Article

Novel immunohistochemical markers differentiate intrahepatic cholangiocarcinoma from benign bile duct lesions

Authors:
  • Protagen Protein Services, Heilbronn, Germany
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Abstract

Aims: The distinction between intrahepatic cholangiocarcinoma (ICC) and benign bile duct lesions can be challenging. Using our previously identified potential biomarkers for ICC, we examined whether these are useful for the differential diagnosis of ICC, bile duct adenoma and reactive bile duct proliferations in an immunohistochemical approach and identified a diagnostic marker panel including known biomarkers. Methods: Subjects included samples from 77 patients with ICC, 33 patients with bile duct adenoma and 47 patients with ductular reactions in liver cirrhosis. Our previously identified biomarkers (stress-induced phosphoprotein 1 (STIP1), SerpinH1, 14-3-3Sigma) were tested immunohistochemically following comparison with candidates from the literature (cluster of differentiation 56, heat shock protein (HSP)27, HSP70, B-cell-lymphoma2, p53, ki67). Results: The expression of SerpinH1 and 14-3-3Sigma was significantly higher in ICC than in bile duct adenomas and ductular reactions (p<0.05), whereas STIP1 expression was significantly higher (p<0.05) in ICC than in ductular reactions, but the difference to the bile duct adenoma group was not significant. A panel of the biomarker SerpinH1, 14-3-3Sigma and ki67 (≥2 marker positive) showed a high diagnostic accuracy (sensitivity 87.8%, specificity 95.9%, accuracy 91.8%) in the differential diagnosis of ICC versus non-malignant bile duct lesions. Conclusions: This suggests that 14-3-3Sigma and SerpinH1 may be useful in the differential diagnosis of malignant, benign and reactive bile duct lesions in addition to ki67 where a cut-off of >5% might be used for the distinction of malignant and non-malignant lesions.

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... Several other immunohistochemical markers have been studied in cholangiocarcinoma, but these are primarily aimed at differentiating between various intrahepatic malignant processes (ie, differentiation of cholangiocarcinoma from hepatocellular carcinoma or metastatic adenocarcinoma) [18,19]. A recent study identified "significant increased expression" of 14-3-3Sigma and SerpinH1 proteins in intrahepatic cholangiocarcinoma, and the authors advocate use of immunohistochemical stains for these proteins, together with the Ki-67 proliferative index (with a 5% proliferative index cutoff), as part of a 3-stain panel for distinction between benign and malignant biliary lesions [20]. However, only moderately to poorly differentiated intrahepatic cholangiocarcinoma cases were studied [20] and immunohistochemical stains for 14-3-3Sigma and SerpinH1 are not widely available for clinical use (and require interpretation of stain intensity); in contrast, Ki-67 proliferative index alone was effective in identifying even well-differentiated cholangiocarcinoma in our study (using a cutoff of 10%), which is the most problematic diagnostic scenario. ...
... A recent study identified "significant increased expression" of 14-3-3Sigma and SerpinH1 proteins in intrahepatic cholangiocarcinoma, and the authors advocate use of immunohistochemical stains for these proteins, together with the Ki-67 proliferative index (with a 5% proliferative index cutoff), as part of a 3-stain panel for distinction between benign and malignant biliary lesions [20]. However, only moderately to poorly differentiated intrahepatic cholangiocarcinoma cases were studied [20] and immunohistochemical stains for 14-3-3Sigma and SerpinH1 are not widely available for clinical use (and require interpretation of stain intensity); in contrast, Ki-67 proliferative index alone was effective in identifying even well-differentiated cholangiocarcinoma in our study (using a cutoff of 10%), which is the most problematic diagnostic scenario. ...
Article
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Differentiation between benign and malignant lesions of the hepatic biliary tree may pose a diagnostic problem, as well-differentiated intrahepatic cholangiocarcinoma may mimic biliary hamartoma, bile duct adenoma, or parenchymal extinction. We evaluated Ki-67 proliferative index and p53 status by immunohistochemical staining to aid in exclusion of cholangiocarcinoma. 14 biliary hamartomas, 21 bile duct adenomas, and 11 livers with parenchymal extinction were compared to 26 intrahepatic cholangiocarcinomas (16 well-differentiated and 10 moderate- or poorly- differentiated tumors). We found an increased proliferative index in intrahepatic cholangiocarcinomas compared to benign biliary proliferations (average 23.0% in cholangiocarcinoma vs. 1.4% in all benign biliary lesions, n 26 vs. 46, P<.001). No difference in average proliferative index was observed between well-differentiated and moderately/poorly differentiated cholangiocarcinomas (average 22.7% vs. 23.3%, n 16 vs. 10, P=.92). Average proliferation indices of benign biliary lesions were uniformly low (biliary hamartoma 1.2%; bile duct adenoma 2%; parenchymal extinction 0.4%). The majority of cholangiocarcinomas (23/26, 88.5%), but none of the benign lesions (0/46, 0%) had proliferative indices greater than 10%. Strong nuclear p53 immunohistochemical staining was only seen in cholangiocarcinomas (9/26, 34.6%) and not in benign biliary lesions (0/46, 0%), though many of the benign lesions showed weak to moderate staining Immunohistochemical staining for Ki67 facilitates distinction between benign and malignant lesions of the intrahepatic biliary tree, whereas p53 immunohistochemical staining, is less helpful.
... To aid in the distinction between iCCA and benign intrahepatic biliary lesions, most studies have evaluated immunohistochemical (IHC) stains [10][11][12]. For instance, an increased Ki-67 proliferation index has been reported in iCCA (mean: 23% versus 1.4% for benign intrahepatic biliary lesions; p < 0.001) [12]. ...
... Well-differentiated iCCA may have overlapping histological features with BDA and BDH, and morphological features suggestive of malignancy (such as irregular glands and stratification of epithelial cells, as well as hyperchromatic and pleomorphic nuclei) can be seen in BDA and BDH. Although IHC stains have been used to facilitate this distinction [10][11][12], more objective ancillary testing may be useful in better distinguishing iCCA from benign intrahepatic biliary lesions. In this regard, we note that aneuploidy detected by DNA flow cytometry may potentially serve as a diagnostic marker of malignancy, as almost half of iCCA samples (47%) showed aneuploidy (versus 0% in its benign morphological mimics) ( Table 1). ...
Article
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The distinction between well-differentiated intrahepatic cholangiocarcinoma (iCCA) from its morphological mimics such as bile duct adenoma (BDA) and hamartoma (BDH) can be challenging, particularly in small biopsies. Although a few cases of BDA and BDH have been reported to undergo malignant transformation into iCCA, their neoplastic versus benign nature remains debated. DNA flow cytometry was performed on 47 formalin-fixed paraffin-embedded samples of iCCA, 14 BDA, and 18 BDH. Aneuploidy was detected in 22 iCCA (47%) but in none of the 32 BDA and BDH samples. Among the 34 iCCA patients who underwent complete resection and were followed up to tumor recurrence, tumor-related death, or at least for 1 year, the overall recurrence or death rates (regardless of flow cytometric results) were 18, 56, and 71% within 1, 3, and 5 years, respectively. The 1-, 3-, and 5-year recurrence or death rates in 18 iCCA patients with aneuploidy were 28, 66, and 66%, respectively, whereas 16 iCCA patients in the setting of normal DNA content had 1-, 3-, and 5-year rates of 6, 44, and 72%, respectively. Although aneuploid tumors were associated with worse outcomes during the first 3 years, this difference was not statistically significant (hazard ratio = 1.4, p = 0.473) in the present sample size. In conclusion, the frequency of aneuploidy was significantly higher in iCCA (47%) than in its benign morphological mimics (0%), suggesting that it may potentially serve as a diagnostic marker of malignancy in challenging situations. Our findings also suggest that most BDAs and BDHs, if not all, are benign entities and may not represent precursor lesions to iCCAs that often harbor aneuploidy. Although a larger cohort will be necessary to further determine the prognostic significance of aneuploidy in iCCA patients after resection, the patients with aneuploid tumors may have a higher risk for tumor progression, especially during the first 3 years.
... C'est également le cas dans le cancer du pancréas comme l'ont rapporté d'autres(328,329). Toutefois, tous ces résultats contrastent avec ceux de Bertram et al(330) qui ont montré une plus forte expression de SFN dans les tissus de CCAi. Néanmoins, le fait que SFN ait pu être considéré pour sses valeurs diagnostiques et pronostiques dans des cancers en première ligne d'exposition aux microbes, tels l'adénocarcinome pulmonaire(325,326) et le cancer du côlon, suggère que les microbes pourrait être associés à l'expression du gène via la dysméthylation comme nous avons pu le discuter ailleurs(331). ...
Thesis
Le cancer est la première cause de mortalité en France. Les Cholangiocarcinomes (CCAs) ou cancers des voies biliaires représentent 3% de l’ensemble des cancers digestifs. Le CCA intrahépatique avec 10 à 20 % de l’ensemble, est derrière l’hépatocarcinome, la deuxième tumeur primitive maligne du foie. La bile est un fluide stérile biologique, synthétisé dans le foie, hébergé dans la vésicule biliaire et libéré dans le duodénum après la prise alimentaire. Les populations microbiennes des différentes parties du tractus gastro-intestinal des mammifères (estomac, intestin grêle, colon, selles) ont été étudiées. Des changements dans le microbiote humain ont été associés au développement de cancer colorectal, du pancréas et du poumon. Dans la plupart des séries étudiées le mécanisme reliant le microbiote à la carcinogenèse semble être de type épigénétique. La carcinogenèse biliaire est mal connue, les caractéristiques génétiques des individus et les facteurs environnementaux y jouent un rôle prépondérant. La caractérisation du microbiote biliaire n’a été abordée que de façon fragmentaire jusqu’à présent. Nous avons caractérisé le microbiote biliaire chez des patients atteints du CCA extra-hépatique et chez des sujets témoins sans tumeurs. Des échantillons de biles ont été prélevés par la Cholangiographie pancréatique rétrograde par voie endoscopique CPRE chez 28 patients atteints de CCA sporadiques et 47 sujets témoins (porteurs de calculs biliaires).La bile a été soumise aux extractions d’ADN bactérien ainsi qu’à la caractérisation de méthylation du génome humain grâce aux séquençage d’ARN16s bactérien et des puces de méthylation (Illumina). De plus, les profils de méthylation des gènes ont été caractérisés dans des échantillons tissulaires d’adénocarcinomes pancréatique et de CCAs.Nous avons pu montrer une dysbiose biliaire associées aux CCAs, identifié une signature népigénétique liée aux CCAs dont nous avons distingué la spécificité versus celle des cellules mononuclées du sang périphérique (PBMC). Deux gènes candidats ont ainsi été sélectionnés à l’issu d’analyses de puces des méthylations et leurs expressions ont été ensuite étudiées par la technique d’immunohistochimie (IHC) dans les tissus tumoraux de CCAs intra- et extra-hépatiques et d’adénocarcinome pancréatique. Nous avons conclu sur une possible relation entre la dysbiose biliaire et le niveau d’expression des deux gènes par le phénomène de dysméthylation. Ces résultats pourraient avoir un impact diagnostique et devenir des cibles thérapeutiques.
... 32,39,40 STIP1 41 is a co-chaperone that is overexpressed in a number of malignancies. 30,31,[42][43][44][45][46][47][48][49] In the present study, we used targeting STIP1 for an anticancer therapeutic strategy as a demonstration of the efficiency, specificity, and clinical potential of protein transfection through the HEPES method. ...
... 32,39,40 STIP1 41 is a co-chaperone that is overexpressed in a number of malignancies. 30,31,[42][43][44][45][46][47][48][49] In the present study, we used targeting STIP1 for an anticancer therapeutic strategy as a demonstration of the efficiency, specificity, and clinical potential of protein transfection through the HEPES method. ...
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The delivery of active proteins into cells (protein transfection) for biological purposes offers considerable potential for clinical applications. Herein we demonstrate that, with a readily available, inexpensive organic agent, the 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) method can be used for simple and efficient protein transfection. By mixing proteins with a pure HEPES solution before they are applied to live cells, proteins with various molecular weights (including antibodies, recombinant proteins, and peptides) were successfully delivered into the cytoplasm of different cell types. The protein transfection efficiency of the HEPES method was not inferior to that of commercially available systems that are both more expensive and time consuming. Studies using endocytotic inhibitors and endosomal markers have revealed that cells internalize HEPES-protein mixtures through endocytosis. Results that HEPES-protein mixtures exhibited a low diffusion coefficient suggest that HEPES might neutralize the charges of proteins and, thus, facilitate their cellular internalization. Upon internalization, the cytosolic antibodies caused the degradation of targeted proteins in TRIM21-expressing cells. In summary, the HEPES method is efficient for protein transfection and has potential for myriad clinical applications.
... The tumor is composed of closely packed small ductules with varying degree of cellular atypia, usually no cystic dilatation and with intervening fibrotic stroma. Sometimes it might be problematic to distinguish BDA from the ductular reaction, a frequent finding in chronic liver diseases and from well differentiated CC, especially in intraoperative frozen sections [43,52,53]. The epithelial cells express CK7 and CK19, epithelial membrane antigen (EMA) and low proliferation activity detected by Ki67 immunoreaction [54]. ...
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Cholangiocarcinoma (CC) is the second most common tumor of the liver, originating from the biliary system with increasing incidence and mortality worldwide. Several new classifications review the significance of tumor localization, site of origin, proliferation and biomarkers in the intrahepatic, perihilar and distal forms of the lesion. Based on growth pattern mass-forming, periductal-infiltrating, intraductal, undefined and mixed types are differentiated. There are further subclassifications which are applied for the histological features, in particular for intrahepatic CC. Recognition of the precursors and early lesions of CC including biliary intraepithelial neoplasia (BilIN), intraductal papillary neoplasm of the bile ducts (IPNB), biliary mucinous cystic neoplasm (MCNB) and the candidate precursors, such as bile duct adenoma and von Meyenburg complex is of increasing significance. In addition to the previously used biliary markers detected by immunohistochemistry, several new markers have been added to the differentiation of both the benign and malignant lesions, which can be used to aid in the subclassification in association with the outcome of CC. Major aspects of biliary carcinogenesis have been revealed, yet, the exact way of this diverse process is still unclear. The factors contributing to molecular cholangiocarcinogenesis include various risk factors, different anatomical localizations, multiple cellular origins, genetic and epigenetic alterations, tumor microenvironment, heterogeneity and clonal evolution. Driver mutations have been identified, implying that they are optimal candidates for targeted therapy. The most promising therapeutic candidates have entered clinical trials.
... Studies of molecular alteration reported that BilIN showed a step-wise increased expression of oncogene-related molecules such as p21, p53, and cyclin D, and a stepwise decreased expression of Smad4 during the progression of the grades of BilIN [13]. In other studies, Ki67 proliferative index was reported to range from 11 to 84.8% in intrahepatic CC [14,15]. In this study, Ki-67 expression was demonstrated in patients with extrahepatic CC and it suggests that BilIN-positive CC shows higher cell proliferation. ...
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Backgrounds and aims: Biliary intraepithelial neoplasia (BilIN) is a precursor of cholangiocarcinoma (CC) and it has been associated with several chronic inflammatory conditions. This study aimed to elucidate the prevalence of BilIN in CC and its clinicopathological significance. Methods: Medical records of 193 patients with histologically confirmed CC were analyzed. We reviewed the pathology findings of 48 patients who underwent curative surgery for CC. Results: Of the 48 patients analyzed, 34 and 14 patients had extrahepatic and intrahepatic CC respectively. BilIN was detected in 28 patients (58%) and showed a significantly higher prevalence in extrahepatic CC (75%) than in intrahepatic CC (21%; p < 0.001). In the subgroup of 34 patients with extrahepatic CC, 25 and 9 patients were BilIN positive and negative respectively. Poor differentiation and T3 stage were significantly more common in the BilIN-negative group than in the BilIN-positive group (p < 0.05). The expression of MUC5AC, p53, and loss of Smad4 showed no difference between BilIN-positive CC and in BilIN-negative CC, but the Ki-67 expression was significantly higher (p < 0.05). Conclusion: BilIN-positive CC showed less invasiveness than negative cases. The Ki-67 expression was significantly higher in BilIN-positive CC.
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Solid pseudopapillary tumor, pancreatoblastoma, undifferentiated carcinoma with osteoclastic-like giant cells, and acinar cell carcinomas are rare pancreatic nonductal neoplasms. Compared to the significant advances in our understanding of the pathogenesis of pancreatic ductal adenocarcinomas in the last decades, the molecular mechanisms underlying pancreatic nonductal neoplasms are poorly understood. In order to elucidate their molecular pathogenesis, we constructed tissue microarrays to study the expression of some novel pancreatic ductal adenocarcinoma-associated tumor markers in these nonductal pancreatic neoplasms. We analyzed nine markers including tumor suppressor gene (14-3-3 sigma), proliferation marker (topoisomerase II alpha), epithelial markers (prostate stem cell antigen, mesothelin and cytokeratin 19), stromal markers (fascin, hsp47 and fibronectin), and gamma-synuclein whose function is not delineated. In addition, we included tumor suppressor gene DPC4 and oncogene Beta-catenin to further confirm their expression in pancreatic nonductal tumors. Our results showed that in contrast to pancreatic ductal adenocarcinomas that show loss of Dpc4 protein in 55% of cases, loss of Dpc4 expression is absent in pancreatic nonductal neoplasms. Expression of 14-3-3 sigma is frequently seen in both pancreatic nonductal neoplasms (25-100%) and ductal adenocarcinomas (89%). Aberrant nuclear expression of beta-catenin is common in pancreatic nonductal neoplasms, specifically in solid pseudopapillary tumors (88%) and pancreatoblastomas (100%) but is rarely seen in pancreatic ductal adenocarcinomas (<5%). Expression of topoisomerase II alpha is not seen in solid pseudopapillary tumors and undifferentiated carcinomas with osteoclastic-like giant cells but is focally seen in pancreatoblastomas (50%) and acinar cell carcinomas (85%). Expression of PSCA and mesothelin was observed in pancreatic nonductal neoplasms but their expression was seen less frequently (0-50%) and weaker than that in pancreatic ductal adenocarcinomas (60-100%). CK19, a marker of pancreatic ductal adenocarcinomas, is not expressed in pancreatic nonductal neoplasms. Expression of gamma-synuclein as well as stromal markers (fascin, hsp47 and fibronectin) is frequently seen in both. Our findings indicate pancreatic nonductal neoplasms have distinctive patterns of protein expression relative to pancreatic ductal adenocarcinomas and suggest that pancreatic nonductal neoplasms have different genetic pathways from the more common pancreatic ductal adenocarcinomas.
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The aim of the present study was to examine the immunohistochemical distribution of the 47-kDa heat shock protein (HSP47) to enhance the understanding of the mechanisms involved in stromal fibrosis, which accompanies cancer infiltration in scirrhous carcinoma of the stomach. In vitro gastric cancer models were prepared by collagen gel cultures using three different human gastric cancer cell lines (KATO-III, MKN-74, MKN-45) and a human fibroblast cell line (TIG-101). Tumor tissues were obtained from ten patients with early gastric cancer (5 scirrhous carcinoma; 5 non-scirrhous carcinoma) and three patients with advanced scirrhous gastric cancer. The gels and the tissues were immunostained by a monoclonal antibody against human HSP47 and then examined by light and electron microscopy. The staining intensity of the fibroblasts was stronger than that of cancer cells in both the culture models and patient tissues. Moreover, the number of fibroblasts in scirrhous gastric cancer was significantly greater than that in non-scirrhous gastric cancer (p = 0.0004). In addition, the discharge of HSP47 was observed in the extracellular matrix as granular deposits of staining. These findings indicate that fibroblasts predominantly produce stromal collagen and may play an important role in the development of stromal change in scirrhous gastric cancer. Further studies are required to elucidate the significance of HSP47 in the development of new clinical approaches for treating scirrhous gastric cancer.
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The most common pathologic form of pulmonary fibrosis arises from excessive deposition of extracellular matrix proteins such as collagen. The 47 kDa heat shock protein 47 (HSP47) is a collagen-specific molecular chaperone that has been shown to play a major role during the processing and/or secretion of procollagen. To determine whether inhibition of HSP47 could have beneficial effects in mitigating bleomycin-induced pulmonary fibrosis in rats. All experiments were performed with 250-300 g male Wistar rats. Animals were randomly divided into five experimental groups that were administered: 1) saline alone, 2) bleomycin alone, 3) antisense HSP47 oligonucleotides alone, 4) bleomycin + antisense HSP47 oligonucleotides, and 5) bleomycin + sense control oligonucleotides. We investigated lung histopathology and performed immunoblot and immunohistochemistry analyses. In rats treated with HSP47 antisense oligonucleotides, pulmonary fibrosis was significantly reduced. In addition, treatment with HSP47 antisense oligonucleotides significantly improved bleomycin-induced morphological changes. Treatment with HSP47 antisense oligonucleotides alone did not produce any significant changes to lung morphology. Immunoblot analyses of lung homogenates confirmed the inhibition of HSP47 protein by antisense oligonucleotides. The bleo + sense group, however, did not exhibit any improvement in lung pathology compared to bleomycin alone groups, and also had no effect on HSP47 expression. These findings suggest that HSP47 antisense oligonucleotide inhibition of HSP47 improves bleomycin-induced pulmonary fibrosis pathology in rats.
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The fifth edition of TNM classification of malignant tumours contains some additions and modifications for classifying tumours of the digestive system. Apart from the tumours of the stomach the most important revisions have been made in tumours of the pancreas. In tumours of other sites of the digestive system only minor changes or modifications have been introduced. A new element of the pN classification is that the number of lymph nodes ordinarily included in a lymphadenectomy specimen is noted at each site. The designation pNO is usually based on this figure.
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BACKGROUND & PURPOSE: Hepatocellular carcinoma (HCC) is a major lethal cancer worldwide. Despite sophisticated diagnostic algorithms, the differential diagnosis of small liver nodules still is difficult. While imaging techniques have advanced, adjuvant protein-biomarkers as glypican3 (GPC3), glutamine-synthetase (GS) and heat-shock protein 70 (HSP70) have enhanced diagnostic accuracy. The aim was to further detect useful protein-biomarkers of HCC with a structured systematic approach using differential proteome techniques, bring the results to practical application and compare the diagnostic accuracy of the candidates with the established biomarkers. METHODS: After label-free and gel-based proteomics (n=18 HCC/corresponding non-tumorous liver tissue (NTLT)) biomarker candidates were tested for diagnostic accuracy in immunohistochemical analyses (n=14 HCC/NTLT). Suitable candidates were further tested for consistency in comparison to known protein-biomarkers in HCC (n=78), hepatocellular adenoma (n=25; HCA), focal nodular hyperplasia (n=28; FNH) and cirrhosis (n=28). RESULTS: Of all protein-biomarkers, 14-3-3Sigma (14-3-3S) exhibited the most pronounced up-regulation (58.8×) in proteomics and superior diagnostic accuracy (73.0%) in the differentiation of HCC from non-tumorous hepatocytes also compared to established biomarkers as GPC3 (64.7%) and GS (45.4%). 14-3-3S was part of the best diagnostic three-biomarker panel (GPC3, HSP70, 14-3-3S) for the differentiation of HCC and HCA which is of most important significance. Exclusion of GS and inclusion of 14-3-3S in the panel (>1 marker positive) resulted in a profound increase in specificity (+44.0%) and accuracy (+11.0%) while sensitivity remained stable (96.0%). CONCLUSIONS: 14-3-3S is an interesting protein biomarker with the potential to further improve the accuracy of differential diagnostic process of hepatocellular tumors. This article is part of a Special Issue entitled: Medical Proteomics.
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Cholangiocarcinoma (CCA) is a lethal malignancy which occurs with relatively high incidence in Thailand. This cancer is often difficult to diagnose and associated with high mortality. The secretome, containing the secreted proteins from cells, are potentially useful as biomarkers of cancers. Since three-dimensional (3D) cell culture may mimic growth characteristics and microenvironment of solid tumors in vivo better than monolayer culture, we have developed culture of CCA in natural collagen-based scaffold, to enable analysis of the secretome by 2DE. Our results indicated that CCA growth in 3D environment alters cell shape significantly and enhances extracellular matrix deposition. Interestingly, more secreted proteins were detected from 3D culture compared to monolayer culture. Secretome analysis using 2DE coupled with LC-MS/MS demonstrated 10 secreted proteins uniquely found in 3D culture. Moreover, 25 proteins were enriched in 3D culture compared to monolayer culture, including 14-3-3 σ, triosephosphate isomerase, phosphoglycerate mutase 1, α-enolase, and L-plastin. Immunoblotting was used to confirm the presence of L-plastin in conditioned media of CCA and of hepatocellular carcinoma (HCC) cell lines. The results revealed that L-plastin, an actin bundling protein, was uniquely expressed only in the CCA cell line and could be a promising biomarker for differential diagnosis of CCA compared to HCC.
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The aim of this study was the identification of novel biomarker candidates for the diagnosis of cholangiocellular carcinoma (CCC) and its immunohistochemical differentiation from benign liver and bile duct cells. CCC is a primary cancer which arises from the epithelial cells of bile ducts and is characterised by high mortality rates due to its late clinical presentation and limited treatment options. Tumorous tissue and adjacent non-tumorous liver tissue from eight CCC patients were analysed by two-dimensional differential in-gel electrophoresis (2D-DIGE) and by mass spectrometry-based label-free proteomics. After data analysis and statistical evaluation of the proteins which were found to be differentially regulated between the two experimental groups (fold change ≥ 1.5; p-value ≤ 0.05), 14 candidate proteins were chosen for determination of the cell type-specific expression profile by immunohistochemistry in a cohort of 14 patients. This confirmed the significant up-regulation of serpin H1, 14-3-3 protein sigma (SFN) and stress-induced phosphoprotein 1 (STIP1) in tumorous cholangiocytes in comparison to normal hepatocytes and non-tumorous cholangiocytes, whereas some proteins were detectable specifically in hepatocytes. Since STIP1 exhibited both sensitivity and specificity of 100%, an immunohistochemical verification examining tissue sections of 60 CCC patients was performed. This resulted in a specificity of 98% and a sensitivity of 64%. We therefore conclude that this protein should be considered as potential diagnostic biomarker for CCC in an immunohistochemical application, possibly in combination with other candidates from this study in the form of a biomarker panel. This could improve the differential diagnosis of CCC and benign bile duct diseases as well as metastatic malignancies in the liver.
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Heat shock protein 47 (HSP47), also known as SERPINH1, is a product of CBP2 gene located at chromosome 11q13.5, a region frequently amplified in human cancers. HSP47 has been demonstrated to effect on limiting tumor invasion and motility. The previous studies showed that HSP47 is overexpressed in many human cancers, including stomach cancer, lung cancer, pancreatic ductal adenocarcinoma, and ulcerative colitis-associated carcinomas. However, the role of HSP47 in human glioma is still unknown. Here, we examined the expression of HSP47 in a group of glioma tumors and matched non-tumor brain tissues using qRT-PCR. We found that HSP47 is significantly overexpressed in glioma tissues and cell lines and associated with glioma tumor grade. Next, we knockdown the expression of HSP47 in the glioma cells using small interfering RNAs. The result showed that knockdown of HSP47 inhibits glioma cell growth, migration and invasion in vitro. We further investigated the posttranscriptional regulation of HSP47 by microRNAs using bioinformatics analysis and experimental validation. The results suggested that the expression of HSP47 is regulated by miR-29a. Finally, stable knockdown of HSP47 using shRNA inhibits glioma tumor growth and induces apoptosis in mice models in vivo. Therefore, our data suggested that HSP47 regulated by miR-29a to enhance glioma tumor growth and invasion. Taken together, HSP47 plays important role in tumor growth and invasion and thus could be a therapeutic target for treating glioma in the future.
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Stress-induced phosphoprotein1 (STIP1) is a candidate biomarker in epithelial ovarian cancer (EOC). In this study, we investigated in detail the expression of STIP1, as well as its functions, in EOC. STIP1 expression was assessed by immunohistochemistry (IHC) and the results were compared with clinicopathologic factors, including survival data. The effects of STIP1 gene silencing via small interfering RNA (siRNA) were examined in EOC cells and a xenograft model. The expression of STIP1 protein in EOC was significantly higher than in the other study groups (P < 0.001), and this increase of expression was significantly associated with tumor stage (P = 0.005), tumor grade (P = 0.029), and lymph node metastasis (P = 0.020). In multivariate analysis, overall survival in EOC was significantly shorter in cases with high STIP1 expression (HR = 2.78 [1.01-7.63], P = 0.047). STIP1 silencing in EOC cells resulted in inhibition of cell proliferation and invasion. In addition, in vivo experiments using STIP1 siRNA clearly showed a strong inhibition of tumor growth and a modulation of expression of prosurvival and apoptotic genes, further suggesting that STIP1 silencing can prevent cell proliferation and invasion. In conclusion, increased STIP1 expression is associated with poor survival outcome in EOC, and STIP1 may represent a useful therapeutic target in EOC patients. © 2014 Wiley Periodicals, Inc.
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Distinction between primary intrahepatic cholangiocarcinoma (ICC) and metastatic pancreatic ductal adenocarcinoma (PDA) on a liver biopsy is essentially impossible histologically but has important clinical implications. In this study, 41 ICCs and 60 PDAs were immunohistochemically evaluated for the expression of S100P, pVHL, IMP3, maspin, MUC5AC, and CK17 proteins. The results showed pVHL expression in 29 (71%) ICCs but in only 3 (5%) PDAs. S100P, MUC5AC, and CK17 were frequently expressed in PDAs, seen in 57 (95%), 40 (67%), and 36 (60%) cases, respectively. In contrast, only 11 (27%), 5 (12%), and 5 (12%) ICC cases expressed these proteins. IMP3 was expressed in 37 (90%) ICC and 54 (90%) PDA cases with equal frequency. All 60 (100%) PDA and 30 (73%) ICC cases showed positive maspin immunoreactivity. A S100P-/pVHL+/MUC5AC-/CK17- staining pattern was essentially indicative of ICC, whereas the S100P+/pVHL-/MUC5AC+/CK17+ and S100P+/pVHL-/MUC5AC-/CK17+ staining patterns were suggestive of PDA. These observations demonstrate that S100P, pVHL, MUC5AC, and CK17 are a useful immunohistochemical panel that may help distinguish primary ICC from metastatic PDA.
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The aim of this study was to detect stress-induced phosphoprotein 1 (STIP1) expression in papillary thyroid carcinoma (PTC) and to analyze its association with prognosis of PTC patients. Immunohistochemistry was performed to detect the expression of STIP1 in 113 PTC tissues and paired adjacent noncancerous tissues. The χ2 test was used to analyze the relationship between STIP1 expression and clinicopathological characteristics. Survival curves were plotted by the Kaplan-Meier method and compared using the log-rank test. Survival data was evaluated using univariate and multivariate Cox regression analysis. We identified abnormally elevated expression of STIP1 protein in PTC tissues compared to paired adjacent noncancerous tissues. Clinicopathological analysis showed that STIP1 expression was significantly correlated with tumor size (P = 0.017), lymph node metastasis (P = 0.007), and TNM stage (P = 0.026). Patients with higher STIP1 expression had shorter overall survival time, whereas those with lower STIP1 expression had longer survival time. Multivariate analysis suggested that STIP1 expression might be an independent prognostic indicator (P < 0.05) for the survival of patients with PTC. In conclusion, our findings provide evidences that positive expression of STIP1 in PTC may be important in the acquisition of an aggressive phenotype, and it is an independent biomarker for poor prognosis of patients with PTC.
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Small-molecule modulation of protein-protein interactions (PPIs) is one of the most exciting but also difficult fields in chemical biology and drug development. As one of the most important "hub" proteins with at least 200 - 300 interaction partners the 14-3-3 proteins are an especially fruitful case for PPI intervention. Here, we summarize recent success stories in small-molecule modulation - both inhibition and stabilization - of 14-3-3 PPIs. The chemical breath of modulators includes natural products such as fusicoccin A and derivatives. But also compounds identified via high throughput- and in silico-screening, which has yielded a toolbox of useful inhibitors and stabilizers for this interesting class of adapter proteins.
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Cholangiocarcinoma often is diagnosed at an advanced stage. Thus, it is necessary to establish sensitive screening methods that would allow cholangiocarcinoma and preferably its precursor lesion [biliary intraepithelial neoplasia (BilIN)] to be detected. We sought to clarify the usefulness of heat shock protein (HSP) 27 and HSP70 as biomarkers of cholangiocarcinoma and have used immunohistochemical analyses of hepatolithiatic livers to characterize HSP27 and HSP70 expression during the multistep cholangiocarcinogenesis process. HSP27 and HSP70 were measured in serum and bile samples via enzyme-linked immunosorbent assay. In hepatolithiatic tissue, the expression of HSP27 and HSP70 was increased in BilIN as well as in invasive cholangiocarcinoma. The serum levels of HSP27 and HSP70 were not significantly different between the hepatolithiatic patients with and without cholangiocarcinoma. In contrast, the bile levels of HSP27 and HSP70 were increased significantly in the patients with cholangiocarcinoma compared with those in the patients with lithiasis. Combining the measurements of the bile levels of HSP27 and HSP70 increased their usefulness as biomarkers, and the sum (HSP27 + HSP70) yielded the best sensitivity (90%) and specificity (100%). These results suggest that HSP27 and HSP70 could be used as biliary biomarkers for the detection of cholangiocarcinoma including BilIN.
Article
Unlabelled: There is limited information on test performance for detecting cholangiocarcinoma in primary sclerosing cholangitis (PSC), particularly when used sequentially. This study aimed to characterize diagnostic performance of serum carbohydrate antigen 19-9 (CA 19-9), ultrasonography, computed tomography, magnetic resonance imaging, cholangiography, and biliary cytologic techniques for detecting cholangiocarcinoma in PSC. All consecutive patients with PSC were screened and followed for development of cholangiocarcinoma from 2000 through 2006. Of 230 patients, 23 developed cytopathologically confirmed cholangiocarcinoma with an annual incidence of 1.2%. The optimal cutoff value for serum CA 19-9 was 20 U/mL, which yielded a sensitivity of 78%, specificity of 67%, positive predictive value (PPV) of 23%, and negative predictive value (NPV) of 96%. Serum CA 19-9 combined with either ultrasonography, computed tomography, or magnetic resonance imaging provided a sensitivity of 91%, 100%, and 96%, specificity of 62%, 38%, and 37%, PPV of 23%, 22%, and 24%, and NPV of 98%, 100%, and 98%, respectively, if at least one method was positive. Subsequent cholangiographic examinations in these patients increased specificity to 69% and PPV to 42% while maintaining sensitivity of 91% and NPV of 96%. Following this group, conventional cytology, aneuploidy detection by digital imaging analysis, and aneusomy detection by fluorescence in situ hybridization in brushing samples of biliary strictures had a sensitivity of 50%, 57%, and 86%, specificity of 97%, 94%, and 83%, PPV of 86%, 89%, and 80%, and NPV of 83%, 74%, and 88%, respectively, for detecting cholangiocarcinoma. Conclusion: Tumor serology combined with cross-sectional liver imaging may be useful as a screening strategy and cholangiography with cytologic examination is helpful for the diagnosis of cholangiocarcinoma in patients with PSC.
Article
Methods of evaluating and comparing the performance of diagnostic tests are of increasing importance as new tests are developed and marketed. When a test is based on an observed variable that lies on a continuous or graded scale, an assessment of the overall value of the test can be made through the use of a receiver operating characteristic (ROC) curve. The curve is constructed by varying the cutpoint used to determine which values of the observed variable will be considered abnormal and then plotting the resulting sensitivities against the corresponding false positive rates. When two or more empirical curves are constructed based on tests performed on the same individuals, statistical analysis on differences between curves must take into account the correlated nature of the data. This paper presents a nonparametric approach to the analysis of areas under correlated ROC curves, by using the theory on generalized U-statistics to generate an estimated covariance matrix.
Article
Eight hepatic atypical adenomatous hyperplasias (AH), 30 hepatocellular carcinomas (HCC) consisting of 11 well-, 13 moderately and six poorly differentiated HCC, and 10 intrahepatic cholangiocarcinomas (CC) were investigated immunohistochemically with anti-alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA), CA19-9, epithelial membrane antigen (EMA), and cytokeratins (CK) 18 and 19 antibodies. Immunostaining was regarded as positive when more than 5% of cells were stained. Alpha-fetoprotein was positive, although focally, in five (17%) of 30 HCC but negative in all AH and CC. Carcinoembryonic antigen (polyclonal antibody) did not stain the cytoplasm of all AH and HCC, but stained two (25%) of eight AH and 10 (33%) of 30 HCC in a bile canalicular staining manner. Carcinoembryonic antigen showed intracytoplasmic or luminal border staining in six (60%) of 10 CC. CA19-9 was negative in all AH and HCC, while six (60%) of 10 CC were positive for CA19-9. Epithelial membrane antigen was positive in one (13%) of eight AH, seven (23%) of 30 HCC and in all 10 cases of CC. Cytokeratin 18 was positive in all AH, HCC and CC. Cytokeratin 19 was negative in both AH and HCC, whereas it stained the cytoplasm of tumor cells in all CC diffusely and intensely. These results suggest that immunostaining of AFP, CEA, CA19-9, EMA, CK18 and CK19 are not useful in the differential diagnosis between AH and well-differentiated HCC, and that CK19 is the most suitable reagent for the differential diagnosis between HCC and CC.
Article
Diagnosis of early cholangiocarcinoma (CC) in patients with primary sclerosing cholangitis with available radiological methods is very difficult. This type of tumor is the second most common cause of mortality after liver failure in these patients. The recognition of CC is important for the selection of patients for, and the results of, liver transplantation (Ltx). In this study our aim was to investigate the value of measuring cancer markers (CA 19-9 and CEA) in patients with PSC for early diagnosis of CC. 72 PSC patients who were followed at our institution for a long period were included in the study; 9 with CC and 63 without CC. Furthermore, nine patients with CC but without concomitant PSC were included, as well as 24 patients with various cholestatic liver diseases. Serum levels of CA 19-9 and CEA were measured, in 39 PSC patients without CC, on multiple occasions. Moreover, bile was collected during a diagnostic ERCP from 20 patients for measurements of CA 19-9 and CEA. In those PSC patients without CC during the follow-up and with more than one year of follow-up, 15 patients had increased values of CA 19-9 (>37 ng/ml) on some of the occasions. Four of them demonstrated large fluctuations (more than 100 ng/ml difference at different occasions) in serum levels of Ca 19-9. A significant correlation between high CA 19-9 values and serum alkaline phosphatase levels was observed in these patients. The sensitivity of CA 19-9 in detecting CC in PSC patients was only 63%. The sensitivity of CEA and the combination of CA 19-9 and CEA (marker product; King's College formula) were still lower (33%) although the specificity was relatively high (85%). Bile levels of the tumor markers did not demonstrate any clinically useful differences between the different patient groups. Tumor markers as a diagnostic tool in diagnosing CC in patients with PSC are unfortunately not as valuable as previously reported. The serum levels of CA 19-9 can rise temporarily in association with a "biochemical relapse" of PSC (increased values of serum alkaline phosphatase). The marker product of CA 19-9 and CEA has a low sensitivity but a relatively high specificity for the detection of CC in PSC patients.
Article
This study was conducted to assess the clinical value of biliary CA 19-9, CA 125 and CEA sampled in different situations for the diagnosis of cholangiocarcinoma. Bile was obtained from patients with bile duct obstruction on the day of biliary drainage and 3 days later separately. The etiology of biliary obstruction included choledocholithiasis (N = 51), hepatolithiasis (N = 19) and cholangiocarcinoma (N = 28). Patients of the former two conditions were all complicated with cholangitis. The sensitivity of biliary CEA, CA 19-9 and CA 125, whenever checked were all less than 70%. The biliary CEA and CA 19-9 were elevated in the presence of cholangitis. In addition, the biliary CEA was also increased in the patients with hepatolithiasis. The specificity of CA 125 was better than those of CEA and CA 19-9 (75.7% vs. 33.3% and 60%, respectively) on the day of biliary drainage. The diagnostic efficiency was slightly improved when combining biliary CA 125 and CEA. As the biliary CA 125 was less affected by inflammation and hepatolithiasis, it is more useful than CEA and CA 19-9 in the diagnosis of cholangiocarcinoma. The single tumor marker test of CA 125 or combined tumor marker test of CEA and CA 125 may be used as a useful complement to other investigative methods to differentiate benign from malignant causes of the bile duct obstruction.
Article
In order to confirm 14-3-3 sigma (sigma) protein distribution in human tissues, immunohistochemistry was performed using various paraffin-embedded human tissues. In normal human tissues, the strongest immunoreactivity for 14-3-3sigma protein was observed in squamous epithelia at various sites, followed by basal cells of the trachea, bronchus and basal or myoepithelial cells of various glands. Moderate to weak 14-3-3sigma immunoreactivity was seen in the epithelial cells of the alimentary tract, gall bladder, urinary tract and endometrium. In the lung, 14-3-3sigma immunoreactivity was also observed in hyperplastic type II alveolar cells and metaplastic squamous cells. Immunohistochemical study using non-small-cell lung cancers revealed that 14-3-3sigma immunoreactivity was stronger in squamous cell carcinomas than in adenocarcinomas. The present study revealed that 14-3-3sigma expression was exclusively present in various epithelial cells and had a tendency to be stronger in cells destined for squamous epithelium or differentiating toward squamous cells in human normal and neoplastic cells.
Article
Surgery remains the only curative treatment option for cholangiocarcinoma (CC). Currently, both early identification of CC in affected individuals at high risk and accurate diagnosis of unexplained biliary strictures are problematic. However, growing insights into biochemical and molecular mechanisms underlying biliary carcinogenesis have suggested serum and bile markers for the diagnosis of CC. These tools include tumor antigens or products (e.g., carbohydrate antigen [CA] 19-9), cytokines (e.g., interleukin-6), metabolic products (e.g., lactate), proteases (e.g., trypsinogen-2), regulatory peptides (e.g., pancreatic polypeptide), and (epi-)genetic lesions (e.g., K- ras and p53 mutations, p16 (INK4a) or p14 (ARF) promoter hypermethylation). In this article we discuss these new potential tumor markers for the diagnosis of CC.
Article
The distinction among inflammatory, benign, and malignant lesions of the biliary tract can at times be difficult. Several methods have been used, including immunohistochemistry (IHC), with variable success. We evaluated a panel of IHC stains to determine their utility in discriminating between bile duct lesions. Formalin-fixed, paraffin-embedded 4-microm sections from 12 inflammatory lesions, 10 bile duct adenomas, and 13 bile duct carcinomas were immunostained using a modified avidin-biotin-complex technique after epitope enhancement using antibodies for p53, Ki-67, and bcl-2. For p53 and bcl-2, greater than 1% of cells staining positive was interpreted as positive. The proliferation index was calculated by determining the number of Ki-67-positive cells in a 1000 cell count. In the inflammatory group, 0 of 12 reacted with anti-p53, 2 of 12 were positive with anti-bcl-2, and the proliferation index with was 22.9% +/- 3.9%. Two of 10 bile duct adenomas showed reactivity with anti-bcl-2, and none were decorated with anti-p53 or Ki-67. In the carcinoma group, 6 of 13 were positive with anti-p53, 9 of 12 were positive with anti-bcl-2, and the proliferation index was 35.3% +/- 5.5%. The proliferation rates differed significantly between groups (P < 0.05). The presence of bcl-2 and p53 immunoreactivity coupled with a high proliferative rate in a biliary tract lesion suggests a malignant process. A panel using these antibodies may be useful in difficult cases.
Article
14-3-3 sigma is an exclusive epithelial marker and data on its expression in different malignancies are very scarce. The aims of the present study are to screen its expression in the most common neoplasms occurring in the urological and gynecological tract and to evaluate its use as a diagnostic marker. A tissue microarray was constructed using 350 samples from 13 different neoplasms. Immunohistochemical analysis using a polyclonal 14-3-3 sigma antibody was performed. Overall, this protein was positive in 141 and negative in 209 tumors. The most frequent expression was seen in squamous cell carcinoma of the cervix and urothelial bladder carcinoma, followed by prostatic and endometrial adenocarcinoma. 14-3-3 sigma was able to distinguish prostate adenocarcinoma from urothelial bladder carcinoma, with an odds ratio of 0.028 (P = 0.001; 95% CI, 0.0003-0.222), and distinguish seminoma from embryonal carcinoma of the testis, with an odds ratio of 0.061 (P = 0.009; 95% CI, 0.007-0.5014). It also has a good value in differentiating renal clear cell carcinoma from papillary carcinoma, with an odds ratio 0.470 (P < 0.001; 95% CI, 0.008-0.261). 14-3-3 sigma seems to have good potential use as an epithelial marker, after confirmation with further targeted studies. Finally, as with all immunohistochemical markers, we can optimize the utility of this protein to distinguish tumor mimics by including it in an appropriate immunohistochemical panel.
Article
CD56 (neuronal cell adhesion molecule, N-CAM) has been reported in neuroendocrine tumours and as a marker of reactive biliary epithelial cells. However, up to date, it is not used to distinguish malignant from non-malignant biliary lesions. In this study, we systematically examined CD56 expression on 98 tumours arising from the biliary tree as well as intrahepatic conditions with reactive neoductules. When neuroendocrine carcinomas are excluded, only 4 of 32 (12.5%) cholangiocarcinomas expressed CD56, 2 of which showed clear cell morphology. Reactive bile ductules adjacent to cirrhotic nodules as well as in focal nodular hyperplasia were CD56 positive. Twelve of 17 (70.5%) bile duct adenomas were CD56 positive, whereas von Meyenburg complexes expressed CD56 only very focally in less than 5% of lesional cells. Bile duct cysts were negative for CD56 with the exception of focally interspersed neuroendocrine cells, similar to that seen in segmental bile ducts. Thus, if van Meyenburg complexes are excluded, CD56 can be used to differentiate intrahepatic non-neoplastic from neoplastic proliferations, which is a helpful diagnostic tool in small liver biopsies.
Article
The major heat shock protein (Hsp) chaperones Hsp70 and Hsp90 both bind the co-chaperone Hop (Hsp70/Hsp90 organizing protein), which coordinates Hsp actions in folding protein substrates. Hop contains three tetratricopeptide repeat (TPR) domains that have binding sites for the conserved EEVD C termini of Hsp70 and Hsp90. Crystallographic studies have shown that EEVD interacts with positively charged amino acids in Hop TPR-binding pockets (called carboxylate clamps), and point mutations of these carboxylate clamp positions can disrupt Hsp binding. In this report, we use circular dichroism to assess the effects of point mutations and Hsp70/Hsp90 peptide binding on Hop conformation. Our results show that Hop global conformation is destabilized by single point mutations in carboxylate clamp positions at pH 5, while the structure of individual TPR domains is unaffected. Binding of peptides corresponding to the C termini of Hsp70 and Hsp90 alters the global conformation of wild-type Hop, whereas peptide binding does not alter conformation of individual TPR domains. These results provide biophysical evidence that Hop-binding pockets are directly involved with domain:domain interactions, both influencing Hop global conformation and Hsp binding, and contributing to proper coordination of Hsp70 and Hsp90 interactions with protein substrates.
Article
The aim of this study was to identify aberrantly expressed proteins in pediatric primitive neuroectodermal tumors (PNETs) and ependymomas. Tumor tissue of 29 PNET and 12 ependymoma patients was subjected to 2-dimensional difference gel electrophoresis. Gel analysis resulted in 79 protein spots being differentially expressed between PNETs and ependymomas (p < 0.01, fold change difference in expression >2). Three proteins, stathmin, annexin A1, and calcyphosine, were chosen for validation by immunohistochemistry. Stathmin was expressed 2.6-fold higher in PNETs than in ependymomas, and annexin A1 and calcyphosine were expressed 2.5- and 37.6-fold higher, respectively, in ependymomas. All PNETs showed strong staining for stathmin, and all ependymomas were strongly positive for annexin A1, whereas control tissues were negative. Calcyphosine immunoreactivity was observed in 59% of the ependymomas and was most profound in ependymoma tissue showing epithelial differentiation. mRNA expression levels of stathmin, annexin A1, and calcyphosine significantly correlated (Rs = 0.65 [p < 0.0001], Rs = 0.50 [p = 0.001], and Rs = 0.72 [p < 0.0001], respectively) with protein expression levels. In conclusion, using a proteome-wide approach, stathmin, annexin A1, and calcyphosine were successfully identified as tumor-specific proteins in pediatric PNETs and ependymomas. Ongoing studies are focused on characterizing the role of these proteins as tumor markers and potential drug targets in pediatric brain tumors.
Article
Difficult diagnoses and a lack of effective therapy complicate biliary tract malignancies. Interleukin-6 (IL-6) is a human bile duct epithelium growth factor correlated with tumor burden. We evaluated the usefulness of serum IL-6 in the diagnosis of primary BDC and measured changes in serum IL-6 levels following photodynamic therapy (PDT). We prospectively measured serum IL-6 levels in patients with BDC (N = 26: 14 patients treated with PDT, 12 with biliary drainage alone), hepatocelluar carcinoma (HCC, N = 26), and healthy adults (N = 23). Serum IL-6 levels were measured by an enzyme-linked immunosorbent assay. Patients with clinical conditions known to increase IL-6 levels were excluded. IL-6 was detected in all patients with BDC and HCC, and in 6 of 23 healthy adults. Median and mean levels of IL-6 were higher in patients with BDC than in both other groups (P < 0.001). Using a 25.8 pg/mL cutoff, IL-6 provided a diagnostic sensitivity of 73% and a specificity of 92%; positive and negative predictive values were 83% and 87%, respectively. Serum levels of IL-6 were correlated with tumor burden in BDC patients. One month after treatment of BDC with PDT, the mean IL-6 level decreased significantly from 282.1 +/- 121.8 to 38.2 +/- 9.9 pg/mL (P= 0.008). Serum IL-6 concentration is a useful addition to the available tests for the differential diagnosis of BDC, and serves as a marker for monitoring the response to treatment of BDC with PDT.
Article
Cruciforms are secondary DNA structures, serving as recognition signals at or near eukaryotic (yeast and mammalian) origins of DNA replication. The cruciform-binding protein is a member of the 14-3-3 protein family and binds to origins of DNA replication in a cell cycle-dependent manner. Five 14-3-3 protein isoforms (beta, gamma, epsilon, zeta and sigma) have been identified as having cruciform binding activity.
Article
To identify novel nasopharyngeal carcinoma (NPC) biomarkers by laser capture microdissection and a proteomic approach. Proteins from pooled microdissected NPC and normal nasopharyngeal epithelial tissues (NNET) were separated by two-dimensional gel electrophoresis, and differential proteins were identified by mass spectrometry. Expression of three differential proteins (stathmin, 14-3-3sigma, and annexin I) in the above two tissues as well as four NPC cell lines was determined by Western blotting. Immunohistochemistry was also done to detect the expression of three differential proteins in 98 cases of primary NPC, 30 cases of NNET, and 20 cases of cervical lymph node metastases, and the correlation of their expression levels with clinicopathologic features and clinical outcomes were evaluated. Thirty-six differential proteins between the NPC and NNET were identified. The expression levels of stathmin, 14-3-3sigma, and annexin I in the two types of tissues were confirmed and related to differentiation degree and/or metastatic potential of the NPC cell lines. Significant stathmin up-regulation and down-regulation of 14-3-3sigma and annexin I were observed in NPC versus NNET, and significant down-regulation of 14-3-3sigma and annexin I was also observed in lymph node metastasis versus primary NPC. In addition, stathmin up-regulation and down-regulation of 14-3-3sigma and annexin I were significantly correlated with poor histologic differentiation, advanced clinical stage, and recurrence, whereas down-regulation of 14-3-3sigma and annexin I was also significantly correlated with lymph node and distant metastasis. Furthermore, survival curves showed that patients with stathmin up-regulation and down-regulation of 14-3-3sigma and annexin I had a poor prognosis. Multivariate analysis revealed that the expression status of stathmin, 14-3-3sigma, and annexin I was an independent prognostic indicator. The data suggest that stathmin, 14-3-3sigma, and annexin I are potential biomarkers for the differentiation and prognosis of NPC, and their dysregulation might play an important role in the pathogenesis of NPC.
Utility of serum tumor markers, imaging, and biliary cytology for detecting cholangiocarcinoma in primary sclerosing cholangitis
  • P Charatcharoenwitthaya
  • Fb Enders
  • Kc Halling
Charatcharoenwitthaya P, Enders FB, Halling KC, et al. Utility of serum tumor markers, imaging, and biliary cytology for detecting cholangiocarcinoma in primary sclerosing cholangitis. Hepatology 2008;48:1106–17.
Liver—Intrahepatic Bile Ducts Classification of malignant tumours
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Wittekind Ch. Liver—Intrahepatic Bile Ducts. In: Sobin LGM, Wittekind C, eds. Classification of malignant tumours. 7th edn. New York: Wiley-Blackwell, 2009:114–17.
[Recommendation for uniform definition of an immunoreactive score (IRS) for immunohistochemical estrogen receptor detection (ER-ICA) in breast cancer tissue]
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Remmele W, Stegner HE. [Recommendation for uniform definition of an immunoreactive score (IRS) for immunohistochemical estrogen receptor detection (ER-ICA) in breast cancer tissue]. Pathologe 1987;8:138-40.
doi:10.1136/jclinpath-2015-203418 Original article group
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Bertram S, et al. J Clin Pathol 2016;0:1–8. doi:10.1136/jclinpath-2015-203418 Original article group.bmj.com on January 14, 2016 -Published by http://jcp.bmj.com/ Downloaded from
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Theise ND. Liver cancer. In: Steward BW, ed. World Cancer Report 2014. Lyon: International Agency for Research on Cancer, 2014:403-12.
WHO classification of tumours of the digestive system
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  • Intrahepatic