Stage IV perihilar cholangiocarcinoma with loss of expression of MSH2 and MSH6: hereditary cancer syndrome?

We present the case of a 69-year-old male diagnosed with stage IV perihilar cholangiocarcinoma with loss of expression of MSH2 and MSH6 proteins, but somatic wild type MSH2 and MSH6 genes with Oncomine Comprehensive Assay (OCA) genomic sequencing panel. In his cancer family history, there was a maternal aunt with sigmoid colon adenocarcinoma also missing MSH2 and MSH6 protein expression. Subsequently, we will discuss whether or not we are facing a hereditary cancer syndrome.

KLF4 suppresses the proliferation of perihilar cholangiocarcinoma by negatively regulating GDF15 and phosphorylating AKT.

Krüppel­like factor 4 (KLF4) is a transcription factor which functions as a tumor suppressor or an oncogene in numerous types of solid tumors. However, its expression levels and function in perihilar cholangiocarcinoma (pCCA) have yet to be elucidated. In the present study, in order to investigate its roles in pCCA, reverse transcription­quantitative PCR (RT­qPCR), western blot analysis and immunohistochemistry were used to detect KLF4 expression in pCCA. The Chi­squared test was used to analyze the associations between KLF4 and the clinicopathological features of patients with pCCA. Univariate and multivariate analyses were subsequently used to analyze the prognostic significance of KLF4. The tumor suppression of KLF4 was investigated for the purposes of illustrating its biological function both in vitro and in vivo. Furthermore, the association between KLF4 and growth/differentiation factor 15 (GDF15) was determined using pCCA tissue microarray (TMA) analysis and RT­qPCR. The underlying molecular mechanisms between KLF4 and GDF15 were subsequently investigated in vitro. In pCCA tissues, KLF4 was found to be downregulated, and this was negatively associated with the histological grade and tumor size. The knockdown of KLF4 was also found to be a prognostic indicator of the poorer survival of patients with pCCA. Based on in vitro and in vivo analyses, KLF4 was found to suppress tumor progression and induce cell apoptosis. Furthermore, it was found that KLF4 executed its tumor suppressive effects via the regulation of the GDF15/AKT signaling pathway. Taken together, the findings of the present study demonstrate that KLF4 may be considered as an independent biomarker of a favorable prognosis of patients with pCCA, and the KLF4/GDF15/AKT signaling pathway may potentially be a novel molecular therapeutic target for patients with pCCA.

NTCP Change in Rats of Hilar Cholangiocarcinoma and Therapeutic Significance.

Background: The study aims to detect the expression of Na+/taurocholate cotransporter polypeptide in hilar cholangiocarcinoma of rat model, to provide a new therapeutic target for gene therapy of hilar cholangiocarcinoma. Methods: 60 male Wistar rats (weighing 190 ± 8 g) were randomly divided into 3 groups (experimental group, control group, and sham operation group; 20 rats in each group). The 3 groups were fed with standard diet. The QBC939 cell suspension of cholangiocarcinoma was injected into the hilar bile duct in the experimental group with a micro syringe. The control group was injected with normal saline, and the sham operation group was not injected with any drugs. Comprehensive behavior score and Basso Beattie Bresnahan were used to evaluate the mental state and exercise of rats every day. At 5 weeks, one rat in the experimental group was killed, and the changes in hilar bile duct were recorded. The procedure was repeated at one and half months. After one and half months, hilar cholangiocarcinoma only occurred in the experimental group. Pathological examination confirmed the formation of tumor, and hilar bile duct tissues were taken from the 3 groups. Na+/taurocholate cotransporter polypeptide expression in hilar bile duct was detected by real-time polymerase chain reaction and immunohistochemistry. Results: After 2 weeks, the rats in experimental group ate less, and their weight was significantly reduced compared with the other 2 groups. One and half months later, hilar cholangiocarcinoma was detected in 16 rats in the experimental group. The levels of alanine aminotransferase and aspartate transaminase in the experimental group were higher than those in the other 2 groups. The ratio of Na+/taurocholate cotransporter polypeptide/GAPDH mRNA in hilar cholangiocarcinoma, control group, and sham operation group was significantly different. Under the light microscope, Na+/taurocholate cotransporter polypeptide protein reacted with anti-Na+/taurocholate cotransporter polypeptide antibody and showed granular expression. Every pathological section included 4800 cells. 3823 positive cells were in the experimental group, 1765 positive cells were in the control group, and 1823 positive cells were in the sham operation group. Conclusions: Na+/taurocholate cotransporter polypeptide expression in hilar cholangiocarcinoma of rats was significantly higher than normal hilar bile duct tissues, suggesting that drugs targeting Na+/taurocholate cotransporter polypeptide may be a new strategy for the treatment of hilar cholangiocarcinoma.

FXR expression in rats of hilar cholangiocarcinoma.

The study objective was to detect the expression of farnesoid X receptor (FXR) in a rat model of hilar cholangiocarcinoma to provide a new therapeutic target for gene therapy in hilar cholangiocarcinoma. Sixty male Wistar rats (weighing 190 ± 8 g) were randomly divided into three groups (experimental group, control group and sham operation group, 20 rats in each group). The three groups were fed a standard diet. The QBC939 cell suspension of cholangiocarcinoma was injected into the hilar bile duct in the experimental group with a microsyringe. The control group was injected with normal saline, and the sham operation group was not injected with any drugs. A modified tail suspension test (TST) was used to evaluate the mental state and physical activity of rats every day. At 5 weeks, one rat in the experimental group was euthanized, and the changes in the hilar bile duct were recorded. The procedure was repeated at one and half months. After one and half months, hilar cholangiocarcinoma only occurred in the experimental group. Pathological examination confirmed the formation of tumours, and hilar bile duct tissues were taken from the three groups. FXR expression in the hilar bile duct was detected by real-time polymerase chain reaction (RT-PCR) and immunohistochemistry. After two weeks, the rats in the experimental group ate less, and their weight was significantly reduced. One and half months later, hilar cholangiocarcinoma was detected in 16 rats in the experimental group. The levels of alanine aminotransferase and aspartate transaminase in the experimental group were higher than those in the other two groups. The ratio of FXR/GAPDH mRNA was significantly different among the hilar cholangiocarcinoma, control and sham operation groups. Under the light microscope, FXR protein reacted with anti-FXR antibody and showed granular expression. Every pathological section included 4800 cells. A total of 1856 positive cells were in the experimental group, 3279 positive cells were in the control group, and 3371 positive cells were in the sham operation group. FXR expression in the hilar cholangiocarcinoma of rats was significantly lower than that in normal hilar bile duct tissues, suggesting that drugs targeting FXR may be a new strategy for the treatment of hilar cholangiocarcinoma.

FOXK1 plays an oncogenic role in the progression of hilar cholangiocarcinoma.

Hilar cholangiocarcinoma (HC) has a poor outcome in terms of survival. Forkhead box K1 (FOXK1) dysregulation is critical in solid tumors, which serves a pivotal role in the biological characteristics, such as invasion and migration, but its expression and functions in HC are unclear. The present study investigated the clinical significance and biological functions of FOXK1 in HC. Tumor microarrays and immunohistochemistry were used to evaluate FOXK1 in HC and its expression was modulated to determine its effects on chemoresistance and tumorigenesis. FOXK1 was highly expressed in HC and cell lines, which was associated with tumor invasion, regional lymph node metastasis, tumor recurrence and poor prognosis. Silencing FOXK1 in HC cells inhibited invasion and migration, upregulated E-cadherin, and downregulated vimentin, matrix metallopeptidase 9 and Twist in HC cells. Sensitivity to 5-fluorouracil and cisplatin was increased, and glutathione S-transferase π, multidrug resistance mutation 1 and P-glycoprotein expression levels were downregulated in RBE cells in vitro following FOXK1 knockdown. These results indicated that FOXK1 plays an oncogenic role in HC progression and can serve as a novel therapeutic target for HC.

PTPN3 suppresses the proliferation and correlates with favorable prognosis of perihilar cholangiocarcinoma by inhibiting AKT phosphorylation.

BACKGROUND: Perihilar cholangiocarcinoma (PHCCA) is the most common type of human cholangiocarcinoma with a very dismal prognosis. Tumor markers and target drugs of PHCCA are desperately needed. Protein phosphatase N3 (PTPN3) has dual roles in the progression of human cancers, but its expression and functions in PHCCA have not been elucidated. MATERIALS AND METHODS: The expression of PTPN3 in PHCCA was detected with western blotting, qRT-PCR and immunohistochemistry. The clinical significance of PTPN3 was identified by analyzing the correlations between its expression and the clinicopathological variables, and the prognostic value was evaluated by univariate and multivariate analyses. The functions of PTPN3 in the progression of PHCCA were estimated with both in vitro and in vivo experiments. RESULTS: PTPN3 expression was down-regulated in PHCCA compared with normal bile duct. Low PTPN3 expression was markedly associated with large tumor size and unfavorable prognosis. After knocking down PTPN3, the percentages of G2/S phase of PHCCA cells were elevated, and the proliferation increased significantly. Moreover, we demonstrated that the phosphorylation of AKT was elevated by PTPN3 knockdown, and it was required in PTPN3-involved proliferation of PHCCA. Within vivo experiments, PTPN3 and AKT inhibitor MK-2206 were demonstrated to suppress tumor size of PHCCA. CONCLUSION: PTPN3 was a favorable prognostic biomarker of PHCCA. PTPN3 suppressed the proliferation of PHCCA by inhibiting AKT phosphorylation and arresting cell cycle. Our results suggested thatpost-operative detection of PTPN3 would be a helpful approach to stratify the PHCCA patients with high-risk.

Transcription factor 7 promotes the progression of perihilar cholangiocarcinoma by inducing the transcription of c-Myc and FOS-like antigen 1.

BACKGROUND: Perihilar cholangiocarcinoma (PHCC) is the most common type of cholangiocarcinoma with the worst prognosis. Radical resection of PHCC is difficult; thus, few effective biomarkers or useful molecular profiles for PHCC have been reported in recent years. Therefore, in this study, we aimed to assess biomarkers for PHCC. METHODS: We screened potential biomarkers for PHCC using exome and transcriptome sequencing with PHCC tissues and paired normal tissues. Transcription factor 7 (TCF7) expression was evaluated using quantitative reverse transcription polymerase chain reaction, western blotting, and immunohistochemistry. The correlations between TCF7 and clinicopathological factors were analyzed with Chi-square test, and the prognostic significance of TCF7 was evaluated with univariate and multivariate analyses. The functions of TCF7 and its main effectors in PHCC cells were investigated in vitro and in vivo. FINDINGS: TCF7 expression was upregulated in PHCC and was an unfavorable prognostic biomarker. c-Myc was a main effector of TCF7 in PHCC cells and modulated TCF7-induced proliferation, invasion, and migration. FOS-like antigen 1 (FOSL1) was identified as a downstream target of TCF7 and was required in TCF7-induced PHCC proliferation. Triple-positive expression of TCF7, c-Myc, and FOSL1 predicted a much worse prognosis in patients with PHCC than TCF7 expression alone. INTERPRETATION: Postoperative detection of TCF7, c-Myc, and FOSL1 may be useful for stratifying patients with a high risk of unfavorable prognosis, and suppressing TCF7 or its downstream effectors may be a promising strategy for the treatment of PHCC.

Patterns of gene mutations in bile duct cancers: is it time to overcome the anatomical classification?

BACKGROUND: Two recent studies based on multi-omics data analysis identified distinct subtypes of bile-duct cancers (BDC) with important implications in terms of disease classification and patients' treatment. METHODS: Patients with mutations in KRAS, NRAS, TP53, and ARID1A genes were classified in KRAS/TP53 group while patients with mutations in IDH1-2, BAP1, and PBRM1 were classified in IDH1-2/BAP1/PBRM1 group. The aim of this study was to define long-term outcomes among patients stratified by patterns of genes mutated. RESULTS: Among 105 patients who underwent surgical resection for BDCs, 71 (68%) patients were classified in two groups based on patterns of genes mutated. While in IDH1-2/BAP1/PBRM1 group there were 58%, 22%, and 10% of patients with intrahepatic-cholangiocarcinoma (ICC), perihilar-cholangiocarcinoma (PHCC), and gallbladder cancer (GBC), in KRAS/TP53 group there were 42%, 78%, and 90% of patients with ICC, PHCC, and GBC (p = 0.003), respectively. Patients in IDH1-2/BAP1/PBRM1 group had a 5-year OS of 40% compared with 13% for KRAS/TP53 group (p = 0.032). In a multivariable model adjusted for margins, lymph-node status, microvascular invasion, and tumor grade, patients in KRAS/TP53 group had a 2.1-fold increased risk of death compared with patients in IDH1-2/BAP1/PBRM1 group (p = 0.028). CONCLUSIONS: Genetic data were able to overcome the clinical based staging system in predicting patients' prognosis.

Histological and molecular characterization of intrahepatic bile duct cancers suggests an expanded definition of perihilar cholangiocarcinoma.

BACKGROUND: Growing evidence has suggested that intrahepatic cholangiocarcinoma (iCCA) can be classified into small- and large-duct types. The present study aimed to elucidate how large-duct iCCA is similar and dissimilar to perihilar cholangiocarcinoma (pCCA). METHODS: The study cohort consisted of iCCA (n = 58) and pCCA (n = 44). After iCCA tumors were separated into small- (n = 36) and large-duct (n = 22) types based on our histologic criteria, genetic statuses of the three types of neoplasms were compared. Locations of iCCA were plotted on a three-dimensional image and their distances from the portal bifurcation were measured. RESULTS: Large-duct iCCA was distinct from small-duct iCCA in terms of frequency of bile duct reconstruction required, perineural infiltration, and survival, with these features more similar to pCCA. Large-duct iCCA and pCCA more frequently had the loss of SMAD4 expression and MDM2 amplifications than small-duct iCCA, whereas the loss of BAP1 expression and IDH1 mutations were mostly restricted to small-duct iCCA. From imaging analysis, most tumors of large-duct iCCA were present around the second branches of the portal vein. CONCLUSION: Large-duct type iCCA shared the molecular features with pCCA, and it may be reasonable to expand the definition of pCCA to include cancers originating from the second bile duct branches.

The Plasma LncRNA Acting as Fingerprint in Hilar Cholangiocarcinoma.

BACKGROUND & AIMS: Current studies have indicated that long non-coding RNAs (lncRNAs) could act as tumor biomarkers for disease diagnosis and prognosis prediction. In this study, we mainly focused on determining the expression of circulating lncRNAs in patients suffering for hilar cholangiocarcinoma (HC), aiming to reveal the potential lncRNA as a fingerprint. METHODS: A total 12 lncRNAs were previously proven to be aberrantly expressed in HC tumor tissues. All of the 12 lncRNAs were selected as candidate targets for subsequent circulating lncRNA assay. The candidate lncRNAs were validated by qRT-PCR arranged in training and validation sets. The risk score analysis was employed. Data was presented with receiver operating characteristic curve (ROC). RESULTS: Circulating PCAT1, MALAT1, and CPS1-IT1 were significantly increased in plasma samples of HC patients in both the training set and validation set. Through ROC analysis, we found that the three plasmatic lncRNAs presented the area under ROC curve value (AUC) as 0.784, 0.860, and 0.677. Further combination with the three factors indicated a higher power (AUC, 0.893; sensitivity, 85.5%; specificity, 93.2%). CONCLUSION: This was the first time to reveal the potential circulating fingerprints for predicting HC. PCAT1, MALAT1, and CPS1-IT1 may act as novel early diagnosis biomarkers for predicting HC.

Emerging Technologies for the Diagnosis of Perihilar Cholangiocarcinoma.

The diagnosis of malignant biliary strictures remains problematic, especially in the perihilar region and in primary sclerosing cholangitis (PSC). Conventional cytology obtained during endoscopic retrograde cholangiography (ERC)-guided brushings of biliary strictures is suboptimal due to limited sensitivity, albeit it remains the gold standard with a high specificity. Emerging technologies are being developed and validated to address this pressing unmet patient need. Such technologies include enhanced visualization of the biliary tree by cholangioscopy, intraductal ultrasound, and confocal laser endomicroscopy. Conventional cytology can be aided by employing complementary and advanced cytologic techniques such as fluorescent in situ hybridization (FISH), and this technique should be widely adapted. Interrogation of bile and serum by examining extracellular vesicle number and cargo, and exploiting next-generation sequencing and proteomic technologies, is also being explored. Examination of circulating cell-free deoxyribonucleic acid (cfDNA) for differentially methylated regions is a promising test which is being rigorously validated. The special expertise required for these analyses has to date hampered their validation and adaptation. Herein, we will review these emerging technologies to inform the reader of the progress made and encourage further studies, as well as adaptation of validated approaches.

ADAM-17 is a poor prognostic indicator for patients with hilar cholangiocarcinoma and is regulated by FoxM1.

BACKGROUND: A-disintegrin and metalloproteinases (ADAMs) are members of a family of multidomain transmembrane and secreted proteins. Specific ADAMs are upregulated in human cancers and correlated with tumor progression and poor outcome, but rarely studied in human hilar cholangiocarcinoma (HC). This study aimed to explore the expression profiles of ADAMs and their potential underlying mechanisms promoting cancer progression. METHODS: mRNA expression of ADAM-9, - 10, - 11, - 12, - 15, - 17, - 28, and - 33 was analyzed in human hilar cholangiocarcinoma (HC) samples. Immunohistochemical (IHC) analysis was used to detect the expression of ADAM-10, - 17, - 28, and FoxM1 in HC. The regulation of ADAM-17 by FoxM1 and their functional study was investigated in vivo and in vitro. RESULTS: ADAM-10, - 17, and - 28 were upregulated in tumors compared with matched non-cancerous tissues. IHC analysis revealed increased expression of ADAM-10, - 17, and - 28 in HC cells, and ADAM17 seems to be an independent prognostic factor. ADAM-17 is regulated by FoxM1. A decrease in the expression of ADAM-17 by silencing FoxM1 led to an inhibition of cell proliferation, tumor growth, and the production of tumor necrosis factor α. IHC analysis showed co-expression of FoxM1 and ADAM-17 in HC specimens. CONCLUSIONS: The findings of the present study show an important role of the cross-talk among FoxM1, ADAM-17, and TNFa in HC development and progression.

Surgical approaches for the treatment of perihilar cholangiocarcinoma.

INTRODUCTION: Since their initial description, perihilar cholangiocarcinoma (pCCA) has remained one of the more clinically challenging scenarios encountered by hepatobiliary surgeons. Surgery remains the only potentially curative therapy, but requires complex, technically demanding operations with high associated morbidity and mortality. Over the last several decades, advances in surgical technique and perioperative management have improved patient outcomes. Areas covered: Achievement of optimal outcomes requires a multidisciplinary approach from a team of providers with expertise in hepatobiliary and oncologic surgery, medical oncology, radiation oncology, and advanced gastroenterology. We herein report a comprehensive review on pCCA with an emphasis on surgical strategies and perioperative management. Expert commentary: Despite incremental improvements from advances in surgical technique and perioperative management, outcomes remain poor due to the aggressive systemic nature of this disease and the tendency for locoregional and distant recurrence. The marginal benefit observed with traditional systemic therapies continues to be a key weakness in current management. However, improved understanding of the genetic alterations and pathways that drive tumorigenesis has the potential to dramatically alter the way in which we care for these patients.

Long Non-Coding RNA MALAT1 Interacts With miR-204 to Modulate Human Hilar Cholangiocarcinoma Proliferation, Migration, and Invasion by Targeting CXCR4.

Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is involved in the development and progression of many types of tumors. An aberrant expression of MALAT1 was observed in many kinds of cancers. However, the exact effects and molecular mechanisms of MALAT1 in human hilar cholangiocarcinoma (HCCA) progression are still unknown. Here, we investigated the role of MALAT1 in human HCCA cell lines and clinical tumor samples in order to determine the function of this lncRNA. In our research, lncRNA-MALAT1 was specifically upregulated in HCCA tissues and cell lines, and was associated with pathological T stage, a larger tumor size, and perineural invasion. Knockdown of MALAT1 inhibited the proliferation, migration, and invasion of human HCCA cell. In addition, chemokine receptor-4 (CXCR4) was involved in MALAT1 induced human HCCA growth, migration, and invasion. By using online tools and a series of mechanistic analysis, we also demonstrated that miR-204-dependent CXCR4 regulation was required in MALAT1 modulating HCCA cell growth, migration and invasion. Taken together, our data indicated that MALAT1 might play an oncogenic role in HCCA through miR-204-dependent CXCR4 regulation, and could be regarded as a therapeutic target in HCCA. J. Cell. Biochem. 118: 3643-3653, 2017. © 2017 Wiley Periodicals, Inc.

Characterizing the activation of the Wnt signaling pathway in hilar cholangiocarcinoma using a tissue microarray approach.

Hilar cholangiocarcinoma (HCCA) is an invasive hepatic malignancy that is difficult to biopsy; therefore, novel markers of HCCA prognosis are needed. Here, the level of canonical Wnt activation in patients with HCCA, intrahepatic cholangiocarcinoma (IHCC), and congenital choledochal cysts (CCC) was compared to understand the role of Wnt signaling in HCCA. Pathology specimens from HCCA (n=129), IHCC (n=31), and CCC (n=45) patients were used to construct tissue microarrays. Wnt2, Wnt3, ß-catenin, TCF4, c-Myc, and cyclin D1 were detected by immunohistochemistry. Parallel correlation analysis was used to analyze differences in protein levels between the HCCA, IHCC, and CCC groups. Univariate and multivariate analyses were used to determine independent predictors of successful resection and prognosis in the HCCA group. The protein levels of Wnt2, ß-catenin, TCF4, c-Myc, and cyclin D1 were significantly higher in HCCA compared to IHHC or CCC. Wnt signaling activation (Wnt2+, Wnt3+, nuclear ß-catenin+, nuclear TCF4+) was significantly greater in HCCA tissues than CCC tissues. Univariable analyses indicated that expression of cyclin D1 as well as Wnt signaling activation, and partial Wnt activation (Wnt2+ or Wnt3+ and nuclear ß-catenin+ or nuclear TCF4+) predicted successful resection, but only cyclin D1 expression remained significant in multivariable analyses. Only partial Wnt activation was an independent predictor of survival time. Proteins in the canonical Wnt signaling pathway were present at higher levels in HCCA and correlated with tumor resecility and patient prognosis. These results suggest that Wnt pathway analysis may be a useful marker for clinical outcome in HCCA.

PKM2 regulates neural invasion of and predicts poor prognosis for human hilar cholangiocarcinoma.

BACKGROUND: The therapeutic and prognostic value of the glycolytic enzymes hexokinase, phosphofructokinase, and pyruvate kinase (PK) has been implicated in a variety of cancers, while their roles in treatment of and prognosis for hilar cholangiocarcinoma (HC) remain unclear. In this study, we determined the expression of PKM2 in and its impact on biology and clinical outcome of human HC. METHODS: The regulation and function of PKM2 in HC pathogenesis was evaluated using human tissues, molecular and cell biology, and animal models, and its prognostic significance was determined according to its impact on patient survival. RESULTS: We found that expression of hexokinase 1 and the M2 splice isoform of PK (PKM2) was upregulated in HC tissues and that this expression correlated with tumor recurrence and outcome. PKM2 expression was increased in HC cases with chronic cholangitis as demonstrated by isobaric tags for relative and absolute quantification. High PKM2 expression was highly correlated with high syndecan 2 (SDC2) expression and neural invasion. PKM2 downregulation led to a decrease in SDC2 expression. Treatment with metformin markedly suppressed PKM2 and SDC2 expression at both the transcriptional and posttranscriptional levels and inhibited HC cell proliferation and tumor growth. CONCLUSIONS: PKM2 regulates neural invasion of HC cells at least in part via regulation of SDC2. Inhibition of PKM2 and SDC2 expression contributes to the therapeutic effect of metformin on HC. Therefore, PKM2 is an independent prognostic factor and potential therapeutic target for human HC.

Expression and clinical significance of PIWIL2 in hilar cholangiocarcinoma tissues and cell lines.

The objective of this study was to explore the relationship between PIWI-like protein 2 (PIWIL2) and clinicopathological charac-teristics and prognosis after radical resection. To accomplish this, we analyzed PIWIL2 expression in hilar cholangiocarcinoma tissues and cell lines. PIWIL2 expression was detected by immunohistochemistry in 41 hilar cholangiocarcinoma samples and 10 control tissues. Western blotting and immunocytofluorescence were used to investigate PIWIL2 expression in the cholangiocarcinoma cell line QBC939 and the bile duct epithelial cell line HIBEpic. Univariate and multivariate surviv-al analyses were performed using the Kaplan-Meier method for hilar cholangiocarcinoma patients who underwent radical resection. PIWIL2 expression was significantly higher in the hilar cholangiocarcinoma tissues and QBC939 cells than in control tissues and HIBEpic cells, respectively (P < 0.05). Poorly and moderately differentiated cholan-giocarcinoma tissues had significantly higher PIWIL2 expression than well-differentiated tissues (P < 0.05). Univariate analysis demonstrated that high PIWIL2 expression was associated with shorter survival time after radical resection (P < 0.05). Multivariate analysis showed that PI-WIL2 expression was an independent prognostic factor after radical re-section of hilar cholangiocarcinoma (P < 0.05). PIWIL2 expression was also associated with tumor-node-metastasis stage and differentiation. PIWIL2 was an independent prognostic factor after radical resection of hilar cholangiocarcinoma.

Metastasis-associated in colon cancer 1 is an independent prognostic biomarker for survival in Klatskin tumor patients.

UNLABELLED: Curative treatment of intrahepatic cholangiocarcinoma (ICC) and hilar cholangiocarcinoma (Klatskin tumors) is limited to surgical resection or orthotopic liver transplantation. However, not all patients benefit from a surgical approach and suffer from early tumor recurrence. Response to chemotherapy is generally poor and, until today, no targeted therapy could be established. Metastasis-associated in colon cancer 1 (MACC1) is a recently discovered regulator of the hepatocyte growth factor (HGF)/Met/mitogen-activated protein kinase pathway, which induces proliferation, migration, and invasion in cell culture, as well as metastasis in mice. MACC1 expression shows a significant correlation with Met expression in colon cancer tissue and is highly prognostic for occurrence of distant metastasis and survival in colon cancer patients. Thus, we aimed to measure the expression of MACC1, Met, and HGF messenger RNA in microdissected tumor tissue and corresponding normal liver tissue of 156 patients with Klatskin tumors (n = 76) and ICC (n = 80) using real-time quantitative reverse-transcriptase polymerase chain reaction. We used immunohistochemical staining to validate the results. MACC1 expression in tumor tissue of both tumor entities was significantly higher than in corresponding normal liver tissue (P < 0.001). Klatskin tumor patients with a history of tumor recurrence had significantly higher MACC1 expression than those without tumor recurrence (P = 0.005). Uni- und multivariate survival analysis showed that Klatskin tumor patients with high MACC1 had a significantly shorter overall (OS) and disease-free survival (DFS; P = 0.001 and P < 0.001, respectively). The multivariate analysis confirmed MACC1 to be an independent factor for overall survival in Klatskin tumor patients (hazard ratio: 2.777; 95% confidence interval: 1.389-5.555; P = 0.004). CONCLUSION: Our study identified MACC1 as a highly prognostic biomarker for OS and DFS in Klatskin tumor patients. MACC1 expression could become an important diagnostic tool and might be a candidate for targeted therapy.

Spindle-cell variant of primary cutaneous follicle center lymphoma spreading to the hepatobiliary tree, mimicking Klatskin tumor.

Primary cutaneous follicle center lymphoma (pcFCL) is an indolent type of primary cutaneous B-cell lymphoma (pcBCL) rarely disseminating to other organs. PcBCL with spindle-cell morphology has been described as a rare variant of pcFCL but the prognosis data of this variant is sparse. We report a rare case of spindle-cell pcFCL with CD20(+), CD79a(+), CD3(+), Bcl-6(+), Mum-1(-) and CD10(-) tumor cells that infiltrated the hepatic hilum, mimicking a Klatskin tumor. On the basis of the sparse published data on spindle-cell morphology of pcBCL, this growth pattern should elicit awareness of an increased risk of systemic involvement in the otherwise indolent pcFCL.

Mutual regulation between microRNA-373 and methyl-CpG-binding domain protein 2 in hilar cholangiocarcinoma.

AIM: To investigate the reciprocal modulation between microRNA (miRNA) and DNA methylation via exploring the correlation between miR-373 and methyl-CpG-binding domain protein (MBD)2. METHODS: MiR-373 expression was examined using the TaqMan miRNA assay. Methylation of miR-373 was investigated using methylation-specific polymerase chain reaction, and recruitment of methyl binding proteins was studied using the chromatin immunoprecipitation assay. Mutation analysis was conducted using the QuikChange™ Site-Directed Mutagenesis kit. The activity of miR-373 gene promoter constructs and targeting at MBD2-three prime untranslated region (3'UTR) by miR-373 were evaluated by a dual-luciferase reporter gene assay. RESULTS: In hilar cholangiocarcinoma, miR-373 decreased and was closely associated with poor cell differentiation, advanced clinical stage, and shorter survival. The promoter-associated CpG island of miR-373 gene was hypermethylated and inhibited expression of miR-373. MBD2 was up-regulated and enriched at the promoter-associated CpG island of miR-373. Methylation-mediated suppression of miR-373 required MBD2 enrichment at the promoter-associated CpG island, and miR-373 negatively regulated MBD2 expression through targeting the 3'UTR. CONCLUSION: MiR-373 behaves as a direct transcriptional target and negative regulator of MBD2 activity through a feedback loop of CpG island methylation.