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1.
Cell Chem Biol ; 27(3): 259-268.e5, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-32017919

RESUMO

Carbamoyl phosphate synthetase 1 (CPS1) catalyzes the first step in the ammonia-detoxifying urea cycle, converting ammonia to carbamoyl phosphate under physiologic conditions. In cancer, CPS1 overexpression supports pyrimidine synthesis to promote tumor growth in some cancer types, while in others CPS1 activity prevents the buildup of toxic levels of intratumoral ammonia to allow for sustained tumor growth. Targeted CPS1 inhibitors may, therefore, provide a therapeutic benefit for cancer patients with tumors overexpressing CPS1. Herein, we describe the discovery of small-molecule CPS1 inhibitors that bind to a previously unknown allosteric pocket to block ATP hydrolysis in the first step of carbamoyl phosphate synthesis. CPS1 inhibitors are active in cellular assays, blocking both urea synthesis and CPS1 support of the pyrimidine biosynthetic pathway, while having no activity against CPS2. These newly discovered CPS1 inhibitors are a first step toward providing researchers with valuable tools for probing CPS1 cancer biology.


Assuntos
Carbamoil-Fosfato Sintase (Amônia)/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Piperidinas/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Tiazóis/farmacologia , Trifosfato de Adenosina/antagonistas & inibidores , Trifosfato de Adenosina/metabolismo , Regulação Alostérica/efeitos dos fármacos , Carbamoil-Fosfato Sintase (Amônia)/genética , Carbamoil-Fosfato Sintase (Amônia)/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Humanos , Hidrólise/efeitos dos fármacos , Modelos Moleculares , Estrutura Molecular , Piperidinas/química , Bibliotecas de Moléculas Pequenas/química , Tiazóis/química
2.
Cell Rep ; 23(1): 282-296.e4, 2018 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-29617667

RESUMO

Hotspot mutations in splicing factor genes have been recently reported at high frequency in hematological malignancies, suggesting the importance of RNA splicing in cancer. We analyzed whole-exome sequencing data across 33 tumor types in The Cancer Genome Atlas (TCGA), and we identified 119 splicing factor genes with significant non-silent mutation patterns, including mutation over-representation, recurrent loss of function (tumor suppressor-like), or hotspot mutation profile (oncogene-like). Furthermore, RNA sequencing analysis revealed altered splicing events associated with selected splicing factor mutations. In addition, we were able to identify common gene pathway profiles associated with the presence of these mutations. Our analysis suggests that somatic alteration of genes involved in the RNA-splicing process is common in cancer and may represent an underappreciated hallmark of tumorigenesis.


Assuntos
Taxa de Mutação , Neoplasias/genética , Fatores de Processamento de RNA/genética , Linhagem Celular Tumoral , Genes Supressores de Tumor , Humanos , Mutação com Perda de Função , Neoplasias/classificação , Oncogenes , Splicing de RNA/genética
3.
Nat Med ; 24(4): 497-504, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29457796

RESUMO

Genomic analyses of cancer have identified recurrent point mutations in the RNA splicing factor-encoding genes SF3B1, U2AF1, and SRSF2 that confer an alteration of function. Cancer cells bearing these mutations are preferentially dependent on wild-type (WT) spliceosome function, but clinically relevant means to therapeutically target the spliceosome do not currently exist. Here we describe an orally available modulator of the SF3b complex, H3B-8800, which potently and preferentially kills spliceosome-mutant epithelial and hematologic tumor cells. These killing effects of H3B-8800 are due to its direct interaction with the SF3b complex, as evidenced by loss of H3B-8800 activity in drug-resistant cells bearing mutations in genes encoding SF3b components. Although H3B-8800 modulates WT and mutant spliceosome activity, the preferential killing of spliceosome-mutant cells is due to retention of short, GC-rich introns, which are enriched for genes encoding spliceosome components. These data demonstrate the therapeutic potential of splicing modulation in spliceosome-mutant cancers.


Assuntos
Neoplasias/tratamento farmacológico , Neoplasias/genética , Piperazinas/farmacologia , Piridinas/farmacologia , Splicing de RNA/genética , Bibliotecas de Moléculas Pequenas/uso terapêutico , Spliceossomos/genética , Administração Oral , Animais , Sequência de Bases , Humanos , Íntrons/genética , Células K562 , Leucemia/genética , Leucemia/patologia , Camundongos , Mutação , Neoplasias/patologia , Piperazinas/administração & dosagem , Piridinas/administração & dosagem , Splicing de RNA/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Nat Commun ; 8: 15522, 2017 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-28541300

RESUMO

Pladienolide, herboxidiene and spliceostatin have been identified as splicing modulators that target SF3B1 in the SF3b subcomplex. Here we report that PHF5A, another component of this subcomplex, is also targeted by these compounds. Mutations in PHF5A-Y36, SF3B1-K1071, SF3B1-R1074 and SF3B1-V1078 confer resistance to these modulators, suggesting a common interaction site. RNA-seq analysis reveals that PHF5A-Y36C has minimal effect on basal splicing but inhibits the global action of splicing modulators. Moreover, PHF5A-Y36C alters splicing modulator-induced intron-retention/exon-skipping profile, which correlates with the differential GC content between adjacent introns and exons. We determine the crystal structure of human PHF5A demonstrating that Y36 is located on a highly conserved surface. Analysis of the cryo-EM spliceosome Bact complex shows that the resistance mutations cluster in a pocket surrounding the branch point adenosine, suggesting a competitive mode of action. Collectively, we propose that PHF5A-SF3B1 forms a central node for binding to these splicing modulators.


Assuntos
Adenosina/química , Processamento Alternativo , Proteínas de Transporte/química , Fosfoproteínas/química , Fatores de Processamento de RNA/química , Proliferação de Células , Sobrevivência Celular , Microscopia Crioeletrônica , Cristalografia por Raios X , Compostos de Epóxi/química , Éxons , Álcoois Graxos/química , Células HCT116 , Humanos , Íntrons , Macrolídeos/química , Espectrometria de Massas , Mutagênese Sítio-Dirigida , Mutação , Proteína de Sequência 1 de Leucemia de Células Mieloides/química , Fosfoproteínas/metabolismo , Ligação Proteica , Conformação Proteica , Piranos/química , Interferência de RNA , Fatores de Processamento de RNA/metabolismo , Proteínas de Ligação a RNA , Proteínas Recombinantes/química , Análise de Sequência de RNA , Compostos de Espiro/química , Spliceossomos/metabolismo , Transativadores
5.
PLoS One ; 12(4): e0176045, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28426752

RESUMO

Gastric cancer, a leading worldwide cause of cancer mortality, shows high geographic and ethnic variation in incidence rates, which are highest in East Asia. The anatomic locations and clinical behavior also differ by geography, leading to the controversial idea that Eastern and Western forms of the disease are distinct. In view of these differences, we investigated whether gastric cancers from Eastern and Western patients show distinct genomic profiles. We used high-density profiling of somatic copy-number aberrations to analyze the largest collection to date of gastric adenocarcinomas and utilized genotyping data to rigorously annotate ethnic status. The size of this collection allowed us to accurately identify regions of significant copy-number alteration and separately to evaluate tumors arising in Eastern and Western patients. Among molecular subtypes classified by The Cancer Genome Atlas, the frequency of gastric cancers showing chromosomal instability was modestly higher in Western patients. After accounting for this difference, however, gastric cancers arising in Easterners and Westerners have highly similar somatic copy-number patterns. Only one genomic event, focal deletion of the phosphatase gene PTPRD, was significantly enriched in Western cases, though also detected in Eastern cases. Thus, despite the different risk factors and clinical features, gastric cancer appears to be a fundamentally similar disease in both populations and the divergent clinical outcomes cannot be ascribed to different underlying structural somatic genetic aberrations.


Assuntos
Adenocarcinoma/genética , Povo Asiático/genética , Variações do Número de Cópias de DNA , Neoplasias Gástricas/genética , População Branca/genética , Humanos
6.
Cell Rep ; 13(5): 1033-45, 2015 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-26565915

RESUMO

Recurrent mutations in the spliceosome are observed in several human cancers, but their functional and therapeutic significance remains elusive. SF3B1, the most frequently mutated component of the spliceosome in cancer, is involved in the recognition of the branch point sequence (BPS) during selection of the 3' splice site (ss) in RNA splicing. Here, we report that common and tumor-specific splicing aberrations are induced by SF3B1 mutations and establish aberrant 3' ss selection as the most frequent splicing defect. Strikingly, mutant SF3B1 utilizes a BPS that differs from that used by wild-type SF3B1 and requires the canonical 3' ss to enable aberrant splicing during the second step. Approximately 50% of the aberrantly spliced mRNAs are subjected to nonsense-mediated decay resulting in downregulation of gene and protein expression. These findings ascribe functional significance to the consequences of SF3B1 mutations in cancer.


Assuntos
Processamento Alternativo , Mutação , Neoplasias/genética , Fosfoproteínas/genética , Ribonucleoproteína Nuclear Pequena U2/genética , Alelos , Sequência de Aminoácidos , Sequência de Bases , Células HEK293 , Humanos , Dados de Sequência Molecular , Taxa de Mutação , Degradação do RNAm Mediada por Códon sem Sentido , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Fatores de Processamento de RNA , Ribonucleoproteína Nuclear Pequena U2/química , Ribonucleoproteína Nuclear Pequena U2/metabolismo
7.
Cell Mol Gastroenterol Hepatol ; 1(6): 598-609.e6, 2015 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-26516633

RESUMO

BACKGROUND & AIMS: Intestinal metaplasia (Barrett's esophagus, BE) is the principal risk factor for esophageal adenocarcinoma (EAC). Study of the basis for BE has centered on intestinal factors, but loss of esophageal identity likely also reflects absence of key squamous-cell factors. As few determinants of stratified epithelial cell-specific gene expression are characterized, it is important to identify the necessary transcription factors. METHODS: We tested regional expression of mRNAs for all putative DNA-binding proteins in the mouse digestive tract and verified esophagus-specific factors in human tissues and cell lines. Integration of diverse data defined a human squamous esophagus-specific transcriptome. We used chromatin immunoprecipitation (ChIP-seq) to locate transcription factor binding sites, computational approaches to profile transcripts in cancer datasets, and immunohistochemistry to reveal protein expression. RESULTS: The transcription factor SOX15 is restricted to esophageal and other murine and human stratified epithelia. SOX15 mRNA levels are attenuated in BE and its depletion in human esophageal cells reduced esophageal transcripts significantly and specifically. SOX15 binding is highly enriched near esophagus-expressed genes, indicating direct transcriptional control. SOX15 and hundreds of genes co-expressed in squamous cells are reactivated in up to 30% of EAC specimens. Genes normally confined to the esophagus or intestine appear in different cells within the same malignant glands. CONCLUSIONS: These data identify a novel transcriptional regulator of stratified epithelial cells and a subtype of EAC with bi-lineage gene expression. Broad activation of squamous-cell genes may shed light on whether EACs arise in the native stratified epithelium or in ectopic columnar cells.

8.
Nat Genet ; 47(9): 1047-55, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26192918

RESUMO

Barrett's esophagus is thought to progress to esophageal adenocarcinoma (EAC) through a stepwise progression with loss of CDKN2A followed by TP53 inactivation and aneuploidy. Here we present whole-exome sequencing from 25 pairs of EAC and Barrett's esophagus and from 5 patients whose Barrett's esophagus and tumor were extensively sampled. Our analysis showed that oncogene amplification typically occurred as a late event and that TP53 mutations often occurred early in Barrett's esophagus progression, including in non-dysplastic epithelium. Reanalysis of additional EAC exome data showed that the majority (62.5%) of EACs emerged following genome doubling and that tumors with genomic doubling had different patterns of genomic alterations, with more frequent oncogenic amplification and less frequent inactivation of tumor suppressors, including CDKN2A. These data suggest that many EACs emerge not through the gradual accumulation of tumor-suppressor alterations but rather through a more direct path whereby a TP53-mutant cell undergoes genome doubling, followed by the acquisition of oncogenic amplifications.


Assuntos
Adenocarcinoma/genética , Esôfago de Barrett/genética , Neoplasias Esofágicas/genética , Exoma , Classe I de Fosfatidilinositol 3-Quinases , Inibidor p16 de Quinase Dependente de Ciclina/genética , Análise Mutacional de DNA , Amplificação de Genes , Frequência do Gene , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Fosfatidilinositol 3-Quinases/genética , Mutação Puntual , Proteína Supressora de Tumor p53/genética
9.
Mol Cancer Res ; 13(4): 689-98, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25537453

RESUMO

UNLABELLED: The tumor suppressor gene MEN1 is frequently mutated in sporadic pancreatic neuroendocrine tumors (PanNET) and is responsible for the familial multiple endocrine neoplasia type 1 (MEN-1) cancer syndrome. Menin, the protein product of MEN1, associates with the histone methyltransferases (HMT) MLL1 (KMT2A) and MLL4 (KMT2B) to form menin-HMT complexes in both human and mouse model systems. To elucidate the role of methylation of histone H3 at lysine 4 (H3K4) mediated by menin-HMT complexes during PanNET formation, genome-wide histone H3 lysine 4 trimethylation (H3K4me3) signals were mapped in pancreatic islets using unbiased chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-seq). Integrative analysis of gene expression profiles and histone H3K4me3 levels identified a number of transcripts and target genes dependent on menin. In the absence of Men1, histone H3K27me3 levels are enriched, with a concomitant decrease in H3K4me3 within the promoters of these target genes. In particular, expression of the insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) gene is subject to dynamic epigenetic regulation by Men1-dependent histone modification in a time-dependent manner. Decreased expression of IGF2BP2 in Men1-deficient hyperplastic pancreatic islets is partially reversed by ablation of RBP2 (KDM5A), a histone H3K4-specific demethylase of the jumonji, AT-rich interactive domain 1 (JARID1) family. Taken together, these data demonstrate that loss of Men1 in pancreatic islet cells alters the epigenetic landscape of its target genes. IMPLICATIONS: Epigenetic profiling and gene expression analysis in Men1-deficient pancreatic islet cells reveals vital insight into the molecular events that occur during the progression of pancreatic islet tumorigenesis.


Assuntos
Epigênese Genética , Tumores Neuroendócrinos/genética , Tumores Neuroendócrinos/patologia , Proteínas Proto-Oncogênicas/genética , Proteínas de Ligação a RNA/genética , Adenoma de Células das Ilhotas Pancreáticas , Animais , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Metilação , Camundongos , Dados de Sequência Molecular
10.
Proc Natl Acad Sci U S A ; 111(50): 18055-60, 2014 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-25453105

RESUMO

DNA damage has been implicated in neurodegenerative disorders, including Alzheimer's disease and other tauopathies, but the consequences of genotoxic stress to postmitotic neurons are poorly understood. Here we demonstrate that p53, a key mediator of the DNA damage response, plays a neuroprotective role in a Drosophila model of tauopathy. Further, through a whole-genome ChIP-chip analysis, we identify genes controlled by p53 in postmitotic neurons. We genetically validate a specific pathway, synaptic function, in p53-mediated neuroprotection. We then demonstrate that the control of synaptic genes by p53 is conserved in mammals. Collectively, our results implicate synaptic function as a central target in p53-dependent protection from neurodegeneration.


Assuntos
Senescência Celular/fisiologia , Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica/fisiologia , Neurônios/fisiologia , Sinapses/genética , Sinapses/metabolismo , Tauopatias/prevenção & controle , Proteína Supressora de Tumor p53/metabolismo , Animais , Western Blotting , Imunoprecipitação da Cromatina , Dano ao DNA/fisiologia , Drosophila , Regulação da Expressão Gênica/genética , Ontologia Genética , Imuno-Histoquímica , Neurônios/citologia , Sinapses/patologia , Tauopatias/metabolismo , Proteínas tau/efeitos adversos
11.
J Clin Invest ; 124(12): 5145-58, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25401468

RESUMO

Patients with gastric and esophageal (GE) adenocarcinoma tumors in which the oncogene ERBB2 has been amplified are routinely treated with a combination of cytotoxic chemotherapy and the ERBB2-directed antibody trastuzumab; however, the addition of trastuzumab, even when tested in a selected biomarker-positive patient population, provides only modest survival gains. To investigate the potential reasons for the modest impact of ERBB2-directed therapies, we explored the hypothesis that secondary molecular features of ERBB2-amplified GE adenocarcinomas attenuate the impact of ERBB2 blockade. We analyzed genomic profiles of ERBB2-amplified GE adenocarcinomas and determined that the majority of ERBB2-amplified tumors harbor secondary oncogenic alterations that have the potential to be therapeutically targeted. These secondary events spanned genes involved in cell-cycle regulation as well as phosphatidylinositol-3 kinase and receptor tyrosine kinase signaling. Using ERBB2-amplified cell lines, we demonstrated that secondary oncogenic events could confer resistance to ERBB2-directed therapies. Moreover, this resistance could be overcome by targeting the secondary oncogene in conjunction with ERBB2-directed therapy. EGFR is commonly coamplified with ERBB2, and in the setting of ERBB2 amplification, higher EGFR expression appears to mark tumors with greater sensitivity to dual EGFR/ERBB2 kinase inhibitors. These data suggest that combination inhibitor strategies, guided by secondary events in ERBB2-amplified GE adenocarcinomas, should be evaluated in clinical trials.


Assuntos
Adenocarcinoma , Anticorpos Monoclonais Humanizados/administração & dosagem , Antineoplásicos/administração & dosagem , Biomarcadores Tumorais , Neoplasias Esofágicas , Amplificação de Genes , Receptor ErbB-2 , Neoplasias Gástricas , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/enzimologia , Adenocarcinoma/genética , Adenocarcinoma/patologia , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/genética , Receptores ErbB/metabolismo , Neoplasias Esofágicas/dietoterapia , Neoplasias Esofágicas/enzimologia , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Feminino , Amplificação de Genes/efeitos dos fármacos , Amplificação de Genes/genética , Humanos , Masculino , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/enzimologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Trastuzumab
12.
PLoS One ; 9(10): e109440, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25350844

RESUMO

ERBB2-directed therapy is now a routine component of therapy for ERBB2-amplified metastatic gastroesophageal adenocarcinomas. However, there is little knowledge of the mechanisms by which these tumors develop acquired resistance to ERBB2 inhibition. To investigate this question we sought to characterize cell line models of ERBB2-amplified gastroesophageal adenocarcinoma with acquired resistance to ERBB2 inhibition. We generated lapatinib-resistant (LR) subclones from an initially lapatinib-sensitive ERBB2-amplified esophageal adenocarcinoma cell line, OE19. We subsequently performed genomic characterization and functional analyses of resistant subclones with acquired lapatinib resistance. We identified a novel, acquired SrcE527K mutation in a subset of LR OE19 subclones. Cells with this mutant allele harbour increased Src phosphorylation. Genetic and pharmacologic inhibition of Src resensitized these subclones to lapatinib. Biochemically, Src mutations could activate both the phosphatidylinositol 3-kinase and mitogen activated protein kinase pathways in the lapatinib-treated LR OE19 cells. Ectopic expression of SrcE527K mutation also was sufficient to induce lapatinib resistance in drug-naïve cells. These results indicate that pathologic activation of Src is a potential mechanism of acquired resistance to ERBB2 inhibition in ERBB2-amplified gastroesophageal cancer. Although Src mutation has not been described in primary tumor samples, we propose that the Src hyperactivation should be investigated in the settings of acquired resistance to ERBB2 inhibition in esophageal and gastric adenocarcinoma.


Assuntos
Adenocarcinoma/genética , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Esofágicas/genética , Genes src , Mutação , Receptor ErbB-2/genética , Neoplasias Gástricas/genética , Adenocarcinoma/tratamento farmacológico , Antineoplásicos/farmacologia , Benzodioxóis/farmacologia , Linhagem Celular Tumoral , Análise Mutacional de DNA , Relação Dose-Resposta a Droga , Neoplasias Esofágicas/tratamento farmacológico , Amplificação de Genes , Expressão Gênica , Inativação Gênica , Humanos , Lapatinib , Inibidores de Proteínas Quinases/farmacologia , Quinazolinas/farmacologia , Interferência de RNA , Neoplasias Gástricas/tratamento farmacológico
13.
Mol Cancer ; 13: 135, 2014 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-24885062

RESUMO

BACKGROUND: KRAS mutations in codons 12 and 13 are established predictive biomarkers for anti-EGFR therapy in colorectal cancer. Previous studies suggest that KRAS codon 61 and 146 mutations may also predict resistance to anti-EGFR therapy in colorectal cancer. However, clinicopathological, molecular, and prognostic features of colorectal carcinoma with KRAS codon 61 or 146 mutation remain unclear. METHODS: We utilized a molecular pathological epidemiology database of 1267 colon and rectal cancers in the Nurse's Health Study and the Health Professionals Follow-up Study. We examined KRAS mutations in codons 12, 13, 61 and 146 (assessed by pyrosequencing), in relation to clinicopathological features, and tumor molecular markers, including BRAF and PIK3CA mutations, CpG island methylator phenotype (CIMP), LINE-1 methylation, and microsatellite instability (MSI). Survival analyses were performed in 1067 BRAF-wild-type cancers to avoid confounding by BRAF mutation. Cox proportional hazards models were used to compute mortality hazard ratio, adjusting for potential confounders, including disease stage, PIK3CA mutation, CIMP, LINE-1 hypomethylation, and MSI. RESULTS: KRAS codon 61 mutations were detected in 19 cases (1.5%), and codon 146 mutations in 40 cases (3.2%). Overall KRAS mutation prevalence in colorectal cancers was 40% (=505/1267). Of interest, compared to KRAS-wild-type, overall, KRAS-mutated cancers more frequently exhibited cecal location (24% vs. 12% in KRAS-wild-type; P < 0.0001), CIMP-low (49% vs. 32% in KRAS-wild-type; P < 0.0001), and PIK3CA mutations (24% vs. 11% in KRAS-wild-type; P < 0.0001). These trends were evident irrespective of mutated codon, though statistical power was limited for codon 61 mutants. Neither KRAS codon 61 nor codon 146 mutation was significantly associated with clinical outcome or prognosis in univariate or multivariate analysis [colorectal cancer-specific mortality hazard ratio (HR) = 0.81, 95% confidence interval (CI) = 0.29-2.26 for codon 61 mutation; colorectal cancer-specific mortality HR = 0.86, 95% CI = 0.42-1.78 for codon 146 mutation]. CONCLUSIONS: Tumors with KRAS mutations in codons 61 and 146 account for an appreciable proportion (approximately 5%) of colorectal cancers, and their clinicopathological and molecular features appear generally similar to KRAS codon 12 or 13 mutated cancers. To further assess clinical utility of KRAS codon 61 and 146 testing, large-scale trials are warranted.


Assuntos
Códon , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Mutação , Proteínas Proto-Oncogênicas/genética , Proteínas ras/genética , Idoso , Classe I de Fosfatidilinositol 3-Quinases , Estudos de Coortes , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/mortalidade , Ilhas de CpG , Metilação de DNA , Receptores ErbB/genética , Feminino , Expressão Gênica , Humanos , Elementos Nucleotídeos Longos e Dispersos/genética , Masculino , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/genética , Modelos de Riscos Proporcionais , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras) , Análise de Sobrevida
14.
J Clin Invest ; 124(4): 1636-45, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24590290

RESUMO

The transcription factor SOX2 is an essential regulator of pluripotent stem cells and promotes development and maintenance of squamous epithelia. We previously reported that SOX2 is an oncogene and subject to highly recurrent genomic amplification in squamous cell carcinomas (SCCs). Here, we have further characterized the function of SOX2 in SCC. Using ChIP-seq analysis, we compared SOX2-regulated gene profiles in multiple SCC cell lines to ES cell profiles and determined that SOX2 binds to distinct genomic loci in SCCs. In SCCs, SOX2 preferentially interacts with the transcription factor p63, as opposed to the transcription factor OCT4, which is the preferred SOX2 binding partner in ES cells. SOX2 and p63 exhibited overlapping genomic occupancy at a large number of loci in SCCs; however, coordinate binding of SOX2 and p63 was absent in ES cells. We further demonstrated that SOX2 and p63 jointly regulate gene expression, including the oncogene ETV4, which was essential for SOX2-amplified SCC cell survival. Together, these findings demonstrate that the action of SOX2 in SCC differs substantially from its role in pluripotency. The identification of the SCC-associated interaction between SOX2 and p63 will enable deeper characterization the downstream targets of this interaction in SCC and normal squamous epithelial physiology.


Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas E1A de Adenovirus/genética , Sítios de Ligação/genética , Linhagem Celular , Linhagem Celular Tumoral , Chromatiaceae/genética , Chromatiaceae/metabolismo , Células-Tronco Embrionárias/metabolismo , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Humanos , Fator 3 de Transcrição de Octâmero/metabolismo , Oncogenes , Células-Tronco Pluripotentes/metabolismo , Ligação Proteica , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-ets , Transcriptoma
15.
Genes Dev ; 27(8): 853-8, 2013 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-23630075

RESUMO

Dosage compensation has arisen in response to the evolution of distinct male (XY) and female (XX) karyotypes. In Drosophila melanogaster, the MSL complex increases male X transcription approximately twofold. X-specific targeting is thought to occur through sequence-dependent binding to chromatin entry sites (CESs), followed by spreading in cis to active genes. We tested this model by asking how newly evolving sex chromosome arms in Drosophila miranda acquired dosage compensation. We found evidence for the creation of new CESs, with the analogous sequence and spacing as in D. melanogaster, providing strong support for the spreading model in the establishment of dosage compensation.


Assuntos
Mecanismo Genético de Compensação de Dose , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Cromossomos Sexuais/genética , Animais , Evolução Molecular , Feminino , Cariótipo , Masculino , Dados de Sequência Molecular , Cromossomos Sexuais/metabolismo
16.
Nat Genet ; 45(5): 478-86, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23525077

RESUMO

The incidence of esophageal adenocarcinoma (EAC) has risen 600% over the last 30 years. With a 5-year survival rate of ~15%, the identification of new therapeutic targets for EAC is greatly important. We analyze the mutation spectra from whole-exome sequencing of 149 EAC tumor-normal pairs, 15 of which have also been subjected to whole-genome sequencing. We identify a mutational signature defined by a high prevalence of A>C transversions at AA dinucleotides. Statistical analysis of exome data identified 26 significantly mutated genes. Of these genes, five (TP53, CDKN2A, SMAD4, ARID1A and PIK3CA) have previously been implicated in EAC. The new significantly mutated genes include chromatin-modifying factors and candidate contributors SPG20, TLR4, ELMO1 and DOCK2. Functional analyses of EAC-derived mutations in ELMO1 identifies increased cellular invasion. Therefore, we suggest the potential activation of the RAC1 pathway as a contributor to EAC tumorigenesis.


Assuntos
Adenocarcinoma/genética , Biomarcadores Tumorais/genética , Neoplasias Esofágicas/genética , Exoma/genética , Genoma Humano/genética , Mutação/genética , Mapeamento Cromossômico , Rearranjo Gênico , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Invasividade Neoplásica
17.
Genes Dev ; 27(2): 197-210, 2013 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-23322301

RESUMO

The NKX2-1 transcription factor, a regulator of normal lung development, is the most significantly amplified gene in human lung adenocarcinoma. To study the transcriptional impact of NKX2-1 amplification, we generated an expression signature associated with NKX2-1 amplification in human lung adenocarcinoma and analyzed DNA-binding sites of NKX2-1 by genome-wide chromatin immunoprecipitation. Integration of these expression and cistromic analyses identified LMO3, itself encoding a transcription regulator, as a candidate direct transcriptional target of NKX2-1. Further cistromic and overexpression analyses indicated that NKX2-1 can cooperate with the forkhead box transcription factor FOXA1 to regulate LMO3 gene expression. RNAi analysis of NKX2-1-amplified cells compared with nonamplified cells demonstrated that LMO3 mediates cell survival downstream from NKX2-1. Our findings provide new insight into the transcriptional regulatory network of NKX2-1 and suggest that LMO3 is a transcriptional signal transducer in NKX2-1-amplified lung adenocarcinomas.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma/fisiopatologia , Regulação Neoplásica da Expressão Gênica , Proteínas com Domínio LIM/metabolismo , Neoplasias Pulmonares/fisiopatologia , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Adenocarcinoma de Pulmão , Linhagem Celular Tumoral , Cromatina/metabolismo , Perfilação da Expressão Gênica , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Humanos , Proteínas Nucleares/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Fator Nuclear 1 de Tireoide , Fatores de Transcrição/metabolismo
18.
Cancer Discov ; 3(3): 294-307, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23274911

RESUMO

N-RAS is one member of a family of oncoproteins that are commonly mutated in cancer. Activating mutations in NRAS occur in a subset of colorectal cancers, but little is known about how the mutant protein contributes to the onset and progression of the disease. Using genetically engineered mice, we find that mutant N-RAS strongly promotes tumorigenesis in the context of inflammation. The protumorigenic nature of mutant N-RAS is related to its antiapoptotic function, which is mediated by activation of a noncanonical mitogen-activated protein kinase pathway that signals through STAT3. As a result, inhibition of MAP-ERK kinase selectively induces apoptosis in autochthonous colonic tumors expressing mutant N-RAS. The translational significance of this finding is highlighted by our observation that NRAS mutation correlates with a less favorable clinical outcome for patients with colorectal cancer. These data show for the first time the important role that N-RAS plays in colorectal cancer.


Assuntos
Apoptose/genética , Colite/genética , Colite/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Proteínas ras/genética , Animais , Linhagem Celular Tumoral , Colite/induzido quimicamente , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/prevenção & controle , Progressão da Doença , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Genes ras , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Proteínas Proto-Oncogênicas c-raf/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Proteínas ras/metabolismo
19.
PLoS Genet ; 8(7): e1002830, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22844249

RESUMO

Sex chromosome dosage compensation in Drosophila provides a model for understanding how chromatin organization can modulate coordinate gene regulation. Male Drosophila increase the transcript levels of genes on the single male X approximately two-fold to equal the gene expression in females, which have two X-chromosomes. Dosage compensation is mediated by the Male-Specific Lethal (MSL) histone acetyltransferase complex. Five core components of the MSL complex were identified by genetic screens for genes that are specifically required for male viability and are dispensable for females. However, because dosage compensation must interface with the general transcriptional machinery, it is likely that identifying additional regulators that are not strictly male-specific will be key to understanding the process at a mechanistic level. Such regulators would not have been recovered from previous male-specific lethal screening strategies. Therefore, we have performed a cell culture-based, genome-wide RNAi screen to search for factors required for MSL targeting or function. Here we focus on the discovery of proteins that function to promote MSL complex recruitment to "chromatin entry sites," which are proposed to be the initial sites of MSL targeting. We find that components of the NSL (Non-specific lethal) complex, and a previously unstudied zinc-finger protein, facilitate MSL targeting and display a striking enrichment at MSL entry sites. Identification of these factors provides new insight into how MSL complex establishes the specialized hyperactive chromatin required for dosage compensation in Drosophila.


Assuntos
Proteínas de Ligação a DNA , Mecanismo Genético de Compensação de Dose , Proteínas de Drosophila , Drosophila melanogaster , Fatores de Transcrição , Cromossomo X/genética , Animais , Linhagem Celular , Cromatina/genética , Cromatina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Feminino , Regulação da Expressão Gênica , Masculino , Proteínas Nucleares/genética , Interferência de RNA , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Transporte Vesicular
20.
Nature ; 486(7403): 405-9, 2012 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-22722202

RESUMO

Breast carcinoma is the leading cause of cancer-related mortality in women worldwide, with an estimated 1.38 million new cases and 458,000 deaths in 2008 alone. This malignancy represents a heterogeneous group of tumours with characteristic molecular features, prognosis and responses to available therapy. Recurrent somatic alterations in breast cancer have been described, including mutations and copy number alterations, notably ERBB2 amplifications, the first successful therapy target defined by a genomic aberration. Previous DNA sequencing studies of breast cancer genomes have revealed additional candidate mutations and gene rearrangements. Here we report the whole-exome sequences of DNA from 103 human breast cancers of diverse subtypes from patients in Mexico and Vietnam compared to matched-normal DNA, together with whole-genome sequences of 22 breast cancer/normal pairs. Beyond confirming recurrent somatic mutations in PIK3CA, TP53, AKT1, GATA3 and MAP3K1, we discovered recurrent mutations in the CBFB transcription factor gene and deletions of its partner RUNX1. Furthermore, we have identified a recurrent MAGI3-AKT3 fusion enriched in triple-negative breast cancer lacking oestrogen and progesterone receptors and ERBB2 expression. The MAGI3-AKT3 fusion leads to constitutive activation of AKT kinase, which is abolished by treatment with an ATP-competitive AKT small-molecule inhibitor.


Assuntos
Neoplasias da Mama/classificação , Neoplasias da Mama/genética , Mutação/genética , Translocação Genética/genética , Algoritmos , Neoplasias da Mama/patologia , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade beta de Fator de Ligação ao Core/genética , Análise Mutacional de DNA , Exoma/genética , Feminino , Fusão Gênica/genética , Humanos , Proteínas de Membrana/genética , México , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Vietnã
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