RESUMEN
The organization of the genome in the nucleus and the interactions of genes with their regulatory elements are key features of transcriptional control and their disruption can cause disease. Here we report a genome-wide method, genome architecture mapping (GAM), for measuring chromatin contacts and other features of three-dimensional chromatin topology on the basis of sequencing DNA from a large collection of thin nuclear sections. We apply GAM to mouse embryonic stem cells and identify enrichment for specific interactions between active genes and enhancers across very large genomic distances using a mathematical model termed SLICE (statistical inference of co-segregation). GAM also reveals an abundance of three-way contacts across the genome, especially between regions that are highly transcribed or contain super-enhancers, providing a level of insight into genome architecture that, owing to the technical limitations of current technologies, has previously remained unattainable. Furthermore, GAM highlights a role for gene-expression-specific contacts in organizing the genome in mammalian nuclei.
Asunto(s)
Cromatina/genética , Cromatina/metabolismo , Mapeo Cromosómico , Elementos de Facilitación Genéticos/genética , Genoma/genética , Animales , Cromatina/química , Epigénesis Genética , Masculino , Ratones , Modelos Genéticos , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/metabolismo , Análisis de Secuencia de ADN , Transcripción Genética/genéticaRESUMEN
BACKGROUND: NRG1 gene fusions may be clinically actionable, since cancers carrying the fusion transcripts can be sensitive to tyrosine kinase inhibitors. The NRG1 gene encodes ligands for the HER2(ERBB2)-ERBB3 heterodimeric receptor tyrosine kinase, and the gene fusions are thought to lead to autocrine stimulation of the receptor. The NRG1 fusion expressed in the breast cancer cell line MDA-MB-175 serves as a model example of such fusions, showing the proposed autocrine loop and exceptional drug sensitivity. However, its structure has not been properly characterised, its oncogenic activity has not been fully explained, and there is limited data on such fusions in breast cancer. METHODS: We analysed genomic rearrangements and transcripts of NRG1 in MDA-MB-175 and a panel of 571 breast cancers. RESULTS: We found that the MDA-MB-175 fusion-originally reported as a DOC4(TENM4)-NRG1 fusion, lacking the cytoplasmic tail of NRG1-is in reality a double fusion, PPP6R3-TENM4-NRG1, producing multiple transcripts, some of which include the cytoplasmic tail. We hypothesise that many NRG1 fusions may be oncogenic not for lacking the cytoplasmic domain but because they do not encode NRG1's nuclear-localised form. The fusion in MDA-MB-175 is the result of a very complex genomic rearrangement, which we partially characterised, that creates additional expressed gene fusions, RSF1-TENM4, TPCN2-RSF1, and MRPL48-GAB2. We searched for NRG1 rearrangements in 571 breast cancers subjected to genome sequencing and transcriptome sequencing and found four cases (0.7%) with fusions, WRN-NRG1, FAM91A1-NRG1, ARHGEF39-NRG1, and ZNF704-NRG1, all splicing into NRG1 at the same exon as in MDA-MB-175. However, the WRN-NRG1 and ARHGEF39-NRG1 fusions were out of frame. We identified rearrangements of NRG1 in many more (8% of) cases that seemed more likely to inactivate than to create activating fusions, or whose outcome could not be predicted because they were complex, or both. This is not surprising because NRG1 can be pro-apoptotic and is inactivated in some breast cancers. CONCLUSIONS: Our results highlight the complexity of rearrangements of NRG1 in breast cancers and confirm that some do not activate but inactivate. Careful interpretation of NRG1 rearrangements will therefore be necessary for appropriate patient management.
Asunto(s)
Biomarcadores de Tumor , Neoplasias de la Mama/genética , Neurregulina-1/genética , Proteínas de Fusión Oncogénica/genética , Empalme Alternativo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica , Reordenamiento Génico , Sitios Genéticos , Humanos , Neurregulina-1/química , Neurregulina-1/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Transducción de Señal , Translocación GenéticaRESUMEN
In cancer, many genes are mutated by genome rearrangement, but our understanding of the functional consequences of this remains rudimentary. Here we report the F-box protein encoded by FBXL17 is disrupted in the region of the gene that encodes its substrate-binding leucine rich repeat (LRR) domain. Truncating Fbxl17 LRRs impaired its association with the other SCF holoenzyme subunits Skp1, Cul1 and Rbx1, and decreased ubiquitination activity. Loss of the LRRs also differentially affected Fbxl17 binding to its targets. Thus, genomic rearrangements in FBXL17 are likely to disrupt SCFFbxl17-regulated networks in cancer cells. To investigate the functional effect of these rearrangements, we performed a yeast two-hybrid screen to identify Fbxl17-interacting proteins. Among the 37 binding partners Uap1, an enzyme involved in O-GlcNAcylation of proteins was identified most frequently. We demonstrate that Fbxl17 binds to UAP1 directly and inhibits its phosphorylation, which we propose regulates UAP1 activity. Knockdown of Fbxl17 expression elevated O-GlcNAcylation in breast cancer cells, arguing for a functional role for Fbxl17 in this metabolic pathway.
Asunto(s)
Neoplasias de la Mama/genética , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Acetilglucosamina/metabolismo , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Roturas del ADN , Femenino , Células HEK293 , Humanos , Fosforilación , Procesamiento Proteico-Postraduccional , Eliminación de Secuencia , Ubiquitina-Proteína Ligasas/metabolismo , UbiquitinaciónRESUMEN
The expansion of repressive epigenetic marks has been implicated in heterochromatin formation during embryonic development, but the general applicability of this mechanism is unclear. Here we show that nuclear rearrangement of repressive histone marks H3K9me3 and H3K27me3 into nonoverlapping structural layers characterizes senescence-associated heterochromatic foci (SAHF) formation in human fibroblasts. However, the global landscape of these repressive marks remains unchanged upon SAHF formation, suggesting that in somatic cells, heterochromatin can be formed through the spatial repositioning of pre-existing repressively marked histones. This model is reinforced by the correlation of presenescent replication timing with both the subsequent layered structure of SAHFs and the global landscape of the repressive marks, allowing us to integrate microscopic and genomic information. Furthermore, modulation of SAHF structure does not affect the occupancy of these repressive marks, nor vice versa. These experiments reveal that high-order heterochromatin formation and epigenetic remodeling of the genome can be discrete events.
Asunto(s)
Cromatina/química , Heterocromatina/química , Histonas/metabolismo , Bromodesoxiuridina/farmacología , Senescencia Celular , Cromosomas/ultraestructura , Epigénesis Genética , Fibroblastos/citología , Regulación del Desarrollo de la Expresión Génica , Silenciador del Gen , Genoma , Estudio de Asociación del Genoma Completo , Histonas/química , Humanos , Citometría de Barrido por Láser/métodos , Microscopía Fluorescente/métodosRESUMEN
BACKGROUND: Mobile elements are active in the human genome, both in the germline and cancers, where they can mutate driver genes. RESULTS: While analysing whole genome paired-end sequencing of oesophageal adenocarcinomas to find genomic rearrangements, we identified three ways in which new mobile element insertions appear in the data, resembling translocation or insertion junctions: inserts where unique sequence has been transduced by an L1 (Long interspersed element 1) mobile element; novel inserts that are confidently, but often incorrectly, mapped by alignment software to L1s or polyA tracts in the reference sequence; and a combination of these two ways, where different sequences within one insert are mapped to different loci. We identified nine unique sequences that were transduced by neighbouring L1s, both L1s in the reference genome and L1s not present in the reference. Many of the resulting inserts were small fragments that include little or no recognisable mobile element sequence. We found 6 loci in the reference genome to which sequence reads from inserts were frequently mapped, probably erroneously, by alignment software: these were either L1 sequence or particularly long polyA runs. Inserts identified from such apparent rearrangement junctions averaged 16 inserts/tumour, range 0-153 insertions in 43 tumours. However, many inserts would not be detected by mapping the sequences to the reference genome, because they do not include sufficient mappable sequence. To estimate total somatic inserts we searched for polyA sequences that were not present in the matched normal or other normals from the same tumour batch, and were not associated with known polymorphisms. Samples of these candidate inserts were verified by sequencing across them or manual inspection of surrounding reads: at least 85 % were somatic and resembled L1-mediated events, most including L1Hs sequence. Approximately 100 such inserts were detected per tumour on average (range zero to approximately 700). CONCLUSIONS: Somatic mobile elements insertions are abundant in these tumours, with over 75 % of cases having a number of novel inserts detected. The inserts create a variety of problems for the interpretation of paired-end sequencing data.
Asunto(s)
Adenocarcinoma/genética , Elementos Transponibles de ADN/genética , Neoplasias Esofágicas/genética , Mutagénesis Insercional/genética , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Femenino , Reordenamiento Génico , Genoma Humano/genética , Humanos , Elementos de Nucleótido Esparcido Largo/genética , Masculino , Persona de Mediana Edad , Polimorfismo Genético/genética , Análisis de Secuencia de ADN/métodosRESUMEN
Cancer genomes are complex, carrying thousands of somatic mutations including base substitutions, insertions and deletions, rearrangements, and copy number changes that have been acquired over decades. Recently, technologies have been introduced that allow generation of high-resolution, comprehensive catalogs of somatic alterations in cancer genomes. However, analyses of these data sets generally do not indicate the order in which mutations have occurred, or the resulting karyotype. Here, we introduce a mathematical framework that begins to address this problem. By using samples with accurate data sets, we can reconstruct relatively complex temporal sequences of rearrangements and provide an assembly of genomic segments into digital karyotypes. For cancer genes mutated in rearranged regions, this information can provide a chronological examination of the selective events that have taken place.
Asunto(s)
Genoma Humano , Modelos Genéticos , Neoplasias/genética , Filogenia , Translocación Genética , Biología Computacional/métodos , Variaciones en el Número de Copia de ADN , Evolución Molecular , Humanos , MutaciónRESUMEN
The majority of human breast cancers exhibit luminal epithelial differentiation. However, most aggressive behavior, including invasion and purported cancer stem cell activity, are considered characteristics of basal-like cells. We asked the following questions: Must luminal-like breast cancer cells become basal-like to initiate tumors or to invade? Could luminally differentiated cells within a basally initiated hierarchy also be tumorigenic? To answer these questions, we used rare and mutually exclusive lineage markers to isolate subsets of luminal-like and basal-like cells from human breast tumors. We enriched for populations with or without prominent basal-like traits from individual tumors or single cell cloning from cell lines and recovered cells with a luminal-like phenotype. Tumor cells with basal-like traits mimicked phenotypic and functional behavior associated with stem cells assessed by gene expression, mammosphere formation and lineage markers. Luminal-like cells without basal-like traits, surprisingly, were fully capable of initiating invasive tumors in NOD SCID gamma (NSG) mice. In fact, these phenotypically pure luminal-like cells generated larger and more invasive tumors than their basal-like counterparts. The tumorigenicity and invasive potential of the luminal-like cancer cells relied strongly on the expression of the gene GCNT1, which encodes a key glycosyltransferase controlling O-glycan branching. These findings demonstrate that basal-like cells, as defined currently, are not a requirement for breast tumor aggressiveness, and that within a single tumor there are multiple "stem-like" cells with tumorigenic potential casting some doubt on the hypothesis of hierarchical or differentiative loss of tumorigenicity.
Asunto(s)
Neoplasias de la Mama/patología , Diferenciación Celular , Neoplasias Mamarias Experimentales/patología , Células Madre Neoplásicas/patología , Adapaleno , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Subunidad gamma Común de Receptores de Interleucina/deficiencia , Subunidad gamma Común de Receptores de Interleucina/genética , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Ratones Noqueados , Ratones Desnudos , Mucina-1/metabolismo , N-Acetilglucosaminiltransferasas/genética , N-Acetilglucosaminiltransferasas/metabolismo , Naftalenos/metabolismo , Invasividad Neoplásica , Células Madre Neoplásicas/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Reciprocal chromosome translocations are often not exactly reciprocal. Most familiar are deletions at the breakpoints, up to megabases in extent. We describe here the opposite phenomenon-duplication of tens or hundreds of kilobases at the breakpoint junction, so that the same sequence is present on both products of a translocation. When the products of the translocation are mapped on the genome, they overlap. We report several of these "overlapping-breakpoint" duplications in breast cancer cell lines HCC1187, HCC1806, and DU4475. These lines also had deletions and essentially balanced translocations. In HCC1187 and HCC1806, we identified five cases of duplication ranging between 46 kb and 200 kb, with the partner chromosome showing deletions between 29 bp and 31 Mb. DU4475 had a duplication of at least 200 kb. Breakpoints were mapped using array painting, i.e., hybridization of chromosomes isolated by flow cytometry to custom oligonucleotide microarrays. Duplications were verified by fluorescent in situ hybridization (FISH), PCR on isolated chromosomes, and cloning of breakpoints. We propose that these duplications are the counterpart of deletions and that they are produced at a replication bubble, comprising two replication forks with the duplicated sequence in between. Both copies of the duplicated sequence would go to one daughter cell, on different products of the translocation, while the other daughter cell would show deletion. These duplications may have been overlooked because they may be missed by FISH and array-CGH and may be interpreted as insertions by paired-end sequencing. Such duplications may therefore be quite frequent.
Asunto(s)
Rotura Cromosómica , Replicación del ADN/genética , Eliminación de Gen , Translocación Genética , Secuencia de Bases , Línea Celular Tumoral , Cromosomas Humanos/genética , Humanos , Modelos Genéticos , Datos de Secuencia Molecular , Alineación de SecuenciaRESUMEN
High-grade serous ovarian carcinoma (HGSOC) is characterized by genomic instability, ubiquitous TP53 loss, and frequent development of platinum resistance. Loss of homologous recombination (HR) is a mutator phenotype present in 50% of HGSOCs and confers hypersensitivity to platinum treatment. We asked which other mutator phenotypes are present in HGSOC and how they drive the emergence of platinum resistance. We performed whole-genome paired-end sequencing on a model of two HGSOC cases, each consisting of a pair of cell lines established before and after clinical resistance emerged, to describe their structural variants (SVs) and to infer their ancestral genomes as the SVs present within each pair. The first case (PEO1/PEO4), with HR deficiency, acquired translocations and small deletions through its early evolution, but a revertant BRCA2 mutation restoring HR function in the resistant lineage re-stabilized its genome and reduced platinum sensitivity. The second case (PEO14/PEO23) had 216 tandem duplications and did not show evidence of HR or mismatch repair deficiency. By comparing the cell lines to the tissues from which they originated, we showed that the tandem duplicator mutator phenotype arose early in progression in vivo and persisted throughout evolution in vivo and in vitro, which may have enabled continual evolution. From the analysis of SNP array data from 454 HGSOC cases in The Cancer Genome Atlas series, we estimate that 12.8% of cases show patterns of aberrations similar to the tandem duplicator, and this phenotype is mutually exclusive with BRCA1/2 carrier mutations.
Asunto(s)
Antineoplásicos/uso terapéutico , Resistencia a Antineoplásicos/genética , Duplicación de Gen , Mutación , Neoplasias Quísticas, Mucinosas y Serosas/genética , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Compuestos de Platino/uso terapéutico , Secuencias Repetidas en Tándem , Proteína BRCA1/genética , Proteína BRCA2/genética , Línea Celular Tumoral , Evolución Molecular , Femenino , Eliminación de Gen , Predisposición Genética a la Enfermedad , Recombinación Homóloga , Humanos , Clasificación del Tumor , Neoplasias Quísticas, Mucinosas y Serosas/patología , Análisis de Secuencia por Matrices de Oligonucleótidos , Neoplasias Ováricas/patología , Fenotipo , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADN , Translocación GenéticaRESUMEN
Rearrangements of the genome can be detected by microarray methods and massively parallel sequencing, which identify copy-number alterations and breakpoint junctions, but these techniques are poorly suited to reconstructing the long-range organization of rearranged chromosomes, for example, to distinguish between translocations and insertions. The single-DNA-molecule technique HAPPY mapping is a method for mapping normal genomes that should be able to analyse genome rearrangements, i.e. deviations from a known genome map, to assemble rearrangements into a long-range map. We applied HAPPY mapping to cancer cell lines to show that it could identify rearrangement of genomic segments, even in the presence of normal copies of the genome. We could distinguish a simple interstitial deletion from a copy-number loss at an inversion junction, and detect a known translocation. We could determine whether junctions detected by sequencing were on the same chromosome, by measuring their linkage to each other, and hence map the rearrangement. Finally, we mapped an uncharacterized reciprocal translocation in the T-47D breast cancer cell line to about 2 kb and hence cloned the translocation junctions. We conclude that HAPPY mapping is a versatile tool for determining the structure of rearrangements in the human genome.
Asunto(s)
Aberraciones Cromosómicas , Mapeo Cromosómico/métodos , Neoplasias/genética , Línea Celular Tumoral , Deleción Cromosómica , Variaciones en el Número de Copia de ADN , Ligamiento Genético , Genoma Humano , Humanos , Translocación GenéticaRESUMEN
Analysis of circulating tumor DNA (ctDNA) to monitor cancer dynamics and detect minimal residual disease has been an area of increasing interest. Multiple methods have been proposed but few studies have compared the performance of different approaches. Here, we compare detection of ctDNA in serial plasma samples from patients with breast cancer using different tumor-informed and tumor-naïve assays designed to detect structural variants (SVs), single nucleotide variants (SNVs), and/or somatic copy-number aberrations, by multiplex PCR, hybrid capture, and different depths of whole-genome sequencing. Our results demonstrate that the ctDNA dynamics and allele fractions (AFs) were highly concordant when analyzing the same patient samples using different assays. Tumor-informed assays showed the highest sensitivity for detection of ctDNA at low concentrations. Hybrid capture sequencing targeting between 1,347 and 7,491 tumor-identified mutations at high depth was the most sensitive assay, detecting ctDNA down to an AF of 0.00024% (2.4 parts per million, ppm). Multiplex PCR targeting 21-47 tumor-identified SVs per patient detected ctDNA down to 0.00047% AF (4.7 ppm) and has potential as a clinical assay.
Asunto(s)
Neoplasias de la Mama , ADN Tumoral Circulante , Humanos , Femenino , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/genética , Biomarcadores de Tumor/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , ADN Tumoral Circulante/genética , MutaciónRESUMEN
BACKGROUND: It has recently emerged that common epithelial cancers such as breast cancers have fusion genes like those in leukaemias. In a representative breast cancer cell line, ZR-75-30, we searched for fusion genes, by analysing genome rearrangements. RESULTS: We first analysed rearrangements of the ZR-75-30 genome, to around 10kb resolution, by molecular cytogenetic approaches, combining array painting and array CGH. We then compared this map with genomic junctions determined by paired-end sequencing. Most of the breakpoints found by array painting and array CGH were identified in the paired end sequencing-55% of the unamplified breakpoints and 97% of the amplified breakpoints (as these are represented by more sequence reads). From this analysis we identified 9 expressed fusion genes: APPBP2-PHF20L1, BCAS3-HOXB9, COL14A1-SKAP1, TAOK1-PCGF2, TIAM1-NRIP1, TIMM23-ARHGAP32, TRPS1-LASP1, USP32-CCDC49 and ZMYM4-OPRD1. We also determined the genomic junctions of a further three expressed fusion genes that had been described by others, BCAS3-ERBB2, DDX5-DEPDC6/DEPTOR and PLEC1-ENPP2. Of this total of 12 expressed fusion genes, 9 were in the coamplification. Due to the sensitivity of the technologies used, we estimate these 12 fusion genes to be around two-thirds of the true total. Many of the fusions seem likely to be driver mutations. For example, PHF20L1, BCAS3, TAOK1, PCGF2, and TRPS1 are fused in other breast cancers. HOXB9 and PHF20L1 are members of gene families that are fused in other neoplasms. Several of the other genes are relevant to cancer-in addition to ERBB2, SKAP1 is an adaptor for Src, DEPTOR regulates the mTOR pathway and NRIP1 is an estrogen-receptor coregulator. CONCLUSIONS: This is the first structural analysis of a breast cancer genome that combines classical molecular cytogenetic approaches with sequencing. Paired-end sequencing was able to detect almost all breakpoints, where there was adequate read depth. It supports the view that gene breakage and gene fusion are important classes of mutation in breast cancer, with a typical breast cancer expressing many fusion genes.
Asunto(s)
Neoplasias de la Mama/genética , Genoma Humano/genética , Proteínas de Fusión Oncogénica/genética , Secuencia de Bases , Línea Celular Tumoral , Mapeo Cromosómico , Clonación Molecular , Hibridación Genómica Comparativa/métodos , Femenino , Humanos , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADNRESUMEN
For many years, it was assumed that gene fusions were a type of mutation confined largely to leukemias and sarcomas. However, fusion genes are now known to be important in several epithelial cancers and a number have been described in breast cancers. In the December 2011 issue of Nature Medicine, Robinson and colleagues reported many more gene fusions -including the first recurrent fusion, SEC16A-NOTCH1 - in breast cancers. Several genes, including members of the MAST (microtubule-associated serine threonine) kinase and Notch gene families, are fused more than once. This finding supports an emerging story that most breast cancers express a number of fusion genes.
Asunto(s)
Neoplasias de la Mama/genética , Fusión Génica , Femenino , Proteínas de Fusión bcr-abl/genética , Humanos , Masculino , Proteínas Asociadas a Microtúbulos/genética , Neoplasias de la Próstata/genética , Proteínas Serina-Treonina Quinasas/genética , Receptor Notch1/genética , Proteínas de Transporte Vesicular/genéticaRESUMEN
OBJECTIVE: To determine hepatitis C (HCV) treatment effectiveness and predictors of response in the "real-world" Australian clinic setting. DESIGN, SETTING AND PARTICIPANTS: Patients with chronic HCV, who were HCV-treatment-naive at enrolment, and were then treated with standard therapy (pegylated interferon-α plus ribavirin), were recruited prospectively through a national network of 24 HCV clinics between April 2008 and December 2009. Patients were interviewed and a medical record review was conducted at enrolment and at routine follow-up clinic visits. MAIN OUTCOME MEASURES: Proportion of patients achieving a sustained virological response (SVR), predictors of SVR, and impact of treatment on biochemical markers of liver disease (alanine aminotransferase levels and aspartate aminotransferase-to-platelet ratio index scores). RESULTS: The SVR by intention to treat was 60% (327/550). Infection with HCV genotype 2 or 3 (compared with genotype 1) was an independent predictor of SVR (odds ratio [OR], 2.45; 95% CI, 1.70-3.52), while HIV coinfection (OR, 0.28; 95% CI, 0.10-0.82), cirrhosis (OR, 0.38; 95% CI, 0.18-0.81), and increased body mass index for ≥ 30 kg/m(2) v ≤ 25 kg/m(2) (OR, 0.58; 95% CI, 0.35-0.96) were independently associated with lower SVR. There was a significant improvement in biochemical markers of liver disease following SVR (P< 0.001). CONCLUSIONS: Our findings are similar to those seen in clinical trials, despite the inclusion of patients with a broad range of comorbid conditions such as injecting drug and alcohol use and psychiatric illness. They suggest that, with appropriate patient and infrastructure support, expansion of treatment services to the broader HCV-infected community is warranted.
Asunto(s)
Antivirales/uso terapéutico , Hepatitis C Crónica/tratamiento farmacológico , Interferón-alfa/uso terapéutico , Polietilenglicoles/uso terapéutico , Ribavirina/uso terapéutico , Adulto , Australia , Índice de Masa Corporal , Coinfección , Quimioterapia Combinada , Femenino , Estudios de Seguimiento , Infecciones por VIH/complicaciones , Hepacivirus/genética , Hepatitis C Crónica/complicaciones , Hepatitis C Crónica/virología , Humanos , Interferón alfa-2 , Modelos Logísticos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Estudios Prospectivos , Proteínas Recombinantes/uso terapéutico , Resultado del TratamientoRESUMEN
Oesophageal adenocarcinoma (OAC) provides an ideal case study to characterize large-scale rearrangements. Using whole genome short-read sequencing of 383 cases, for which 214 had matched whole transcriptomes, we observed structural variations (SV) with a predominance of deletions, tandem duplications and inter-chromosome junctions that could be identified as LINE-1 mobile element (ME) insertions. Complex clusters of rearrangements resembling breakage-fusion-bridge cycles or extrachromosomal circular DNA accounted for 22% of complex SVs affecting known oncogenes. Counting SV events affecting known driver genes substantially increased the recurrence rates of these drivers. After excluding fragile sites, we identified 51 candidate new drivers in genomic regions disrupted by SVs, including ETV5, KAT6B and CLTC. RUNX1 was the most recurrently altered gene (24%), with many deletions inactivating the RUNT domain but preserved the reading frame, suggesting an altered protein product. These findings underscore the importance of identification of SV events in OAC with implications for targeted therapies.
Asunto(s)
Adenocarcinoma , Neoplasias Esofágicas , Adenocarcinoma/genética , Neoplasias Esofágicas/genética , Genoma Humano , Histona Acetiltransferasas/genética , Humanos , Secuenciación Completa del GenomaRESUMEN
It has been known for 25 years that fusion genes play a central role in leukaemias and sarcomas but they have been neglected in the common carcinomas, largely because of technical limitations of cytogenetics. In the last few years it has emerged that gene fusions, caused by chromosome translocations, inversions, deletions, etc., are important in the common epithelial cancers, such as prostate and lung carcinoma. Most prostate cancers, for example, have an androgen-regulated fusion of one of the ETS transcription factor gene family. Early results of genome-wide searches for gene fusions in breast and other epithelial cancers suggest that most individual tumours will have several fused genes. Fusion genes are exceptionally powerful mutations. In their simplest form they can turn on expression by promoter insertion but they can also, for example, force dimerization of a protein or change its subcellular location. They are correspondingly important clinically, in classification and management and as targets for therapy. This review surveys what we know of fusion genes in the carcinomas, summarizes the technical advances that now make it possible to search systematically for such genes, and concludes by putting fusion genes into the current picture of mutation in cancers.
Asunto(s)
Fusión Génica/genética , Genes Relacionados con las Neoplasias/genética , Neoplasias Glandulares y Epiteliales/genética , Translocación Genética/genética , Estudio de Asociación del Genoma Completo , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodosRESUMEN
Each year, the Canadian health education system graduates thousands of health professionals who have the best intentions of practising to their full scope of knowledge and skills to help improve the patient care experience in this country. However, a recent research study points to the fact that members of the healthcare team may be practising in a challenging environment in which only a limited number of their skills are actually being used. The Michener Institute for Applied Health Sciences believes that these issues, which include increased role specialization, limited scopes of practice, rapidly advancing technology, and challenges transitioning from hospital to community settings, have broader health education and health system implications that need to be addressed by policy makers, educators, and healthcare system leaders in order to enhance health professional education as well as patient care.
Asunto(s)
Técnicos Medios en Salud , Especialización , Canadá , HumanosRESUMEN
Homozygous deletions in cancer cells have been thought to harbor tumor suppressor genes. We show that the 25 and 50 kb homozygous deletions in WWOX in the colon cancer cell line HCT116 result from a complex set of heterozygous deletions, some of which overlap to give homozygous loss. One of the heterozygous deletions has removed exons 6-8 of one allele of WWOX, and there is also a third copy of the distal region of WWOX in an unbalanced translocation. The exon 6-8 deletion results in allele-specific expression of a deleted transcript, which seems likely to be the main biological consequence of the deletions, since similar transcripts are found in other tumors. We show that such a complex set of deletions could form in a single exchange event between two homologous chromosomes, so that the selective advantage of such rearrangements need not be within the homozygous deletion. We conclude that homozygous deletions can be markers of complex rearrangements that have targets outside the homozygous deletion itself and that the target of deletions in the FRA16D region is indeed WWOX, the common outcome being the removal of particular WWOX exons. This article contains supplementary material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat.
Asunto(s)
Neoplasias del Colon/genética , Exones , Eliminación de Gen , Homocigoto , Mutación , Oxidorreductasas/genética , Proteínas Supresoras de Tumor/genética , Secuencia de Bases , Línea Celular Tumoral , Cromosomas Humanos Par 16 , Cartilla de ADN , Humanos , Reacción en Cadena de la Polimerasa , ARN Mensajero/genética , Translocación Genética , Oxidorreductasa que Contiene Dominios WWRESUMEN
BACKGROUND: Rearrangement of the short arm of chromosome 8 (8p) is very common in epithelial cancers such as breast cancer. Usually there is an unbalanced translocation breakpoint in 8p12 (29.7 Mb - 38.5 Mb) with loss of distal 8p, sometimes with proximal amplification of 8p11-12. Rearrangements in 8p11-12 have been investigated using high-resolution array CGH, but the first 30 Mb of 8p are less well characterised, although this region contains several proposed tumour suppressor genes. METHODS: We analysed the whole of 8p by array CGH at tiling-path BAC resolution in 32 breast and six pancreatic cancer cell lines. Regions of recurrent rearrangement distal to 8p12 were further characterised, using regional fosmid arrays. FISH, and quantitative RT-PCR on over 60 breast tumours validated the existence of similar events in primary material. RESULTS: We confirmed that 8p is usually lost up to at least 30 Mb, but a few lines showed focal loss or copy number steps within this region. Three regions showed rearrangements common to at least two cases: two regions of recurrent loss and one region of amplification. Loss within 8p23.3 (0 Mb - 2.2 Mb) was found in six cell lines. Of the genes always affected, ARHGEF10 showed a point mutation of the remaining normal copies in the DU4475 cell line. Deletions within 12.7 Mb - 19.1 Mb in 8p22, in two cases, affected TUSC3. A novel amplicon was found within 8p21.3 (19.1 Mb - 23.4 Mb) in two lines and one of 98 tumours. CONCLUSION: The pattern of rearrangements seen on 8p may be a consequence of the high density of potential targets on this chromosome arm, and ARHGEF10 may be a new candidate tumour suppressor gene.
Asunto(s)
Neoplasias de la Mama/genética , Aberraciones Cromosómicas , Cromosomas Humanos Par 8/genética , Análisis Citogenético , Hibridación Genética , Distribución de Chi-Cuadrado , Deleción Cromosómica , Pintura Cromosómica , Femenino , Genes Supresores de Tumor , Humanos , Hibridación Fluorescente in Situ , Pérdida de Heterocigocidad , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Polimorfismo Genético , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Human chromosome 8p is prone to recurrent rearrangements with inv dup del(8p) being most common. Each of these recurrent rearrangements is associated with different clinical manifestations. Some of these recurrent rearrangements at 8p are mediated by an 8p submicroscopic paracentric inversion between the olfactory gene clusters present in one of the parents. However, recent reports have shown that some of the rearrangements are unique and complex and are mediated by other repetitive elements within 8p. Here, we report on a unique and complex 8p rearrangement with seizures as the major presenting feature in the patient. Extensive fluorescence in situ hybridization and microarray analyses with tiling path 8p array showed that the rearrangement is unique in that the 8p duplication is a direct tandem duplication and, unlike the more common inv dup del(8p), is not derived from parental submicroscopic inversion. Also unlike the inv dup del(8p), the phenotype in our case is milder with no central nervous system malformations or cardiac defects.