RESUMO
The fusion oncoprotein BCR-ABL1 exhibits aberrant tyrosine kinase activity and it has been proposed that it deregulates signaling networks involving both transcription factors and non-coding microRNAs that result in chronic myeloid leukemia (CML). Previously, microRNA expression profiling showed deregulated expression of miR-150 and miR-155 in CML. In this study, we placed these findings into the broader context of the MYC/miR-150/MYB/miR-155/PU.1 oncogenic network. We propose that up-regulated MYC and miR-155 in CD34+ leukemic stem and progenitor cells, in concert with BCR-ABL1, impair the molecular mechanisms of myeloid differentiation associated with low miR-150 and PU.1 levels. We revealed that MYC directly occupied the -11.7 kb and -0.35 kb regulatory regions in the MIR150 gene. MYC occupancy was markedly increased through BCR-ABL1 activity, causing inhibition of MIR150 gene expression in CML CD34+ and CD34- cells. Furthermore, we found an association between reduced miR-150 levels in CML blast cells and their resistance to tyrosine kinase inhibitors (TKIs). Although TKIs successfully disrupted BCR-ABL1 kinase activity in proliferating CML cells, this treatment did not efficiently target quiescent leukemic stem cells. The study presents new evidence regarding the MYC/miR-150/MYB/miR-155/PU.1 leukemic network established by aberrant BCR-ABL1 activity. The key connecting nodes of this network may serve as potential druggable targets to overcome resistance of CML stem and progenitor cells.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Fusão bcr-abl/genética , Genes myc/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , MicroRNAs/genética , Adulto , Idoso , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Regulação para Baixo/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Células HL-60 , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Masculino , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Inibidores de Proteínas Quinases/farmacologiaRESUMO
OBJECTIVE: A prominent, safe and efficient therapy for patients with chronic myeloid leukemia (CML) is inhibiting oncogenic protein BCR::ABL1 in a targeted manner with imatinib, a tyrosine kinase inhibitor. A substantial part of patients treated with imatinib report skeletomuscular adverse events affecting their quality of life. OCTN2 membrane transporter is involved in imatinib transportation into the cells. At the same time, the crucial physiological role of OCTN2 is cellular uptake of carnitine which is an essential co-factor for the mitochondrial ß-oxidation pathway. This work investigates the impact of imatinib treatment on carnitine intake and energy metabolism of muscle cells. METHODS: HTB-153 (human rhabdomyosarcoma) cell line and KCL-22 (CML cell line) were used to study the impact of imatinib treatment on intracellular levels of carnitine and vice versa. The energy metabolism changes in cells treated by imatinib were quantified and compared to changes in cells exposed to highly specific OCTN2 inhibitor vinorelbine. Mouse models were used to test whether in vitro observations are also achieved in vivo in thigh muscle tissue. The analytes of interest were quantified using a Prominence HPLC system coupled with a tandem mass spectrometer. RESULTS: This work showed that through the carnitine-specific transporter OCTN2, imatinib and carnitine intake competed unequally and intracellular carnitine concentrations were significantly reduced. In contrast, carnitine preincubation did not influence imatinib cell intake or interfere with leukemia cell targeting. Blocking the intracellular supply of carnitine with imatinib significantly reduced the production of most Krebs cycle metabolites and ATP. However, subsequent carnitine supplementation rescued mitochondrial energy production. Due to specific inhibition of OCTN2 activity, the influx of carnitine was blocked and mitochondrial energy metabolism was impaired in muscle cells in vitro and in thigh muscle tissue in a mouse model. CONCLUSIONS: This preclinical experimental study revealed detrimental effect of imatinib on carnitine-mediated energy metabolism of muscle cells providing a possible molecular background of the frequently occurred side effects during imatinib therapy such as fatigue, muscle pain and cramps.
RESUMO
ABSTRACT: Venetoclax (VEN), a B-cell lymphoma 2 (BCL2) inhibitor, has a promising single-agent activity in mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), and large BCLs, but remissions were generally short, which call for rational drug combinations. Using a panel of 21 lymphoma and leukemia cell lines and 28 primary samples, we demonstrated strong synergy between VEN and A1155463, a BCL-XL inhibitor. Immunoprecipitation experiments and studies on clones with knockout of expression or transgenic expression of BCL-XL confirmed its key role in mediating inherent and acquired VEN resistance. Of note, the VEN and A1155463 combination was synthetically lethal even in the cell lines with lack of expression of the proapoptotic BCL2L11/BIM and in the derived clones with genetic knockout of BCL2L11/BIM. This is clinically important because BCL2L11/BIM deletion, downregulation, or sequestration results in VEN resistance. Immunoprecipitation experiments further suggested that the proapoptotic effector BAX belongs to principal mediators of the VEN and A1155463 mode of action in the BIM-deficient cells. Lastly, the efficacy of the new proapoptotic combination was confirmed in vivo on a panel of 9 patient-derived lymphoma xenografts models including MCL (n = 3), B-ALL (n = 2), T-ALL (n = 1), and diffuse large BCL (n = 3). Because continuous inhibition of BCL-XL causes thrombocytopenia, we proposed and tested an interrupted 4 days on/3 days off treatment regimen, which retained the desired antitumor synergy with manageable platelet toxicity. The proposed VEN and A1155463 combination represents an innovative chemotherapy-free regimen with significant preclinical activity across diverse BCL2+ hematologic malignancies irrespective of the BCL2L11/BIM status.
Assuntos
Proteína 11 Semelhante a Bcl-2 , Compostos Bicíclicos Heterocíclicos com Pontes , Resistencia a Medicamentos Antineoplásicos , Sulfonamidas , Proteína bcl-X , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Humanos , Proteína bcl-X/metabolismo , Proteína bcl-X/antagonistas & inibidores , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Proteína 11 Semelhante a Bcl-2/metabolismo , Proteína 11 Semelhante a Bcl-2/genética , Animais , Camundongos , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto , Apoptose/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Sinergismo Farmacológico , Isoquinolinas , BenzotiazóisRESUMO
Membrane transporters are important determinants of drug bioavailability. Their expression and activity affect the intracellular drug concentration in leukemic cells impacting response to therapy. Pharmacogenomics represents genetic markers that reflect allele arrangement of genes encoding drug transporters associated with treatment response. In previous work, we identified SNP rs460089 located in the promotor of SLC22A4 gene encoding imatinib transporter OCTN1 as influential on response of patients with chronic myeloid leukemia treated with imatinib. Patients with rs460089-GC pharmacogenotype had significantly superior response to first-line imatinib treatment compared to patients with rs460089-GG. This study investigated whether pharmacogenotypes of rs460089 are associated with sustainability of treatment-free remission (TFR) in patients from the EUROpean Stop Kinase Inhibitor (EURO-SKI) trial. In the learning sample, 176 patients showed a significantly higher 6-month probability of molecular relapse free survival (MRFS) in patients with GC genotype (73%, 95% CI: 60-82%) compared to patients with GG (51%, 95% CI: 41-61%). Also over time, patients with GC genotype had significantly higher MRFS probabilities compared with patients with GG (HR: 0.474, 95% CI: 0.280-0.802, p = 0.0054). Both results were validated with data on 93 patients from the Polish STOP imatinib study. In multiple regression models, in addition to the investigated genotype, duration of TKI therapy (EURO-SKI trial) and duration of deep molecular response (Polish study) were identified as independent prognostic factors. The SNP rs460089 was found as an independent predictor of TFR.
Assuntos
Antineoplásicos , Leucemia Mielogênica Crônica BCR-ABL Positiva , Humanos , Mesilato de Imatinib/uso terapêutico , Prognóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Antineoplásicos/efeitos adversos , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas de Membrana Transportadoras/uso terapêutico , Resultado do TratamentoRESUMO
Elevated levels of microRNA miR-155 represent a candidate pathogenic factor in chronic B-lymphocytic leukemia (B-CLL). In this study, we present evidence that MYB (v-myb myeloblastosis viral oncogene homolog) is overexpressed in a subset of B-CLL patients. MYB physically associates with the promoter of miR-155 host gene (MIR155HG, also known as BIC, B-cell integration cluster) and stimulates its transcription. This coincides with the hypermethylated histone H3K4 residue and spread hyperacetylation of H3K9 at MIR155HG promoter. Our data provide evidence of oncogenic activities of MYB in B-CLL that include its stimulatory role in MIR155HG transcription.
Assuntos
Leucemia Linfocítica Crônica de Células B/genética , MicroRNAs/genética , Proteínas Oncogênicas v-myb/fisiologia , Cromatina/química , Cromatina/genética , Cromatina/metabolismo , Análise por Conglomerados , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Células HeLa , Humanos , Leucemia Linfocítica Crônica de Células B/metabolismo , Análise em Microsséries , Proteínas Oncogênicas v-myb/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Transcrição Gênica/fisiologia , Transfecção , Células Tumorais CultivadasRESUMO
BACKGROUND/AIMS: Podocytes are typically cultured on collagen I; however, collagen I is absent from healthy glomerular basement membranes. Erythropoietin (EPO) is thought to protect podocytes in vivo. Here, we studied how various types of extracellular matrix (ECM) proteins and EPO affect podocytes in culture. METHODS: Primary rat podocytes were replated on collagen I, collagen IV, whole ECM extract, laminin, or bare plastic. Cellular adhesion (8 hours after plating), proliferation (5 days, 10 % serum), and resistance to serum deprivation (3 days, 0.5 % serum) were assessed. BrdU incorporation and expression of podocyte-specific markers were employed as measures of cellular proliferation and differentiation, respectively. qPCR was used to verify expression of EPO receptor in cultured podocytes. RESULTS: Cellular adhesion was similar on all ECM proteins and unaffected by EPO. Proliferation was accelerated by laminin and the ECM extract, but the final cell density was similar on all ECM surfaces. Collagen IV supported the serum-deprived cells better than the other ECM proteins. EPO (2-20 ng/ml) improved viability of serum-deprived podocytes on collagen I, collagen IV, and ECM, but not on laminin or bare plastic. The cells expressed mRNA for EPO receptor. CONCLUSION: The physiological ECM proteins are more supportive of primary podocytic cultures compared with collagen I. The protective effects of EPO during serum deprivation are modulated by the cultivation surface.
Assuntos
Eritropoetina/farmacologia , Proteínas da Matriz Extracelular/fisiologia , Glomérulos Renais/efeitos dos fármacos , Podócitos/efeitos dos fármacos , Animais , Células Cultivadas , Corantes , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Cultura Primária de Células , Ratos , Receptores da Eritropoetina/biossíntese , Receptores da Eritropoetina/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Sais de Tetrazólio , TiazóisRESUMO
Somatic mutations are a common molecular mechanism through which chronic myeloid leukemia (CML) cells acquire resistance to tyrosine kinase inhibitors (TKIs) therapy. While most of the mutations in the kinase domain of BCR-ABL1 can be successfully managed, the recurrent somatic mutations in other genes may be therapeutically challenging. Despite the major clinical relevance of mutation-associated resistance in CML, the mechanisms underlying mutation acquisition in TKI-treated leukemic cells are not well understood. This work demonstrated de novo acquisition of mutations on isolated single-cell sorted CML clones growing in the presence of imatinib. The acquisition of mutations was associated with the significantly increased expression of the LIG1 and PARP1 genes involved in the error-prone alternative nonhomologous end-joining pathway, leading to genomic instability, and increased expression of the UNG, FEN and POLD3 genes involved in the base-excision repair (long patch) pathway, allowing point mutagenesis. This work showed in vitro and in vivo that de novo acquisition of resistance-associated mutations in oncogenes is the prevalent method of somatic mutation development in CML under TKIs treatment.
Assuntos
Crise Blástica , Imidazóis , Leucemia Mielogênica Crônica BCR-ABL Positiva , Mutação , Piridazinas , Piridazinas/uso terapêutico , Piridazinas/administração & dosagem , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Imidazóis/uso terapêutico , Imidazóis/administração & dosagem , Humanos , Crise Blástica/genética , Crise Blástica/tratamento farmacológico , Crise Blástica/patologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Camundongos , Proteínas de Fusão bcr-abl/genética , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Niacinamida/análogos & derivados , PirazóisRESUMO
BACKGROUND: Through high-throughput next-generation sequencing of promoters of solute carrier and ATP-binding cassette genes, which encode drug transporters, we aimed to identify SNPs associated with the response to imatinib administered for first-line treatment of patients with chronic myeloid leukemia. METHODS: In silico analysis using publicly available databases was done to select the SLC and ABC genes and their promoters for the next-generation sequencing. SNPs associated with the imatinib response were identified using Fisher's exact probability tests and subjected to the linkage disequilibrium analyses with regulatory loci of concerned genes. We analyzed cumulative achievement of major molecular response and probability of event free survival in relation to identified SNP genotypes in 129 CML patients and performed multivariate analysis for determination of genotypes as independent predictors of outcome. Gene expression analysis of eight cell lines naturally carrying different genotypes was performed to outline an impact of genotypes on the gene expression. RESULTS: We observed significant differences in the frequencies of the rs460089-GC and rs460089-GG (SLC22A4) genotypes among rs2631365-TC (SLC22A5) genotype carriers that were associated with optimal and non-optimal responses, respectively. Loci rs460089 and rs2631365 were in highly significant linkage disequilibrium with 12 regulatory loci in introns of SLC22A4 and SLC22A5 encoding imatinib transporters. Genotype association analysis with the response to imatinib indicated that rs460089-GC carriers had a significantly higher probability of achieving a stable major molecular response (BCR-ABL1 transcript level below or equal to 0.1% in the international scale). In contrast, the rs460089-GG represented a risk factor for imatinib failure, which was significantly higher in rs460089-GG_rs2631365-TC carriers. CONCLUSIONS: This exploratory study depicted potentially important genetic markers predicting outcome of imatinib treatment, which may be helpful for tailoring therapy in clinical practice.
Assuntos
Antineoplásicos/administração & dosagem , Mesilato de Imatinib/administração & dosagem , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Proteínas de Transporte de Cátions Orgânicos/genética , Polimorfismo de Nucleotídeo Único , Membro 5 da Família 22 de Carreadores de Soluto/genética , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Feminino , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Mesilato de Imatinib/uso terapêutico , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Desequilíbrio de Ligação , Masculino , Taxa de Mutação , Regiões Promotoras Genéticas , Análise de Sequência de DNA/métodos , Simportadores , Resultado do TratamentoRESUMO
GATA-1 and PU.1 are two important hematopoietic transcription factors that mutually inhibit each other in progenitor cells to guide entrance into the erythroid or myeloid lineage, respectively. PU.1 controls its own expression during myelopoiesis by binding to the distal URE enhancer, whose deletion leads to acute myeloid leukemia (AML). We herein present evidence that GATA-1 binds to the PU.1 gene and inhibits its expression in human AML-erythroleukemias (EL). Furthermore, GATA-1 together with DNA methyl Transferase I (DNMT1) mediate repression of the PU.1 gene through the URE. Repression of the PU.1 gene involves both DNA methylation at the URE and its histone H3 lysine-K9 methylation and deacetylation as well as the H3K27 methylation at additional DNA elements and the promoter. The GATA-1-mediated inhibition of PU.1 gene transcription in human AML-EL mediated through the URE represents important mechanism that contributes to PU.1 downregulation and leukemogenesis that is sensitive to DNA demethylation therapy.
Assuntos
Fator de Transcrição GATA1/genética , Leucemia Eritroblástica Aguda/genética , Leucemia Mieloide Aguda/genética , Proteínas Proto-Oncogênicas/genética , Transativadores/genética , Diferenciação Celular/genética , DNA (Citosina-5-)-Metiltransferase 1 , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/genética , Elementos Facilitadores Genéticos , Fator de Transcrição GATA1/metabolismo , Regulação Leucêmica da Expressão Gênica , Histonas/genética , Humanos , Leucemia Eritroblástica Aguda/patologia , Leucemia Mieloide Aguda/patologia , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/metabolismo , Transativadores/biossíntese , Transativadores/metabolismo , Transcrição GênicaRESUMO
Bone marrow transplantation or ponatinib treatment are currently recommended strategies for management of patients with chronic myeloid leukemia (CML) harboring the T315I mutation and compound or polyclonal mutations. However, in some individual cases, these treatment scenarios cannot be applied. We used an alternative treatment strategy with interferon-α (IFN-α) given solo, sequentially or together with TKI in a group of 6 cases of high risk CML patients, assuming that the TKI-independent mechanism of action may lead to mutant clone repression. IFN-α based individualized therapy decreases of T315I or compound mutations to undetectable levels as assessed by next-generation deep sequencing, which was associated with a molecular response in 4/6 patients. Based on the observed results from immune profiling, we assumed that the principal mechanism leading to the success of the treatment was the immune activation induced with dasatinib pre-treatment followed by restoration of immunological surveillance after application of IFN-α therapy. Moreover, we showed that sensitive measurement of mutated BCR-ABL1 transcript levels augments the safety of this individualized treatment strategy.
Assuntos
Proteínas de Fusão bcr-abl/genética , Interferon-alfa/administração & dosagem , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Feminino , Proteínas de Fusão bcr-abl/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Interferon-alfa/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Masculino , Pessoa de Meia-Idade , Medicina de Precisão , Inibidores de Proteínas Quinases/farmacologia , Resultado do Tratamento , Adulto JovemAssuntos
Análise Mutacional de DNA/métodos , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Mutação , Reação em Cadeia da Polimerase/métodos , Inibidores de Proteínas Quinases/farmacologia , Alelos , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , PrognósticoRESUMO
CCCTC-binding factor (CTCF) can both activate as well as inhibit transcription by forming chromatin loops between regulatory regions and promoters. In this regard, Ctcf binding on non-methylated DNA and its interaction with the Cohesin complex results in differential regulation of the H19/Igf2 locus. Similarly, a role for CTCF has been established in normal hematopoietic development; however its involvement in leukemia remains elusive. Here, we show that Ctcf binds to the imprinting control region of H19/Igf2 in AML blasts. We also demonstrate that Smarca5, which also associates with the Cohesin complex, facilitates Ctcf binding to its target sites on DNA. Furthermore, Smarca5 supports Ctcf functionally and is needed for enhancer-blocking effect at ICR. We next asked whether CTCF and SMARCA5 control the expression of key hematopoiesis regulators. In normally differentiating myeloid cells both CTCF and SMARCA5 together with members of the Cohesin complex are recruited to the SPI1 gene, a key hematopoiesis regulator and leukemia suppressor. Due to DNA methylation, CTCF binding to the SPI1 gene is blocked in AML blasts. Upon AZA-mediated DNA demethylation of human AML blasts, CTCF and SMARCA5 are recruited to the -14.4 Enhancer of SPI1 gene and block its expression. Our data provide new insight into complex SPI1 gene regulation now involving additional key epigenetic factors, CTCF and SMARCA5 that control PU.1 expression at the -14.4 Enhancer.
Assuntos
Adenosina Trifosfatases/genética , Proteínas Cromossômicas não Histona/genética , Epigênese Genética , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/genética , Transativadores/genética , Doença Aguda , Adenosina Trifosfatases/metabolismo , Animais , Azacitidina/farmacologia , Fator de Ligação a CCCTC , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/metabolismo , Metilação de DNA/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Impressão Genômica , Células HeLa , Humanos , Immunoblotting , Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/metabolismo , Células K562 , Leucemia Eritroblástica Aguda/genética , Leucemia Eritroblástica Aguda/metabolismo , Leucemia Eritroblástica Aguda/patologia , Leucemia Mieloide/genética , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Microscopia Confocal , Ligação Proteica , Proteínas Proto-Oncogênicas/metabolismo , Interferência de RNA , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Proteínas Repressoras/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transativadores/metabolismoRESUMO
Hematopoietic transcription factors GATA-1 and PU.1 bind each other on DNA to block transcriptional programs of undesired lineage during hematopoietic commitment. Murine erythroleukemia (MEL) cells that coexpress GATA-1 and PU.1 are blocked at the blast stage but respond to molecular removal (downregulation) of PU.1 or addition (upregulation) of GATA-1 by inducing terminal erythroid differentiation. To test whether GATA-1 blocks PU.1 in MEL cells, we have conditionally activated a transgenic PU.1 protein fused with the estrogen receptor ligand-binding domain (PUER), resulting in activation of a myeloid transcriptional program. Gene expression arrays identified components of the PU.1-dependent transcriptome negatively regulated by GATA-1 in MEL cells, including CCAAT/enhancer binding protein alpha (Cebpa) and core-binding factor, beta subunit (Cbfb), which encode two key hematopoietic transcription factors. Inhibition of GATA-1 by small interfering RNA resulted in derepression of PU.1 target genes. Chromatin immunoprecipitation and reporter assays identified PU.1 motif sequences near Cebpa and Cbfb that are co-occupied by PU.1 and GATA-1 in the leukemic blasts. Significant derepression of Cebpa and Cbfb is achieved in MEL cells by either activation of PU.1 or knockdown of GATA-1. Furthermore, transcriptional regulation of these loci by manipulating the levels of PU.1 and GATA-1 involves quantitative increases in a transcriptionally active chromatin mark: acetylation of histone H3K9. Collectively, we show that either activation of PU.1 or inhibition of GATA-1 efficiently reverses the transcriptional block imposed by GATA-1 and leads to the activation of a myeloid transcriptional program directed by PU.1.