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1.
Medicine (Baltimore) ; 99(35): e22047, 2020 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-32871963

RESUMO

BACKGROUND: We identified the hub genes and pathways dysregulated in acute myeloid leukemia and the potential molecular mechanisms involved. METHODS: We downloaded the GSE15061 gene expression dataset from the Gene Expression Omnibus database and used weighted gene co-expression network analysis to identify hub genes. Differential expression of the genes was evaluated using the limma package in R software. Subsequently, we built a protein-protein interaction network followed by functional enrichment analysis. Then, the prognostic significance of gene expression was explored in terms of overall survival. Finally, transcription factor-mRNA (ribonucleic acid) and microRNA-mRNA interaction analysis was also explored. RESULTS: We identified 100 differentially expressed hub genes. Functional enrichment analysis indicated that the genes were principally involved in immune system regulation, host defense, and negative regulation of apoptosis and myeloid cell differentiation. We identified 4 hub genes, the expression of which was significantly correlated with overall survival. Finally, 26 key regulators for hub genes and 38 microRNA-mRNA interactions were identified. CONCLUSION: We performed a comprehensive bioinformatics analysis of hub genes potentially involved in acute myeloid leukemia development. Further molecular biological experiments are required to confirm the roles played by these genes.


Assuntos
Leucemia Mieloide Aguda/metabolismo , Biologia Computacional , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidade , MicroRNAs/metabolismo , Mapas de Interação de Proteínas , Análise de Sobrevida , Fatores de Transcrição/metabolismo , Transcriptoma
2.
Nat Commun ; 11(1): 4056, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792483

RESUMO

Autophagy has been associated with oncogenesis with one of its emerging key functions being its contribution to the metabolism of tumors. Therefore, deciphering the mechanisms of how autophagy supports tumor cell metabolism is essential. Here, we demonstrate that the inhibition of autophagy induces an accumulation of lipid droplets (LD) due to a decrease in fatty acid ß-oxidation, that leads to a reduction of oxidative phosphorylation (OxPHOS) in acute myeloid leukemia (AML), but not in normal cells. Thus, the autophagic process participates in lipid catabolism that supports OxPHOS in AML cells. Interestingly, the inhibition of OxPHOS leads to LD accumulation with the concomitant inhibition of autophagy. Mechanistically, we show that the disruption of mitochondria-endoplasmic reticulum (ER) contact sites (MERCs) phenocopies OxPHOS inhibition. Altogether, our data establish that mitochondria, through the regulation of MERCs, controls autophagy that, in turn finely tunes lipid degradation to fuel OxPHOS supporting proliferation and growth in leukemia.


Assuntos
Autofagia/fisiologia , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia/metabolismo , Mitocôndrias/metabolismo , Animais , Autofagia/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Proliferação de Células/fisiologia , Citometria de Fluxo , Humanos , Leucemia/genética , Leucemia Mieloide Aguda/patologia , Metabolismo dos Lipídeos/genética , Metabolismo dos Lipídeos/fisiologia , Lipogênese/genética , Lipogênese/fisiologia , Camundongos , Mitocôndrias/genética , Oxirredução , Fosforilação Oxidativa
3.
Proc Natl Acad Sci U S A ; 117(33): 20117-20126, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32747558

RESUMO

t(8;21)(q22;q22) acute myelogenous leukemia (AML) is morphologically characterized by a continuum of heterogeneous leukemia cells from myeloblasts to differentiated myeloid elements. Thus, t(8;21) AML is an excellent model for studying heterogeneous cell populations and cellular evolution during disease progression. Using integrative analyses of immunophenotype, RNA-sequencing (RNA-seq), and single-cell RNA-sequencing (scRNA-seq), we identified three distinct intrapatient leukemic cell populations that were arrested at different stages of myeloid differentiation: CD34+CD117dim blasts, CD34+CD117bri blasts, and abnormal myeloid cells with partial maturation (AM). CD117 is also known as c-KIT protein. CD34+CD117dim cells were blocked in the G0/G1 phase at disease onset, presenting with the regular morphology of myeloblasts showing features of granulocyte-monocyte progenitors (GMP), and were drug-resistant to chemotherapy. Genes associated with cell migration and adhesion (LGALS1, EMP3, and ANXA 2) were highly expressed in the CD34+CD117dim population. CD34+CD117bri blasts were blocked a bit later than the CD34+CD117dim population in the hematopoietic differentiation stage and displayed high proliferation ability. AM cells, which bear abnormal myelocyte morphology, especially overexpressed granule genes AZU1, ELANE, and PRTN3 and were sensitive to chemotherapy. scRNA-seq at different time points identified CD34+CD117dim blasts as an important leukemic cluster that expanded at postrelapse refractory stage after several cycles of chemotherapy. Patients with t(8;21) AML with a higher proportion of CD34+CD117dim cells had significantly worse clinical outcomes than those with a lower CD34+CD117dim proportion. Univariate and multivariate analyses identified CD34+CD117dim proportion as an independent factor for poor disease outcome. Our study provides evidence for the multidimensional heterogeneity of t(8;21)AML and may offer new tools for future disease stratification.


Assuntos
Leucemia Mieloide Aguda/classificação , Leucemia Mieloide Aguda/patologia , Subpopulações de Linfócitos T/classificação , Subpopulações de Linfócitos T/metabolismo , Adulto , Regulação Neoplásica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/metabolismo , Transcriptoma
4.
Nat Commun ; 11(1): 4147, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32811837

RESUMO

Mutated receptor tyrosine kinases (MT-RTKs) such as internal tandem duplication of FMS-like tyrosine kinase 3 (FLT3 ITD) and a point mutation KIT D816V are driver mutations for acute myeloid leukemia (AML). Clathrin assembly lymphoid myeloid leukemia protein (CALM) regulates intracellular transport of RTKs, however, the precise role for MT-RTKs remains elusive. We here show that CALM knock down leads to severely impaired FLT3 ITD- or KIT D814V-dependent cell growth compared to marginal influence on wild-type FLT3- or KIT-mediated cell growth. An antipsychotic drug chlorpromazine (CPZ) suppresses the growth of primary AML samples, and human CD34+CD38- AML cells including AML initiating cells with MT-RTKs in vitro and in vivo. Mechanistically, CPZ reduces CALM protein at post transcriptional level and perturbs the intracellular localization of MT-RTKs, thereby blocking their signaling. Our study presents a therapeutic strategy for AML with MT-RTKs by altering the intracellular localization of MT-RTKs using CPZ.


Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Clorpromazina/farmacologia , Leucemia Mieloide Aguda/genética , Proteínas Monoméricas de Montagem de Clatrina/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Feminino , Células HL-60 , Humanos , Leucemia Mieloide Aguda/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Proteínas Monoméricas de Montagem de Clatrina/genética , Mutação Puntual , Transdução de Sinais/efeitos dos fármacos , Sequências de Repetição em Tandem/genética , Transplante Heterólogo , Adulto Jovem
5.
Nat Commun ; 11(1): 3639, 2020 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-32686665

RESUMO

Integrated analysis of genomes, transcriptomes, proteomes and drug responses of cancer cell lines (CCLs) is an emerging approach to uncover molecular mechanisms of drug action. We extend this paradigm to measuring proteome activity landscapes by acquiring and integrating quantitative data for 10,000 proteins and 55,000 phosphorylation sites (p-sites) from 125 CCLs. These data are used to contextualize proteins and p-sites and predict drug sensitivity. For example, we find that Progesterone Receptor (PGR) phosphorylation is associated with sensitivity to drugs modulating estrogen signaling such as Raloxifene. We also demonstrate that Adenylate kinase isoenzyme 1 (AK1) inactivates antimetabolites like Cytarabine. Consequently, high AK1 levels correlate with poor survival of Cytarabine-treated acute myeloid leukemia patients, qualifying AK1 as a patient stratification marker and possibly as a drug target. We provide an interactive web application termed ATLANTiC (http://atlantic.proteomics.wzw.tum.de), which enables the community to explore the thousands of novel functional associations generated by this work.


Assuntos
Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Proteoma/metabolismo , Adenilato Quinase/metabolismo , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Biologia Computacional , Simulação por Computador , Citarabina/metabolismo , Citarabina/farmacologia , Desenvolvimento de Medicamentos , Genômica , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Neoplasias/metabolismo , Proteoma/genética , Proteômica , Cloridrato de Raloxifeno/metabolismo , Cloridrato de Raloxifeno/farmacologia , Receptores de Progesterona/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
6.
Life Sci ; 257: 118021, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32621919

RESUMO

AIMS: Tribbles homolog 3 (TRIB3) is emerging as a multifunctional oncoprotein associated with various cellular events in different tumors. However, the regulatory mechanism of TRIB3 in acute myeloid leukemia (AML) remains unknown. This study aims to investigate the molecular mechanisms and uncover the functions of TRIB3 in AML. METHODS: Western blotting and quantitative real-time PCR were used to analyze the expression levels of TRIB3, peroxisome proliferator-activated receptor α (PPARα), apoptosis markers and autophagy markers in AML cells. Flow cytometry was used to assess cell apoptosis. The interaction of TRIB3 and PPARα was evaluated by immunofluorescence, coimmunoprecipitation, and in vivo ubiquitination assays. KEY FINDINGS: We demonstrated that downregulating TRIB3 in leukemic cells effectively induced apoptosis and autophagy by regulating the degradation of PPARα. Mechanistically, TRIB3 interacted with PPARα and contributed to its destabilization by promoting its ubiquitination. When PPARα was activated by its specific agonist clofibrate, the apoptosis and autophagy of AML cells were significantly enhanced. These results were confirmed by rescue experiments. Blocking PPARα expression using the PPARα inhibitor GW6471 reversed the functional influence of TRIB3 on AML cells. SIGNIFICANCE: The aim of this study is to provide evidence of the degradation of PPARα by TRIB3 via ubiquitin-dependent proteasomal degradation. This process meditates the progression of AML and prolongs the survival of leukemic cells. As a result, these data indicate that TRIB3 is a novel and promising therapeutic target for AML treatment.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Leucemia Mieloide Aguda/genética , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Apoptose/fisiologia , Autofagia/fisiologia , Bases de Dados Genéticas , Humanos , Leucemia Mieloide Aguda/metabolismo , PPAR alfa/genética , PPAR alfa/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteostase/fisiologia , Transdução de Sinais/fisiologia , Ubiquitina/metabolismo , Ubiquitinação
7.
Life Sci ; 257: 118041, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32622945

RESUMO

AIM: Transcription factor CCAAT/Enhancer binding protein alpha (C/EBPα) is a key regulator of myeloid differentiation, granulopoiesis in particular. Although CEBPA mutations are found in more than 10% in AML, functional inhibition of C/EBPα protein is also widely observed in AML. Here, we sought to examine if SKP2, an aberrantly enhanced E3 ubiquitin ligase in primary AMLs inhibits C/EBPα stability to induce differentiation block. MAIN METHODS: Here we employed cell based assays such as transfections, immunoblotting, co-immunoprecipitation, luciferase and gel shift assays along with differentiation assays to investigate SKP2 regulated C/EBPα protein stability in acute myeloid leukemia. KEY FINDINGS: Here we discovered that oncogenic E3 ubiquitin ligase SCFskp2 ubiquitinates and destabilizes C/EBPα in a proteasome-dependent manner. Our data demonstrates that SKP2 physically interacts with C-terminal of C/EBPα and promotes its K48-linked ubiquitination-mediated degradation leading to its reduced transactivation potential, DNA binding ability and cellular functions. We further show that while overexpression of SKP2 inhibits both ectopic as well as endogenous C/EBPα in heterologous (HEK293T) as well as myeloid leukemia cells respectively, SKP2 depletion restores endogenous C/EBPα leading to reduced colony formation and enhanced myeloid differentiation of myeloid leukemia cells. Using Estradiol-inducible K562-C/EBPα-ER cells as yet another model of granulocytic differentiation, we further confirmed that SKP2 overexpression indeed inhibits granulocytic differentiation by mitigating C/EBPα stability. SIGNIFICANCE: Our findings identify SKP2 as a potential negative regulator of C/EBPα stability and function in AML which suggests that SKP2 can be potentially targeted in AML to restore C/EBPα and overcome differentiation block.


Assuntos
Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Células HEK293 , Humanos , Complexo de Endopeptidases do Proteassoma/metabolismo , Fatores de Transcrição/metabolismo , Transfecção , Células U937 , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
8.
Proc Natl Acad Sci U S A ; 117(25): 14331-14341, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32513686

RESUMO

Acute myeloid leukemia (AML) is the most common acute leukemia in adults, with approximately four new cases per 100,000 persons per year. Standard treatment for AML consists of induction chemotherapy with remission achieved in 50 to 75% of cases. Unfortunately, most patients will relapse and die from their disease, as 5-y survival is roughly 29%. Therefore, other treatment options are urgently needed. In recent years, immune-based therapies have led to unprecedented rates of survival among patients with some advanced cancers. Suppression of T cell function in the tumor microenvironment is commonly observed and may play a role in AML. We found that there is a significant association between T cell infiltration in the bone marrow microenvironment of newly diagnosed patients with AML and increased overall survival. Functional studies aimed at establishing the degree of T cell suppression in patients with AML revealed impaired T cell function in many patients. In most cases, T cell proliferation could be restored by blocking the immune checkpoint molecules PD-1, CTLA-4, or TIM3. Our data demonstrate that AML establishes an immune suppressive environment in the bone marrow, in part through T cell checkpoint function.


Assuntos
Medula Óssea/metabolismo , Leucemia Mieloide Aguda/metabolismo , Linfócitos T/metabolismo , Microambiente Tumoral/fisiologia , Medula Óssea/imunologia , Antígeno CTLA-4/metabolismo , Proliferação de Células , Citocinas/metabolismo , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Humanos , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/terapia , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/imunologia
9.
Nat Commun ; 11(1): 3021, 2020 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-32541670

RESUMO

The caudal-related homeobox transcription factor CDX2 is expressed in leukemic cells but not during normal blood formation. Retroviral overexpression of Cdx2 induces AML in mice, however the developmental stage at which CDX2 exerts its effect is unknown. We developed a conditionally inducible Cdx2 mouse model to determine the effects of in vivo, inducible Cdx2 expression in hematopoietic stem and progenitor cells (HSPCs). Cdx2-transgenic mice develop myelodysplastic syndrome with progression to acute leukemia associated with acquisition of additional driver mutations. Cdx2-expressing HSPCs demonstrate enrichment of hematopoietic-specific enhancers associated with pro-differentiation transcription factors. Furthermore, treatment of Cdx2 AML with azacitidine decreases leukemic burden. Extended scheduling of low-dose azacitidine shows greater efficacy in comparison to intermittent higher-dose azacitidine, linked to more specific epigenetic modulation. Conditional Cdx2 expression in HSPCs is an inducible model of de novo leukemic transformation and can be used to optimize treatment in high-risk AML.


Assuntos
Fator de Transcrição CDX2/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Leucemia Mieloide Aguda/metabolismo , Síndromes Mielodisplásicas/metabolismo , Animais , Fator de Transcrição CDX2/genética , Transformação Celular Neoplásica , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/fisiopatologia
10.
Cytogenet Genome Res ; 160(5): 255-263, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32544910

RESUMO

Fusions of the Runt-related transcription factor 1 (RUNX1) with different partner genes have been associated with various hematological disorders. Interestingly, the C-terminally truncated form of RUNX1 and RUNX1 fusion proteins are similarly considered important contributors to leukemogenesis. Here, we describe a 59-year-old male patient who was initially diagnosed with acute myeloid leukemia, inv(16)(p13;q22)/CBFB-MYH11 (FAB classification M4Eo). He achieved complete remission and negative CBFB-MYH11 status with daunorubicin/cytarabine combination chemotherapy but relapsed 3 years later. Cytogenetic analysis of relapsed leukemia cells revealed CBFB-MYH11 negativity and complex chromosomal abnormalities without inv(16)(p13;q22). RNA-seq identified the glutamate receptor, ionotropic, kinase 2 (GRIK2) gene on 6q16 as a novel fusion partner for RUNX1 in this case. Specifically, the fusion of RUNX1 to the GRIK2 antisense strand (RUNX1-GRIK2as) generated multiple missplicing transcripts. Because extremely low levels of wild-type GRIK2 were detected in leukemia cells, RUNX1-GRIK2as was thought to drive the pathogenesis associated with the RUNX1-GRIK2 fusion. The truncated RUNX1 generated from RUNX1-GRIK2as induced the expression of the granulocyte colony-stimulating factor (G-CSF) receptor on 32D myeloid leukemia cells and enhanced proliferation in response to G-CSF. In summary, the RUNX1-GRIK2as fusion emphasizes the importance of aberrantly truncated RUNX1 in leukemogenesis.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , DNA Antissenso/genética , Fusão Gênica/genética , Fator Estimulador de Colônias de Granulócitos/farmacologia , Leucemia Mieloide Aguda/genética , Receptores de Ácido Caínico/genética , Deleção de Sequência/genética , Translocação Genética/genética , Proliferação de Células/efeitos dos fármacos , Humanos , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , RNA Mensageiro/genética , Receptores de Fator Estimulador de Colônias de Granulócitos/biossíntese , Receptores de Fator Estimulador de Colônias de Granulócitos/metabolismo
11.
Gene ; 755: 144889, 2020 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-32534056

RESUMO

Ferroptosis, a newly discovered form of non-apoptotic cell death, is induced by an excessive degree of iron-dependent lipid peroxide. ATPR, a novel all-trans retinoic acid (ATRA) derivative, has been extensively developed to show superior anticancer effect than ATRA in acute myeloid leukemia (AML). However, whether ferroptosis exists during ATPR treatment of AML remains unclear. Herein, we found that ferroptosis occurred in an AML xenograft mouse model of ATPR treatment. In vitro, ATPR was verified to induce ferroptosis in a dose-dependent manner by proferroptotic protein marker, lipid peroxidation, and lipid ROS, which could be significantly reversed by ferrostatin-1. Using lysosomal inhibitor chloroquine and iron chelator desferrioxamine, we further revealed that ATPR-induced ferroptosis was regulated by autophagy via iron homeostasis, especially Nrf2. Furthermore, targeting ferroptosis contributes to ATPR-induced AML differentiation. In conclusion, these results indicated that ferroptosis play an important role in ATPR-induced differentiation, and suggested that ATPR would provide a potential therapeutic value for AML treatment.


Assuntos
Ferroptose/efeitos dos fármacos , Leucemia Mieloide Aguda/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Retinoides/farmacologia , Animais , Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Homeostase , Humanos , Ferro/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Nus , Transdução de Sinais/efeitos dos fármacos , Tretinoína/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Mol Immunol ; 123: 7-17, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32387766

RESUMO

The identification of T cell epitopes derived from tumour specific antigens remains a significant challenge for the development of peptide-based vaccines and immunotherapies. The use of mass spectrometry-based approaches (immunopeptidomics) can provide powerful new avenues for the identification of such epitopes. In this study we report the use of complementary peptide antigen enrichment methods and a comprehensive mass spectrometric acquisition strategy to provide in-depth immunopeptidome data for the THP-1 cell line, a cell line used widely as a model of human leukaemia. To accomplish this, we combined robust experimental workflows that incorporated ultrafiltration or off-line reversed phase chromatography to enrich peptide ligand as well as a multifaceted data acquisition strategy using an Orbitrap Fusion LC-MS instrument. Using the combined datasets from the two ligand enrichment methods we gained significant depth in immunopeptidome coverage by identifying a total of 41,816 HLA class I peptides from THP-1 cells, including a significant number of peptides derived from different oncogenes or over expressed proteins associated with cancer. The physicochemical properties of the HLA-bound peptides dictated their recovery using the two ligand enrichment approaches and their distribution across the different precursor charge states considered in the data acquisition strategy. The data highlight the complementarity of the two enrichment procedures, and in cases where sample is not limiting, suggest that the combination of both approaches will yield the most comprehensive immunopeptidome information.


Assuntos
Antígenos de Neoplasias/análise , Mineração de Dados/métodos , Epitopos de Linfócito T/imunologia , Leucemia Mieloide Aguda/metabolismo , Peptídeos/análise , Proteoma/análise , Antígenos de Neoplasias/química , Antígenos de Neoplasias/imunologia , Células Cultivadas , Cromatografia Líquida/métodos , Bases de Dados de Proteínas , Humanos , Imunoterapia/métodos , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/patologia , Leucemia Mieloide Aguda/terapia , Ligantes , Espectrometria de Massas/métodos , Biblioteca de Peptídeos , Peptídeos/química , Proteoma/química , Proteômica/métodos , Células THP-1
13.
J Cancer Res Ther ; 16(1): 23-27, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32362605

RESUMO

Objective: Growth factor independence 1 (GFI1), a transcriptional repressor, is required for hematopoietic stem cell maintenance and self-renewal in addition to controlling differentiation and proliferation of myeloid cells. As murine studies have demonstrated that this transcription factor has a notable role in the initiation and progression of acute myeloid leukemia (AML) disease, the aim of the current study was to investigate and review the influence of GFI1 in human AML cells. Methods: GFI1 expression levels were measured by means of real-time polymerase chain reaction in 96 primary AML samples which were then compared to gene expression levels observed in 18 healthy subjects. Moreover, GFI1 expression patterns were analyzed based on specific AML subtypes including acute promyelocytic leukemia (APL). Finally, leukemic cells were stained to measure levels of myeloperoxidase (MPO) activity. Results: This study reports that AML patients have significantly higher GFI1 mRNA levels in comparison to healthy subjects and that, when considering AML subtypes, patients with APL have higher GFI1 expression than non-APL patients. Conclusion: It is also concluded that GFI1 overexpression in patients with high MPO levels, such as those of the APL subtype, is correlated with favorable disease prognosis as supported by other studies which demonstrate that increased peroxide activity and GFI1 are independently correlated with a favorable prognosis.


Assuntos
Biomarcadores Tumorais/metabolismo , Proteínas de Ligação a DNA/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia Promielocítica Aguda/metabolismo , Fatores de Transcrição/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Criança , Pré-Escolar , Proteínas de Ligação a DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Fatores de Transcrição/genética , Adulto Jovem
14.
J Cancer Res Ther ; 16(1): 105-109, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32362618

RESUMO

Background: Aberrant phenotypes in acute leukemia have been reported with varying frequencies in independent studies and their association with prognostic factors is still a matter of debate. Aim: This study aims to identify the frequency of aberrant immunophenotypes in de novo acute myeloid leukemia (AML) and to evaluate their association with initial clinical and hematological features. Materials and Methods: A total of 181 patients of de novo AML were included during the time (July 2010-June 2012). The immunophenotype of all cases of AML was studied by using flow cytometry. Results: Aberrant lymphoid antigen expression was seen in 43.1% cases. Most frequent aberrant lymphoid antigen was CD7, seen in 26.5% cases. All French-American-British (FAB) subtypes except AML-M3 expressed aberrant lymphoid antigens. The expression was most common in AML-M4 in the current study. CD34 expression in AMLs was significantly associated with the expression of aberrant lymphoid antigens. Lymphoid antigen expression in adult AML was significantly associated with higher white blood cell (WBC) count (>50,000/mm3) and higher number of peripheral blasts (>70%). Conclusion: In summary, CD7 is the most common aberrant lymphoid antigen expressed in AML. FAB subtype AML-M3 is usually not associated with aberrant lymphoid antigen expression. AML cases with CD34 positivity are more likely to express aberrant lymphoid markers. The current study also supports that aberrant lymphoid antigen expression in adult AML is associated with adverse presenting hematological features (WBC count >50,000/mm3, peripheral blasts >70%). Pediatric Ly + AML cases are not associated with adverse presenting clinical and biological features.


Assuntos
Antígenos de Neoplasias/imunologia , Biomarcadores Tumorais/imunologia , Imunofenotipagem/métodos , Leucemia Mieloide Aguda/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD34/imunologia , Antígenos CD34/metabolismo , Antígenos CD7/imunologia , Antígenos CD7/metabolismo , Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/metabolismo , Masculino , Pessoa de Meia-Idade , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Prognóstico , Estudos Prospectivos , Centros de Atenção Terciária , Adulto Jovem
15.
Gene ; 752: 144758, 2020 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-32422235

RESUMO

Drugs targeting chromatin-modifying enzymes have entered clinical trials for myeloid malignancies, including INCB059872, a selective irreversible inhibitor of Lysine-Specific Demethylase 1 (LSD1). While initial studies of LSD1 inhibitors suggested these compounds may be used to induce differentiation of acute myeloid leukemia (AML), the mechanisms underlying this effect and dose-limiting toxicities are not well understood. Here, we used precision nuclear run-on sequencing (PRO-seq) and ChIP-seq in AML cell lines to probe for the earliest regulatory events associated with INCB059872 treatment. The changes in nascent transcription could be traced back to a loss of CoREST activity and activation of GFI1-regulated genes. INCB059872 is in phase I clinical trials, and we evaluated a pre-treatment bone marrow sample of a patient who showed a clinical response to INCB059872 while being treated with azacitidine. We used single-cell RNA-sequencing (scRNA-seq) to show that INCB059872 caused a shift in gene expression that was again associated with GFI1/GFI1B regulation. Finally, we treated mice with INCB059872 and performed scRNA-seq of lineage-negative bone marrow cells, which showed that INCB059872 triggered accumulation of megakaryocyte early progenitor cells with gene expression hallmarks of stem cells. Accumulation of these stem/progenitor cells may contribute to the thrombocytopenia observed in patients treated with LSD1 inhibitors.


Assuntos
Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Histona Desmetilases/antagonistas & inibidores , Leucemia Mieloide Aguda/metabolismo , Animais , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Camundongos , Camundongos Endogâmicos C57BL , RNA-Seq , Análise de Célula Única/métodos , Células-Tronco/metabolismo , Células THP-1 , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Sequenciamento Completo do Exoma/métodos
16.
Proc Natl Acad Sci U S A ; 117(24): 13670-13679, 2020 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-32471953

RESUMO

Acute myeloid leukemia (AML) is a deadly hematologic malignancy with poor prognosis, particularly in the elderly. Even among individuals with favorable-risk disease, approximately half will relapse with conventional therapy. In this clinical circumstance, the determinants of relapse are unclear, and there are no therapeutic interventions that can prevent recurrent disease. Mutations in the transcription factor CEBPA are associated with favorable risk in AML. However, mutations in the growth factor receptor CSF3R are commonly co-occurrent in CEBPA mutant AML and are associated with an increased risk of relapse. To develop therapeutic strategies for this disease subset, we performed medium-throughput drug screening on CEBPA/CSF3R mutant leukemia cells and identified sensitivity to inhibitors of lysine-specific demethylase 1 (LSD1). Treatment of CSF3R/CEBPA mutant leukemia cells with LSD1 inhibitors reactivates differentiation-associated enhancers driving immunophenotypic and morphologic differentiation. LSD1 inhibition is ineffective as monotherapy but demonstrates synergy with inhibitors of JAK/STAT signaling, doubling median survival in vivo. These results demonstrate that combined inhibition of JAK/STAT signaling and LSD1 is a promising therapeutic strategy for CEBPA/CSF3R mutant AML.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/genética , Inibidores Enzimáticos/administração & dosagem , Histona Desmetilases/antagonistas & inibidores , Janus Quinase 2/antagonistas & inibidores , Leucemia Mieloide Aguda/tratamento farmacológico , Receptores de Fator Estimulador de Colônias/genética , Fatores de Transcrição STAT/metabolismo , Animais , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Feminino , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Receptores de Fator Estimulador de Colônias/metabolismo , Fatores de Transcrição STAT/antagonistas & inibidores , Fatores de Transcrição STAT/genética , Transdução de Sinais/efeitos dos fármacos
17.
Proc Natl Acad Sci U S A ; 117(22): 12332-12340, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32424097

RESUMO

Double knockout of the two miR-15/16 loci in mouse resulted in the development of acute myeloid leukemia (AML). This result suggested that, at least, a fraction of human AMLs could be due to a similar mechanism. We analyzed the role of the two miR-15/16 clusters in 93 myelodysplastic syndrome (MDS) patients divided in three subgroups: patients with MDS, patients with MDS before transforming into AML (MDS-T), and patients with AML evolving from MDS (MDS-AML). Then, we tested 139 AML cases and 14 different AML cell lines by assessing microRNA (miRNA) expression, target protein expression, genetic loss, and silencing. MDS-T and MDS-AML patients show a reduction of the expression of miR-15a/-15b/-16 compared to MDS patients. Each miRNA can significantly predict MDS and MDS-T groups. Then, 79% of primary AMLs show a reduced expression of miR-15a and/or miR-15b. The expression of miR-15a/-15b/-16 significantly stratified AML patients in two prognostic classes. Furthermore, 40% of AML cell lines showed a combined loss of the expression of miR-15a/-15b and overexpression of their direct/indirect targets. As potential mechanisms involved in the silencing of the two miR-15/16 loci, we identified a genetic loss of miR-15a and miR-15b and silencing of these two loci by methylation. We identified a potential driver oncogenic role in the loss of expression of both miR-15/16 clusters in the progression of MDS into AML and in AML pathogenesis. The stratification of AML patients, based on miR-15/16 expression, can lead to targeted and combination therapies for the treatment of this incurable disease.


Assuntos
Leucemia Mieloide Aguda/genética , MicroRNAs/genética , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Progressão da Doença , Feminino , Regulação Leucêmica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Masculino , MicroRNAs/metabolismo , Pessoa de Meia-Idade
18.
Am J Hematol ; 95(7): 799-808, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32249963

RESUMO

This multi-institutional study retrospectively evaluated clinicopathologic and genetic characteristics in 351 patients with core-binding-factor acute myeloid leukemia (CBF-AML), comprising 69 therapy-related (t-CBF-AML) and 282 de novo cases. The T-CBF-AML patients were older, had lower WBC counts, and slightly higher hemoglobin than patients with de novo disease. Secondary cytogenetic abnormalities were more frequent in patients with de novo disease than t-CBF-AML (57.1% vs 41.1%, P = .026). Patients with secondary cytogenetic abnormalities had longer overall survival (OS) than those without abnormalities (median 190 vs 87 months, P = .021); trisomy 8, trisomy 22, and loss of the X or Y chromosome were associated with longer OS. In the 165 cases performed of targeted gene sequencing, pathogenic mutations were detected in 75.7% of cases, and were more frequent in de novo than in therapy-related disease (P = .013). Mutations were found in N/KRAS (37.0%), FLT3 (27.8%), KIT (17.2%), TET2 (4.9%), and ASXL1 (3.9%). The TET2 mutations were associated with shorter OS (P = .012) while N/KRAS mutation was associated with longer OS in t(8;21) AML patients (P = .001). The KIT mutation did not show prognostic significance in this cohort. Although they received similar therapy, t-CBF-AML patients had shorter OS than de novo patients (median 69 vs 190 months, P = .038). In multivariate analysis of all patients, older age and absence of any secondary cytogenetic abnormalities were significant predictors of shorter OS. Among the t-CBF-AML subset, age and hemoglobin were significant on multivariate analysis. This study demonstrated that although de novo and t-CBF-AML patients share many features, t-CBF-AML patients have worse clinical outcome than de novo patients.


Assuntos
Aberrações Cromossômicas , Fatores de Ligação ao Core , Leucemia Mieloide Aguda , Proteínas de Neoplasias , Adulto , Fatores de Ligação ao Core/genética , Fatores de Ligação ao Core/metabolismo , Intervalo Livre de Doença , Feminino , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/mortalidade , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Estudos Retrospectivos , Taxa de Sobrevida
19.
Nat Cell Biol ; 22(6): 689-700, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32313104

RESUMO

Leukaemia stem cells (LSCs) underlie cancer therapy resistance but targeting these cells remains difficult. The Wnt-ß-catenin and PI3K-Akt pathways cooperate to promote tumorigenesis and resistance to therapy. In a mouse model in which both pathways are activated in stem and progenitor cells, LSCs expanded under chemotherapy-induced stress. Since Akt can activate ß-catenin, inhibiting this interaction might target therapy-resistant LSCs. High-throughput screening identified doxorubicin (DXR) as an inhibitor of the Akt-ß-catenin interaction at low doses. Here we repurposed DXR as a targeted inhibitor rather than a broadly cytotoxic chemotherapy. Targeted DXR reduced Akt-activated ß-catenin levels in chemoresistant LSCs and reduced LSC tumorigenic activity. Mechanistically, ß-catenin binds multiple immune-checkpoint gene loci, and targeted DXR treatment inhibited expression of multiple immune checkpoints specifically in LSCs, including PD-L1, TIM3 and CD24. Overall, LSCs exhibit distinct properties of immune resistance that are reduced by inhibiting Akt-activated ß-catenin. These findings suggest a strategy for overcoming cancer therapy resistance and immune escape.


Assuntos
Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Leucemia Mieloide Aguda/patologia , Células-Tronco Neoplásicas/patologia , PTEN Fosfo-Hidrolase/fisiologia , Proteínas Wnt/fisiologia , beta Catenina/fisiologia , Animais , Antibióticos Antineoplásicos/farmacologia , Apoptose , Proliferação de Células , Feminino , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Masculino , Camundongos , Camundongos Knockout , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Adv Exp Med Biol ; 1219: 403-411, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32130711

RESUMO

This chapter provides a brief overview of the methods to study and modulate the metabolic phenotype of the tumor microenvironment, including own research work to demonstrate the impact that metabolic shifts in the host have on cancer. Firstly, we briefly discuss the relevance of using animal models to address this topic, and also the importance of acknowledging that animals have diverse metabolic phenotypes according to species, and even with strain, age or sex. We also present original data to highlight the impact that changes in metabolic phenotype of the microenvironment have on tumor progression. Using an acute leukemia mouse xenograft model and high-fat diet we show that a shift in the host metabolic phenotype, induced by high-fat feeding, significantly impacts on tumor progression. The mechanism through which this occurs involves a direct effect of the increased levels of circulating lipoproteins in both tumor and non-neoplastic cells.


Assuntos
Leucemia Mieloide Aguda/metabolismo , Microambiente Tumoral , Animais , Dieta Hiperlipídica , Modelos Animais de Doenças , Leucemia Mieloide Aguda/patologia , Camundongos , Fenótipo
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