4 and promotes ATL cell growth. Among the identified potential target genes of FRA-2/JUND was SOX4. Here, we examine the expression and function of SOX4 in ATL. SOX4 was indeed consistently expressed in primary ATL cells. FRA-2/JUND efficiently activated the SOX4 promoter via an AP-1 site. Knockdown of SOX4 expression by small interfering RNA (siRNA) strongly suppressed cell growth of ATL cell lines. Microarray analyses revealed that SOX4 knockdown reduced the expression of genes such as germinal center kinase related (GCKR), NAK-associated protein 1 (NAP1), and histone deacetylase 8 (HDAC8). We confirmed consistent expression of GCKR, NAP1, and HDAC8 in primary ATL cells. We also showed direct activation of the HDAC8 promoter by SOX4. Furthermore, siRNA knockdown of GCKR, NAP1, and HDAC8 each significantly suppressed cell growth of ATL cell lines. Taken together, we have revealed an important oncogenic cascade involving FRA-2/JUND and SOX4 in ATL, which leads to the expression of genes such as GCKR, NAP1, and HDAC8.