RESUMO
In search for novel markers for breast cancer, we aimed to identify and validate novel serum protein profiles specific for breast cancer, and assess the influence of clinical (subjects age) and pre-analytical (sample storage duration) variables on the constructed classifiers. To this end, sera of breast cancer patients (n=152) and healthy controls (n=129), randomly divided into a training and test set, were analysed by surface-enhanced laser desorption/ionisation time-of-flight mass spectrometry (SELDI-TOF MS). In the training set, 14 peak clusters were found to differ significantly in expression between cases and controls. None of the peak clusters were influenced by subjects age and sample storage duration. Ten peak clusters were also found significantly discriminative in the test set. Peak clusters were structurally identified as C3a des-arginine anaphylatoxin, (tentative) inter-alpha-trypsin inhibitor heavy chain 4 fragments and a fibrinogen fragment. Logistic regression analyses on the training set yielded a classification model with a moderate performance on the test set, corresponding to those reported in previously performed validation studies. Most likely originating from the highly heterogeneous nature of breast cancer, selection of breast cancer subgroups for comparison with healthy controls is expected to improve results of future diagnostic SELDI-TOF MS studies.
Assuntos
Biomarcadores Tumorais/sangue , Neoplasias da Mama/diagnóstico , Proteínas de Neoplasias/sangue , Análise Serial de Proteínas/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Adulto , Fatores Etários , Idoso , Proteínas Sanguíneas , Neoplasias da Mama/sangue , Estudos de Casos e Controles , Análise por Conglomerados , Complemento C3a/análise , Feminino , Produtos de Degradação da Fibrina e do Fibrinogênio/análise , Glicoproteínas/sangue , Humanos , Modelos Logísticos , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Proteínas Secretadas Inibidoras de Proteinases/sangue , Reprodutibilidade dos Testes , Manejo de EspécimesRESUMO
AIM: Novel diagnostic breast cancer markers have been extensively searched for in the proteome, using, among others, surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS). Thus far, the majority of SELDI-TOF MS studies have investigated samples originating from biorepositories, which hampers biomarker discovery as they likely suffer from variable adherence to collection protocols. MATERIAL AND METHODS: We investigated breast cancer (n=75) and control (n=26) serum and tissue samples, collected prospectively by rigorous adherence to a strictly defined protocol. Sera were collected preoperatively and postoperatively, and serum and tissue samples were analyzed by SELDI-TOF MS using the IMAC30 Ni and Q10 pH 8 array. RESULTS: Three serum peaks were significantly associated with breast cancer, while in tissue, 27 discriminative peaks were detected. Several peak clusters gradually increased or decreased in intensity from healthy to benign to cancer, or with increasing cancer stage. The constructed classification trees had a tenfold cross-validated performance of 67% to 87%. Two tissue peaks were identified as N-terminal albumin fragments. These are likely to have been generated by (breast) cancer-specific proteolytic activity in the tumor microenvironment. CONCLUSIONS: These albumin fragment scan potentially provide insights into the pathophysiological mechanisms associated with, or underlying, breast cancer, and aid in improving breast cancer diagnosis.
Assuntos
Proteínas Sanguíneas/análise , Neoplasias da Mama/sangue , Neoplasias da Mama/diagnóstico , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Idoso , Neoplasias da Mama/patologia , Grupos Diagnósticos Relacionados , Feminino , Humanos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Ovarianas/sangue , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/patologiaRESUMO
INTRODUCTION: PolycombGroup (PcG) proteins maintain gene repression through histone modifications and have been implicated in stem cell regulation and cancer. EZH2 is part of Polycomb Repressive Complex 2 (PRC2) and trimethylates H3K27. This histone mark recruits the BMI1-containing PRC1 that silences the genes marked by PRC2. Based on their role in stem cells, EZH2 and BMI1 have been predicted to contribute to a poor outcome for cancer patients. METHODS: We have analysed the expression of EZH2 and BMI1 in a well-characterised dataset of 295 human breast cancer samples. RESULTS: Interestingly, although EZH2 overexpression correlates with a poor prognosis in breast cancer, BMI1 overexpression correlates with a good outcome. Although this may reflect transformation of different cell types, we also observed a functional difference. The PcG-target genes INK4A and ARF are not expressed in tumours with high BMI1, but they are expressed in tumours with EZH2 overexpression. ARF expression results in tumour protein P53 (TP53) activation, and we found a significantly higher proportion of TP53 mutations in tumours with high EZH2. This may explain why tumours with high EZH2 respond poorly to therapy, in contrast to tumours with high BMI1. CONCLUSIONS: Overall, our data highlight that whereas EZH2 and BMI1 may function in a 'linear' pathway in normal development, their overexpression has different functional consequences for breast tumourigenesis.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteínas de Ligação a DNA/metabolismo , Mutação/genética , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/genética , Fator 1 de Ribosilação do ADP/metabolismo , Neoplasias da Mama/patologia , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste , Feminino , Humanos , Técnicas Imunoenzimáticas , Complexo Repressor Polycomb 1 , Complexo Repressor Polycomb 2 , Reação em Cadeia da Polimerase , Prognóstico , Taxa de Sobrevida , Proteína Supressora de Tumor p53/metabolismoRESUMO
OBJECTIVE: To determine the value of gene-expression profiling as a predictor of the status of the regional nodes in patients with penile carcinoma. PATIENTS AND METHODS: Tumour samples of 56 patients with penile squamous cell carcinoma were analysed for the gene expression on 35 k oligoarrays; 32 were from patients with histopathologically confirmed lymph node metastases and 24 from those with no lymph node involvement. The 56 patients were divided into a training and validation set. For the training set, 15 patients with histopathologically confirmed nodal metastases and 15 without were selected. The validation set consisted of the remaining 26 patients, containing 17 node-positive and nine with no nodal metastases. RESULTS: A 44-probe classifier had the best performance within the training set; this classifier correctly assigned 29 of 30 specimens in the training set to the two outcome groups. In the validation set of 26 tumours, the classifier correctly assigned 14 of the 26 (54%) specimens to the two outcome groups. Of the 17 specimens with histologically confirmed nodal involvement, 12 were classified as node-positive and five as node-negative, resulting in a sensitivity of 71%. Of the nine specimens from node-negative patients, two were correctly classified as node-negative and seven as node positive, resulting in a specificity of 22%. CONCLUSIONS: In this series, gene expression profiling did not produce a useful classifier to predict nodal involvement in patients with penile carcinoma.
Assuntos
Carcinoma de Células Escamosas/secundário , Perfilação da Expressão Gênica/métodos , Linfonodos/patologia , Neoplasias Penianas , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia/métodos , Carcinoma de Células Escamosas/genética , Humanos , Metástase Linfática/genética , Masculino , Análise em Microsséries , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Penianas/genética , Neoplasias Penianas/patologiaRESUMO
Genome-wide mRNA expression measurements can identify molecular signatures of cancer and are anticipated to improve patient management. Such expression profiles are currently being critically evaluated based on an apparent instability in gene composition and the limited overlap between signatures from different studies. We have recently identified a primary tumor signature for detection of lymph node metastasis in head and neck squamous cell carcinomas. Before starting a large multicenter prospective validation, we have thoroughly evaluated the composition of this signature. A multiple training approach was used for validating the original set of predictive genes. Based on different combinations of training samples, multiple signatures were assessed for predictive accuracy and gene composition. The initial set of predictive genes is a subset of a larger group of 825 genes with predictive power. Many of the predictive genes are interchangeable because of a similar expression pattern across the tumor samples. The head and neck metastasis signature has a more stable gene composition than previous predictors. Exclusion of the strongest predictive genes could be compensated by raising the number of genes included in the signature. Multiple accurate predictive signatures can be designed using various subsets of predictive genes. The absence of genes with strong predictive power can be compensated by including more genes with lower predictive power. Lack of overlap between predictive signatures from different studies with the same goal may be explained by the fact that there are more predictive genes than required to design an accurate predictor.
Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Perfilação da Expressão Gênica , Humanos , Metástase Linfática , RNA Mensageiro/biossíntese , RNA Mensageiro/genéticaRESUMO
PURPOSE: The goal of the present study was to improve prediction of outcome after chemoradiation in advanced head and neck cancer using gene expression analysis. MATERIALS AND METHODS: We collected 92 biopsies from untreated head and neck cancer patients subsequently given cisplatin-based chemoradiation (RADPLAT) for advanced squamous cell carcinomas (HNSCC). After RNA extraction and labeling, we performed dye swap experiments using 35k oligo-microarrays. Supervised analyses were performed to create classifiers to predict locoregional control and disease recurrence. Published gene sets with prognostic value in other studies were also tested. RESULTS: Using supervised classification on the whole series, gene sets separating good and poor outcome could be found for all end points. However, when splitting tumors into training and validation groups, no robust classifiers could be found. Using Gene Set Enrichment analysis, several gene sets were found to be enriched in locoregional recurrences, although with high false-discovery rates. Previously published signatures for radiosensitivity, hypoxia, proliferation, "wound," stem cells, and chromosomal instability were not significantly correlated with outcome. However, a recently published signature for HNSCC defining a "high-risk" group was shown to be predictive for locoregional control in our dataset. CONCLUSION: Gene sets can be found with predictive potential for locoregional control after combined radiation and chemotherapy in HNSCC. How treatment-specific these gene sets are needs further study.
Assuntos
Carcinoma de Células Escamosas/genética , Perfilação da Expressão Gênica , Adulto , Idoso , Antineoplásicos/uso terapêutico , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/radioterapia , Cisplatino/uso terapêutico , Terapia Combinada/métodos , Feminino , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/radioterapia , Humanos , Neoplasias Laríngeas/tratamento farmacológico , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/radioterapia , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/tratamento farmacológico , Neoplasias Bucais/genética , Neoplasias Bucais/radioterapia , Recidiva Local de Neoplasia/genética , Neoplasias Faríngeas/tratamento farmacológico , Neoplasias Faríngeas/genética , Neoplasias Faríngeas/radioterapia , Radiossensibilizantes/uso terapêutico , Resultado do TratamentoRESUMO
UNLABELLED: A pathologic complete response to neoadjuvant chemotherapy (NAC) containing platinum is a strong prognostic determinant for patients with muscle-invasive bladder cancer (MIBC). Despite comprehensive molecular characterization of bladder cancer, associations of molecular alterations with treatment response are still largely unknown. We selected pathologic complete responders (ypT0N0; n=38) and nonresponders (higher than ypT2; n=33) from a cohort of high-grade MIBC patients treated with NAC. DNA was isolated from prechemotherapy tumor tissue and used for next-generation sequencing of 178 cancer-associated genes (discovery cohort) or targeted sequencing (validation cohort). We found that 9 of 38 complete responders had erb-b2 receptor tyrosine kinase 2 (ERBB2) missense mutations, whereas none of 33 nonresponders had ERBB2 mutations (p=0.003). ERBB2 missense mutations in complete responders were mostly confirmed activating mutations. ERCC2 missense mutations, recently found associated with response to NAC, were more common in complete responders; however, this association did not reach statistical significance in our cohort. We conclude that ERBB2 missense mutations characterize a subgroup of MIBC patients with an excellent response to NAC. PATIENT SUMMARY: In this report we looked for genetic alterations that can predict the response to neoadjuvant chemotherapy (NAC) in bladder cancer. We found that mutations in the gene ERBB2 are exclusively present in patients responding to NAC.
Assuntos
Biomarcadores Tumorais/genética , Mutação de Sentido Incorreto , Terapia Neoadjuvante , Receptor ErbB-2/genética , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/genética , Adulto , Idoso , Quimioterapia Adjuvante , Análise Mutacional de DNA , Intervalo Livre de Doença , Feminino , Predisposição Genética para Doença , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Seleção de Pacientes , Fenótipo , Medicina de Precisão , Valor Preditivo dos Testes , Estudos Prospectivos , Fatores de Tempo , Resultado do Tratamento , Neoplasias da Bexiga Urinária/patologiaRESUMO
BACKGROUND: Issues have been raised concerning the robustness and validity of alleged serum markers discovered by surface-enhanced laser desorption/ionisation time-of-flight mass spectrometry (SELDI-TOF MS). Pre-analytical variables have been shown to exert a profound effect on protein profiles, irrespective of true biological variation. However, little is known about the possible effects of sample storage duration on protein profiles. We, therefore, aimed to investigate the effects of extended storage duration on the serum protein profile. METHODS: Archival sera from 140 breast cancer patients, stored at -30 degrees C for 1-11 years, were profiled by SELDI-TOF MS using immobilised metal affinity capture (IMAC) arrays, a condition applied in the majority of breast cancer biomarker discovery studies. RESULTS: Fourteen peak clusters, structurally identified as C3a des-arginine anaphylatoxin and multiple fragments of albumin and fibrinogen, were found to be significantly associated with sample storage duration by five distinct patterns. These proteins have been described previously as potential cancer markers, rendering them specific to both disease and sample handling issues. CONCLUSIONS: To prevent experimental variation being interpreted erroneously as disease associated variation, assessment of potential confounding pre-analytical parameters is a pre-requisite in biomarker discovery and validation studies. In addition, with respect to the different (non-)linear patterns observed in the current study, simply performing linear corrections to account for sample storage duration will not necessarily suffice.
Assuntos
Biomarcadores Tumorais/sangue , Proteínas Sanguíneas/análise , Proteômica/métodos , Manejo de Espécimes/métodos , Idoso , Sequência de Aminoácidos , Neoplasias da Mama/diagnóstico , Feminino , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Estabilidade Proteica , Proteômica/normas , Manejo de Espécimes/normas , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Fatores de TempoRESUMO
BACKGROUND: Results of gene expression profiling studies from different institutes often lack consistency. This could be due to the use of different microarray platforms and protocols, or to intratumoral heterogeneity in mRNA expression. The aim of our study was to quantify intratumoral heterogeneity in head and neck cancer. METHODS: Forty-four fresh frozen biopsies were taken from 22 patients, 2 per tumor. RNA was extracted, tested for quality, amplified, labeled, and hybridized to a 35k oligoarray. RESULTS: Unsupervised clustering analyses using all genes, the most variable genes, or random gene sets showed that 80% to 90% of biopsy pairs clustered together. A within-pair-between-pair scatter ratio analysis showed that the similarity between matching pairs was significantly greater than that between random pairs (p <.00001). CONCLUSIONS: Two biopsies from the same tumor show far greater similarity in gene expression than biopsies from different tumors, supporting the use of 1 biopsy for expression profiling.