RESUMO
Histone modifications play crucial roles in transcriptional activation, and aberrant epigenetic changes are associated with oncogenesis. Lysine (K) acetyltransferases 5 (TIP60, also known as KAT5) is reportedly implicated in cancer development and maintenance, although its function in lung cancer remains controversial. Here we demonstrate that TIP60 knockdown in non-small cell lung cancer cell lines decreased tumor cell growth, migration, and invasion. Furthermore, analysis of a mouse lung cancer model with lung-specific conditional Tip60 knockout revealed suppressed tumor formation relative to controls, but no apparent effects on normal lung homeostasis. RNA-seq and ChIP-seq analyses of inducible TIP60 knockdown H1975 cells relative to controls revealed transglutaminase enzyme (TGM5) as downstream of TIP60. Investigation of a connectivity map database identified several candidate compounds that decrease TIP60 mRNA, one that suppressed tumor growth in cell culture and in vivo. In addition, TH1834, a TIP60 acetyltransferase inhibitor, showed comparable antitumor effects in cell culture and in vivo. Taken together, suppression of TIP60 activity shows tumor-specific efficacy against lung cancer, with no overt effect on normal tissues. Our work suggests that targeting TIP60 could be a promising approach to treating lung cancer.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Camundongos , Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Transformação Celular Neoplásica/genética , Histona Acetiltransferases/genética , Histona Acetiltransferases/metabolismo , Neoplasias Pulmonares/genética , HumanosRESUMO
The variable clinical course of follicular lymphoma (FL) is determined by the molecular heterogeneity of tumor cells and complex interactions within the tumor microenvironment (TME). IL-4 producing follicular helper T cells (TFH) are critical components of the FL TME. Binding of IL-4 to IL-4R on FL cells activates JAK/STAT signaling. We identified STAT6 mutations (STAT6MUT) in 13% of FL (N = 33/258), all clustered within the DNA binding domain. Gene expression data and immunohistochemistry showed upregulation of IL-4/STAT6 target genes in STAT6MUT FL, including CCL17, CCL22, and FCER2 (CD23). Functionally, STAT6MUT was gain-of-function by serial replating phenotype in pre-B CFU assays. Expression of STAT6MUT enhanced IL-4 induced FCER2/CD23, CCL17 and CCL22 expression and was associated with nuclear accumulation of pSTAT6. RNA sequencing identified PARP14 -a transcriptional switch and co-activator of STAT6- among the top differentially upregulated genes in IL-4 stimulated STAT6MUT lymphoma cells and in STAT6MUT primary FL cells. Quantitative chromatin immunoprecipitation (qChIP) demonstrated binding of STAT6MUT but not STAT6WT to the PARP14 promotor. Reporter assays showed increased IL-4 induced transactivation activity of STAT6MUT at the PARP14 promotor, suggesting a self-reinforcing regulatory circuit. Knock-down of PARP14 or PARP-inhibition abrogated the STAT6MUT gain-of-function phenotype. Thus, our results identify PARP14 as a novel therapeutic target in STAT6MUT FL.
Assuntos
Linfoma de Células B , Linfoma Folicular , Humanos , Imuno-Histoquímica , Interleucina-4 , Poli(ADP-Ribose) Polimerases , Fator de Transcrição STAT6 , Ativação Transcricional , Microambiente TumoralRESUMO
Relapsed follicular lymphoma (FL) can arise from common progenitor cells (CPCs). Conceptually, CPC-defining mutations are somatic alterations shared by the initial and relapsed tumours, mostly B-cell leukaemia/lymphoma 2 (BCL2)/immunoglobulin heavy locus (IGH) translocations and other recurrent gene mutations. Through complementary approaches for highly sensitive mutation detection, we do not find CPC-defining mutations in highly purified BCL2/IGH-negative haematopoietic progenitor cells in clinical remission samples from three patients with relapsed FL. Instead, we find cells harbouring the same BCL2/IGH translocation but lacking CREB binding protein (CREBBP), lysine methyltransferase 2D (KMT2D) and other recurrent gene mutations. Thus, (i) the BCL2/IGH translocation can precede CPC-defining mutations in human FL, and (ii) BCL2/IGH-translocated cells can persist in clinical remission.
Assuntos
Linfoma de Células B , Linfoma Folicular , Células-Tronco Hematopoéticas/metabolismo , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Linfoma de Células B/genética , Linfoma Folicular/patologia , Mutação , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Translocação GenéticaRESUMO
Hematopoietic stem cells (HSCs) have the potential to replenish the blood system for the lifetime of the organism. Their 2 defining properties, self-renewal and differentiation, are tightly regulated by the epigenetic machineries. Using conditional gene-knockout models, we demonstrated a critical requirement of lysine acetyltransferase 5 (Kat5, also known as Tip60) for murine HSC maintenance in both the embryonic and adult stages, which depends on its acetyltransferase activity. Genome-wide chromatin and transcriptome profiling in murine hematopoietic stem and progenitor cells revealed that Tip60 colocalizes with c-Myc and that Tip60 deletion suppress the expression of Myc target genes, which are associated with critical biological processes for HSC maintenance, cell cycling, and DNA repair. Notably, acetylated H2A.Z (acH2A.Z) was enriched at the Tip60-bound active chromatin, and Tip60 deletion induced a robust reduction in the acH2A.Z/H2A.Z ratio. These results uncover a critical epigenetic regulatory layer for HSC maintenance, at least in part through Tip60-dependent H2A.Z acetylation to activate Myc target genes.
Assuntos
Autorrenovação Celular/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Lisina Acetiltransferase 5/genética , Transativadores/genética , Animais , Biomarcadores , Ciclo Celular , Diferenciação Celular/genética , Dano ao DNA , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Histonas/metabolismo , Lisina Acetiltransferase 5/metabolismo , Camundongos , Transporte Proteico , Transativadores/metabolismoRESUMO
Tumor cells orchestrate their microenvironment. Here, we provide biochemical, structural, functional, and clinical evidence that Cathepsin S (CTSS) alterations induce a tumor-promoting immune microenvironment in follicular lymphoma (FL). We found CTSS mutations at Y132 in 6% of FL (19/305). Another 13% (37/286) had CTSS amplification, which was associated with higher CTSS expression. CTSS Y132 mutations lead to accelerated autocatalytic conversion from an enzymatically inactive profrom to active CTSS and increased substrate cleavage, including CD74, which regulates major histocompatibility complex class II (MHC class II)-restricted antigen presentation. Lymphoma cells with hyperactive CTSS more efficiently activated antigen-specific CD4+ T cells in vitro. Tumors with hyperactive CTSS showed increased CD4+ T cell infiltration and proinflammatory cytokine perturbation in a mouse model and in human FLs. In mice, this CTSS-induced immune microenvironment promoted tumor growth. Clinically, patients with CTSS-hyperactive FL had better treatment outcomes with standard immunochemotherapies, indicating that these immunosuppressive regimens target both the lymphoma cells and the tumor-promoting immune microenvironment.
Assuntos
Apresentação de Antígeno/imunologia , Catepsinas/metabolismo , Linfoma Folicular/metabolismo , Microambiente Tumoral/imunologia , Animais , Antígenos de Diferenciação de Linfócitos B/metabolismo , Citocinas/metabolismo , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Terapia de Imunossupressão , Linfoma Folicular/patologia , CamundongosRESUMO
Acute Myeloid Leukemia (AML) with MLL gene rearrangements demonstrate unique gene expression profiles driven by MLL-fusion proteins. Here, we identify the circadian clock transcription factor SHARP1 as a novel oncogenic target in MLL-AF6 AML, which has the worst prognosis among all subtypes of MLL-rearranged AMLs. SHARP1 is expressed solely in MLL-AF6 AML, and its expression is regulated directly by MLL-AF6/DOT1L. Suppression of SHARP1 induces robust apoptosis of human MLL-AF6 AML cells. Genetic deletion in mice delays the development of leukemia and attenuated leukemia-initiating potential, while sparing normal hematopoiesis. Mechanistically, SHARP1 binds to transcriptionally active chromatin across the genome and activates genes critical for cell survival as well as key oncogenic targets of MLL-AF6. Our findings demonstrate the unique oncogenic role for SHARP1 in MLL-AF6 AML.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Leucemia Mieloide Aguda/metabolismo , Fatores de Transcrição/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Carcinogênese , Transformação Celular Neoplásica , Feminino , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Masculino , Camundongos , Camundongos Knockout , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Fatores de Transcrição/genéticaAssuntos
Linfoma Folicular/genética , Linfoma Folicular/terapia , Mutação , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais Murinos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Ciclofosfamida/uso terapêutico , Doxorrubicina/uso terapêutico , Feminino , Humanos , Linfoma Folicular/mortalidade , Masculino , Pessoa de Meia-Idade , Prednisona/uso terapêutico , Rituximab , Análise de Sobrevida , Resultado do Tratamento , Vincristina/uso terapêutico , Adulto JovemRESUMO
The Bruton tyrosine kinase (BTK) inhibitor ibrutinib induces responses in 70% of patients with relapsed and refractory mantle cell lymphoma (MCL). Intrinsic resistance can occur through activation of the nonclassical NF-κB pathway and acquired resistance may involve the BTK C481S mutation. Outcomes after ibrutinib failure are dismal, indicating an unmet medical need. We reasoned that newer heat shock protein 90 (HSP90) inhibitors could overcome ibrutinib resistance by targeting multiple oncogenic pathways in MCL. HSP90 inhibition induced the complete degradation of both BTK and IκB kinase α in MCL lines and CD40-dependent B cells, with downstream loss of MAPK and nonclassical NF-κB signaling. A proteome-wide analysis in MCL lines and an MCL patient-derived xenograft identified a restricted set of targets from HSP90 inhibition that were enriched for factors involved in B-cell receptor and JAK/STAT signaling, the nonclassical NF-κB pathway, cell-cycle regulation, and DNA repair. Finally, multiple HSP90 inhibitors potently killed MCL lines in vitro, and the clinical agent AUY922 was active in vivo against both patient-derived and cell-line xenografts. Together, these findings define the HSP90-dependent proteome in MCL. Considering the disappointing clinical activity of HSP90 inhibitors in other contexts, trials in patients with MCL will be essential for defining the efficacy of and mechanisms of resistance after ibrutinib failure.
Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Isoxazóis/farmacologia , Linfoma de Célula do Manto/tratamento farmacológico , Pirazóis/farmacologia , Pirimidinas/farmacologia , Resorcinóis/farmacologia , Adenina/análogos & derivados , Tirosina Quinase da Agamaglobulinemia , Substituição de Aminoácidos , Animais , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Linfoma de Célula do Manto/genética , Linfoma de Célula do Manto/metabolismo , Linfoma de Célula do Manto/patologia , Camundongos , Mutação de Sentido Incorreto , Piperidinas , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
CCAAT/enhancer-binding protein alpha (C/EBPα) is an essential transcription factor for myeloid lineage commitment. Here we demonstrate that acetylation of C/EBPα at lysine residues K298 and K302, mediated at least in part by general control non-derepressible 5 (GCN5), impairs C/EBPα DNA-binding ability and modulates C/EBPα transcriptional activity. Acetylated C/EBPα is enriched in human myeloid leukaemia cell lines and acute myeloid leukaemia (AML) samples, and downregulated upon granulocyte-colony stimulating factor (G-CSF)- mediated granulocytic differentiation of 32Dcl3 cells. C/EBPα mutants that mimic acetylation failed to induce granulocytic differentiation in C/EBPα-dependent assays, in both cell lines and in primary hematopoietic cells. Our data uncover GCN5 as a negative regulator of C/EBPα and demonstrate the importance of C/EBPα acetylation in myeloid differentiation.
Assuntos
Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Regulação Neoplásica da Expressão Gênica , Granulócitos/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide/genética , Mielopoese/genética , Fatores de Transcrição de p300-CBP/genética , Acetilação , Diferenciação Celular/genética , Linhagem Celular Tumoral , Cromatografia Líquida , Ensaio de Desvio de Mobilidade Eletroforética , Fator Estimulador de Colônias de Granulócitos , Granulócitos/citologia , Células HEK293 , Humanos , Immunoblotting , Imunoprecipitação , Leucemia Mieloide/metabolismo , Leucemia Mieloide Aguda/metabolismo , Espectrometria de Massas , Fatores de Transcrição de p300-CBP/metabolismoRESUMO
Leukemic cells disrupt normal patterns of blood cell formation, but little is understood about the mechanism. We investigated whether leukemic cells alter functions of normal hematopoietic stem and progenitor cells. Exposure to chronic myelogenous leukemia (CML) caused normal mouse hematopoietic progenitor cells to divide more readily, altered their differentiation, and reduced their reconstitution and self-renewal potential. Interestingly, the normal bystander cells acquired gene expression patterns resembling their malignant counterparts. Therefore, much of the leukemia signature is mediated by extrinsic factors. Indeed, IL-6 was responsible for most of these changes. Compatible results were obtained when human CML were cultured with normal human hematopoietic progenitor cells. Furthermore, neutralization of IL-6 prevented these changes and treated the disease.
Assuntos
Citocinas/antagonistas & inibidores , Células-Tronco Hematopoéticas/citologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Animais , Técnicas de Cocultura , Citocinas/metabolismo , Humanos , Interleucina-6/farmacologia , Interleucina-6/uso terapêutico , Camundongos , Células Tumorais CultivadasRESUMO
Dendritic cells (DCs) are master regulators of the immune system, but molecular regulation of early DC differentiation has been poorly understood. Here, we report that the transcription factor C/EBPα coordinates the development of progenitor cells required for production of multiple categories of DCs. C/EBPα was needed for differentiation from stem/progenitor cells to common DC progenitors (CDPs), but not for transition of CDP to mature DCs. C/EBPα deletion in mature DCs did not affect their numbers or function, suggesting that this transcription factor is not needed for maintenance of DCs in lymphoid tissues. ChIP-seq and microarrays were used to identify candidate genes regulated by C/EBPα and required for DC formation. Genes previously shown to be critical for DC formation were bound by C/EBPα, and their expression was decreased in the earliest hematopoietic compartments in the absence of C/EBPα. These data indicate that C/EBPα is important for the earliest stages of steady-state DC differentiation.
Assuntos
Proteína alfa Estimuladora de Ligação a CCAAT/fisiologia , Diferenciação Celular/genética , Células Dendríticas/fisiologia , Células-Tronco/fisiologia , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Diferenciação Celular/imunologia , Células Cultivadas , Análise por Conglomerados , Células Dendríticas/metabolismo , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Tecido Linfoide/citologia , Tecido Linfoide/imunologia , Camundongos , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos , Células-Tronco/metabolismoRESUMO
The combination of sulfadoxine-pyrimethamine (SP) is used as a second line of therapy for the treatment of uncomplicated chloroquine-resistant Plasmodium falciparum malaria. Resistance to SP arises due to certain point mutations in the genes for the dihydrofolate reductase (DHFR) and dihydropteroate synthetase (DHPS) enzymes of the parasite. We have analyzed these mutations in 312 field isolates of P. falciparum collected from different parts of India to assess the effects of drug pressure. The rate of mutation in the gene for DHFR was found to be higher than that in the gene for DHPS, although the latter had mutations in more alleles. There was a temporal rise in the number of isolates with double dhfr mutations and single dhps mutations, resulting in an increased total number of mutations in the loci for DHFR and DHPS combined over a 5-year period. During these 5 years, the number of isolates with drug-sensitive genotypes decreased and the number of isolates with drug-resistant genotypes (double DHFR mutations and a single DHPS mutation) increased significantly. The number of isolates with the triple mutations in each of the genes for the two enzymes (for a total of six mutations), however, remained very low, coinciding with the very low rate of SP treatment failure in the country. There was a regional bias in the mutation rate, as isolates from the northeastern region (the state of Assam) showed higher rates of mutation and more complex genotypes than isolates from the other regions. It was concluded that even though SP is prescribed as a second line of treatment in India, the mutations associated with SP resistance continue to be progressively increasing.