Molecular heterogeneity in intrahepatic cholangiocarcinoma

Intrahepatic cholangiocarcinoma (iCCA) is a heterogeneous primary liver cancer, and currently there exist only a few options of targeted therapy. Histopathologically, iCCA is sub-classified according to morphology (mass forming type, periductal infiltrating type, and intraductal growing type) and histology (small duct type and large duct type). According to different histopathological types, clinical features such as risk factors and prognosis vary. Recent developments in genomic profiling have revealed several molecular markers for poor prognosis and activation of oncogenic pathways. Exploration of molecular characteristics of iCCA in each patient is a major challenge in a clinical setting, and there is no effective molecular-based targeted therapy. However, several recent studies suggested molecular-based subtypes with corresponding clinical and pathological features. Even though the subtypes have not yet been validated, it is possible that molecular features can be predicted based on clinicopathological characteristics and that this could be used for a more rational approach to integrative clinical and molecular subclassification and targeted therapy. In this review, we explored the genomic landscape of iCCA and attempted to find relevance between clinicopathologic and molecular features in molecular subtypes in several published studies. The results reveal future directions that may lead to a rational approach to the targeted therapy.     

胆管癌浸润转移途径及其机制

胆管癌的浸润和转移具有其自身的特点,经淋巴转移是胆管癌的主要转移方式之一,以局部淋巴结转移为主,较少发生远处转移.由于胆管的解剖部位的特点以及其与周围神经、包括多种神经肽的密切关系,其神经周围浸润转移途径近年来越来越受到人们的重视.除浸润转移、种植转移、淋巴道及血道转移等途径之外,经神经周围浸润和转移是胆管癌的一种重要病理特征,对胆管癌细胞与神经纤维的相互作用机制以及胆管癌神经周围浸润转移过程的调控因子的研究是近年的研究热点之一.     

Molecular mechanism of cholangiocarcinoma carcinogenesis

Cholangiocarcinoma (CCA) is a highly malignant cancer of the biliary tract with a poor prognosis, which often arises from conditions causing long‐term inflammation, injury, and reparative biliary epithelial cell proliferation. Several conditions are known to be major risk factors for cancer in the biliary tract or gallbladder, including primary sclerosing cholangitis, liver fluke infection, pancreaticobiliary maljunction, and chemical exposure in proof‐printing workers. Abnormalities in various signaling cascades, molecules, and genetic mutations are involved in the pathogenesis of CCA. CCA is characterized by a series of highly recurrent mutations in genes, including KRAS, BRF, TP53, Smad, and p16^INK4a. Cytokines that are affected by inflammatory environmental conditions, such as interleukin‐6 (IL‐6), transforming growth factor‐β (TGF‐β), tumor necrosis factor‐α (TNF‐α), and platelet‐derived growth factor (PDGF), play an important role in cancer pathogenesis. Prominent signaling pathways important in carcinogenesis include TGF‐β/Smad, IL‐6/STAT‐3, PI3K/AKT, Wnt, RAF/MEK/MAPK, and Notch. Additionally, some microRNAs regulate targets in critical pathways of CCA development and progression. This review article provides the understanding of the genetic and epigenetic mechanism(s) of carcinogenesis in CCA, which leads to the development of new therapeutic targets for the prevention and treatment of this devastating cancer.     

Strong expression of CD133 is associated with increased cholangiocarcinoma progression

AIM:To determine the role of CD133 in cholangiocarcinoma progression. METHODS:CD133 protein expression was evaluated by immunohistochemistry in 34 cholangiocarcinoma specimens.In addition,proliferation,chemoresistance and invasive properties of CD133-enriched(CD133 + ) and CD133-depleted(CD133 )RMCCA1 cholangiocarcinoma cells were studied and compared. RESULTS:Strong CD133 expression was observed in 67.6%(23/34)of the cholangiocarcinoma specimens. Strong expression of CD133 was significantly associated with...     

Molecular aspects of cholangiocarcinoma

Novel targets for therapeutic or chemopreventive approaches against cholangiocarcinoma (CCA) are urgently needed. In this review article, we discuss the molecular aspects of CCA including the role of erbB receptor tyrosine kinases (RTKs), downstream signaling pathways of these erbB RTKs, inflammatory mediators during gallbladder carcinogenesis and bile acids based on our study using a mouse model for human CCA (BK5.erbB2 mice) as well as additional information in the literature.     

WWOX基因转染对胆管癌细胞增殖、凋亡及侵袭的影响

目的:探讨WWOX基因转染胆管癌细胞株QBC939后对其增殖、凋亡与侵袭性的影响.方法:用脂质体转染法将WWOX重组真核表达质粒转染QBC939细胞,建立稳定表达WWOX基因的细胞株.将其分为以下3组:QBC939组,QBC939/con组和QBC939/WWOX组.荧光定量RT-PCR和Western blot法检测...     

胰岛β细胞凋亡的分子机制

胰岛β细胞凋亡在糖尿病的发病中扮演重要角色,1、2型糖尿病β细胞凋亡的分子机制有所不同。在1型糖尿病中,胰岛β细胞主要通过死亡受体介导的信号转导途径及颗粒酶B途径发生凋亡,而在2型糖尿病中,线粒体途径是胰岛β细胞凋亡的主要信号转导途径。多种细胞因子通过激活核转录因子调节相应基因表达,进而调控胰岛β细胞的凋亡。     

三苯氧胺抗胆管癌机制的研究进展

胆管癌(cholangiocarcinoma,CCA)是一种来源于肝内或肝外胆管上皮细胞的恶性肿瘤.早期诊断困难,临床疗效差和5年生存率低等特点使之成为临床重点攻坚肿瘤.因此,需要研究和开发新型有效的化疗药物.三苯氧胺(tamoxifen,TAM)是一种非甾体类抗雌激素药物,临床上主要用于乳腺癌的治疗,对其他肿瘤包括CCA也有抑制活性,但具体机制尚不完全清楚.本文主要阐述TAM抗CCA机制的研究进展.     

Establishment and characterization of a cholangiocarcinoma cell line （RMCCA-1） from a Thai patient

INTRODUCTIONCholangiocarcinoma is a highly malignant epithelial neoplasm that arises within the intrahepatic and extrahepatic biliary tract[1].The pathogenesis of this disease has been strongly associated with chronic inflammation and cellular injury with…     

Proteasome inhibition-induces endoplasmic reticulum dysfunction and cell death of human cholangiocarcinoma cells

AIM： To determine if proteasome inhibition induces apoptosis in human cholangiocarcinoma cells, and if so, to elucidate the cellular mechanisms.
METHODS： Studies were performed in the human KMCH, KMBC, and Mz-ChA-1 cholangiocarcinoma, and normal rat cell lines. MG132, a peptide aldehyde, which inhibits the chymotrypsin-like activity of the proteaosome was employed for this study. Apoptosis was assessed morphologically by 4＇-6-Diamidino-2-phenylindole （DAPI） nuclear staining and fluorescence microscopy. Mitochondrial membrane potential was examined using a fluorescent unquenching assay. Ultrastructural changes during cell death were examined using transmission electron microscopy （TEM）. Caspase 3/7 activity was assessed using an enzymatic-based fluorescent assay. Cytosolic-free calcium concentrations were measured using Fura-2 and digitized fluorescent microscopy.
RESULTS： MG132, a proteasome inhibitor, induced apoptosis in all the cholangiocarcinoma cell lines examined. In contrast, minimal cytotoxicity was observed in normal rat cholangiocytes. Apoptosis was time- and -concentration-dependent. There was no change in the mitochondrial membrane potential between treated and untreated cells. Ultrastructural examination by transmission electron microscopy displayed the classic features of apoptosis, but in addition, there was also dramatic vacuolization of the endoplasmic reticulum （ER）. Unexpectedly, no increase in caspase 3/7 activity was observed in MG132 treated cells, nor did the pancaspase inhibitor, Q-VD-OPh prevent cell death. The protein synthesis inhibitor, cycloheximide, blocked apoptosis induced by proteosome inhibitor indicating that ER dysfunction was dependent upon the formation of new proteins.
CONCLUSION： Proteosome inhibition induces ER dysfunction and caspase-independent cell death selectively in human cholangiocarcinoma cells. Proteasome inhibitors warrant evaluation as anticancer agents for the treatment of human cholangiocarcinoma.     

基于GEO数据库对胆管癌潜在分子靶点的筛选及生物信息学分析

背景胆管癌恶性程度高,预后差.靶向治疗是胆管癌的重要研究方向,探索新的分子靶点对于胆管癌靶向治疗至关重要.目的用生物信息学分析方法挖掘胆管癌的枢纽基因,为胆管癌的靶向治疗提供潜在分子靶点.方法从GEO数据库中下载2组胆管癌表达谱芯片数据,采用GEO2R在线分析工具筛选胆管癌肿瘤组织与正常组织差异表达基因,对差异表达基因作GO富集分析、KEGG通路分析、蛋白质相互作用网络分析,利用Cytoscape软件筛选枢纽基因.使用GEPIA数据库对枢纽基因在胆管癌组织中的表达量进行验证.结果共得到共同差异表达基因158个.GO富集分析结果显示,差异基因主要参与细胞对锌离子反应、细胞增殖与粘附、代谢以及蛋白质聚合等生物学过程,主要存在外泌体、胞外区、弹性纤维等区域,主要分子功能与结合肝素、半胱氨酸型内肽酶抑制剂活性、蛋白质同源二聚化、受体结合及磷酸吡哆醛结合等相关.KEGG通路分析结果显示,差异基因主要参与矿物质吸收、代谢、PPAR信号通路及脂肪酸降解等过程.基于String数据库构建蛋白质相互作用网络图,Cytoscape软件CytoHubba插件筛选枢纽基因,皆为上调基因.GEPIA数据库验证枢纽基因在胆管癌组织中表达量显著高于正常组织.结论本研究获取了8个与胆管癌相关的枢纽基因,分别是NUSAP1,TOP2A,RAD51AP1,MCM4,KIAA0101,CDCA5,TYMS,ZWINT.这些基因为深入研究胆管癌的靶向治疗提供了新思路,有望成为新的分子治疗靶点.     

Molecular pathogenesis of intrahepatic cholangiocarcinoma

Cholangiocarcinoma (CCA) is an orphan cancer of the hepatobiliary tract, the incidence of which has increased in the past decade. The molecular pathogenesis of this treatment‐refractory disease is poorly understood. Desmoplasia is a key causal feature of CCA; however, a majority of tumors develop with no apparent etiological background. The impact of the stromal compartment on tumor progression as well as resistance to therapy is in vogue, and the epithelial‐stromal crosstalk may present a target for novel treatment strategies. As such, the complexity of tumor cellularity and the molecular mechanisms underlying the diversity of growth patterns of this malignancy remain a clinical concern. It is crucial to advance our present understanding of the molecular pathogenesis of CCA to improve current clinical strategies and patient outcome. This will facilitate the delineation of patient subsets and individualization for precision therapies. Many questions persevere as to the evolutionary process and cellular origin of the initial transforming event, the context of intratumoral plasticity and the causal driver action. Next‐generation sequencing has begun to underline the persistent alterations, which may be the trigger of acquired drug resistance, and the cause of metastasis and disease recurrence. A complex issue that remains is to account for the heterogeneous pool of “backseat” aberrations, which in chromosomal proximity to the causative variant are likely to influence, for example, drug response. This review explores the recent advances in defining the molecular pathways implicated in the development of this devastating disease and, which present putative clinical strategies.     

Epidemiology, risk factors, and pathogenesis of cholangiocarcinoma

Cholangiocarcinoma (CCA) is a fatal cancer of the biliary epithelium, arising either within the liver (intrahepatic, ICC) or in the extrahepatic bile ducts (extrahepatic ECC). Globally, CCA is the second most common primary hepatic malignancy. Several recent epidemiological studies have shown that the incidence and mortality rates of ICC are increasing. This review of the literature on the international epidemiological rates of CCA, both intra- and extrahepatic, explores possible explanations for the trends found. The possible role of epidemiological artifact in the findings is discussed and the known risk factors for CCA are summarized. These include primary sclerosing cholangitis, liver fluke infestation, congenital fibropolycystic liver, bile duct adenomas, and biliary papillomatosis, hepatolithiasis, chemical carcinogens such as nitrosamines, Thorotrast, chronic viral hepatitis, cirrhosis, chronic non-alcoholic liver disease and obesity. Potential pathways involved in the molecular pathogenesis of CCA are also summarized.     

Effect of mutant p27kip1 gene on human cholangiocarcinoma cell line, QBC939

AIM:To investigate the effects of exogenously mutated p27kip1 (p27) on proliferation and apoptosis of human cholangiocarcinoma cell line,QBC939 in vivo.METHODS:Adenviral vectors were used to transfect mutated p27 cDNA into human QBC939 cell line.Expression of p27 was detected by RT-PCR.Western blot.Cell growth,morphological change,cell cycle,apoptosis and cloning formation were determined by MTT assay and flow cytometry.RESULTS:The expression of p27 protein and mRNA was increased significantly in QBC939 cell line transfected with Ad-p27mt.The transfer of Adp27mt could significantly inhibit the growth of QBC939cells,decrease the cloning formation rate and induce apoptosis,p27 over expression caused cell cycle arrest at G0/G1 phase 72 h after infection with Adp27mt.CONCLUSION:p27 may cause cell cycle arrest at G0/G1 phase and subsequently lead to apoptosis.Recombinant adenovirus expressing mutant p27 may be potentially useful in gene therapy for cholangiocarcinoma.     

MUC1 and MUC5AC mucin expression in liver fluke-associated intrahepatic cholangiocarcinoma

AIM: To investigate the expressions of MUC1 and MUC5AC in intrahepatic cholangiocarcinoma (ICC). Association of expressions of mucins MUC1 and MUC5AC with clinical findings, metastasis, and survival of the liver fluke-associated ICC patients was determined. METHODS: The expressions of MUC1 and MUC5AC mucins were examined by immunohistochemical staining in 87 cases of histologically-proven ICC. The expressions of mucins in relationship between clinicopathological significance and prognosis of the patients were evaluated. RESULTS: Fifty-two patients (60%) exhibited both MUC1 and MUC5AC expressions, whereas 31% expressed either MUC1 or MUC5AC, and 9% expressed neither. High MUC1 immunoreactivity displayed a significant correlation with tumor progression as reflected by vascular invasion (P<0.01), whereas high expression of MUC5AC significantly correlated with neural invasion (P = 0.022) and advanced ICC stage (P= 0.008). Patients with high expression of MUC1 had a significantly shorter survival (P = 0.0002). According to multivariate analyses, MUC1 reactivity (P = 0.026), histological grading and stage of tumor represented the least probability of survival. CONCLUSION: MUC1 is overexpressed in liver fluke-associated cholangiocarcinoma and relates to vascular invasion and poor prognosis, whereas MUC5AC mucin is neoexpressed and relates to neural invasion and advanced ICC stage. High MUC1 expression in tumor may be useful for predicting the poor outcome of ICC patients.     

Staging cholangiocarcinoma by cholangioscopy

Peroral cholangioscopy (POCS) and percutaneous transhepatic cholangioscopy (PTCS) were first developed in the 1970s, and technical developments and clinical applications have taken place gradually ever since. POCS is used to diagnose small mucosal biliary lesions in non-icteric patients and early malignant changes in patients with persistent primary sclerosing cholangitis (PSC). Although PTCS is a more invasive diagnostic procedure than POCS, it has the advantage of precise diagnosis with mapping biopsy in defining the proximal and distal extension of superficially spreading cholangiocarcinoma (CCA) or mucin-producing CCA, which is predominantly found in papillary type CCA. POCS is significantly superior to ERCP in distinguishing between malignant and benign dominant bile duct stenoses in patients with PSC. The positive rate of PTCS biopsy for CCA is 96%, while morbidity and mortality of PTCS are 9% and 0%, respectively. Although magnetic resonance (MR) cholangiography may replace PTCS in determining the longitudinal spread of infiltrating type hilar CCA, the accuracy of MR cholangiography in papillary type hilar CCA is significantly lower than that of PTCS.     

Involvement of PI3K and ERK1/2 pathways in hepatocyte growth factor-induced cholangiocarcinoma cell invasion

AIM:To investigate the role of hepatocyte growth factor(HGF) in cholangiocarcinoma(CCA) cell invasiveness and the mechanisms underlying such cellular responses. METHODS:Effects of HGF on cell invasion and motility were investigated in two human CCA cell lines,HuCCA-1 and KKU-M213,using Transwell in vitro assay.Levels of proteins of interest and their phosphorylated forms were determined by Western blotting.Localization of E-cadherin was analyzed by immunofluorescence staining and visualized under confocal m...     

Effect of mutant p27(kipl) gene on human cholangiocarcinoma cell line, QBC(939)

AIM：To investigate the effects of exogenously mutated p27^kip1 （p27） on proliferation and apoptosis of human cholangiocarcinoma cell line, QBC939 in vivo.METHODS： Adenviral vectors were used to transfect mutated p27 cDNA into human QBC939 cell line. Expression of p27 was detected by RT-PCR. Western blot. Cell growth, morphological change, cell cycle, apoptosis and cloning formation were determined by MTT assay and flow cytometry.RESULTS： The expression of p27 protein and mRNA was increased signifi cantly in QBC939 cell line transfected with Ad-p27mt. The transfer of Ad-p27mt could signifi cantly inhibit the growth of QBC939 cells, decrease the cloning formation rate and induce apoptosis. p27 over expression caused cell cycle arrest at G0/G1 phase 72 h after infection with Ad-p27mt.CONCLUSION： p27 may cause cell cycle arrest at G0/G1 phase and subsequently lead to apoptosis. Recombinant adenovirus expressing mutant p27 may be potentially useful in gene therapy for cholangiocarcinoma.     

Molecular signaling mechanisms of apoptosis in hereditary non-polyposis colorectal cancer

Colorectal cancer is the second most leading cause of cancer related deaths in the western countries. One of the forms of colorectal cancer is hereditary non-polyposis colorectal cancer (HNPCC), also known as "Lynch syndrome". It is the most common hereditary form of cancer accounting for 5%-10% of all colon cancers. HNPCC is a dominant autosomal genetic disorder caused by germ line mutations in mismatch repair genes. Human mismatch repair genes play a crucial role in genetic stability of DNA, the inactivation of which results in an increased rate of mutation and often a loss of mismatch repair function. Recent studies have shown that certain mismatch repair genes are involved in the regulation of key cellular processes including apoptosis. Thus, differential expression of mismatch repair genes particularly the contributions of MLH1 and MSH2 play important roles in therapeutic resistance to certain cytotoxic drugs such as cisplatin that is used normally as chemoprevention. An understanding of the role of mismatch repair genes in molecular signaling mechanism of apoptosis and its involvement in HNPCC needs attention for further work into this important area of cancer research, and this review article is intended to accomplish that goal of linkage of apoptosis with HNPCC. The current review was not intended to provide a comprehensive enumeration of the entire body of literature in the area of HNPCC or mismatch repair system or apoptosis; it is rather intended to focus primarily on the current state of knowledge of the role of mismatch repair proteins in molecular signaling mechanism of apoptosis as it relates to understanding of HNPCC.