RESUMO
BACKGROUND: Despite the fact that metastases are the leading cause of colorectal cancer deaths, little is known about the underlying molecular changes in these advanced disease stages. Few have studied the overall gene expression levels in metastases from colorectal carcinomas, and so far, none has investigated the peritoneal carcinomatoses by use of DNA microarrays. Therefore, the aim of the present study is to investigate and compare the gene expression patterns of primary carcinomas (n = 18), liver metastases (n = 4), and carcinomatoses (n = 4), relative to normal samples from the large bowel. RESULTS: Transcriptome profiles of colorectal cancer metastases independent of tumor site, as well as separate profiles associated with primary carcinomas, liver metastases, or peritoneal carcinomatoses, were assessed by use of Bayesian statistics. Gains of chromosome arm 5p are common in peritoneal carcinomatoses and several candidate genes (including PTGER4, SKP2, and ZNF622) mapping to this region were overexpressed in the tumors. Expression signatures stratified on TP53 mutation status were identified across all tumors regardless of stage. Furthermore, the gene expression levels for the in vivo tumors were compared with an in vitro model consisting of cell lines representing all three tumor stages established from one patient. CONCLUSION: By statistical analysis of gene expression data from primary colorectal carcinomas, liver metastases, and carcinomatoses, we are able to identify genetic patterns associated with the different stages of tumorigenesis.
Assuntos
Carcinoma/metabolismo , Neoplasias do Colo/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Peritoneais/metabolismo , Linhagem Celular Tumoral , Análise por Conglomerados , Perfilação da Expressão Gênica , Genes p53 , Predisposição Genética para Doença , Humanos , Neoplasias Hepáticas/secundário , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Peritoneais/secundário , Análise Serial de ProteínasRESUMO
To identify chromosomal aberrations that differentiate among the Dukes' stages of colorectal cancer (CRC) as well as those that are responsible for the progression into liver metastases, we performed a meta-analysis of data obtained from 31 comparative genomic hybridization (CGH) studies comprising a total of 859 CRCs. Individual copy number profiles for 373 primary tumors and 102 liver metastases were recorded and several statistical analyses, such as frequency, multivariate logistic regression, and trend tests, were performed. In addition, time of occurrence analysis was applied for the first time to copy number changes identified by CGH, and each genomic imbalance was thereby classified as an early or late event in colorectal tumorigenesis. By combining data from the different statistical tests, we present a novel genetic pathway for CRC progression that distinguishes the Dukes' stages and identifies early and late events in both primary carcinomas and liver metastases. Results from the combined analyses suggest that losses at 17p and 18 and gains of 8q, 13q, and 20 occur early in the establishment of primary CRCs, whereas loss of 4p is associated with the transition from Dukes' A to B-D. Deletion of 8p and gains of 7p and 17q are correlated with the transition from primary tumor to liver metastasis, whereas losses of 14q and gains of 1q, 11, 12p, and 19 are late events. We supplement these findings with a list of potential target genes for the specific alterations from a publicly available microarray expression dataset of CRC.
Assuntos
Aberrações Cromossômicas , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Dosagem de Genes , Análise por Conglomerados , Humanos , Neoplasias Hepáticas/secundário , Masculino , Análise Multivariada , Estadiamento de Neoplasias , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologiaRESUMO
Molecular markers that could stratify prostate cancer patients according to risk of disease progression would allow a significant improvement in the management of this clinically heterogeneous disease. In the present study, we analyzed the genetic profile of a consecutive series of 51 clinically confined prostate carcinomas and 27 benign prostatic hyperplasias using comparative genomic hybridization (CGH). We then added our findings to the existing literature data in order to perform a meta-analysis on a total of 294 prostate cancers with detailed CGH and clinicopathological information, using multivariate statistical methods that included principal component, hierarchical clustering, time of occurrence, and regression analyses. Whereas several genomic imbalances were shared by organ-confined, locally invasive, and metastatic prostate cancers, 6q and 10q losses and 7q and 8q gains were significantly more frequent in patients with extra-prostatic disease. Regression analysis indicated that 8q gain and 13q loss were the best predictors of locally invasive disease, whereas 8q gain and 6q and 10q losses were associated with metastatic disease. We propose a genetic pathway of prostate carcinogenesis with two distinct initiating events, namely, 8p and 13q losses. These primary imbalances are then preferentially followed by 8q gain and 6q, 16q, and 18q losses, which in turn are followed by a set of late events that make recurrent and metastatic prostate cancers genetically more complex. We conclude that significant differences exist in the genetic profile of organ-confined, locally invasive, and advanced prostate cancer and that genetic features may carry prognostic information independently of Gleason grade.
Assuntos
Neoplasias da Próstata/genética , Idoso , Humanos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Hibridização de Ácido Nucleico , Hiperplasia Prostática/genéticaRESUMO
TP53 mutations play an important role in the development of several cancers and are present in 20-40% of all breast carcinomas, contributing to increased genomic instability. In order to address the relationship of mutated TP53 to genomic complexity, the present study analysed 61 breast carcinomas for TP53 mutations and compared mutation status with the pattern of genomic imbalances as assessed by comparative genomic hybridization (CGH). Twenty per cent of the present series of breast carcinomas harboured TP53 mutations. An increasing number of abnormalities, as identified by CGH (higher genomic complexity), correlated significantly with mutant TP53. Among the chromosome arms most commonly altered (in more than 20% of the tumours), loss of 8p and gain of 8q were associated with TP53 mutations, whereas loss of 16q was associated with wild-type TP53. By performing supervised hierarchical clustering analysis of the CGH data, a cluster of chromosome imbalances was observed that showed differences between wild-type and mutant TP53 cases. Among these, loss of chromosome arm 5q revealed the strongest correlation with altered TP53. To investigate further the most commonly deleted region of 5q, gene expression patterns from two publicly available microarray data sets of breast carcinomas were evaluated statistically. The expression data sets identified potential target genes, including genes involved in ubiquitination and the known TP53 target CSPG2. The genomic complexity of breast carcinomas as assessed by CGH is associated with TP53 mutation status; breast cancers with TP53 mutations display more complex genomes than do those with wild-type TP53. The pattern of genomic imbalances associated with mutant TP53 is non-random, with loss of chromosome arm 5q being particularly closely associated with TP53 mutations.
Assuntos
Neoplasias da Mama/genética , Carcinoma Ductal de Mama/genética , Genes p53/genética , Mutação , Aberrações Cromossômicas , Cromossomos Humanos Par 5 , Análise por Conglomerados , Análise Mutacional de DNA/métodos , DNA de Neoplasias/genética , Regulação para Baixo , Feminino , Humanos , Hibridização de Ácido NucleicoRESUMO
An unselected series of 310 colorectal carcinomas, stratified according to microsatellite instability (MSI) and DNA ploidy, was examined for mutations and/or promoter hypermethylation of five components of the WNT signaling cascade [APC, CTNNB1 (encoding beta-catenin), AXIN2, TCF4, and WISP3] and three genes indirectly affecting this pathway [CDH1 (encoding E-cadherin), PTEN, and TP53]. APC and TP53 mutations were each present more often in microsatellite-stable (MSS) tumors than in those with MSI (P < .001 for both). We confirmed that the aneuploid MSS tumors frequently contained TP53 mutations (P < .001), whereas tumors with APC mutations and/or promoter hypermethylation revealed no associations to ploidy. Mutations in APC upstream of codons 1020 to 1169, encoding the beta-catenin binding site, were found in 15/144 mutated tumors and these patients seemed to have poor clinical outcome (P = .096). Frameshift mutations in AXIN2, PTEN, TCF4, and WISP3 were found in 20%, 17%, 46%, and 28% of the MSI tumors, respectively. More than half of the tumors with heterozygote mutations in AXIN2 were concurrently mutated in APC. The present study showed that more than 90% of all samples had alteration in one or more of the genes investigated, adding further evidence to the vital importance of activated WNT signaling in colorectal carcinogenesis.
Assuntos
Biomarcadores Tumorais/metabolismo , Instabilidade Cromossômica , Neoplasias Colorretais/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Repetições de Microssatélites , Mutação/genética , Transdução de Sinais , Proteína da Polipose Adenomatosa do Colo/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteína Axina , Proteínas de Sinalização Intercelular CCN , Caderinas/genética , Proteínas do Citoesqueleto/genética , Metilação de DNA , Proteínas de Ligação a DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , PTEN Fosfo-Hidrolase , Monoéster Fosfórico Hidrolases/genética , Ploidias , Fatores de Transcrição TCF , Transativadores/genética , Proteína 2 Semelhante ao Fator 7 de Transcrição , Fatores de Transcrição/genética , Proteína Supressora de Tumor p53/genética , Proteínas Supressoras de Tumor/genética , Proteínas Wnt , beta CateninaRESUMO
In cancer biology, cell lines are often used instead of primary tumors because of their widespread availability and close reflection of the in vivo state. Cancer is a genetic disease, commonly caused by small- and large-scale DNA rearrangements. Therefore, it is essential to know the genomic profiles of tumor cell lines to enable their correct and efficient use as experimental tools. Here, we present a comprehensive study of the genomic profiles of 20 colon cancer cell lines combining conventional karyotyping (G-banding), comparative genomic hybridization (CGH), and multicolor fluorescence in situ hybridization (M-FISH). Major differences between the microsatellite instability (MSI) and chromosome instability (CIN) cell lines are shown; the CIN cell lines exhibited complex karyotypes involving many chromosomes (mean: 8.5 copy number changes), whereas the MSI cell lines showed considerably fewer aberrations (mean: 2.6). The 3 techniques complement each other to provide a detailed picture of the numerical and structural chromosomal changes that characterize cancer cells. Therefore, 7 of the cell lines (Colo320, EB, Fri, IS2, IS3, SW480, and V9P) are here completely karyotyped for the first time and, among these, 5 have not previously been cytogenetically described. By hierarchical cluster analysis, we show that the cell lines are representative models for primary carcinomas at the genome level. We also present the genomic profiles of an experimental model for tumor progression, including 3 cell lines (IS1, IS2, and IS3) established from a primary carcinoma, its corresponding liver- and peritoneal metastasis from the same patient. To address the question of clonality, we compared the genome of 3 common cell lines grown in 2 laboratories. Finally, we compared all our results with previously published CGH data and karyotypes of colorectal cell lines. In conclusion, the large variation in genetic complexity of the cell lines highlights the importance of a comprehensive reference of genomic profiles for investigators engaged in functional studies using these research tools.
Assuntos
Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Genoma Humano , Linhagem Celular Tumoral , Instabilidade Cromossômica , Aberrações Cromossômicas , Bandeamento Cromossômico , Células Clonais , Análise por Conglomerados , Neoplasias do Colo/patologia , Perfilação da Expressão Gênica , Variação Genética , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Repetições de Microssatélites , Metástase Neoplásica , Hibridização de Ácido Nucleico , Neoplasias Peritoneais/genética , Neoplasias Peritoneais/secundário , PloidiasRESUMO
BACKGROUND: Colorectal cancer (CRC) is one of the most common causes of cancer-related deaths in the Western world, and despite the fact that metastases are usually the ultimate cause of deaths, the knowledge of the genetics of advanced stages of this disease is limited. In order to identify potential genetic abnormalities underlying the development of local and distant metastases in CRC patients, we have, by comparative genomic hybridization, compared the DNA copy number profiles of 10 primary carcinomas, 14 local recurrences, 7 peritoneal carcinomatoses, and 42 liver metastases from 61 CRC patients. RESULTS: The median number of aberrations among the primary carcinomas, local recurrences, carcinomatoses, and liver metastases was 10, 6, 13, and 14, respectively. Several genetic imbalances, such as gains of 7, 8q, 13q, and 20, and losses of 4q, 8p, 17p, and 18, were common in all groups. In contrast, gains of 5p and 12p were more common in the carcinomatoses than in other stages of the disease. With hierarchical cluster analysis, liver metastases could be divided into two main subgroups according to clusters of chromosome changes. CONCLUSIONS: Each stage of CRC progression is characterized by a particular genetic profile, and both carcinomatoses and liver metastases are more genetically complex than local recurrences and primary carcinomas. This is the first genome profiling of local recurrences and carcinomatoses, and gains of 5p and 12p seem to be particularly important for the spread of the CRC cells within the peritoneal cavity.
Assuntos
Carcinoma/patologia , Neoplasias Colorretais/patologia , Genoma Humano , Neoplasias Hepáticas/secundário , Carcinoma/genética , Aberrações Cromossômicas , Análise por Conglomerados , Neoplasias Colorretais/genética , Humanos , Neoplasias Hepáticas/genética , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Hibridização de Ácido Nucleico/métodosRESUMO
PURPOSE: To examine several genetic changes in primary colorectal carcinomas (CRCs) from patients with 10 years of follow-up and associate the findings with clinicopathologic variables. MATERIAL AND METHODS: DNA from 220 CRCs were analyzed for allelic imbalances at 12 loci on chromosome arms 1p, 14q, 17p, 18q, and 20q, and the microsatellite instability (MSI) status was determined. The clinical significance of the tumor protein 53 (TP53) mutations was re-evaluated. RESULTS: Patients with tumors containing 17p or 18q deletions had shorter survival than those without these alterations (P =.021, P =.008, respectively). This was also significant for the Dukes' B group (P =.025, P =.010, respectively). Furthermore, patients with tumors showing losses of both chromosome arms revealed an even poorer disease outcome than those with either 17p or 18q loss. Patients with low increase in 20q copy number in their tumors had longer survival compared with those without changes (P =.009) or those with a high increase of copy number (P =.037). This was also evident for the Dukes' C group (P =.018, P =.030, respectively). MSI was seemingly a beneficial marker for survival (P =.071). A significant association between mutations affecting the L3 zinc-binding domain of TP53 and survival was confirmed in this cohort after 10 years of follow-up, and also was found to apply for patients in the Dukes' B group. Several associations were found among genetic and pathologic data. CONCLUSION: The present study indicates that 17p, 18q, and 20q genotypes, and TP53 mutation status add information in the subclassification of Dukes' B and C patients and may have impact on the choice of treatment.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , DNA de Neoplasias/análise , Marcadores Genéticos/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 14/genética , Cromossomos Humanos Par 17/genética , Cromossomos Humanos Par 18/genética , Cromossomos Humanos Par 20/genética , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Feminino , Humanos , Perda de Heterozigosidade , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Mutação , Estadiamento de Neoplasias , Prognóstico , Estudos Prospectivos , Fatores de Risco , Taxa de Sobrevida , Proteína Supressora de Tumor p53/genéticaRESUMO
The majority of genetic studies of colorectal carcinogenesis have focused on changes found in primary tumors. Despite the fact that liver metastases are a leading cause of colorectal cancer deaths, the molecular genetic basis of the advanced disease stages remains poorly understood. We performed comparative genomic hybridization (CGH) on 17 liver metastases from colorectal carcinomas and compared the quantitative profile with the qualitative profile previously obtained with chromosome banding. An average of 12.6 aberrations per tumor was found by CGH. Chromosome 18 and chromosome arms 4q, 8p, and 17p were most frequently lost, whereas chromosomes 7 and 20 and chromosome arms 6p, 8q, and 13q were most frequently gained. We compared the chromosome banding and CGH data after converting the karyotypes into net copy number gains and losses. Ten tumors showed agreement between the findings of the two techniques, whereas five tumors did not (in two cases, no mitotic cells were obtained for banding analysis). All five discordant cases had a "simple" abnormal or normal karyotype, but revealed multiple changes by CGH. A likely explanation for this discrepancy is that in vitro growth before G-banding selected against the cancer cells. Interestingly, by comparing the CGH profiles of the "complex" vs. the "simple"/normal karyotype groups, deletion of 8p and gain of 16q were seen more frequently in the former group. The liver metastases had the same aberrations as seen in primary colorectal carcinomas, summarized in a literature survey. However, these aberrations were seen more frequently in liver metastases, which may be attributable to increased genetic instability.