RESUMO
BACKGROUND: The aim of this study was to investigate the potential of cell-free DNA (cfDNA) as a disease biomarker in oesophageal squamous cell carcinoma (ESCC) that can be used for treatment response evaluation and early detection of tumour recurrence. METHODS: Matched tumour tissue, pre- and post-surgery plasma and WBCs obtained from 17 ESCC patients were sequenced using a panel of 483 cancer-related genes. RESULTS: Somatic mutations were detected in 14 of 17 tumour tissues. Putative harmful mutations were observed in genes involved in well-known cancer-related pathways, including PI3K-Akt/mTOR signalling, Proteoglycans in cancer, FoxO signalling, Jak-STAT signalling, Chemokine signalling and Focal adhesion. Forty-six somatic mutations were found in pre-surgery cfDNA in 8 of 12 patients, with mutant allele frequencies (MAF) ranging from 0.24 to 4.91%. Three of the 8 patients with detectable circulating tumour DNA (ctDNA) had stage IIA disease, whereas the others had stage IIB-IIIB disease. Post-surgery cfDNA somatic mutations were detected in only 2 of 14 patients, with mutant allele frequencies of 0.28 and 0.36%. All other somatic mutations were undetectable in post-surgery cfDNA, even in samples collected within 3-4 h after surgery. CONCLUSION: Our study shows that somatic mutations can be detected in pre-surgery cfDNA in stage IIA to IIIB patients, and at a lower frequency in post-surgery cfDNA. This indicates that cfDNA could potentially be used to monitor disease load, even in low disease-stage patients.
Assuntos
DNA Tumoral Circulante/genética , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas do Esôfago/genética , Adulto , Idoso , Sequência de Bases/genética , Biomarcadores Tumorais/genética , Estudos de Coortes , Neoplasias Esofágicas/sangue , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/cirurgia , Carcinoma de Células Escamosas do Esôfago/sangue , Carcinoma de Células Escamosas do Esôfago/patologia , Carcinoma de Células Escamosas do Esôfago/cirurgia , Feminino , Seguimentos , Frequência do Gene/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Recidiva Local de Neoplasia/genética , Estadiamento de Neoplasias , Período Pós-Operatório , Período Pré-OperatórioRESUMO
Coronin 1B has been shown to be critical for cell motility and various actin-dependent processes. To understand its role more extensively, the expression and transcriptional regulation of Coro1b gene during mouse development were explored. Coronin 1B is ubiquitously expressed in the whole embryo but nevertheless shows distinct expression pattern in developing heart. In addition to the localization in endocardium, Coronin 1B is specifically expressed in the endocardial cushion and epicardium where cardiac EMT processes take place as the heart develops. Promoter deletion analysis identified the positions between -1038 and -681 is important for Coro1b basal promoter activity. In addition to a correlation of Coronin 1B localization with Wt1 expression in the epicardium, we also identified putative Wt1 binding sequences within Coro1b promoter. Direct binding of Wt1 to GC-rich sequences within the Coro1b promoter is required for the regulation of Coro1b gene expression. In accordance with the motility defect found in Coronin 1B-knockdown cells, a modest decrease in expression of Coronin 1B in the remaining epicardium of Wt1(EGFPCre/EGFPCre) mutant embryos was observed. These findings seem to shed light on the role of Wt1 during cell migration and suggest that, at least in part, this involves transcriptional control of Coro1b gene expression.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Proteínas dos Microfilamentos/metabolismo , Pericárdio/metabolismo , Proteínas WT1/metabolismo , Sequência de Aminoácidos , Animais , Movimento Celular , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário , Sequência Rica em GC , Genes Reporter , Células HeLa , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Células NIH 3T3 , Especificidade de Órgãos , Pericárdio/embriologia , Regiões Promotoras Genéticas , Ligação Proteica , Ativação Transcricional , Proteínas WT1/genéticaRESUMO
Cancer-initiating cells [cancer stem cells (CSCs)] in colon cancer cells can be selectively suppressed when they are cultured on Pluronic (nanosegment)-grafted dishes, whereas CSCs are maintained on conventional tissue culture dishes and extracellular matrix-coated dishes. CSCs persist in tumors as a distinct population and cause relapse and metastasis by giving rise to new tumorigenic clones. The purification or depletion (suppression) of CSCs should be useful for analyzing CSC characteristics and for clinical application. CSCs can be selectively suppressed from colon cancer cells containing adipose-derived stem cells (ADSCs) on Pluronic-grafted dishes, while ADSCs remain on the dishes. ADSCs on Pluronic-grafted dishes after the suppression of the CSCs can differentiate into osteoblasts, chondrocytes, adipocytes, cardiomyocytes, and neuronal cells. The CSCs and ADSCs exhibited different characteristics. The selection of ADSCs was possible on Pluronic-grafted dishes that suppressed the CSCs from the fat tissues of cancer patients (i.e., cell-sorting dishes), which was explained by specific biomedical characteristics of Pluronic.
Assuntos
Adipócitos/fisiologia , Materiais Biocompatíveis , Células-Tronco Neoplásicas/fisiologia , Células-Tronco/fisiologia , Idoso , Idoso de 80 Anos ou mais , Animais , Diferenciação Celular , Linhagem Celular Tumoral , Células Cultivadas , Neoplasias do Colo/patologia , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Imuno-Histoquímica , Camundongos , Pessoa de Meia-Idade , Transplante de Neoplasias , Poloxâmero , PoliestirenosRESUMO
PURPOSE: We evaluated the higher levels of carcinoembryonic antigen (CEA) secreted by the LoVo human colon carcinoma cells in a medium containing anticancer drugs. Drug-resistant LoVo cells were analyzed by subcutaneously xenotransplanting them into mice. The aim of this study was to evaluate whether the drug-resistant cells isolated in this study were cancer-initiating cells, known also as cancer stem cells (CSCs). METHODS: The production of CEA was investigated in LoVo cells that were cultured with 0-10 mM of anticancer drugs, and we evaluated the increase in CEA production by the LoVo cells that were stimulated by anticancer drug treatment. The expression of several CSC markers in LoVo cells treated with anticancer drugs was also evaluated. Following anticancer drug treatment, LoVo cells were injected subcutaneously into the flanks of severe combined immunodeficiency mice in order to evaluate the CSC fraction. RESULTS: Production of CEA by LoVo cells was stimulated by the addition of anticancer drugs. Drug-resistant LoVo cells expressed lower levels of CSC markers, and LoVo cells treated with any of the anticancer drugs tested did not generate tumors within 8 weeks from when the cells were injected subcutaneously into severe combined immunodeficiency mice. These results suggest that the drug-resistant LoVo cells have a smaller population of CSCs than the untreated LoVo cells. CONCLUSION: Production of CEA by LoVo cells can be stimulated by the addition of anticancer drugs. The drug-resistant subpopulation of LoVo colon cancer cells could stimulate the production of CEA, but these cells did not act as CSCs in in vivo tumor generation experiments.