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1.
BMC Cancer ; 19(1): 831, 2019 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-31443698

RESUMO

BACKGROUND: Metastasis is responsible for the majority of deaths in a variety of cancer types, including breast cancer. Although several factors or biomarkers have been identified to predict the outcome of patients with breast cancer, few studies have been conducted to identify metastasis-associated biomarkers. METHODS: Quantitative iTRAQ proteomics analysis was used to detect differentially expressed proteins between lymph node metastases and their paired primary tumor tissues from 23 patients with metastatic breast cancer. Immunohistochemistry was performed to validate the expression of two upregulated (EpCAM, FADD) and two downregulated (NDRG1, αB-crystallin) proteins in 190 paraffin-embedded tissue samples. These four proteins were further analyzed for their correlation with clinicopathological features in 190 breast cancer patients. RESULTS: We identified 637 differentially regulated proteins (397 upregulated and 240 downregulated) in lymph node metastases compared with their paired primary tumor tissues. Data are available via ProteomeXchange with identifier PXD013931. Furthermore, bioinformatics analysis using GEO profiling confirmed the difference in the expression of EpCAM between metastases and primary tumors tissues. Two upregulated (EpCAM, FADD) and two downregulated (NDRG1, αB-crystallin) proteins were associated with the progression of breast cancer. Obviously, EpCAM plays a role in the metastasis of breast cancer cells to the lymph node. We further identified αB-crystallin as an independent biomarker to predict lymph node metastasis and the outcome of breast cancer patients. CONCLUSION: We have identified that EpCAM plays a role in the metastasis of breast cancer cells to the lymph node. αB-crystallin, a stress-related protein that has recently been shown to be important for cell invasion and survival, was identified as a potential prognostic biomarker to predict the outcome of breast cancer patients.


Assuntos
Biomarcadores Tumorais , Neoplasias da Mama/etiologia , Neoplasias da Mama/patologia , Molécula de Adesão da Célula Epitelial/genética , Cadeia B de alfa-Cristalina/genética , Neoplasias da Mama/mortalidade , Biologia Computacional/métodos , Molécula de Adesão da Célula Epitelial/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Metástase Neoplásica , Estadiamento de Neoplasias , Modelos de Riscos Proporcionais , Proteoma , Proteômica/métodos , Cadeia B de alfa-Cristalina/metabolismo
2.
BMC Urol ; 19(1): 67, 2019 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-31324239

RESUMO

BACKGROUND: Epithelial cell adhesion molecule (EpCAM) expression has been reported in many types of cancer, including prostate cancer (PCa). However, the role of EpCAM expression remains inconsistent. We conducted a meta-analysis to assess the clinicopathological and prognostic significance of EpCAM expression in PCa. METHODS: Publications were searched online using electronic databases. The available data were obtained from The Cancer Genome Atlas (TCGA). The odds ratios (ORs) or hazard ratios (HRs) with their 95% confidence intervals (CIs) were calculated. RESULTS: We identified seven studies in which immunohistochemistry was used and that included 871 prostatic tissue samples. EpCAM expression was significantly higher in PCa samples than in benign and normal tissue samples (OR = 77.93, P = 0.002; OR = 161.61, P <  0.001; respectively). No correlation of EpCAM overexpression with pT stage and lymph node metastasis was observed; however, EpCAM overexpression showed a significant correlation with Gleason score (OR = 0.48, P = 0.012) and bone metastasis (OR = 145.80, P <  0.001). Furthermore, TCGA data showed that EpCAM overexpression was not closely correlated with age, pT stage, lymph node metastasis, number of lymph node, prostate-specific antigen level, Gleason score, biochemical recurrence, and overall survival. Based on multivariate Cox proportional-hazards regression analysis, a significant correlation was observed between EpCAM overexpression and 5-year worse biochemical recurrence free-survival. CONCLUSIONS: EpCAM overexpression may be correlated with the development of bone metastasis and worse biochemical recurrence free-survival of PCa. Further studies are needed to verify these findings.


Assuntos
Bases de Dados Genéticas , Molécula de Adesão da Célula Epitelial/genética , Regulação Neoplásica da Expressão Gênica , Genoma/genética , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/genética , Molécula de Adesão da Célula Epitelial/biossíntese , Humanos , Masculino , Gradação de Tumores , Prognóstico , Neoplasias da Próstata/metabolismo , Fatores de Risco
3.
Nat Commun ; 10(1): 3350, 2019 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-31350390

RESUMO

The liver parenchyma is composed of hepatocytes and bile duct epithelial cells (BECs). Controversy exists regarding the cellular origin of human liver parenchymal tissue generation during embryonic development, homeostasis or repair. Here we report the existence of a hepatobiliary hybrid progenitor (HHyP) population in human foetal liver using single-cell RNA sequencing. HHyPs are anatomically restricted to the ductal plate of foetal liver and maintain a transcriptional profile distinct from foetal hepatocytes, mature hepatocytes and mature BECs. In addition, molecular heterogeneity within the EpCAM+ population of freshly isolated foetal and adult human liver identifies diverse gene expression signatures of hepatic and biliary lineage potential. Finally, we FACS isolate foetal HHyPs and confirm their hybrid progenitor phenotype in vivo. Our study suggests that hepatobiliary progenitor cells previously identified in mice also exist in humans, and can be distinguished from other parenchymal populations, including mature BECs, by distinct gene expression profiles.


Assuntos
Fígado/citologia , Transcrição Genética , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Molécula de Adesão da Célula Epitelial/genética , Molécula de Adesão da Célula Epitelial/metabolismo , Feto/metabolismo , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Análise de Célula Única , Células-Tronco/citologia , Células-Tronco/metabolismo
4.
Int J Clin Oncol ; 24(9): 999-1011, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31273487

RESUMO

Lynch syndrome is a cancer-predisposing syndrome inherited in an autosomal-dominant manner, wherein colon cancer and endometrial cancer develop frequently in the family, it results from a loss-of-function mutation in one of four different genes (MLH1, MSH2, MSH6, and PMS2) encoding mismatch repair proteins. Being located immediately upstream of the MSH2 gene, EPCAM abnormalities can affect MSH2 and cause Lynch syndrome. Mismatch repair proteins are involved in repairing of incorrect pairing (point mutations and deletion/insertion of simple repetitive sequences, so-called microsatellites) that can arise during DNA replication. MSH2 forms heterodimers with MSH6 or MSH3 (MutSα, MutSß, respectively) and is involved in mismatch-pair recognition and initiation of repair. MLH1 forms a complex with PMS2, and functions as an endonuclease. If the mismatch repair system is thoroughly working, genome integrity is maintained completely. Lynch syndrome is a state of mismatch repair deficiency due to a monoallelic abnormality of any mismatch repair genes. The phenotype indicating the mismatch repair deficiency can be frequently shown as a microsatellite instability in tumors. Children with germline biallelic mismatch repair gene abnormalities were reported to develop conditions such as gastrointestinal polyposis, colorectal cancer, brain cancer, leukemia, etc., and so on, demonstrating the need to respond with new concepts in genetic counseling. In promoting cancer genome medicine in a new era, such as by utilizing immune checkpoints, it is important to understand the genetic and genomic molecular background, including the status of mismatch repair deficiency.


Assuntos
Neoplasias Colorretais Hereditárias sem Polipose/genética , Reparo de Erro de Pareamento de DNA/fisiologia , Neoplasias Encefálicas/genética , Criança , Reparo de Erro de Pareamento de DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Neoplasias do Endométrio/genética , Molécula de Adesão da Célula Epitelial/genética , Feminino , Aconselhamento Genético , Testes Genéticos , Humanos , Instabilidade de Microssatélites , Endonuclease PMS2 de Reparo de Erro de Pareamento/genética , Endonuclease PMS2 de Reparo de Erro de Pareamento/metabolismo , Proteína 1 Homóloga a MutL/genética , Proteína 1 Homóloga a MutL/metabolismo , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , Mutação
5.
Cancer Sci ; 110(8): 2590-2599, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31169336

RESUMO

Liquid biopsy of circulating tumor cells (CTC) and circulating tumor DNA (ctDNA) is gaining attention as a method for real-time monitoring in cancer patients. Conventional methods based upon epithelial cell adhesion molecule (EpCAM) expression have a risk of missing the most aggressive CTC subpopulations due to epithelial-mesenchymal transition and may, thus, underestimate the total number of actual CTC present in the bloodstream. Techniques utilizing a label-free inertial microfluidics approach (LFIMA) enable efficient capture of CTC without the need for EpCAM expression. In this study, we optimized a method for analyzing genetic alterations using next-generation sequencing (NGS) of extracted ctDNA and CTC enriched using an LFIMA as a first-phase examination of 30 patients with head and neck cancer, esophageal cancer, gastric cancer and colorectal cancer (CRC). Seven patients with advanced CRC were enrolled in the second-phase examination to monitor the emergence of alterations occurring during treatment with epidermal growth factor receptor (EGFR)-specific antibodies. Using LFIMA, we effectively captured CTC (median number of CTC, 14.5 cells/mL) from several types of cancer and detected missense mutations via NGS of CTC and ctDNA. We also detected time-dependent genetic alterations that appeared during anti-EGFR therapy in CTC and ctDNA from CRC patients. The results of NGS analyses indicated that alterations in the genomic profile revealed by the liquid biopsy could be expanded by using a combination of assays with CTC and ctDNA. The study was registered with the University Hospital Medical Information Network Clinical Trials Registry (ID: UMIN000014095).


Assuntos
DNA Tumoral Circulante/genética , DNA de Neoplasias/genética , Neoplasias/genética , Neoplasias/patologia , Células Neoplásicas Circulantes/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Molécula de Adesão da Célula Epitelial/genética , Transição Epitelial-Mesenquimal/genética , Receptores ErbB/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Biópsia Líquida/métodos , Masculino , Pessoa de Meia-Idade , Mutação/genética
6.
Sensors (Basel) ; 19(8)2019 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-31010258

RESUMO

A new electrochemical immunosensor for cancer cell detection based on a specific interaction between the metastasis-related antigen of epithelial cell adhesion molecule (EpCAM) on the cell membrane and its monoclonal antibody (Anti-EpCAM) immobilized on a gold electrode has been developed. The amino-terminated polyamidoamine dendrimer (G6 PAMAM) was first covalently attached to the 3-mercaptopropionic acid (MPA)-functionalized gold electrode to obtain a thin film, and then completely carboxylated by succinic anhydride (SA). Next, the Anti-EpCAM was covalently bound with the G6 PAMAM to obtain a stable recognition layer. In the presence of the EpCAM expressing hepatocellular carcinomas cell line of HepG2, the specific immune recognition (Anti-EpCAM/EpCAM) led to an obvious change of the electron transfer ability. The properties of the layer-by-layer assembly process was examined by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The final determination of HepG2 cells was performed in the presence of the reversible [Fe(CN)6]3-/4- redox couple using impedance technique. Based on the advantages of PAMAM nanomaterial and immune reaction, a linear response to HepG2 cells ranging from 1 × 104 to 1 × 106 cells mL-1 with a calculated detection limit of 2.1 × 103 cells mL-1 was obtained. We expect this method can provide a potential tool for cancer cell monitoring and protein expression analysis.


Assuntos
Técnicas Biossensoriais , Molécula de Adesão da Célula Epitelial/genética , Imunoensaio , Neoplasias Hepáticas/genética , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Membrana Celular/química , Membrana Celular/imunologia , Dendrímeros/química , Espectroscopia Dielétrica , Molécula de Adesão da Célula Epitelial/química , Molécula de Adesão da Célula Epitelial/imunologia , Células Hep G2 , Humanos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/imunologia , Nanoestruturas/química , Poliaminas/química
7.
ACS Appl Mater Interfaces ; 11(18): 16347-16356, 2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-31032616

RESUMO

We report new surface coatings that adhesively distinguish three breast epithelial cell lines (MCF-10A, MCF-7, and TMX2-28) when cell suspensions in buffer or breast milk are flowed over the coatings. We also report the selective capture of epithelial cells and rejection of Jurkat lymphocytes, with average selectivities exceeding 60 and captured cell purities often exceeding >99%. The surfaces achieve the dual goals of selective cell capture and resistance to fouling by proteins and other components of breast milk. The coatings do not rely on antibody targeting of cell surface markers but instead contain polycation chains embedded within a layer of end-tethered poly(ethylene glycol) (PEG) chains. The PEG, somewhat shielding the polycations, prevents surface fouling by proteins, nondesired cells, and other milk components, while the polycations produce electrostatic attractions that are heterogeneous on nanoscopic length scales. These electrostatic heterogeneities on the engineered coating, shown to produce curvature-selective particle capture in other studies, produce cell selectivity here. The ability of the engineered surfaces to discriminate these cell lines via an electrostatic driving force is remarkable, as the cells are of very similar surface charge as evidenced by their nearly identical ζ-potentials. The current surfaces, which likely distinguish cells based on their electrostatic surface landscape combined with other factors, adhesively distinguish cell lines that may differ only slightly in their expression of a surface marker, or cancer cells that minimally express EpCAM but which have different distributions of electrostatic charge on their surfaces. These surfaces are among the first to be documented for the compatibility of a polymer brush with human breast milk and may find use in technologies that capture cells from human breast milk or other complex fluids for cancer risk assessment.


Assuntos
Adesivos/farmacologia , Mama/efeitos dos fármacos , Linfócitos/efeitos dos fármacos , Leite Humano/química , Adesivos/química , Mama/citologia , Tampões (Química) , Molécula de Adesão da Célula Epitelial/genética , Células Epiteliais/química , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Células Jurkat , Células MCF-7 , Polietilenoglicóis/química , Polímeros/química , Polímeros/farmacologia , Propriedades de Superfície
8.
Mol Genet Genomic Med ; 7(5): e587, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30916491

RESUMO

BACKGROUND: Inherited epimutations of Mismatch Repair (MMR) genes are responsible for Lynch Syndrome (LS) in a small, but well defined, subset of patients. Methylation of the MSH2 promoter consequent to the deletion of the upstream EPCAM gene is found in about 1%-3% of the LS patients and represents a classical secondary, constitutional and tissue-specific epimutation. Several different EPCAM deletions have been reported worldwide, for the most part representing private variants caused by an Alu-mediated recombination. METHODS: 712 patients with suspected LS were tested for MMR mutation in our Institute. EPCAM deletions were detected by multiplex ligation-dependent probe amplification (MLPA) and then defined by Long-Range polymerase chain reaction (PCR)/Sanger sequencing. A comprehensive molecular characterization of colorectal cancer (CRC) tissues was carried out by immunohistochemistry of MMR proteins, Microsatellite Instability (MSI) assay, methylation specific MLPA and transcript analyses. In addition, somatic deletions and/or variants were investigated by MLPA and next generation sequencing (NGS). RESULTS: An EPCAM deletion was found in five unrelated probands in Italy: variants c.556-490_*8438del and c.858+1193_*5826del are novel; c.859-1430_*2033del and c.859-670_*530del were previously reported. All probands were affected by CRC at young age; tumors showed MSI and abnormal MSH2/MSH6 proteins expression. MSH2 promoter methylation, as well as aberrant in-frame or out-of-frame EPCAM/MSH2 fusion transcripts, were detected in CRCs and normal mucosae. CONCLUSION: An EPCAM deletion was the causative variant in about 2% of our institutional series of 224 LS patients, consistent with previously estimated frequencies. Early age and multiple CRCs was the main clinical feature of this subset of patients.


Assuntos
Neoplasias Colorretais Hereditárias sem Polipose/genética , Molécula de Adesão da Célula Epitelial/genética , Deleção de Genes , Frequência do Gene , Adulto , Metilação de DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , Fenótipo
9.
J Biol Chem ; 294(19): 7769-7786, 2019 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-30926604

RESUMO

Mesenchymal stem cells (MSCs) are widely considered to be an attractive cell source for regenerative therapies, but maintaining multipotency and self-renewal in cultured MSCs is especially challenging. Hence, the development and mechanistic description of strategies that help promote multipotency in MSCs will be vital to future clinical use. Here, using an array of techniques and approaches, including cell biology, RT-quantitative PCR, immunoblotting, immunofluorescence, flow cytometry, and ChIP assays, we show that the extracellular domain of epithelial cell adhesion molecule (EpCAM) (EpEX) significantly increases the levels of pluripotency factors through a signaling cascade that includes epidermal growth factor receptor (EGFR), signal transducer and activator of transcription 3 (STAT3), and Lin-28 homolog A (LIN28) and enhances the proliferation of human bone marrow MSCs. Moreover, we found that EpEX-induced LIN28 expression reduces the expression of the microRNA LET7 and up-regulates that of the transcription factor high-mobility group AT-hook 2 (HMGA2), which activates the transcription of pluripotency factors. Surprisingly, we found that EpEX treatment also enhances osteogenesis of MSCs under differentiation conditions, as evidenced by increases in osteogenic markers, including Runt-related transcription factor 2 (RUNX2). Taken together, our results indicate that EpEX stimulates EGFR signaling and thereby context-dependently controls MSC states and activities, promoting cell proliferation and multipotency under maintenance conditions and osteogenesis under differentiation conditions.


Assuntos
Molécula de Adesão da Célula Epitelial/metabolismo , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Proteínas de Ligação a RNA/biossíntese , Transdução de Sinais , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Molécula de Adesão da Célula Epitelial/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Regulação da Expressão Gênica , Proteína HMGA2/genética , Proteína HMGA2/metabolismo , Humanos , Células-Tronco Mesenquimais/citologia , MicroRNAs/genética , Proteínas de Ligação a RNA/genética , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
10.
Analyst ; 144(8): 2574-2583, 2019 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-30821313

RESUMO

Label free sorting of circulating tumor cells (CTCs) often remains a challenge due to their rarity in peripheral blood and identical morphology to white blood cells. We present a novel label-free passive microfluidic technique for isolation of cancer cells (EpCAM+ and CD45-) from peripheral blood mononuclear cells (PBMCs) (CD45+ and EpCAM-) in aqueous two-phase system (ATPS). Our technique involves non-inertial lift induced lateral cell migration across liquid-liquid interface that is employed for sorting cells of different size and stiffness. The interplay between lift force and interfacial tension (IFT) force governs cell migration phenomena. We estimate the order of magnitude of the lift force and find it to be higher than the IFT for cancer cells above a critical strain rate parameter ([small gamma, Greek, dot above]/h). The effect of spreading parameter and viscoelastic force was found to have negligible effect on lateral migration of cells. We demonstrated isolation of two different types of cancer cells, namely, MCF-7 and MDA-MB-231 from PBMCs and quantify our sorting results by tagging the cells with EpCAM and CD45 and using fluorescence imaging. With 102-104 cancer cells in 105-107 PBMCs, we achieved a processing rate of >25 000 cells per min at a sorting efficiency of ∼99%. Moreover, we demonstrated that cancer cells isolated from PBMCs using the proposed technique remain viable and can be cultured for downstream analysis.


Assuntos
Separação Celular/métodos , Leucócitos Mononucleares/citologia , Técnicas Analíticas Microfluídicas/métodos , Células Neoplásicas Circulantes/patologia , Linhagem Celular Tumoral , Movimento Celular , Sobrevivência Celular , Dextranos/química , Módulo de Elasticidade , Molécula de Adesão da Célula Epitelial/genética , Humanos , Antígenos Comuns de Leucócito/genética , Células Neoplásicas Circulantes/química , Polietilenoglicóis/química , Tensão Superficial
11.
Tumour Biol ; 41(2): 1010428318823361, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30808252

RESUMO

The objective of the present feasibility study was to transfer single cell line cells to either microscopy slides for downstream immune characterization or to polymerase chain reaction tubes for downstream DNA quantitation. Tumour cell lines, SKBR3 and MCF7 and trophoblast cell line JEG-3 were spiked in healthy donor blood. The CytoTrack system was used to scan the spiked blood samples to identify target cells. Individual target cells were identified, picked by use of a CytoPicker and deposited to either a microscopic slide or a polymerase chain reaction tube (PCR). Single tumour cells on microscopic slides were further immunostained with human epidermal growth factor receptor 2 (Her2) and epithelial cell adhesion molecule (EpCAM). From the picked cells in polymerase chain reaction tubes, DNA was amplified, quantified and used for Short Tandem Repeat genotyping. Depositing rare cells to microscopy slides was laborious with only five cells per hour. In this study with a trained operator, the picked cells had an 80.5% recovery rate. Depositing single trophoblast cells in PCR tubes was a faster process with 10 cells in 5 min. Immunostaining of isolated cells by both Her2 and EpCAM was possible but showed varying staining intensity. Presence of trophoblasts and contaminating white blood cells in PCR tubes after cell picking was confirmed based on DNA yield and mixed Short Tandem Repeat profiles in five out of eight samples. Using the CytoPicker tool, single tumour and trophoblast cells were successfully isolated and moved from blood samples, allowing subsequent immunostaining or Short Tandem Repeat genotyping.


Assuntos
Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Células Neoplásicas Circulantes/patologia , Análise de Célula Única/métodos , Linhagem Celular Tumoral , Separação Celular/métodos , DNA/genética , Molécula de Adesão da Célula Epitelial/genética , Feminino , Humanos , Células MCF-7 , Reação em Cadeia da Polimerase/métodos , Receptor ErbB-2/genética , Trofoblastos/patologia
12.
Mol Med Rep ; 19(3): 2317-2322, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30747214

RESUMO

Epithelial cell adhesion molecule (EpCAM), which is expressed in the majority of epithelial tissues, exhibits tumor growth promoting abilities and is overexpressed in human epithelial ovarian cancer. Therefore, EpCAM is considered to be a promising target for specific immune­based therapies. The present study evaluated the role of IL­6 and IL­8 in the expression of EpCAM in the A2780 human ovarian cancer cell line. Furthermore, the cellular localization of the EpCAM protein in A2780 cells was determined and the effect of EpCAM inhibition on the proliferation of the A2780 cells was investigated. An MTT assay demonstrated that blocking EpCAM with anti­EPCAM antibodies had no effect on cellular metabolic activity (proliferation). Gene expression analysis revealed that IL­8 increased EpCAM expression, whereas IL­6 and the combination of IL­6/IL­8 had no effect on EpCAM expression. Immunofluorescence analysis confirmed that EpCAM is expressed on A2780 cell membranes. The present results demonstrated that IL­8 increased EpCAM expression at the mRNA level in ovarian cancer cells and suggested a potential role of IL­6 as an inhibitor of IL­8­stimulated EpCAM expression.


Assuntos
Molécula de Adesão da Célula Epitelial/genética , Regulação Neoplásica da Expressão Gênica , Interleucina-6/imunologia , Interleucina-8/imunologia , Neoplasias Ovarianas/genética , Linhagem Celular Tumoral , Molécula de Adesão da Célula Epitelial/imunologia , Feminino , Humanos , Neoplasias Ovarianas/imunologia
13.
Medicine (Baltimore) ; 98(6): e14345, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30732163

RESUMO

This study aimed to identify modules associated with breast cancer (BC) development by constructing a gene co-expression network, and mining hub genes that may serve as markers of invasive breast cancer (IBC).We downloaded 2 gene expression datasets from the Gene Expression Omnibus (GEO) database, and used weighted gene co-expression network analysis (WGCNA) to dynamically study the changes of co-expression genes in normal breast tissues, ductal carcinoma in situ (DCIS) tissues, and IBC tissues. Modules that highly correlated with BC development were carried out functional enrichment analysis for annotation, visualization, and integration discovery. The hub genes detected by WGCNA were also confirmed using the Oncomine dataset.We detected 17 transcriptional modules in total and 4 - namely tan, greenyellow, turquoise, and brown - were highly correlated with BC development. The functions of these 4 modules mainly concerned cell migration (tan module, P = 3.03 × 10), the cell cycle (greenyellow module, P = 3.08 × 10), cell-cell adhesion (turquoise module, P = .002), and the extracellular exosome (brown module, P = 1.38 × 10). WGCNA also mined the hub genes, which were highly correlated with the genes in the same module and with BC development. The Oncomine database confirmed that the expressions levels of 6 hub genes were significantly higher in BC tissues than in normal tissues, with fold changes larger than 2 (all P < .05). Apart from the 2 well-known genes EPCAM and MELK, during the development of BC, KRT8, KRT19, KPNA2, and ECT2 also play key roles, and may be used as new targets for the detection or treatment of BC.In summary, our study demonstrated that hub genes such as EPCAM and MELK are highly correlated with breast cancer development. However, KRT8, KRT19, KPNA2, and ECT2 may also have potential as diagnostic and prognostic biomarkers of IBC.


Assuntos
Neoplasias da Mama/genética , Redes Reguladoras de Genes/genética , Biomarcadores Tumorais , Carcinoma Ductal/genética , Adesão Celular , Ciclo Celular , Movimento Celular , Molécula de Adesão da Célula Epitelial/genética , Exossomos , Perfilação da Expressão Gênica , Humanos , Queratinas/genética , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas/genética , alfa Carioferinas/genética
14.
J Surg Res ; 239: 14-21, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30782542

RESUMO

BACKGROUND: Adjuvant therapy for early-stage colorectal cancer improves survival. Biologic agents have shown promise as adjuncts to chemotherapy in metastatic colon cancer, but the effect on earlier stage cancer remains unclear. MATERIALS AND METHODS: We conducted a systematic review and meta-analysis of the additive effect of biologic agents to adjuvant chemotherapy on survival in colorectal cancer (all comers and subpopulations defined by microsatellite instability, BRAF and KRAS status, and stage). Only randomized controlled trials published between 2002 and 2017 in MEDLINE, EMBASE, and CENTRAL were included. The control arm: chemotherapy alone, the intervention arm: chemotherapy with biologic agents. OUTCOMES: overall survival (OS) and disease-free survival. RESULTS: Six trials including 10,754 patients were included. OS (hazard ratio [HR] 2.55, 95% confidence interval [CI] 2.15-3.03) and disease-free survival (HR 2.54, 95% CI 2.25-2.87) were significantly worse in the intervention arm. High heterogeneity was explained by subgroup analysis of different biologic agents (bevacizumab versus others); however, results still showed harm in the intervention arm across subgroups. Bevacizumab was associated with improved OS in patients with microsatellite instability (HR 0.58, 95% CI 0.36-0.92); this was the only indication of benefit for a biomarker-defined subpopulation. Analyses by tumor stage failed to demonstrate advantage with use of a biologic agent; however, it explained heterogeneity. CONCLUSIONS: The addition of biologic agents to adjuvant chemotherapy in the treatment of high-risk stage II and III colorectal cancer is associated with worse survival outcomes. The only subgroup of patients that may benefit from the addition of bevacizumab to adjuvant chemotherapy is those with microsatellite unstable tumors.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Colectomia , Neoplasias Colorretais/terapia , Protectomia , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Bevacizumab/farmacologia , Bevacizumab/uso terapêutico , Produtos Biológicos/farmacologia , Produtos Biológicos/uso terapêutico , Quimioterapia Adjuvante/métodos , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Intervalo Livre de Doença , Molécula de Adesão da Célula Epitelial/antagonistas & inibidores , Molécula de Adesão da Célula Epitelial/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Humanos , Instabilidade de Microssatélites , Estadiamento de Neoplasias , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Análise de Sobrevida
15.
BMC Cancer ; 18(1): 1092, 2018 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-30419852

RESUMO

BACKGROUND: Development of chemo-/radioresistance is a major challenge for the current prostate cancer (CaP) therapy. We have previously demonstrated that epithelial cell adhesion molecule (EpCAM) is associated with CaP growth and therapeutic resistance in vitro, however, the role of EpCAM in CaP in vivo is not fully elucidated. Here, we aimed to investigate how expression of EpCAM is involved in CaP growth and chemo-/radiotherapy response in NOD/SCID mouse models in vivo and to validate its role as a therapeutic target for CaP therapy. METHODS: EpCAM was knocked down in PC-3 CaP cell line using short hairpin RNA (shRNA). The effect of EpCAM-knockdown (KD) on tumour growth, chemo-/radiotherapy response and animal survival was evaluated on subcutaneous (s.c) and orthotopic mouse models. RESULTS: We found that KD of EpCAM significantly inhibited tumour growth, increased xenograft sensitivity to chemotherapy/radiotherapy, and prolonged the survival of tumour-bearing mice. In addition, we demonstrated that KD of EpCAM is associated with downregulation of the PI3K/Akt/mTOR pathway. CONCLUSIONS: In conclusion, our data confirms that CaP growth and chemo-/radioresistance in vivo is associated with over-expression of EpCAM, which serves both a functional biomarker and promising therapeutic target.


Assuntos
Molécula de Adesão da Célula Epitelial/genética , Neoplasias da Próstata/genética , Animais , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Quimioterapia Adjuvante , Modelos Animais de Doenças , Molécula de Adesão da Célula Epitelial/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Fosfatidilinositol 3-Quinases/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Interferente Pequeno/genética , Radioterapia Adjuvante , Transdução de Sinais , Serina-Treonina Quinases TOR , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Pathology ; 50(7): 737-741, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30389218

RESUMO

The mutational landscape of adenoid cystic carcinoma (ACC) is currently being revealed, but further studies are needed to identify biomarkers as therapeutic targets or prognostic factors of ACC. In this study, we investigated the expression of epithelial cell adhesion molecule (EpCAM) in ACCs. We retrospectively collected 83 cases of surgically resected ACCs. Using tissue microarray, we conducted immunohistochemical staining using the anti-EpCAM antibody. EpCAM expression was analysed by intensity score and the total immunostaining score. The positivity was 97.6% (81/83 cases), regardless of the intensity score. A higher histological grade (p = 0.006) and specific tumour location (non-salivary gland origin, p = 0.02) showed a correlation with higher EpCAM intensity. Higher EpCAM expression by total immunostaining score was associated with histological grade (p = 0.004), distant metastasis (p = 0.004) and poorer prognosis (overall survival p = 0.015 and progression-free survival p = 0.033). We suggest EpCAM as a candidate prognostic marker and a putative therapeutic target in ACC. Also, ACCs arising from salivary gland and non-salivary gland sites, respectively, might display different pathophysiologies in which EpCAM could play a role.


Assuntos
Carcinoma Adenoide Cístico/patologia , Molécula de Adesão da Célula Epitelial/metabolismo , Neoplasias das Glândulas Salivares/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/metabolismo , Carcinoma Adenoide Cístico/diagnóstico , Carcinoma Adenoide Cístico/metabolismo , Molécula de Adesão da Célula Epitelial/genética , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Prognóstico , Intervalo Livre de Progressão , Estudos Retrospectivos , Neoplasias das Glândulas Salivares/diagnóstico , Neoplasias das Glândulas Salivares/metabolismo , Glândulas Salivares/metabolismo , Glândulas Salivares/patologia , Análise Serial de Tecidos , Adulto Jovem
17.
Epigenetics Chromatin ; 11(1): 70, 2018 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-30445998

RESUMO

BACKGROUND: Epithelial mesenchymal transition (EMT) is tightly regulated by a network of transcription factors (EMT-TFs). Among them is the nuclear factor ZEB2, a member of the zinc-finger E-box binding homeobox family. ZEB2 nuclear localization has been identified in several cancer types, and its overexpression is correlated with the malignant progression. ZEB2 transcriptionally represses epithelial genes, such as E-cadherin (CDH1), by directly binding to the promoter of the genes it regulates and activating mesenchymal genes by a mechanism in which there is no full agreement. Recent studies showed that EMT-TFs interact with epigenetic regulatory enzymes that alter the epigenome, thereby providing another level of control. The role of epigenetic regulation on ZEB2 function is not well understood. In this study, we aimed to characterize the epigenetic effect of ZEB2 repressive function on the regulation of a small Rab GTPase RAB25. RESULTS: Using cellular models with conditional ZEB2 expression, we show a clear transcriptional repression of RAB25 and CDH1. RAB25 contributes to the partial suppression of ZEB2-mediated cell migration. Furthermore, a highly significant reverse correlation between RAB25 and ZEB2 expression in several human cancer types could be identified. Mechanistically, ZEB2 binds specifically to E-box sequences on the RAB25 promoter. ZEB2 binding is associated with the local increase in DNA methylation requiring DNA methyltransferases as well as histone deacetylation (H3K9Ac) depending on the activity of SIRT1. Surprisingly, SIRT1 and DNMTs did not interact directly with ZEB2, and while SIRT1 inhibition decreased the stability of long-term repression, it did not prevent down-regulation of RAB25 and CDH1 by ZEB2. CONCLUSIONS: ZEB2 expression is resulting in drastic changes at the chromatin level with both clear DNA hypermethylation and histone modifications. Here, we revealed that SIRT1-mediated H3K9 deacetylation helps to maintain gene repression but is not required for the direct ZEB2 repressive function. Targeting epigenetic enzymes to prevent EMT is an appealing approach to limit cancer dissemination, but inhibiting SIRT1 activity alone might have limited effect and will require drug combination to efficiently prevent EMT.


Assuntos
Epigênese Genética , Transição Epitelial-Mesenquimal/fisiologia , Sirtuína 1/metabolismo , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo , Proteínas rab de Ligação ao GTP/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Metilação de DNA , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , Molécula de Adesão da Célula Epitelial/genética , Molécula de Adesão da Célula Epitelial/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Regiões Promotoras Genéticas , Sirtuína 1/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/genética , Proteínas rab de Ligação ao GTP/metabolismo
18.
Bull Cancer ; 105(10): 907-917, 2018 Oct.
Artigo em Francês | MEDLINE | ID: mdl-30268633

RESUMO

INTRODUCTION: Next generation sequencing allows the simultaneous analysis of large panel of genes for families or individuals with a strong suspicion of hereditary breast and/or ovarian cancer (HBOC). Because of lack of guidelines, several panels of genes potentially involved in HBOC were designed, with large disparities not only in their composition but also in medical care offered to mutation carriers. Then, homogenization in practices is needed. METHODS: The French Genetic and Cancer Group (GGC) - Unicancer conducted an exhaustive bibliographic work on 18 genes of interest. Only publications with unbiased risk estimates were retained. RESULTS: The expertise of each 18 genes was based on clinical utility criteria, i.e. a relative risk of cancer of 4 and more, available medical tools for screening and prevention of mutation carriers, and pre-symptomatic genetic tests for relatives. Finally, 13 genes were selected to be included in a HBOC diagnosis gene panel: BRCA1, BRCA2, PALB2, TP53, CDH1, PTEN, RAD51C, RAD51D, MLH1, MSH2, MSH6, PMS2, EPCAM. The reasons for excluding NBN, RAD51B, CHEK2, STK11, ATM, BARD1, BRIP1 from the HBOC diagnosis panel are presented. Screening, prevention and genetic counselling guidelines were detailed for each of the 18 genes. DISCUSSION: Due to the rapid increase in knowledge, the GGC has planned a yearly update of the bibliography to take into account new findings. Furthermore, genetic-epidemiological studies are being initiated to better estimate the cancer risk associated with genes which are not yet included in the HBOC diagnosis panel.


Assuntos
Neoplasias da Mama/genética , Predisposição Genética para Doença/genética , Neoplasias Ovarianas/genética , Antígenos CD , Caderinas , Proteínas de Ligação a DNA/genética , Molécula de Adesão da Célula Epitelial/genética , Proteína do Grupo de Complementação N da Anemia de Fanconi/genética , Feminino , França , Genes BRCA1 , Genes BRCA2 , Genes p53 , Marcadores Genéticos/genética , Humanos , Endonuclease PMS2 de Reparo de Erro de Pareamento/genética , Proteína 1 Homóloga a MutL/genética , Proteína 2 Homóloga a MutS/genética , PTEN Fosfo-Hidrolase/genética
19.
PLoS One ; 13(10): e0204957, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30304739

RESUMO

Epithelial cell adhesion molecule (EpCAM) is a glycoprotein on the surface of epithelial cells that is essential for intestinal epithelial integrity and expressed at high levels in many epithelial derived cancers and circulating tumor cells. Here we show the effect of EpCAM levels on migration of Madin-Darby-Canine Kidney (MDCK) epithelial cells. MDCK cells depleted of EpCAM show increased activation of extracellular signal-regulated kinase (ERK) and of myosin, and increased cell spreading and epithelial sheet migration into a gap. In contrast, over-expression of EpCAM inhibits ERK and myosin activation, and slows epithelial sheet migration. Loss of EpCAM is rescued by EpCAM-YFP mutated in the extracellular domain required for cis-dimerization whereas EpCAM-YFP with a mutation that inhibits Claudin-7 interaction cannot rescue increased ERK, myosin activation, and increased migration in EpCAM-depleted cells. In summary, these results indicate that interaction of EpCAM and Claudin-7 at the cell surface negatively regulates epithelial migration by inhibiting ERK and actomyosin contractility.


Assuntos
Adesão Celular/fisiologia , Claudinas/metabolismo , Molécula de Adesão da Célula Epitelial/metabolismo , Actinas/metabolismo , Animais , Movimento Celular/efeitos dos fármacos , Claudinas/química , Dimerização , Cães , Molécula de Adesão da Célula Epitelial/antagonistas & inibidores , Molécula de Adesão da Célula Epitelial/genética , Células Epiteliais/citologia , Células Epiteliais/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fator de Crescimento de Hepatócito/farmacologia , Células Madin Darby de Rim Canino/citologia , Células Madin Darby de Rim Canino/metabolismo , Microscopia Confocal , Miosinas/antagonistas & inibidores , Miosinas/metabolismo , Fosforilação/efeitos dos fármacos , Domínios Proteicos , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Imagem com Lapso de Tempo
20.
PLoS One ; 13(10): e0205297, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30296284

RESUMO

BACKGROUND: Human ovarian surface epithelial (HOSE) cells are a critical cell source for ovarian cancer research; however, they are difficult to obtain and maintain under standard laboratory conditions in large quantities. The aim of this study was to generate immortalized HOSE (IHOSE) cells with maintained properties to the original cell source, thereby guaranteeing a sufficiently large cell quantity for ovarian cancer research. METHODS: HOSE cells isolated from four non-cancer patients and five IHOSE cell lines were established by induction of HPV-E6/E7 expression or SV40 large T antigen using a lenti-viral system. Each of IHOSE cells was confirmed to be distinct by STR profiling. RNA-sequencing was used to compare gene expression profiles in HOSE, IHOSE and ovarian cancer cells. RESULTS: RNA-sequencing results revealed a stronger linear correlation in gene expression between IHOSE and HOSE cells (R2 = 0.9288) than between IHOSE or HOSE cells and ovarian cancer cells (R2 = 0.8562 and R2 = 0.7982, respectively). The gene expression pattern of 319 differentially expressed genes revealed minimal differences between HOSE and IHOSE cells, while a strong difference between ovarian cancer cells and HOSE or IHOSE cells was observed. Furthermore, the five IHOSE cell lines displayed morphological characteristics typical of epithelial cells but showed a lower level of EpCAM, CD133 and E-cadherin, as cancer stem marker, than ovarian cancer cells. Moreover, unlike cancer cells, IHOSE cells could not form colonies in the anchorage-independent soft agar growth assay. CONCLUSION: These findings demonstrate that five newly established IHOSE cell lines have characteristics of progenitor HOSE cells while exhibiting continuous growth, and thus, should be highly useful as control cells for ovarian cancer research.


Assuntos
Antígenos Transformantes de Poliomavirus/genética , Biomarcadores Tumorais/genética , Células Epiteliais/metabolismo , Efeito Fundador , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/genética , Proteínas Repressoras/genética , Antígeno AC133/genética , Antígeno AC133/metabolismo , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos Transformantes de Poliomavirus/metabolismo , Biomarcadores Tumorais/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Transformada , Linhagem Celular Tumoral , Proliferação de Células , Molécula de Adesão da Célula Epitelial/genética , Molécula de Adesão da Célula Epitelial/metabolismo , Células Epiteliais/citologia , Feminino , Expressão Gênica , Humanos , Proteínas Oncogênicas Virais/metabolismo , Ovário/citologia , Ovário/metabolismo , Proteínas E7 de Papillomavirus/metabolismo , Proteínas Repressoras/metabolismo , Análise de Sequência de RNA
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