RESUMO
B lymphocyte development and selection are central to adaptive immunity and self-tolerance. These processes require B cell receptor (BCR) signaling and occur in bone marrow, an environment with variable hypoxia, but whether hypoxia-inducible factor (HIF) is involved is unknown. We show that HIF activity is high in human and murine bone marrow pro-B and pre-B cells and decreases at the immature B cell stage. This stage-specific HIF suppression is required for normal B cell development because genetic activation of HIF-1α in murine B cells led to reduced repertoire diversity, decreased BCR editing and developmental arrest of immature B cells, resulting in reduced peripheral B cell numbers. HIF-1α activation lowered surface BCR, CD19 and B cell-activating factor receptor and increased expression of proapoptotic BIM. BIM deletion rescued the developmental block. Administration of a HIF activator in clinical use markedly reduced bone marrow and transitional B cells, which has therapeutic implications. Together, our work demonstrates that dynamic regulation of HIF-1α is essential for normal B cell development.
Assuntos
Linfócitos B/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Linfopoese/genética , Animais , Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/metabolismo , Linfócitos B/citologia , Linfócitos B/imunologia , Biomarcadores , Regulação da Expressão Gênica , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Cadeias Leves de Imunoglobulina/genética , Imunofenotipagem , Camundongos , Camundongos Knockout , Edição de RNA , Receptores de Antígenos de Linfócitos B/metabolismo , Transdução de Sinais , Ativação TranscricionalRESUMO
HOXA9 is commonly upregulated in acute myeloid leukemia (AML), in which it confers a poor prognosis. Characterizing the protein interactome of endogenous HOXA9 in human AML, we identified a chromatin complex of HOXA9 with the nuclear matrix attachment protein SAFB. SAFB perturbation phenocopied HOXA9 knockout to decrease AML proliferation, increase differentiation and apoptosis in vitro, and prolong survival in vivo. Integrated genomic, transcriptomic, and proteomic analyses further demonstrated that the HOXA9-SAFB (H9SB)-chromatin complex associates with nucleosome remodeling and histone deacetylase (NuRD) and HP1γ to repress the expression of factors associated with differentiation and apoptosis, including NOTCH1, CEBPδ, S100A8, and CDKN1A. Chemical or genetic perturbation of NuRD and HP1γ-associated catalytic activity also triggered differentiation, apoptosis, and the induction of these tumor-suppressive genes. Importantly, this mechanism is operative in other HOXA9-dependent AML genotypes. This mechanistic insight demonstrates the active HOXA9-dependent differentiation block as a potent mechanism of disease maintenance in AML that may be amenable to therapeutic intervention by targeting the H9SB interface and/or NuRD and HP1γ activity.
Assuntos
Leucemia Mieloide Aguda , Proteínas de Ligação à Região de Interação com a Matriz , Humanos , Proteômica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Fatores de Transcrição/genética , Proteínas Associadas à Matriz Nuclear , Cromatina , Receptores de Estrogênio/genética , Receptores de Estrogênio/uso terapêutico , Proteínas de Ligação à Região de Interação com a Matriz/genéticaRESUMO
The accessibility of cell surface proteins makes them tractable for targeting by cancer immunotherapy, but identifying suitable targets remains challenging. Here we describe plasma membrane profiling of primary human myeloma cells to identify an unprecedented number of cell surface proteins of a primary cancer. We used a novel approach to prioritize immunotherapy targets and identified a cell surface protein not previously implicated in myeloma, semaphorin-4A (SEMA4A). Using knock-down by short-hairpin RNA and CRISPR/nuclease-dead Cas9 (dCas9), we show that expression of SEMA4A is essential for normal myeloma cell growth in vitro, indicating that myeloma cells cannot downregulate the protein to avoid detection. We further show that SEMA4A would not be identified as a myeloma therapeutic target by standard CRISPR/Cas9 knockout screens because of exon skipping. Finally, we potently and selectively targeted SEMA4A with a novel antibody-drug conjugate in vitro and in vivo.
Assuntos
Mieloma Múltiplo , Semaforinas , Membrana Celular/metabolismo , Humanos , Fatores Imunológicos , Imunoterapia , Proteínas de Membrana , Mieloma Múltiplo/genética , Mieloma Múltiplo/terapia , Proteômica , Semaforinas/genética , Semaforinas/metabolismoRESUMO
Haematopoietic stem cells drive blood production, but their population size and lifetime dynamics have not been quantified directly in humans. Here we identified 129,582 spontaneous, genome-wide somatic mutations in 140 single-cell-derived haematopoietic stem and progenitor colonies from a healthy 59-year-old man and applied population-genetics approaches to reconstruct clonal dynamics. Cell divisions from early embryogenesis were evident in the phylogenetic tree; all blood cells were derived from a common ancestor that preceded gastrulation. The size of the stem cell population grew steadily in early life, reaching a stable plateau by adolescence. We estimate the numbers of haematopoietic stem cells that are actively making white blood cells at any one time to be in the range of 50,000-200,000. We observed adult haematopoietic stem cell clones that generate multilineage outputs, including granulocytes and B lymphocytes. Harnessing naturally occurring mutations to report the clonal architecture of an organ enables the high-resolution reconstruction of somatic cell dynamics in humans.
Assuntos
Células Sanguíneas/citologia , Células Sanguíneas/metabolismo , Linhagem da Célula/genética , Análise Mutacional de DNA , Mutação , Células-Tronco Adultas/citologia , Teorema de Bayes , Contagem de Células , Divisão Celular , Células Clonais/citologia , Células Clonais/metabolismo , Desenvolvimento Embrionário/genética , Genoma Humano/genética , Granulócitos/citologia , Granulócitos/metabolismo , Hematopoese/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Linfócitos/citologia , Linfócitos/metabolismo , Masculino , Pessoa de Meia-Idade , Fatores de TempoRESUMO
Cohesin complex disruption alters gene expression, and cohesin mutations are common in myeloid neoplasia, suggesting a critical role in hematopoiesis. Here, we explore cohesin dynamics and regulation of hematopoietic stem cell homeostasis and differentiation. Cohesin binding increases at active regulatory elements only during erythroid differentiation. Prior binding of the repressive Ets transcription factor Etv6 predicts cohesin binding at these elements and Etv6 interacts with cohesin at chromatin. Depletion of cohesin severely impairs erythroid differentiation, particularly at Etv6-prebound loci, but augments self-renewal programs. Together with corroborative findings in acute myeloid leukemia and myelodysplastic syndrome patient samples, these data suggest cohesin-mediated alleviation of Etv6 repression is required for dynamic expression at critical erythroid genes during differentiation and how this may be perturbed in myeloid malignancies.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Regulação Leucêmica da Expressão Gênica , Mutação , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/metabolismo , Biomarcadores Tumorais , Linhagem Celular Tumoral , Feminino , Dosagem de Genes , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/patologia , Histonas/metabolismo , Humanos , Masculino , Transtornos Mieloproliferativos/diagnóstico , Gradação de Tumores , Ligação Proteica , Proteínas Proto-Oncogênicas c-ets/metabolismo , Sequências Reguladoras de Ácido Nucleico , Proteínas Repressoras/metabolismo , Coesinas , Variante 6 da Proteína do Fator de Translocação ETSRESUMO
Bromodomain and extra terminal protein (BET) inhibitors are first-in-class targeted therapies that deliver a new therapeutic opportunity by directly targeting bromodomain proteins that bind acetylated chromatin marks. Early clinical trials have shown promise, especially in acute myeloid leukaemia, and therefore the evaluation of resistance mechanisms is crucial to optimize the clinical efficacy of these drugs. Here we use primary mouse haematopoietic stem and progenitor cells immortalized with the fusion protein MLL-AF9 to generate several single-cell clones that demonstrate resistance, in vitro and in vivo, to the prototypical BET inhibitor, I-BET. Resistance to I-BET confers cross-resistance to chemically distinct BET inhibitors such as JQ1, as well as resistance to genetic knockdown of BET proteins. Resistance is not mediated through increased drug efflux or metabolism, but is shown to emerge from leukaemia stem cells both ex vivo and in vivo. Chromatin-bound BRD4 is globally reduced in resistant cells, whereas the expression of key target genes such as Myc remains unaltered, highlighting the existence of alternative mechanisms to regulate transcription. We demonstrate that resistance to BET inhibitors, in human and mouse leukaemia cells, is in part a consequence of increased Wnt/ß-catenin signalling, and negative regulation of this pathway results in restoration of sensitivity to I-BET in vitro and in vivo. Together, these findings provide new insights into the biology of acute myeloid leukaemia, highlight potential therapeutic limitations of BET inhibitors, and identify strategies that may enhance the clinical utility of these unique targeted therapies.
Assuntos
Benzodiazepinas/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Proteínas Nucleares/antagonistas & inibidores , Fatores de Transcrição/antagonistas & inibidores , Animais , Azepinas/farmacologia , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Células Cultivadas , Cromatina/metabolismo , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Células Clonais/patologia , Resistencia a Medicamentos Antineoplásicos/genética , Epigênese Genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genes myc/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Terapia de Alvo Molecular , Células-Tronco Neoplásicas/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica/efeitos dos fármacos , Triazóis/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/metabolismoRESUMO
FLT3 internal tandem duplication (FLT3ITD) mutations are common in acute myeloid leukemia (AML) associated with poor patient prognosis. Although new-generation FLT3 tyrosine kinase inhibitors (TKI) have shown promising results, the outcome of FLT3ITD AML patients remains poor and demands the identification of novel, specific, and validated therapeutic targets for this highly aggressive AML subtype. Utilizing an unbiased genome-wide clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 screen, we identify GLS, the first enzyme in glutamine metabolism, as synthetically lethal with FLT3-TKI treatment. Using complementary metabolomic and gene-expression analysis, we demonstrate that glutamine metabolism, through its ability to support both mitochondrial function and cellular redox metabolism, becomes a metabolic dependency of FLT3ITD AML, specifically unmasked by FLT3-TKI treatment. We extend these findings to AML subtypes driven by other tyrosine kinase (TK) activating mutations and validate the role of GLS as a clinically actionable therapeutic target in both primary AML and in vivo models. Our work highlights the role of metabolic adaptations as a resistance mechanism to several TKI and suggests glutaminolysis as a therapeutically targetable vulnerability when combined with specific TKI in FLT3ITD and other TK activating mutation-driven leukemias.
Assuntos
Glutamina/metabolismo , Leucemia Mieloide Aguda , Mutação , Inibidores de Proteínas Quinases/farmacologia , Tirosina Quinase 3 Semelhante a fms , Sistemas CRISPR-Cas , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Estudo de Associação Genômica Ampla , Glutamina/genética , Humanos , Células K562 , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/enzimologia , Leucemia Mieloide Aguda/genética , Células THP-1 , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/metabolismoRESUMO
PURPOSE OF REVIEW: The field of acute myeloid leukemia (AML) has been revolutionized in recent years by the advent of high-throughput techniques, such as next-generation sequencing. In this review, we will discuss some of the recently identified mutations that have defined a new molecular landscape in this disease, as well as their prognostic, predictive, and therapeutic implications. RECENT FINDINGS: Recent studies have shown how many cases of AML evolve from a premalignant period of latency characterized by the accumulation of several mutations and the emergence of one or multiple dominant clones. The pattern of co-occurring mutations and cytogenetic abnormalities at diagnosis defines risk and can determine therapeutic approaches to induce remission. Besides the genetic landscape at diagnosis, the continued presence of particular gene mutations during or after treatment carries prognostic information that should further influence strategies to maintain remission in the long term. The recent progress made in AML research is a seminal example of how basic science can translate into improving clinical practice. Our ability to characterize the genomic landscape of individual patients has not only improved our ability to diagnose and prognosticate but is also bringing the promise of precision medicine to fruition in the field.
Assuntos
Leucemia Mieloide Aguda/genética , Aberrações Cromossômicas , Humanos , Leucemia Mieloide Aguda/mortalidade , Leucemia Mieloide Aguda/terapia , Mutação , Proteínas Nucleares/genética , Nucleofosmina , Prognóstico , Tirosina Quinase 3 Semelhante a fms/genéticaRESUMO
Over the last decade transcriptional dysregulation and altered epigenetic programs have emerged as a hallmark in the majority of hematological cancers. Several epigenetic regulators are recurrently mutated in many hematological malignancies. In addition, in those cases that lack epigenetic mutations, altered function of epigenetic regulators has been shown to play a central role in the pathobiology of many hematological neoplasms, through mechanisms that are becoming increasingly understood. This, in turn, has led to the development of small molecule inhibitors of dysregulated epigenetic pathways as novel targeted therapies for hematological malignancies. In this review, we will present the most recent advances in our understanding of the role played by dysregulated epigenetic programs in the development and maintenance of hematological neoplasms. We will describe novel therapeutics targeting altered epigenetic programs and outline their mode of action. We will then discuss their use in specific conditions, identify potential limitations and putative toxicities while also providing an update on their current clinical development. Finally, we will highlight the opportunities presented by epigenetically targeted therapies in hematological malignancies and introduce the challenges that need to be tackled by both the research and clinical communities to best translate these novel therapies into clinical practice and to improve patient outcomes.
Assuntos
Metilação de DNA , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hematológicas/genética , Animais , Neoplasias Hematológicas/patologia , HumanosRESUMO
Recent major advances in understanding the molecular basis of acute myeloid leukemia (AML) provide a double-edged sword. Although defining the topology and key features of the molecular landscape are fundamental to development of novel treatment approaches and provide opportunities for greater individualization of therapy, confirmation of the genetic complexity presents a huge challenge to successful translation into routine clinical practice. It is now clear that many genes are recurrently mutated in AML; moreover, individual leukemias harbor multiple mutations and are potentially composed of subclones with differing mutational composition, rendering each patient's AML genetically unique. In order to make sense of the overwhelming mutational data and capitalize on this clinically, it is important to identify (1) critical AML-defining molecular abnormalities that distinguish biological disease entities; (2) mutations, typically arising in subclones, that may influence prognosis but are unlikely to be ideal therapeutic targets; (3) mutations associated with preleukemic clones; and (4) mutations that have been robustly shown to confer independent prognostic information or are therapeutically relevant. The reward of identifying AML-defining molecular lesions present in all leukemic populations (including subclones) has been exemplified by acute promyelocytic leukemia, where successful targeting of the underlying PML-RARα oncoprotein has eliminated the need for chemotherapy for disease cure. Despite the molecular heterogeneity and recognizing that treatment options for other forms of AML are limited, this review will consider the scope for using novel molecular information to improve diagnosis, identify subsets of patients eligible for targeted therapies, refine outcome prediction, and track treatment response.
Assuntos
Antineoplásicos/uso terapêutico , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/tratamento farmacológico , Terapia de Alvo Molecular , Adulto , Humanos , Prognóstico , Adulto JovemRESUMO
Recurrent chromosomal translocations involving the mixed lineage leukaemia (MLL) gene initiate aggressive forms of leukaemia, which are often refractory to conventional therapies. Many MLL-fusion partners are members of the super elongation complex (SEC), a critical regulator of transcriptional elongation, suggesting that aberrant control of this process has an important role in leukaemia induction. Here we use a global proteomic strategy to demonstrate that MLL fusions, as part of SEC and the polymerase-associated factor complex (PAFc), are associated with the BET family of acetyl-lysine recognizing, chromatin 'adaptor' proteins. These data provided the basis for therapeutic intervention in MLL-fusion leukaemia, via the displacement of the BET family of proteins from chromatin. We show that a novel small molecule inhibitor of the BET family, GSK1210151A (I-BET151), has profound efficacy against human and murine MLL-fusion leukaemic cell lines, through the induction of early cell cycle arrest and apoptosis. I-BET151 treatment in two human leukaemia cell lines with different MLL fusions alters the expression of a common set of genes whose function may account for these phenotypic changes. The mode of action of I-BET151 is, at least in part, due to the inhibition of transcription at key genes (BCL2, C-MYC and CDK6) through the displacement of BRD3/4, PAFc and SEC components from chromatin. In vivo studies indicate that I-BET151 has significant therapeutic value, providing survival benefit in two distinct mouse models of murine MLL-AF9 and human MLL-AF4 leukaemia. Finally, the efficacy of I-BET151 against human leukaemia stem cells is demonstrated, providing further evidence of its potent therapeutic potential. These findings establish the displacement of BET proteins from chromatin as a promising epigenetic therapy for these aggressive leukaemias.
Assuntos
Cromatina/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Proteína de Leucina Linfoide-Mieloide/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Animais , Linhagem Celular Tumoral , Cromatina/genética , Imunoprecipitação da Cromatina , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Compostos Heterocíclicos de 4 ou mais Anéis/uso terapêutico , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Modelos Moleculares , Complexos Multiproteicos/química , Complexos Multiproteicos/metabolismo , Ligação Proteica/efeitos dos fármacos , Proteômica , Transcrição Gênica/efeitos dos fármacosRESUMO
Chronic myeloid leukemia (CML) is initiated and maintained by the tyrosine kinase BCR-ABL which activates a number of signal transduction pathways, including PI3K/AKT signaling and consequently inactivates FOXO transcription factors. ABL-specific tyrosine kinase inhibitors (TKIs) induce minimal apoptosis in CML progenitor cells, yet exert potent antiproliferative effects, through as yet poorly understood mechanisms. Here, we demonstrate that in CD34+ CML cells, FOXO1 and 3a are inactivated and relocalized to the cytoplasm by BCR-ABL activity. TKIs caused a decrease in phosphorylation of FOXOs, leading to their relocalization from cytoplasm (inactive) to nucleus (active), where they modulated the expression of key FOXO target genes, such as Cyclin D1, ATM, CDKN1C, and BCL6 and induced G1 arrest. Activation of FOXO1 and 3a and a decreased expression of their target gene Cyclin D1 were also observed after 6 days of in vivo treatment with dasatinib in a CML transgenic mouse model. The over-expression of FOXO3a in CML cells combined with TKIs to reduce proliferation, with similar results seen for inhibitors of PI3K/AKT/mTOR signaling. While stable expression of an active FOXO3a mutant induced a similar level of quiescence to TKIs alone, shRNA-mediated knockdown of FOXO3a drove CML cells into cell cycle and potentiated TKI-induced apoptosis. These data demonstrate that TKI-induced G1 arrest in CML cells is mediated through inhibition of the PI3K/AKT pathway and reactivation of FOXOs. This enhanced understanding of TKI activity and induced progenitor cell quiescence suggests that new therapeutic strategies for CML should focus on manipulation of this signaling network.
Assuntos
Fatores de Transcrição Forkhead/biossíntese , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Animais , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Dasatinibe/farmacologia , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Fase G1/efeitos dos fármacos , Perfilação da Expressão Gênica , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Fosforilação , Transdução de Sinais , TransfecçãoRESUMO
Despite their known transforming properties, the effects of leukemogenic FLT3-ITD mutations on hematopoietic stem and multipotent progenitor cells and on hematopoietic differentiation are not well understood. We report a mouse model harboring an ITD in the murine Flt3 locus that develops myeloproliferative disease resembling CMML and further identified FLT3-ITD mutations in a subset of human CMML. These findings correlated with an increase in number, cell cycling, and survival of multipotent stem and progenitor cells in an ITD dose-dependent manner in animals that exhibited alterations within their myeloid progenitor compartments and a block in normal B cell development. This model provides insights into the consequences of constitutive signaling by an oncogenic tyrosine kinase on hematopoietic progenitor quiescence, function, and cell fate.
Assuntos
Proliferação de Células , Células-Tronco Hematopoéticas/metabolismo , Leucemia Mielomonocítica Crônica/metabolismo , Células-Tronco Multipotentes/metabolismo , Mutação , Transtornos Mieloproliferativos/metabolismo , Tirosina Quinase 3 Semelhante a fms/metabolismo , Animais , Diferenciação Celular , Sobrevivência Celular , Células Cultivadas , Éxons , Regulação Neoplásica da Expressão Gênica , Genótipo , Células-Tronco Hematopoéticas/patologia , Humanos , Estimativa de Kaplan-Meier , Leucemia Experimental/metabolismo , Leucemia Experimental/patologia , Leucemia Mielomonocítica Crônica/genética , Leucemia Mielomonocítica Crônica/mortalidade , Leucemia Mielomonocítica Crônica/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Multipotentes/patologia , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/patologia , Fenótipo , Transdução de Sinais , Tirosina Quinase 3 Semelhante a fms/genéticaRESUMO
Many cancers seem to depend on a small population of 'cancer stem cells' for their continued growth and propagation. The leukaemia stem cell (LSC) was the first such cell to be described. The origins of these cells are controversial, and their biology - like that of their normal-tissue counterpart, the haematopoietic stem cell (HSC) - is still not fully elucidated. However, the LSC is likely to be the most crucial target in the treatment of leukaemias, and a thorough understanding of its biology - particularly of how the LSC differs from the HSC - might allow it to be selectively targeted, improving therapeutic outcome.
Assuntos
Leucemia/patologia , Células-Tronco Neoplásicas/fisiologia , Animais , Biomarcadores Tumorais , Divisão Celular , Sistemas de Liberação de Medicamentos , Células-Tronco Hematopoéticas/fisiologia , História do Século XIX , História do Século XX , Humanos , Leucemia/genética , Leucemia/história , Camundongos , Modelos Biológicos , Pesquisa , Transcrição GênicaRESUMO
CML is readily treatable with tyrosine kinase inhibitors (TKIs); however, resistance occurs, with the disease curable in only â¼15%-20% of patients. Using integrated functional genomics, Adnan Awad et al.1 identify agents effective against CML stem cells and describe mechanisms underlying TKI resistance.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Genômica , Leucemia Mielogênica Crônica BCR-ABL Positiva , Inibidores de Proteínas Quinases , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Genômica/métodosRESUMO
Myeloproliferative neoplasms (MPNs) are associated with recurrent activating mutations of signaling proteins such as Janus kinase 2 (JAK2). However, the actual downstream signaling events and how these alter myeloid homeostasis are poorly understood. We developed an assay to measure basal levels of phosphorylated signaling intermediates by flow cytometry during myeloid differentiation in MPN patients. Our study provides the first systematic demonstration of specific signaling events and their comparison with disease phenotype and JAK2 mutation status. We demonstrate increased basal signaling in MPN patients, which occurs in both early and later stages of myeloid differentiation. In addition, the pattern of signaling is not correlated with JAK2 mutation status and signaling intensity is poorly correlated with mutant JAK2 allele burden. In contrast, signaling differences are detected between different MPN disease phenotypes. Finally, we demonstrate that signaling can be inhibited by a JAK2-selective small molecule, but that this inhibition is not JAK2 V617F specific, because MPN patients with mutant JAK2, wild-type JAK2, and control patients were inhibited to a similar degree. Our data suggest that, in addition to JAK2 mutations, other factors contribute significantly to the MPN phenotype, results that are relevant to both the pathogenesis and therapy of MPN.
Assuntos
Janus Quinase 2/genética , Mutação , Transtornos Mieloproliferativos/genética , Transdução de Sinais , Idoso , Western Blotting , Linhagem Celular Tumoral , Eritropoetina/farmacologia , Feminino , Citometria de Fluxo , Genótipo , Células-Tronco Hematopoéticas/metabolismo , Humanos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/metabolismo , Masculino , Pessoa de Meia-Idade , Transtornos Mieloproliferativos/enzimologia , Transtornos Mieloproliferativos/metabolismo , Fenótipo , Fosfoproteínas/metabolismo , Fosforilação/efeitos dos fármacos , Policitemia Vera/enzimologia , Policitemia Vera/genética , Policitemia Vera/metabolismo , Mielofibrose Primária/enzimologia , Mielofibrose Primária/genética , Mielofibrose Primária/metabolismo , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Trombocitemia Essencial/enzimologia , Trombocitemia Essencial/genética , Trombocitemia Essencial/metabolismoRESUMO
The JAK2 V617F mutation is present in the majority of patients with a myeloproliferative neoplasm (MPN) and is sufficient to recapitulate an MPN in murine models. However, the consequences of JAK2 mutations for myeloid differentiation are poorly understood. After systematic analyses of a large cohort of JAK2-mutated MPN patients, we demonstrate in vivo that JAK2 mutations do not alter hematopoietic stem and progenitor cell com-partment size or in vitro behavior but generate expansion of later myeloid differentiation compartments, where homozygous expression of the mutation confers an added proliferative advantage at the single-cell level. In addition, we demonstrate that these findings may be partially explained by the expression pattern of JAK2, which markedly increases on myeloid differentiation. Our findings have potential clinical relevance, as they predict that JAK2 inhibitors may control myeloproliferation, but may have limited efficacy in eradicating the leukemic stem cells that sustain the human MPN.