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1.
Article in Chinese | WPRIM | ID: wpr-927849

ABSTRACT

Objective To investigate the expression and the potential roles of long non-coding RNA(lncRNA)cancer susceptibility candidate 2(CASC2)and imprinted gene H19 in extrahepatic cholangiocarcinoma(ECC). Methods Four samples from patients with ECC were collected for high-throughput sequencing which was conducted to reveal the transcriptomic profiles of lncRNA CASC2 and H19.Bioinformatics tools were employed to predict the potential roles of the two genes.Another 22 ECC tissue samples and the cholangiocarcinoma cell lines(RBE,QBC939,HuH-28,and HuCCT1)with different degrees of differentiation were selected for validation.The para-carcinoma tissue and normal human intrahepatic biliary epithelial cell(HIBEC)were used as the control groups.The expression levels of lncRNA CASC2 and H19 in carcinoma tissue,para-carcinoma tissue,and cell lines were determined by real-time quantitative polymerase chain reaction(qRT-PCR).The correlation analysis was carried out for the clinical indicators of patients with the expression levels of the target genes. Results The two target genes showed significantly different expression between carcinoma tissue and para-carcinoma tissue(all P<0.05).Specifically,CASC2 had higher expression level in the carcinoma tissue than in the para-carcinoma tissue(t=1.262,P=0.025),whereas the expression of H19 showed an opposite trend(t=1.285,P=0.005).The expression levels of CASC2 in QBC939(t=8.114,P=0.015)and HuH-28(t=9.202,P=0.012)cells were significantly higher than that in the control group.The expression levels of H19 were significantly lower in RBE(t=-10.244,P<0.001),QBC939(t=-10.476,P<0.001),HuH-28(t=-19.798,P<0.001),and HuCCT1(t=-16.193,P=0.004)cells than in the control group.Bioinformatics analysis showed that CASC2 was mainly involved in the metabolic process and H19 in the development of multicellular organisms.Both CASC2 and H19 were related to catalytic activity.The expression level of lncRNA CASC2 was correlated with pathological differentiation(χ 2=6.222,P=0.022)and lymph node metastasis(χ2=5.455,P=0.020),and that of lncRNA H19 with pathological differentiation(χ2=1.174,P=0.029)and tumor size(χ2=-0.507,P=0.037). Conclusions In the case of ECC,lncRNA CASC2 and H19 have transcription disorders.lncRNA CASC2 is generally up-regulated in the carcinoma tissue,while H19 is down-regulated.Both genes have the potential to become new molecular markers for ECC.


Subject(s)
Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic/metabolism , Cholangiocarcinoma/genetics , Gene Expression Regulation, Neoplastic , Humans , RNA, Long Noncoding/genetics , Tumor Suppressor Proteins/genetics
2.
Article in Chinese | WPRIM | ID: wpr-880764

ABSTRACT

OBJECTIVE@#To investigate the antitumor effect of ponatinib on the growth of cholangiocarcinoma xenograft derived from a clinical patient in a mouse model expressing FGFR2-CCDC6 fusion protein.@*METHODS@#Lung metastatic tumor tissue was collected from a patient with advanced intrahepatic cholangiocarcinoma and implanted subcutaneously a NOD/SCID/ Il2rg-knockout (NSG) mouse. The tumor tissues were harvested and transplanted in nude mice to establish mouse models bearing patient-derived xenograft (PDX) of cholangiocarcinoma expressing FGFR2-CCDC6 fusion protein. The PDX mouse models were divided into 4 groups for treatment with citrate buffer (control group), intragastric administration of 20 mg/kg ponatinib dissolved in citrate buffer (ponatinib group), weekly intraperitoneal injections of 50 mg/kg gemcitabine and 2.5 mg/ kg cisplatin (gemcitabine group), or ponatinib combined with gemcitabine and cisplatin at the same doses (10 mice in each group, and 9 mice were evaluated in ponatinib group). The expressions of p-FGFR, p-FRS2, p-AKT, p-ERK, CD31, and Ki-67 in the xenografts were evaluated with immunohistochemistry, and cell apoptosis was analyzed with cleaved caspase-3 (CC3) staining and TUNEL staining. Western blotting was used to detect the expressions of FGFR2, p-FGFR, AKT, p-AKT, ERK, p-ERK, FRS2 and p-FRS2 in the tumor tissues.@*RESULTS@#Compared with those in the control group, the mice in ponatinib group showed a significantly reduced tumor volume (@*CONCLUSIONS@#Ponatinib can regulate FGFR signaling to inhibit the proliferation and induce apoptosis of tumor cells in mice bearing patient-derived cholangiocarcinoma xenograft with FGFR2 fusion. FGFR inhibitor can serve as a treatment option for patients with cholangiocarcinoma with FGFR2 fusion.


Subject(s)
Animals , Bile Duct Neoplasms/genetics , Cell Line, Tumor , Cell Proliferation , Cholangiocarcinoma/genetics , Cytoskeletal Proteins , Heterografts , Humans , Imidazoles , Mice , Mice, Inbred NOD , Mice, Nude , Mice, SCID , Pyridazines , Receptor, Fibroblast Growth Factor, Type 2 , Xenograft Model Antitumor Assays
3.
J. bras. patol. med. lab ; 50(3): 216-220, May-Jun/2014. tab, graf
Article in English | LILACS | ID: lil-715616

ABSTRACT

Introduction: Cholangiocarcinoma is the second most common malignant neoplasm of the hepatobiliary system. During cholangiocarcinogenesis phenotypic changes occur in the ductal epithelium, including the expression of mucins (MUC). However, the evaluating studies of the expression of mucins in the different stages of cholangiocarcinogenesis are scarce. CD56 has also contributed in differentiating benign ductal proliferation and cholangiocarcinoma; however, its expression has not been evaluated in dysplastic epithelium of the bile duct yet. Objective: To assess immunohistochemical profile of (MUC) 1, 2, 5, 6, and CD56 in cholangiocarcinoma, pre-neoplastic and reactive lesions in the epithelium of intrahepatic bile ducts. Material and methods: Immunohistochemical expression of MUC 1, 2, 5, 6, and CD56 were studied for 11 cases of cholangiocarcinoma and 83 intrahepatic bile ducts (67 reactive and 16 dysplastic). Variables were considered significant when p < 0.05. Results: The expression of MUC1 occurred in about 90% of the cholangiocarcinomas, contrasting with the low frequency of positive cases in reactive and dysplastic bile ducts (p < 0.001). However, there was no statistically significant difference in the expression of MUC5, MUC6 and CD56 between the reactive or dysplastic lesions and cholangiocarcinoma. The anti-MUC2 antibody was negative in all cases. Conclusions: MUC1 contributed for the differential diagnosis between cholangiocarcinoma and pre-neoplastic and reactive/regenerative lesions of intrahepatic bile ducts, and it should compose the antibodies panel aiming at improvement of these differential diagnoses. In contrast, MUC2, MUC5, MUC6 and CD56 were not promising in differentiating all the phases of cholangiocarcinogenesis...


Introdução: O colangiocarcinoma é a segunda neoplasia maligna mais comum do sistema hepatobiliar. Durante a colangiocarcinogênese podem ocorrer alterações fenotípicas do epitélio ductal, incluindo a expressão de mucinas. Entretanto, os estudos que avaliam a expressão das mucinas nas diferentes etapas da colangiocarcinogênese são escassos. O CD56, apesar de contribuir na diferenciação entre as proliferações ductais benignas e o colangiocarcinoma, ainda não teve a sua expressão avaliada no epitélio displásico dos ductos biliares. Objetivos: Analisar o perfil das mucinas (MUC) 1, 2, 5, 6 e do CD56 no colangiocarcinoma, nas lesões pré-neoplásicas e reacionais de ductos biliares intra-hepáticos. Material e métodos: A expressão imuno-histoquímica da MUC 1, 2, 5, 6 e do CD56 foram avaliadas em 11 colangiocarcinomas e 83 ductos biliares intra-hepáticos (67 reativos e 16 displásicos). As variáveis foram consideradas como significativas quando p < 0,05. Resultados: A expressão da MUC1 ocorreu em cerca de 90% dos colangiocarcinomas, contrastando com a baixa frequência de casos positivos nos ductos biliares reativos ou displásicos (p < 0,001). Não houve diferença estatisticamente significativa na expressão de MUC5, MUC6 e CD56 entre as lesões reativas, displásicas e o colangiocarcinoma. O anticorpo anti-MUC2 foi negativo em todos os casos. Conclusão: A MUC1 contribuiu no diagnóstico diferencial entre o colangiocarcinoma e as lesões pré-neoplásicas e reacionais/regenerativas dos ductos biliares intra-hepáticos, e deve compor o painel de anticorpos a ser empregado visando o aprimorando destes diagnósticos diferenciais. Contrariamente, a MUC2, MUC5, MUC6 e o CD56 não se mostraram promissoras na diferen...


Subject(s)
Humans , /genetics , Cholangiocarcinoma/genetics , Mucins/genetics , Bile Ducts, Intrahepatic/pathology , Immunohistochemistry , Bile Duct Neoplasms/genetics
4.
Braz. j. med. biol. res ; 44(5): 483-488, May 2011. ilus
Article in English | LILACS | ID: lil-586515

ABSTRACT

Cholangiocarcinomas (CCs) are malignant tumors that originate from epithelial cells lining the biliary tree and gallbladder. Ras correlative C3 creotoxin substrate 1 (Rac1), a small guanosine triphosphatase, is a critical mediator of various aspects of endothelial cell functions. The objective of the present investigation was to study the effect of blocking Rac1 expression in CCs. Seventy-four extrahepatic CC (ECC) specimens and matched adjacent normal mucosa were obtained from the Department of Pathology, Inner Mongolia Medicine Hospital, between 2007 and 2009. Our results showed that the expression of Rac1 was significantly higher (53.12 percent) in tumor tissues than in normal tissues. Western blotting data indicated a significant reduction in Rac1-miRNA cell protein levels. Rac1-miRNA cell growth rate was significantly different at 24, 48, and 72 h after transfection. Flow cytometry analysis showed that Rac1-miRNA cells undergo apoptosis more effectively than control QBC939 cells. Blocking Rac1 expression by RNAi effectively inhibits the growth of CCs. miRNA silencing of the Rac1 gene suppresses proliferation and induces apoptosis of QBC939 cells. These results suggest that Rac1 may be a new gene therapy target for CC. Blocking Rac1 expression in CC cells induces apoptosis of these tumor cells and may thus represent a new therapeutic approach.


Subject(s)
Humans , Apoptosis , Cholangiocarcinoma/metabolism , Gene Silencing , RNA, Small Interfering/metabolism , rac1 GTP-Binding Protein/metabolism , Cell Line, Tumor , Cell Proliferation , Cholangiocarcinoma/genetics , Cholangiocarcinoma/pathology , Flow Cytometry , Immunohistochemistry , RNA, Small Interfering/genetics , Transfection , rac1 GTP-Binding Protein/genetics
5.
Article in English | WPRIM | ID: wpr-103078

ABSTRACT

Cholangiocarcinoma (CC) is an intrahepatic bile duct carcinoma with a high mortality rate and a poor prognosis. Sarcomatous change/epithelial mesenchymal transition (EMT) of CC frequently leads to aggressive intrahepatic spread and metastasis. The aim of this study was to identify the genetic alterations and gene expression pattern that might be associated with the sarcomatous change in CC. Previously, we established 4 human CC cell lines (SCK, JCK1, Cho-CK, and Choi-CK). In the present study, we characterized a typical sarcomatoid phenotype of SCK, and classified the other cell lines according to tumor cell differentiation (a poorly differentiated JCK, a moderately differentiated Cho-CK, and a well differentiated Choi-CK cells), both morphologically and immunocytologically. We further analyzed the genetic alterations of two tumor suppressor genes (p53 and FHIT) and the expression of Fas/FasL gene, well known CC-related receptor and its ligand, in these four CC cell lines. The deletion mutation of p53 was found in the sarcomatoid SCK cells. These cells expressed much less Fas/FasL mRNAs than did the other ordinary CC cells. We further characterize the gene expression pattern that is involved in the sarcomatous progression of CC, using cDNA microarrays that contained 18,688 genes. Comparison of the expression patterns between the sarcomatoid SCK cells and the differentiated Choi-CK cells enabled us to identify 260 genes and 247 genes that were significantly over-expressed and under-expressed, respectively. Northern blotting of the 14 randomly selected genes verified the microarray data, including the differential expressions of the LGALS1, TGFBI, CES1, LDHB, UCHL1, ASPH, VDAC1, VIL2, CCND2, S100P, CALB1, MAL2, GPX1, and ANXA8 mRNAs. Immunohistochemistry also revealed in part the differential expressions of these gene proteins. These results suggest that those genetic and gene expression alterations may be relevant to the sarcomatous change/EMT in CC cells.


Subject(s)
Acid Anhydride Hydrolases/genetics , Animals , Cell Line, Tumor , Cholangiocarcinoma/genetics , Female , Gene Expression Profiling , Humans , Mice , Mice, Inbred BALB C , Mutation , Neoplasm Proteins/genetics , Oligonucleotide Array Sequence Analysis , Sarcoma/genetics , Tumor Suppressor Protein p53/genetics
6.
Article in English | IMSEAR | ID: sea-37776

ABSTRACT

BACKGROUND/AIMS: KRAS oncogene and TP53 tumor suppressor gene have been known as common genes involving in many cancers including cholangiocarcinoma (CCC). Activation of these genes could lead to uncontrolled proliferation and cancer ultimately. The aim of this study was to investigate mutation of KRAS exon 1 and TP53 exon 5-8 in Opisthorchis viverrini (OV)-induced cholangiocarcinoma (CCA) in a hamster model. METHODS: Twenty-seven CCAs were obtained from Syrian golden hamsters induced by OV infection and N-nitrosodimethylnitrosamine (N-NDDM) administration. The tumor tissues were processed for histopathology. Genomic DNA extracted from paraffin sections by microdissection was amplified for KRAS exon 1 and TP53 exon 5-8 mutations by PCR-direct sequencing. RESULTS: Histopathologically, the tumors were classified into tubular (81.5%, 22/27), papillary (3.7%, 1/27), mucinous (3.7%, 1/27) and mixed types (11.1%, 3/27). Of the 27 CCAs, PCR-direct sequencing of KRAS showed G[see text]A transition at codon 37 exon 1 in one CCA sample (3.70%). Point mutations of p53 exon 6 (G[see text]C transversion at codon 119 and 218 and A[see text]C transversion at codon 217) were found in 3 CCA samples (11.1%). CONCLUSIONS: The results suggest that mutation of TP53 particularly at exon 6 may be involved in cholangiocarcinogenesis and a novel mutation of KRAS exon 1 was firstly reported in OV-induced hamster CCA.


Subject(s)
Adenocarcinoma/genetics , Adenocarcinoma, Mucinous/genetics , Animals , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic/parasitology , Carcinoma, Papillary/genetics , Cholangiocarcinoma/genetics , Cricetinae , DNA Primers/chemistry , Exons/genetics , Genes, ras/genetics , Male , Mesocricetus , Mutation/genetics , Opisthorchiasis/genetics , Opisthorchis/pathogenicity , Polymerase Chain Reaction , Tumor Suppressor Protein p53/genetics
7.
Article in Korean | WPRIM | ID: wpr-50299

ABSTRACT

BACKGROUND/AIMS: Intrahepatic cholangiocarcinoma is the second most common intrahepatic neoplasm. Carcinogenesis is believed to be a multistage process that occurs as a result of mutations in oncogenes and tumor suppressor genes. Loss of heterozygosity (LOH) is the phenotype of genetic instability which has been used as a tool for detecting genetic phenotype alterations. Large number of the molecular alterations have been described in human cancer. Among them, that of p53 is quite common. The aim of this study was to determine the frequency of LOH at chromosome 17p related with p53. METHODS: Twenty cases who underwent hepatic resection due to intrahepatic cholangiocarcinoma, were included. LOH was analysed with four microsatellite markers by PCR. For the clinicopathologic parameters, tumor size, differentiation, and metastasis were evaluated. RESULTS: Fifteen patients (75%) showed LOH at one of the loci at the least. Five patients were LOH-high and 10 were LOH-low. The highest frequency of LOH was observed at D17S5 by 38.9%. Those of TP53, D17S796 and D17S513 were 29.4%, 21.4% and 35.3%, respectively. In addition, LOH tended to be more frequent when the tumor is mass forming type, poorly differentiated, or has tumor emboli or vascular invasion. CONCLUSIONS: This study showed that LOH was positive in 75% on chromosome 17p in intrahepatic cholangiocarcinoma which was relatively frequent at D17S5.


Subject(s)
Aged , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic , Cholangiocarcinoma/genetics , Chromosomes, Human, Pair 17 , Female , Gene Frequency , Genes, p53 , Genetic Markers , Humans , Loss of Heterozygosity , Male , Middle Aged
8.
Article in English | WPRIM | ID: wpr-60323

ABSTRACT

Intrahepatic cholangiocarcinoma (ICC), a malignant neoplasm of the biliary epithelium, is usually fatal because of difficulty in early diagnosis and lack of availability of effective therapy. The genetic mechanisms involved in the development of ICC are not well understood and only a few cytogenetic studies of ICC have been published. Recently, technique of degenerate oligonucleotide primed (DOP)-PCR comparative genomic hybridization (CGH) permits genetic imbalances screening of the entire genome using only small amounts of tumor DNA. In this study chromosomal aberrations in 33 Korean ICC were investigated by DOP-PCR CGH. The common sites of copy number increases were 20q (67%), 17 (61%), 11q11-q13 (42%), 8p12-qter (39%), 18p (39%), 15q22-qter (36%), 16p (36%), 6p21 (30%), 3q25-qter (27%), 1q41-qter (24%), and 5p14-q11.2 (24%). DNA amplification was identified in 16 carcinomas (48%). The frequent sites of amplification were 20q, 17p, 17q23-qter, and 7p. The most frequent sites of copy number decreases were 1p32-pter (21%) and 4q (21%). The recurrent chromosomal aberrations identified in this study provide candidate regions involved in the tumorigenesis and progression of ICC.


Subject(s)
Adult , Aged , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic , Cholangiocarcinoma/genetics , Chromosome Aberrations , DNA Primers , Female , Gene Dosage , Humans , Male , Middle Aged , Nucleic Acid Hybridization/methods , Oligonucleotides , Polymerase Chain Reaction/methods
9.
Asian Pac J Allergy Immunol ; 2000 Sep; 18(3): 173-6
Article in English | IMSEAR | ID: sea-36955

ABSTRACT

Cholangiocarcinoma (CCA) constitutes carcinoma of the bile duct found at a high prevalence in northeastern Thailand. In the present study, we examined the sera of altogether 82 Thai CCA patients for the presence of anti-p53 antibodies in order to investigate a role of the tumor suppressor gene, p53 in the carcinogenesis. Our results revealed anti-p53 antibodies in 7.3% of the cases tested, which conforms to the prevalence rate of p53 gene mutation recently reported at 5% among Thai patients. With limited number of the patients, anti-p53 antibodies were rapidly detected more frequently among patients with peripheral tumors than those with central tumors. However, further studies is required to establish significance and prognostic value of the antibodies in the context of CCA.


Subject(s)
Adult , Aged , Analysis of Variance , Antibodies, Antinuclear/blood , Antibodies, Neoplasm/blood , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic , Cholangiocarcinoma/genetics , Enzyme-Linked Immunosorbent Assay , Female , Genes, p53 , Humans , Male , Middle Aged , Mutation , Prevalence , Prognosis , Retrospective Studies , Serum Albumin/metabolism , Survival Analysis , Tumor Suppressor Protein p53/genetics , alpha-Fetoproteins/metabolism
10.
Article in English | IMSEAR | ID: sea-40215

ABSTRACT

In the study, to analyse a K-ras oncogene mutated at codon 12 in 24 patients with cholangiocarcinomas, four (16.67%) of them contained this point mutation. One of 4 was peripheral and the others were hilar tumors. There was no significant relationship between mutation and clinical features in terms of age, sex, endemic area, tumor location, tumor grading and pathological features. In our study, the incidence of K-ras codon 12 mutation in Thai patients with cholangiocarcinoma was lower than that found in British and Japanese patients. The discrepancy of incidence and type of the mutations, in different races and environment probably indicates that there is/are different etiologic mechanism(s) in the pathogenesis of cholangiocarcinoma.


Subject(s)
Adult , Aged , Aged, 80 and over , Base Sequence , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic , Cholangiocarcinoma/genetics , Codon/genetics , Confidence Intervals , DNA, Neoplasm/analysis , Female , Genes, ras/genetics , Humans , Male , Middle Aged , Molecular Sequence Data , Point Mutation/genetics , Polymerase Chain Reaction , Sensitivity and Specificity
11.
Article in English | IMSEAR | ID: sea-32390

ABSTRACT

Paraffin embedded tissues from twenty Thai patients with intrahepatic cholangiocarcinomas were studied for K-ras gene mutations at codon 12, 13 and 61 and for p53 gene mutations in exon 5 to 8 using polymerase chain reaction and thermal cycle sequencing. Results showed that point mutations at these regions in K-ras oncogene were not present in all the samples. One case harbored a p53 gene mutation in codon 282 in exon 8, CGG (arginine) to TGG (tryptophan), but the mutation was not found in other patient's tissues with similar histological features.


Subject(s)
Adult , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic/pathology , Cholangiocarcinoma/genetics , Female , Genes, p53/genetics , Genes, ras/genetics , Humans , Male , Middle Aged , Mutation , Polymerase Chain Reaction , Thailand
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