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Introduction: Lysophosphatidylcholine acyltransferase 1 (LPCAT1) has repeatedly been suggested to be associated with tumorigenesis. To evaluate the role of LPCAT1 in esophageal cancer, LPCAT1 immunostaining was analyzed on a tissue microarray containing samples from esophageal cancer patients., Results: In benign esophageal tissue, LPCAT1 staining was detectable in low intensities. LPCAT1 staining was increased in malignant as compared to benign esophageal tissue and was found in high intensity in 26.4% of 288 interpretable esophageal adenocarcinomas (EACs) and in 23.2% of 211 squamous cell carcinomas (ESCCs). Increased LPCAT1 staining was linked to undifferentiated tumor grading in both subtypes of EACs and ESCCs (p = 0.0273 and p = 0.0085)., Conclusion: However, LPCAT1 was not associated with prognosis of EAC and ESCC patients (p = 0.6838 and p = 0.4695) and thus cannot be considered a prognostic biomarker in esophageal cancers., (© 2021. The Author(s).)

Data on Mesothelin (MSLN) expression in human normal and cancerous tissues is controversial. We employed immunohistochemistry (IHC) on a tissue microarray (TMA) from 599 pancreatic cancers and 12 large tissue sections of pancreatitis. MSLN expression was highest in pancreatic adenocarcinomas (89%) and adenocarcinomas of the ampulla Vateri (79%), infrequent in pancreatitis and absent in 6 acinus cell carcinomas and normal pancreas. MSLN expression was unrelated to pathological tumor stage, grade, metastasis, and tumor-infiltrating CD8+ lymphocytes. In conclusion, pancreatic cancer may be ideally suited for putative anti- MSLN therapies, and MSLN may represent a suitable biomarker for pancreatic cancer diagnosis, especially on small biopsies.

Lung cancer continues to be the leading cause for cancer-related deaths in men and women worldwide. Sufficient screening tools enabling early diagnosis are essential to improve patient outcomes. The aim of this study was to evaluate serum midkine (S-MK) both as a diagnostic and prognostic biomarker in non-small cell lung cancer (NSCLC). This single-center analysis included 59 NSCLC patients counting 30 squamous cell cancers and 29 adenocarcinomas. Preoperative S-MK concentration was determined using ELISA. Patients were followed up to five years. S-MK was found to be significantly overexpressed in patients with NSCLC compared to healthy controls (p < 0.001). The discriminative power of S-MK to differentiate NSCLC subjects from controls was fairly high with an area under the receiver operating characteristic curve of 0.83 (p < 0.001). Optimal sensitivity of 92% and reasonable specificity of 68% was reached at a threshold of 416 pg/ml S-MK. Patients with high S-MK concentration showed a significantly shorter overall survival compared to patients with low S-MK expression (p < 0.05). In conclusion, S-MK is overexpressed in patients with NSCLC and serves as an independent prognostic factor for overall survival. S-MK may thus be considered as an additional non-invasive biomarker not only for NSCLC screening but also for outcome prediction., (© 2021. The Author(s).)

Colorectal cancer (CRC) is one of the leading causes of cancer death worldwide. There is an urgent need to identify prognostic markers for patients undergoing curative resection of CRC. The detection of circulating tumor cells in peripheral blood is a promising approach to identify high-risk patients with disseminated disease in colorectal cancer. This study aims to evaluate the prognostic relevance of preoperative CTCs using the Cellsearch® system (CS) in patients, who underwent resection with curative intent of different stages (UICC I-IV) of colorectal cancer. Out of 91 Patients who underwent colorectal resection, 68 patients were included in this study. CTC analysis was performed in patients with CRC UICC stages I-IV immediately before surgery. Data were correlated with clinicopathological parameters and patient outcomes. One or more CTCs/7.5 mL were detected in 45.6% (31/68) of patients. CTCs were detected in all stages of the Union of International Cancer Control (UICC), in stage I (1/4, 25%), in stage II (4/12, 33.3%), in stage III (5/19, 26.3%) and in stage IV (21/33, 63.6%). The detection of ≥ 1 CTCs/ 7.5ml correlated to the presence of distant overt metastases (p = 0.014) as well as with shorter progression-free (p = 0.008) and overall survival (p = 0.008). Multivariate analyses showed that the detection of ≥ 1 CTCs/ 7.5ml is an independent prognostic indicator for overall survival (HR, 3.14; 95% CI, 1.18-8.32; p = 0.021). The detection of CTCs is an independent and strong prognostic factor in CRC, which might improve the identification of high-risk patients in future clinical trials., Competing Interests: The authors have declared that no competing interests exist.

Genome-wide association studies (GWAS) of esophageal adenocarcinoma (EAC) and its precursor, Barrett's esophagus (BE), have uncovered significant genetic components of risk, but most heritability remains unexplained. Targeted assessment of genetic variation in biologically relevant pathways using novel analytical approaches may identify missed susceptibility signals. Central obesity, a key BE/EAC risk factor, is linked to systemic inflammation, altered hormonal signaling and insulin-like growth factor (IGF) axis dysfunction. Here, we assessed IGF-related genetic variation and risk of BE and EAC. Principal component analysis was employed to evaluate pathway-level and gene-level associations with BE/EAC, using genotypes for 270 single-nucleotide polymorphisms (SNPs) in or near 12 IGF-related genes, ascertained from 3295 BE cases, 2515 EAC cases and 3207 controls in the Barrett's and Esophageal Adenocarcinoma Consortium (BEACON) GWAS. Gene-level signals were assessed using Multi-marker Analysis of GenoMic Annotation (MAGMA) and SNP summary statistics from BEACON and an expanded GWAS meta-analysis (6167 BE cases, 4112 EAC cases, 17 159 controls). Global variation in the IGF pathway was associated with risk of BE (P = 0.0015). Gene-level associations with BE were observed for GHR (growth hormone receptor; P = 0.00046, false discovery rate q = 0.0056) and IGF1R (IGF1 receptor; P = 0.0090, q = 0.0542). These gene-level signals remained significant at q < 0.1 when assessed using data from the largest available BE/EAC GWAS meta-analysis. No significant associations were observed for EAC. This study represents the most comprehensive evaluation to date of inherited genetic variation in the IGF pathway and BE/EAC risk, providing novel evidence that variation in two genes encoding cell-surface receptors, GHR and IGF1R, may influence risk of BE., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objective: Anoctamin 7 (ANO7) is a calcium 2+ -dependent chloride ion channel protein. Its expression is restricted to prostate epithelial cells. The exact function is unknown. This study aimed to analyze ANO7 expression and its clinical significance in prostate cancer (PCa)., Methods: ANO7 expression was assessed by immunohistochemistry in 17,747 clinical PCa specimens., Results: ANO7 was strongly expressed in normal prostate glandular cells but often less abundant in cancer cells. ANO7 staining was interpretable in 13,594 cancer tissues and considered strong in 34.4%, moderate in 48.7%, weak in 9.3%, and negative in 7.6%. Reduced staining was tightly linked to adverse tumor features [high classical and quantitative Gleason grade, lymph node metastasis, advanced tumor stage, high Ki67 labeling index, positive surgical margin, and early biochemical recurrence ( P < 0.0001 each)]. The univariate Cox hazard ratio for prostate-specific antigen (PSA) recurrence after prostatectomy in patients with negative vs. strong ANO7 expression was 2.98 (95% confidence interval 2.61-3.38). The prognostic impact was independent of established pre- or postoperatively available parameters ( P < 0.0001). Analysis of annotated molecular data showed that low ANO7 expression was linked to TMPRSS2:ER G fusions ( P < 0.0001), elevated androgen receptor expression ( P < 0.0001), as well as presence of 9 of 11 chromosomal deletions ( P < 0.05 each). A particularly strong association of low ANO7 expression with phosphatase and tensin homolog (PTEN) deletion may indicate a functional relationship with the PTEN/AKT pathway., Conclusions: These data identify reduced ANO7 protein expression as a strong and independent predictor of poor prognosis in PCa. ANO7 measurement, either alone or in combination, might provide clinically useful prognostic information in PCa., Competing Interests: Conflict of interest statement No potential conflicts of interest are disclosed., (Copyright: © 2021, Cancer Biology & Medicine.)

Background & Aims: Esophageal adenocarcinoma (EA) and its premalignant lesion, Barrett's esophagus (BE), are characterized by a strong and yet unexplained male predominance (with a male-to-female ratio in EA incidence of up to 6:1). Genome-wide association studies (GWAS) have identified more than 20 susceptibility loci for these conditions. However, potential sex differences in genetic associations with BE/EA remain largely unexplored., Methods: Given strong genetic overlap, BE and EA cases were combined into a single case group for analysis. These were compared with population-based controls. We performed sex-specific GWAS of BE/EA in 3 separate studies and then used fixed-effects meta-analysis to provide summary estimates for >9 million variants for male and female individuals. A series of downstream analyses were conducted separately in male and female individuals to identify genes associated with BE/EA and the genetic correlations between BE/EA and other traits., Results: We included 6758 male BE/EA cases, 7489 male controls, 1670 female BE/EA cases, and 6174 female controls. After Bonferroni correction, our meta-analysis of sex-specific GWAS identified 1 variant at chromosome 6q11.1 (rs112894788, KHDRBS2-MTRNR2L9, P BONF  = .039) that was statistically significantly associated with BE/EA risk in male individuals only, and 1 variant at chromosome 8p23.1 (rs13259457, PRSS55-RP1L1, P BONF  = 0.057) associated, at borderline significance, with BE/EA risk in female individuals only. We also observed strong genetic correlations of BE/EA with gastroesophageal reflux disease in male individuals and obesity in female individuals., Conclusions: The identified novel sex-specific variants associated with BE/EA could improve the understanding of the genetic architecture of the disease and the reasons for the male predominance., (Copyright © 2020 AGA Institute. All rights reserved.)

Secreted frizzled-related protein 4 (SFRP4) controls WNT signaling and is thought to play a role for tumor aggressiveness. Here, we analyzed a tissue microarray containing 11,152 prostate cancers with pathological, clinical and molecular data by immunohistochemistry. SFRP4 expression was higher in cancer than in non-neoplastic acinar cells. SFRP4 staining was seen in 64.9% of tumors and classified as weak in 33.2%, moderate in 23.9% and strong in 7.8% of cancers. SFRP4 overexpression was linked to advanced tumor stage, high classical/quantitative Gleason grade (p < 0.0001 each), lymph node metastasis (p = 0.0002), and a positive surgical margin (p = 0.0017). SFRP4 positivity was markedly more frequent in ERG positive (77.4%) than in ERG negative cancers (57.4% p < 0.0001). Subset analyses in 2725 cancers with and 3592 cancers without TMPRSS2:ERG fusion revealed that associations with tumor phenotype and patient outcome were largely driven by the subset of ERG negative tumors. In a multivariate analysis including various postoperative and prognostic clinico-pathological features, SFRP4 protein expression emerged as an independent prognostic parameter in ERG negative cancers. SFRP4 immunostaining was significantly linked with 10 of 11 previously analyzed chromosomal deletions (p < 0.05 each). In conclusion, high SFRP4 immunostaining is associated with poor prognosis and genomic instability in ERG negative prostate cancers.

Colorectal carcinoma (CRC) is one of the most common carcinomas worldwide. Early detection is crucial for reducing morbidity and mortality. Several promising studies described the use of midkine (MK) as a tumor marker. This study aimed to investigate a larger collective to ascertain if the preoperative serum midkine level (S-MK) is suitable as a marker for screening and if S-MK correlates with tumor progression and localization. It was also investigated for the first time whether patients with high S-MK show poor survival. This prospective single-center study included 299 patients with CRC. The preoperative serum midkine level (S-MK) was determined using ELISA. Established tumor markers Carcinoembryonic antigen (CEA) and Carbohydrate antigen 19-9 (CA 19-9) were collected for comparison. The median follow-up period was 65 months. S-MK was significantly elevated in patients with CRC (P < .001). The receiver operation characteristic (ROC) curve has an area under the curve (AUC) of 0.868 (P < .001). A cut-off value of 56.42 pg/mL results in a sensitivity of 84.3% and a specificity of 75.4%. In the one-way analysis of variance (ANOVA), there were no significant correlations between S-MK and tumor progression, localization. Furthermore, no significant correlation to CEA und CA 19-9 could be found. Kaplan-Meier survival analysis was able to show for the first time that patients with S-MK of more than 225 pg/mL have a significantly shorter survival. Multivariate Cox regression showed that only CEA was an independent prognostic factor for survival. S-MK helps estimate the prognosis for CRC and is a valuable component for developing a multimarker panel for screening and surveillance., (© 2020 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Survivin is an inhibitor of apoptosis. Aberrant survivin expression occurs in malignant tumors and has often been linked to unfavorable patient outcome. Here we analyzed 12 432 prostate cancers by immunohistochemistry. Survivin immunostaining was regularly expressed at high levels in normal prostate epithelium but expression was often reduced in prostate cancers. Among 9492 evaluable prostate cancers, 9% expressed survivin strongly, 19% moderately, 28% weakly, and 44% lacked it. Loss of cytoplasmic survivin was seen in advanced tumor stage, higher Gleason score, preoperative PSA levels, and Ki-67 labeling index, and associated with earlier PSA recurrence (P < .0001). Survivin loss was significantly more common in cancers carrying TMPRSS2:ERG fusions (61% survivin negative) than in ERG wild-type cancers (32% survivin negative; P < .0001). Multivariate analysis revealed that reduced cytoplasmic survivin expression predicted poor prognosis independent from Gleason score, pT, pN, and serum PSA level. This was valid for ERG-positive and ERG-negative cancers. Survivin expression loss even retained its prognostic impact in 1020 PTEN deleted cancers, a group that is already characterized by dismal patient prognosis. In conclusion, reduced survivin expression is associated with more aggressive tumors and inferior prognosis in prostate cancer., (© 2019 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Syndecan-1 (CD138) is a transmembrane proteoglycan known to be expressed in various normal and malignant tissues. It is of interest because of a possible prognostic role of differential expression in tumors and its role as a target for indatuximab, a monoclonal antibody coupled with a cytotoxic agent. To comprehensively analyze CD138 in normal and neoplastic tissues, we used tissue microarrays (TMAs) for analyzing immunohistochemically detectable CD138 expression in 2,518 tissue samples from 85 different tumor entities and 76 different normal tissue types. The data showed that CD138 expression is abundant in tumors. At least an occasional weak CD138 immunostaining could be detected in 71 of 82 (87%) different tumor types, and 58 entities (71%) had at least one tumor with a strong positivity. In normal tissues, a particularly strong expression was found in normal squamous epithelium of various organs, goblet and columnar cells of the gastrointestinal tract, and in hepatocytes. The highly standardized analysis of most human cancer types resulted in a ranking order of tumors according to the frequency and levels of CD138 expression. CD138 immunostaining was highest in squamous cell carcinomas such as from the esophagus (100%), cervix uteri (79.5%), lung (85.7%), vagina (89.7%) or vulva (73.3%), and in invasive urothelial cancer (76.2%). In adenocarcinomas, CD138 was also high in lung (82.9%) and colorectal cancer (85.3%) but often lower in pancreas (73.3%), stomach (54.2% in intestinal type), or prostate carcinomas (16.3%). CD138 expression was usually low or absent in germ cell tumors, sarcomas, endocrine tumors including thyroid cancer, and neuroendocrine tumors. In summary, the preferential expression in squamous cell carcinomas of various sites makes these cancers prime targets for anti-CD138 treatments once these might become available. Abundant expression in many different normal tissues might pose obstacles to exploiting CD138 as a therapeutic target, however., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2019 Simon Kind et al.)

Background: The cell adhesion molecule close homologue of L1 (CHL1) is a potential tumour suppressor and was recently detected in non-small cell lung cancer (NSCLC) specimens. The expression pattern, prognostic, and functional role of CHL1 in NSCLCs is unknown., Methods: We evaluated the protein expression of CHL1 by immunohistochemistry in 2161 NSCLC patients based on a tissue microarray. The results were correlated with clinical, histopathological, and patient survival data (Chi square test, t test, and log-rank test, respectively). A multivariate analysis (Cox regression) was performed to validate its impact on patients' survival., Results: CHL1 was expressed in NSCLC patients and was significantly overexpressed in lung adenocarcinomas and squamous cell carcinomas compared to neuroendocrine and large cell carcinomas of the lung (p < 0.001). CHL1 expression was associated with the T stage in adenocarcinomas (p = 0.011) and with metastatic lymph node status and UICC stage in squamous cell carcinomas (p = 0.034 and p = 0.035, respectively). Increased CHL1 expression was associated with improved survival in univariate (p = 0.031) and multivariate analyses (odds ratio 0.797, 95% confidence interval 0.677-0.939, p = 0.007)., Conclusion: The prognostic significance of CHL1 makes it a potential prognostic and therapeutic target and underlines its role as a tumour suppressor. Further validation studies and functional analyses are needed to investigate its potential role in tumourigenesis and dissemination.

Expression of the retinoic acid-induced protein 3 (RAI3) has been suggested to predict clinical outcome in a variety of malignancies. However, its role in esophageal cancers remains unclear. Immunohistochemical RAI3 staining was analyzed on tissue microarrays containing 359 esophageal adenocarcinomas (EAC) and 254 esophageal squamous cell carcinomas (ESCC). RAI3 immunostaining was typically absent or weakly detectable in the membranes in benign esophageal tissues. RAI3 staining was higher in malignant than in benign esophagus epithelium. High-levels of RAI3 staining were found in 79.2% of interpretable EACs and 55.9% of ESCCs. In EACs, strong RAI3 staining was associated with advanced pathological tumor stage (p < .0001), high UICC stage (p < .0001), high tumor grade (p = .0133), and positive lymph nodal status (p = .0002). Additionally, high RAI3 staining predicted shortened overall survival of EAC and ESCC patients (p = .0298 and p = .0227). RAI3 overexpression is associated with poor prognosis in esophageal cancers. We propose that RAI3 overexpression might play a biologically relevant role of RAI3 in esophageal cancers., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background/aim: The clinical significance of circulating tumor cells (CTC) in non-metastatic esophageal cancer (EC) remains controversial and the cellular and molecular characteristics of CTCs are poorly understood. Especially the frequency and oncological impact of HER2 expression in CTCs in patients with EC have not been evaluated until now., Materials and Methods: In this single-center, prospective study, peripheral blood samples were obtained preoperatively from 45 patients who were diagnosed with resectable EC. CTC detection and HER2 expression were performed using the CellSearch System. Data were correlated with clinicopathological parameters and patient outcomes., Results: The study included 13 patients with squamous cell carcinomas (SCC) and 32 patients with adenocarcinomas (AC). HER2 gene amplification in the primary tumor was detected in 9.1% of patients. One or more CTCs were detected in 15.6% (SCC 1/13; AC 6/32) of the patients. None of the detected CTCs showed HER2 expression. Patients with CTCs showed significantly shorter relapse-free (p<0.001) and overall survival (p=0.015) than CTC-negative patients., Conclusion: This is the first study analyzing HER2 expression and the clinical significance of CTCs in patients with non-metastatic EC using an automated immunomagnetic detection system. HER2 expression in CTCs is very rare in patients with non-metastatic EC and seems to have a low clinical and oncological impact., (Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Pancreatic ductal adenocarcinoma (PDAC) is a very aggressive tumor with a five-year survival of less than 6%. Chronic pancreatitis (CP), an inflammatory process in of the pancreas, is a strong risk factor for PDAC. Several genetic polymorphisms have been discovered as susceptibility loci for both CP and PDAC. Since CP and PDAC share a consistent number of epidemiologic risk factors, the aim of this study was to investigate whether specific CP risk loci also contribute to PDAC susceptibility. We selected five common SNPs (rs11988997, rs379742, rs10273639, rs2995271 and rs12688220) that were identified as susceptibility markers for CP and analyzed them in 2,914 PDAC cases, 356 CP cases and 5,596 controls retrospectively collected in the context of the international PANDoRA consortium. We found a weak association between the minor allele of the PRSS1-PRSS2-rs10273639 and an increased risk of developing PDAC (OR homozygous  = 1.19, 95% CI 1.02-1.38, p = 0.023). Additionally all the SNPs confirmed statistically significant associations with risk of developing CP, the strongest being PRSS1-PRSS2-rs10273639 (OR heterozygous  = 0.51, 95% CI 0.39-0.67, p = 1.10 × 10 -6 ) and MORC4-rs 12837024 (OR homozygous  = 2.07 (1.55-2.77, p trend  = 0.7 × 10 -11 ). Taken together, the results from our study do not support variants rs11988997, rs379742, rs10273639, rs2995271 and rs12688220 as strong predictors of PDAC risk, but further support the role of these SNPs in CP susceptibility. Our study suggests that CP and PDAC probably do not share genetic susceptibility, at least in terms of high frequency variants., (© 2017 UICC.)

FAM13C, a gene with unknown function is included in several mRNA signatures for prostate cancer aggressiveness. To understand the impact of FAM13C on prognosis and its relationship to molecularly defined subsets, we analyzed FAM13C expression by immunohistochemistry on a tissue microarray containing 12,400 prostate cancer specimens. Results were compared to phenotype, ERG status, genomic deletions of 3p, 5q, 6q and PTEN, and biochemical recurrence. FAM13C was detectable in cell nuclei of cancerous and non-neoplastic prostate cells. 67.5% of 9,633 interpretable cancers showed FAM13C expression: strong in 28.3%, moderate in 24.6% and weak in 14.6%. Strong FAM13C expression was linked to advanced pT stage, high Gleason grade, positive lymph node status, and early biochemical recurrence (p < 0.0001 each). FAM13C expression was associated with TMPRSS2:ERG fusions. It was present in 85% of ERG positive but in only 54% of ERG negative cancers (p < 0.0001), and in 91.1% of PTEN deleted but in only 69.2% of PTEN non-deleted cancers (p < 0.0001). The prognostic role of FAM13C expression was independent of classical and quantitative Gleason grade, pT stage, pN stage, surgical margin status and preoperative PSA. In conclusion, the results of our study demonstrate that expression of FAM13C is an independent prognostic marker in prostate cancer. Finding FAM13C also in non-neoplastic prostate tissues highlights the importance of properly selecting cancer-rich areas for RNA-based FAM13C expression analysis.

Background/aim: Activated leukocyte cell adhesion molecule (ALCAM/CD166) as a member of the 'immunoglobulin superfamily' is known to be involved in cancer cell proliferation and migration. The aim of this study was to investigate the expression of ALCAM in neuroblastoma tissues., Materials and Methods: ALCAM expression was analyzed in primary neuroblastoma specimens by immunohistochemistry on microarray sections. Histopathological and clinical data were correlated with ALCAM expression and survival analysis was performed., Results: Sixty-six children were included in the study. Strong expression of ALCAM was detected in 52 (79%) of the samples. Weak expression was significantly correlated with the International Neuroblastoma Staging System (INSS) stage (p=0.024) and positive n-MYC amplification (p=0.019). Recurrence-free survival (RFS) and overall survival (OS) were significantly shorter if ALCAM was expressed weakly (p=0.032 and p=0.001)., Conclusion: Weak ALCAM expression was significantly correlated with established markers for poor prognosis, as well as shorter RFS and OS. ALCAM might be considered as a prognostic marker for infantile neuroblastoma., (Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Background: Tenascin-C is overexpressed in the stroma of most solid malignancies and may function as a diagnostic tumor marker. This study was conducted to evaluate the potential significance of Tenascin-C as a predictive marker for tumor progression in the sera of non-small cell lung cancer (NSCLC) patients., Results: Serum concentration of Tenascin-C is significantly elevated in NSCLC patients compared to healthy controls (p=0.013). The sensitivity of Tenascin-C in detecting NSCLC was 74% at a specificity of 57%. Elevated Tenascin-C serum values are associated with larger tumor size and lymph node involvement (p=0.022 and p=0.036, respectively). The Kaplan-Meyer-curves showed a significant association of Tenascin-C with the patient's overall survival (p=0.004), but not with the recurrence-free survival (p=0.328)., Methods: We quantified Tenascin-C in the sera of 103 NSCLC patients and 76 healthy blood donors by enzyme-linked immune-absorbance assay tests. Prognostic significance was determined by area under the curve analysis and Youden-index. The results were correlated with clinical, histopathological, and patient survival data (Chi-square test, Kaplan-Meier analysis, log-rank test, multivariate Cox-regression analysis)., Conclusion: Although significantly elevated in patients with NSCLC, the sensitivity and specificity of the Tenascin-C serum quantification test was low. However, although failing to be an independent prognosticator in multivariate analysis, the results implicate Tenascin-C as a predictive prognostic marker for NSCLC patients. The data must be further validated in future prospective trials with larger patient cohorts.

SALL4 is a transcription factor originally identified as a homeotic gene essential for organ development. Early studies suggested that SALL4 is a useful marker to identify testicular and ovarian germ cell tumors. The aim of the study was to evaluate the diagnostic potential of SALL4 immunohistochemistry. Immunohistochemical staining was performed on a tissue microarray (TMA) with 3966 samples from 94 different tumor types and on a further TMA with 492 esophagus carcinomas. SALL4 immunostaining was by far most prevalent and most intensive in testicular tumors with a positivity rate of 93.1% in seminomas, 80% in mixed germ cell tumors (embryonic carcinomas/yolk sac tumors), and 18.5% in teratomas, respectively. However, SALL4 expression is not specific to germ cell tumors. We observed SALL4 positivity in non-germ cell tumors as carcinomas of the kidney (28.9% of chromophobe, 34.4% of clear cell carcinoma), in intestinal type adenocarcinoma of the stomach (10.9%), in adenocarcinoma (10.5%) and squamous cell carcinoma (7.2%) of the esophagus, and in malignant melanoma (8.1%) and invasive urothelial bladder carcinoma (20%). SALL4 expression was not found in lymphomas, in soft tissue tumors or breast tumors. At analysis of esophagus carcinoma TMA, no significant association was seen between SALL4 expression and overall survival in adenocarcinoma. However, SALL4 expression was strongly associated with worse overall survival in squamous cell carcinoma. SALL4 expression can be found at relevant frequencies in various tumors of different primary sites. SALL4 expression in squamous cell carcinoma of the esophagus may constitute a sign of dedifferentiation leading to poor patient prognosis.

Background: A genome-wide association study (GWAS) suggested inherited genetic single-nucleotide polymorphisms (SNPs) affecting overall survival (OS) in advanced pancreatic cancer. To identify robust clinical biomarkers, we tested the strongest reported candidate loci in an independent patient cohort, assessed cellular drug sensitivity, and evaluated molecular effects., Methods: This study comprised 381 patients with histologically verified pancreatic ductal adenocarcinoma treated with gemcitabine-based chemotherapy. The primary outcome was the relationship between germline polymorphisms and OS. Functional assays addressed pharmacological dose-response effects in lymphoblastoid cell lines (LCLs) and pancreatic cancer cell lines (including upon RNAi), gene expression analyses, and allele-specific transcription factor binding. All statistical tests were two-sided., Results: The A allele (26% in Caucasians) at SNP rs11644322 in the putative tumor suppressor gene WWOX conferred worse prognosis. Median OS was 14 months (95% confidence interval [CI] = 12 to 15 months), 13 months (95% CI = 11 to 15 months), and nine months (95% CI = 7 to 12 months) for the GG, GA, and AA genotypes, respectively (P trend < .001 for trend in univariate log-rank assuming a codominant mode of inheritance; advanced disease subgroup P trend < .001). Mean OS was 25 months (95% CI = 21 to 29 months), 19 months (95% CI = 15 to 22 months), and 13 months (95% CI = 10 to 16 months), respectively. This effect held true after adjustment for age, performance status according to Eastern Cooperative Oncology Group classification, TNM, grading, and resection status and was comparable with the strongest established prognostic factors in multivariable analysis. Consistently, reduced responsiveness to gemcitabine, but not 5-fluorouracil, along with lower WWOX expression was demonstrated in LCLs harboring the AA genotype. Likewise, RNAi-mediated WWOX knockdown in pancreatic cancer cells confirmed differential cytostatic drug sensitivity. In electrophoretic mobility shift assays, the A allele exhibited weaker binding of Sp family members Sp1/Sp3., Conclusions: WWOX rs11644322 represents a major predictive factor in gemcitabine-treated pancreatic cancer. Decreased WWOX expression may interfere with gemcitabine sensitivity, and allele-specific binding at rs11644332 might be a causative molecular mechanism behind the observed clinical associations., (© The Author 2016. Published by Oxford University Press.)

Aims: RNA-binding motif protein 3 (RBM3) has recently been suggested as a prognostic biomarker in an array of human cancers. This study aimed to examine its effects in colorectal cancers., Methods and Results: RBM3 expression was analysed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers (CRCs). Nuclear RBM3 immunohistochemical staining was found in 95.9% of all interpretable CRCs. Loss of RBM3 expression was linked to advanced tumour stage (P < 0.0001), right-sided tumour localization (P < 0.0001), and poor prognosis (P = 0.0003). In a multivariable analysis including RBM3 staining, tumour grade, tumour stage, and nodal status, only tumour stage and nodal status proved to be independent prognostic markers (P < 0.0001 each), whereas the prognostic impact of RBM3 staining was not significant (P = 0.2655)., Conclusions: Our observations indicate that loss of RBM3 expression is an unfavourable prognostic marker in CRC, and is linked to right-sided tumour localization., (© 2015 John Wiley & Sons Ltd.)

To correlate H2Bub1 expression with outcome in colorectal cancer, H2Bub1 expression was analyzed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers. Results were compared to clinicopathological parameters.H2Bub1 IHC was seen in 1256 (79.3%) of 1584 interpretable CRC and was considered weak in 26.2% and strong in 53.1% of cancers. H2Bub1 expression was completely lost in 20.7% of the cases. Loss of H2Bub1 expression was associated with high tumor grade (p = 0.0211), high tumor stage (p = 0.0003), positive nodal status (p = 0.0139) and histological tumor type (p = 0.0202). No link was found between H2Bub1 expression and tumor localization (p = 0.1262), peritumoral lymphocytic infiltration (p = 0.2523) or vascular invasion (p = 0.5970).Loss of H2Bub1 expression in CRC was strongly associated with poor patient survival (p = 0.0006). This observation held true also in a subset survival analysis of nodal negative (N0) and nodal positive (N1) cancers (p = 0.0296 and p = 0.0197, respectively). In the subgroup of p53 negative cancers no prognostic impact of H2Bub1 staining was seen (p = 0.1924), whereas in p53 positive CRC H2Bub1 expression loss was associated with poor prognosis (p = 0.0031). Strikingly worsened outcome was found for nodal negative cancers presenting with accumulation of p53 when H2Bub1 expression was lost (p = 0.0006).Our data demonstrate that a reduced H2Bub1 expression is a strong prognostic biomarker both in nodal negative and nodal positive CRC. H2Bub1 expression measurement might help to select nodal negative CRC patients that may benefit from adjuvant therapy.

Background: Cdc7 is a widely expressed protein kinase implicated in cell division, cell cycle checkpoint mechanisms and cancer progression. Recently, it has been suggested as a target for anti-cancer therapy., Methods: To determine the relationship of Cdc7 protein expression with tumor phenotype, molecular features and prognosis, 1800 colorectal carcinomas were analyzed by immunohistochemistry on a tissue microarray., Results: Cdc7 expression was considered negative in 33.6%, weak in 57.2% and strong in 9.2% of 1711 interpretable CRCs. Loss of Cdc7 expression was significantly associated with high tumor stage (p < 0.0001) and high tumor grade (p = 0.0077), but was unrelated to the nodal status (p = 0.5957). Moreover, a link between Cdc7 expression and the tubular histological tumor type was seen (p < 0.0001). p53 and Cdc7 expression were significantly linked to each other (p = 0.0013). In a multivariate survival analysis, strong Cdc7 expression of CRC was an independent marker of improved patient survival (p = 0.0031)., Conclusion: Our data show that Cdc7 is highly expressed in CRC and a potential therapeutic target in a subset of cancers with high p53 expression. Moreover, our findings strongly argue for a clinical utility of Cdc7 immunostaining as an independent prognostic biomarker in colorectal cancer enabling to select patients for adjuvant treatment.

Immunologic profiling of colorectal cancer (CRC) may help to predict the tumors metastatic potential and patients with an aggressive tumor, although not yet metastasized at time of surgery might benefit from adjuvant therapy. In this study we evaluated the prognostic significance of FOXP3(+) T regulatory cells (Tregs), CD3(+) and CD8(+) lymphocyte densities and conventional histopathologic features in nodal negative (n = 820, UICC stage II) CRC. Immunohistochemical studies showed that high expression of FOXP3(+) Tregs is significantly linked to a better clinical outcome (P = 0.0001). In multivariate analysis including tumor stage, tumor grade, type of tumor invasion margin (pushing vs. infiltrating type), lymphovascular invasion (absent vs. present), CD3(+), CD8(+) and FOXP3(+) Tregs expression, only low tumor stage, absence of lymphovascular invasion and high Foxp3 Tregs density showed prognostic significance (P = 0.0132, P = 0.0022 AND P = 0.0234, respectively). Our findings argue towards a clinical utility of FOXP3(+) Tregs immunostaining as an independent good prognostic biomarker in stage II colorectal cancers. FOXP3(+) Tregs immunoscoring, assessment of tumor stage and lymphovascular invasion may help to define stage II cancers with a potentially aggressive behavior and CRC patients who might benefit from adjuvant therapy. A two-scale immunosore related to the median count of FOXP3(+) Tregs proved to be easy and quick to perform.

Purpose: To investigate the significance of mammalian target of rapamycin (mTOR) in colorectal cancers. mTOR has recently been suggested as a prognostic biomarker and therapeutic target in an array of human cancers., Findings: phospho-mTOR (p-mTOR) expression was analyzed by immunohistochemistry (IHC) on a tissue microarray containing 1800 colorectal cancers (CRC). Clinical follow-up data were available from all cancer patients. Positive p-mTOR immunostaining was seen in 83.5% of 1640 interpretable CRC and was considered weak in 862 (52.5%) and strong in 508 cases (31.0%). Matching clinico-pathological parameters were available in 1580 cases. p-mTOR staining was more frequent in tubular adenocarcinomas than in the less common histological subtypes (mucinous, medullary, signet cell; P=0.0163) and significantly linked to carcinomas of the left-sided colon and rectum as compared to right-sided CRC (P=0.0066). There was no significant association between p-mTOR expression and patients' gender, tumor stage, tumor grade or nodal status. In a survival analysis, p-mTOR IHC status of all CRC was unrelated to patient survival (P=0.702). In a multivariate analysis including pT, pN, tumor grade, tumor localization and p-mTOR expression, only pT, pN (both P<0.0001) and grade (P=0.0001) showed prognostic impact, but not tumor localization (P=0.9472) or p-mTOR expression (P=0.8879)., Conclusion: Our observations indicate that p-mTOR overexpression is abundant in CRC and linked to left-sided tumor localization. The high frequency and overexpression of p-mTOR is providing further rationale for targeting this pathway therapeutically in CRC patients. However, a prognostic role of p-mTOR overexpression in CRC could not be confirmed.

Histone deacetylases (HDACs) play an important role in tumor development and progression by modifying histone and non-histone proteins. In the current study we analyzed prevalence and prognostic impact of HDAC1 in prostate cancer. HDAC1 expression was analyzed by immunohistochemistry on a tissue microarray containing more than 12,400 prostate cancer specimens. Results were compared to tumor phenotype, biochemical recurrence, and molecular subtypes defined by ERG status as well as genomic deletions of 3p, 5q, 6q and PTEN. HDAC1 immunostaining was detectable in 75.4% of 9744 interpretable cancers and considered strong in 15.4%, moderate in 39.4% and weak in 20.7% of cases. High HDAC1 expression was associated with high Gleason grade (p<0.0001), advanced pathological tumor stage (p<0.0001), positive nodal status (p=0.0010), elevated preoperative PSA-level (p=0.0127), early PSA recurrence (p<0.0001) and increased cell proliferation (p<0.0001). Moreover, high-level HDAC1 staining was associated with TMPRSS2:ERG rearrangement and ERG expression in prostate cancers (p<0.0001) and was linked to deletions of PTEN (p<0.0001), 6q (p<0.0001) and 5q (p=0.0028) in ERG-negative cancers. The prognostic impact of HDAC1 was independent of established clinicopathological parameters and was mostly driven by ERG-negative cancers as revealed by subgroup analyses. HDAC1 has strong prognostic impact in prostate cancer and might contribute to the development of a fraction of genetically instable and particularly aggressive prostate cancers. HDAC1 measurement might therefore be of clinical value for risk stratification of prostate cancer and should be further evaluated in this regard., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background and Objective: Cyclin D1 is an important regulator protein for the G1-S cell cycle phase transition. The aim of this trial was to evaluate the impact of the CCND1 polymorphism G870A and corresponding protein expression and CCND1 amplification on the survival of the patients., Methods: 425 patients with ductal pancreatic adenocarcinoma who underwent resection were included after histopathological confirmation. DNA was analyzed for Cyclin D1 polymorphisms, immunhistochemical examination and fluorescence in situ hybridization analysis of the tumor were performed., Results: Overall, the mean survival was 22.9 months (20.5-25.3). The survival in patients with Cyclin D1 G870A polymorphism Adenine/Adenine was 15.1 months (95% CI 11.3-18.9), 21.5 months (17.4-25.6) for Adenine/Guanine, and 29.4 months (95% CI 23.8-35.0) for Guanine/Guanine (P = 0.003). A shorter survival was associated with strong/moderate protein expression in immunohistochemistry (IHC) compared to weak/no expression (P = 0.028). Additionally, a significant coherency between unfavourable polymorphism (AA/AG) and increased protein expression was detected (P = 0.005)., Conclusions: A strong impact on survival of Cyclin D1 G870A polymorphism and the detected corresponding protein expression was found. The biological mechanism of CCND1 in carcinogenesis has not been fully examined; but at present Cyclin D1 seems to be an interesting biomarker for the prognosis of ductal adenocarcinoma., (© 2014 Wiley Periodicals, Inc.)

This study was performed to assess the prognostic relevance of genomic aberrations at chromosome 4q in NSCLC patients. We have previously identified copy number changes at 4q12-q32 to be significantly associated with the early hematogenous dissemination of non-small cell lung cancer (NSCLC), and now aim to narrow down potential hot-spots within this 107 Mb spanning region. Using eight microsatellite markers at position 4q12-35, allelic imbalance (AI) analyses were performed on a preliminary study cohort (n = 86). Positions indicating clinicopathological and prognostic associations in AI analyses were further validated in a larger study cohort using fluorescence in situ hybridization (FISH) in 209 NSCLC patients. Losses at positions 4q21.23 and 4q22.1 were shown to be associated with advanced clinicopathological characteristics as well as with shortened disease free (DFS) and overall survival (OS) (DFS: P = 0.019; OS: P = 0.002). Multivariate analyses identified the losses of 4q21.23-22.1 to be an independent prognostic marker for both DFS and OS in NSCLC (HR 1.64-2.20, all P<0.04), and especially in squamous cell lung cancer (P<0.05). A case report study of a lung cancer patient further revealed a loss of 4q21.23 in disseminated tumor cells (DTCs). Neither gains at the latter positions, nor genomic aberrations at 4q12, 4q31.2 and 4q35.1, indicated a prognostic relevance. In conclusion, our data indicate that loss at 4q21.23-22.1 in NSCLC is of prognostic relevance in NSCLC patients and thus, includes potential new tumor suppressor genes with clinical relevance.

Background: Aim of this study was to assess the biological function in tumor progression and metastatic process carcinoembryonic antigen-related cell adhesion molecules (CEACAM) 1, 5 and 6 in pancreatic adenocarcinoma (PDAC)., Experimental Design: CEACAM knock down cells were established and assessed in vitro and in a subcutaneous and intraperitoneal mouse xenograft model. Tissue and serum expression of patients with PDAC were assessed by immunohistochemistry (IHC) and by enzyme linked immunosorbent assays., Results: Presence of lymph node metastasis was correlated with CEACAM 5 and 6 expression (determined by IHC) and tumor recurrence exclusively with CEACAM 6. Patients with CEACAM 5 and 6 expression showed a significantly shortened OS in Kaplan-Meier survival analyses. Elevated CEACAM6 serum values showed a correlation with distant metastasis and. Survival analysis revealed a prolonged OS for patients with low serum CEACAM 1 values. In vitro proliferation and migration capacity was increased in CEACAM knock down PDAC cells, however, mice inoculated with CEACAM knock down cells showed a prolonged overall-survival (OS). The number of spontaneous pulmonary metastasis was increased in the CEACAM knock down group., Conclusion: The effects mediated by CEACAM expression in PDAC are complex, though overexpression is correlated with loco-regional aggressive tumor growth. However, loss of CEACAM can be considered as a part of epithelial-mesenchymal transition and is therefore of rather importance in the process of distant metastasis.

Recently, the activated leukocyte cell adhesion molecule (CD166) was identified as an "inert" cancer stem cell (CSC) marker for non-small cell lung cancer (NSCLC). Few data exist regarding the clinical relevance of CD166 expression in NSCLC. We evaluated the expression of CD166 using immunohistochemistry in a large cohort of NSCLC patients (n = 1,910) on a tissue microarray basis. Expression was inversely associated with tumor size and lymph node status. Grading slightly failed to be significantly inversely associated, and survival analysis revealed no significant survival benefit of CD166-positive patients. Due to the results of this study, the theory of CD166 as a CSC marker for NSCLC must be questioned. The association of CD166 with smaller tumors and no nodal metastases does not make it a typical CSC marker. Further studies are required to investigate the functional role of CD166 in NSCLC., (© 2014 AlphaMed Press.)

Increased levels of karyopherin α2 (KPNA2) expression have been described to be linked to poor prognosis in a variety of malignancies. This study was undertaken to evaluate the clinical impact of KPNA2 expression and its association with key genomic alterations in prostate cancers. A tissue microarray containing samples from 11 152 prostate cancers was analyzed for KPNA2 expression by immunohistochemistry. Results were compared with oncological follow-up data and genomic alterations such as TMPRSS2-ERG fusions and deletions of PTEN, 5q21, 6q15 or 3p13. KPNA2 expression was absent or weak in benign prostatic glands and was found to be in weak, moderate or strong intensities in 68.4% of 7964 interpretable prostate cancers. KPNA2 positivity was significantly linked to the presence of ERG rearrangement (P<0.0001). In ERG-negative and -positive prostate cancers, KPNA2 immunostaining was significantly associated with advanced pathological tumor stage (pT3b/pT4), high Gleason grade and early biochemical recurrence (P<0.0001 each). Multivariate analysis including all established prognostic criteria available after surgery revealed that the prognostic role of KPNA2 (P=0.001) was independent of high Gleason grade, advanced pathological tumor stage, high preoperative prostate-specific antigen level and positive surgical margin status (P<0.0001 each). The comparison of KPNA2 expression with deletions of PTEN, 5q21, 6q15 and 3p13 in ERG-positive and -negative cancers revealed a strong link to PTEN deletions in both subgroups (P<0.0001). In conclusion, the strong independent prognostic impact of KPNA2 expression raises the possibility that measurement of KPNA2 expression alone or in combination with other molecular parameters might possibly result in clinically useful information. The data also emphasize a critical role of the functionality of the nuclear import machinery for prostate cancer biology.

Lysophosphatidylcholine acyltransferase 1 (LPCAT1) has been suggested to play a role in cancer. To assess its role in prostate cancer, LPCAT1 expression was analyzed on a tissue microarray containing samples from 11,152 prostate cancer patients. In benign prostate glands, LPCAT1 immunostaining was absent or weak. In prostate cancer, LPCAT1 positivity was found in 73.8% of 8786 interpretable tumors including 29.2% with strong expression. Increased LPCAT1 expression was associated with advanced tumor stage (pT3b/T4) (p < 0.0001), high Gleason score (≥4 + 4) (p < 0.0001), positive nodal involvement (p = 0.0002), positive surgical margin (p = 0.0005), and early PSA recurrence (p < 0.0001). High LPCAT1 expression was strongly linked to ERG-fusion type prostate cancer. Strong LPCAT1 staining was detected in 45.3% of ERG positive but in only 16.7% of ERG negative tumors (p < 0.0001). Within ERG negative cancers, LPCAT1 staining was strongly increased within the subgroup of PTEN deleted cancers (p < 0.0001). Further subgroup analyses revealed that associations of high LPCAT1 expression with PSA recurrence and unfavorable tumor phenotype were largely driven by ERG negative cancers (p < 0.0001) while these effects were substantially mitigated in ERG positive cancers (p = 0.0073). The prognostic impact of LPCAT1 expression was independent of histological and clinical parameters. It is concluded, that LPCAT1 measurement, either alone or in combination, may be utilized for better clinical decision-making. These data also highlight the potentially important role of lipid metabolism in prostate cancer biology., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Background: Human epidermal growth factor receptor 2 (HER2) is an important prognostic factor in several types of solid tumours. Although HER2 seems not to influence survival in esophageal carcinomas, an impact of the HER2 status of disseminated tumour cells (DTCs) on survival has been shown. The aim of our study was to investigate the significance of the HER2 status in primary esophageal carcinomas and matched lymph node metastases., Materials and Methods: The HER2 status of primary tumours and matched lymph node metastases were analysed for 158 patients with esophageal carcinoma using immunohistochemistry (IHC) and fluorescence in situ hybridisation (FISH)., Results: The study specimen included 90 adenocarcinomas (AC) and 68 squamous cell carcinomas (SCC). HER2 amplification was found in 12% and overexpression in 8.9% of all primary tumours. HER2 amplification was identical in the primary tumour and lymph node metastases in all AC and in 75% of SCC. Discordant-positive HER2 lymph node status and negative primary tumour status was found in 4.4% of AC and 1.5% of SCC in FISH analyses. No significant associations were found between HER2 amplification/overexpression and overall survival., Conclusion: HER2 gene status remains highly conserved in metastatic esophageal carcinoma. Discrepancies occur rarely between primary tumour and lymph node metastases and might be due to heterogeneity of the HER2 status of the primary tumour. This could be the reason for heterogeneity of DTCs and may result in metastasis of only a subset of tumour cells.

Colorectal cancer is the third most common malignancy worldwide. Anti-epidermal growth factor receptor (EGFR)-targeted therapy shows clinical evidence in this malignancy and improves outcome. The tumor suppressor gene phosphatase and tensin homologue (PTEN) is considered a potential predictor of nonresponse to anti-EGFR agents. The purpose of this study was to assess whether associations between PTEN alterations (PTEN gene deletion or PTEN gene disruption) and clinical outcome could be caused by a prognostic (and not predictive) effect of PTEN inactivation. Therefore, we analyzed 404 colorectal cancers not previously treated with anti-EGFR drugs in a tissue microarray format. PTEN deletion and PTEN gene rearrangements were analyzed by fluorescence in situ hybridization. Heterogeneity analysis of all available large tissue sections was performed in 6 cases with genomic PTEN alteration. Twenty-seven (8.8%) of 307 analyzable colorectal cancer spots showed genomic PTEN alterations including 24 hemizygous and 1 homozygous deletion as well as 2 PTEN gene disruptions. Genomic PTEN alterations were associated with reduced patient survival in rectal cancer in univariate and multivariate analyses (P = .012; hazard ratio, 2.675; 95% confidence interval, 1.242-5.759) but not in colon cancer. Large-section evaluation revealed a homogeneous distribution pattern in all 4 analyzed cases with PTEN deletion and in both cases with a PTEN gene disruption. In conclusion, genomic PTEN gene alterations caused by deletion or gene disruption characterize a fraction of rectal cancers with particularly poor outcome., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background and Objective: E- and P-selectins expressed on the luminal surface of mesodermally derived endothelial cells play a crucial role in the formation of haematogenous metastases in a number of malignancies. As peritoneal mesothelial cells are also derived form the mesoderm, it was hypothesised that selectins are also of importance in peritoneal tumour spread., Methods: Immunohistochemistry was used to identify selectin expression on normal human peritoneum and isolated mesothelial cells. E- and P-selectin interactions with human pancreatic adenocarcinoma cells were investigated in dynamic flow assays and flow cytometry; the latter was also used to determine the main selectin ligands on pancreatic adenocarcinoma cell lines PaCa 5061, BxPC-3 and PaCa 5072, and selectin expression on human mesothelial cells. All cell lines were xenografted into the peritoneum of E- and P-selectin-deficient pfp/rag2 mice and selectin wild-type controls. Peritoneal carcinomatosis was quantified using MRI or a scoring system., Results: E- and P-selectin were constitutively expressed on human mesothelial and endothelial cells in the peritoneum. PaCa 5061 and BxPC-3 cells interacted with E- and P-selectins in dynamic flow assays and flow cytometry, with CA19-9 (Sialyl Lewis a) being the main E-selectin ligand. For xenografted PaCa 5061 and BxPC-3 cells, peritoneal metastasis was significantly reduced in E- and P-selectin double knockout mice compared with wild-type pfp/rag2 animals. In contrast, PaCa 5072 cells were almost devoid of selectin binding sites and no intraperitoneal tumour growth was observed., Conclusion: Interactions of tumour cells with peritoneal selectins play an important role in the peritoneal spread of pancreatic adenocarcinoma.

Next to EGFR mutation, EGFR gene copy number evaluated by fluorescence in situ hybridization (FISH) emerged as a potential predictive marker for sensitivity to EGFR tyrosine kinase inhibitors, although controversial data exist. As the diagnostic accuracy of predictive biomarkers can be substantially limited by regional differences within tumors, heterogeneity of EGFR gene copy gain in NSCLC was assessed in this study. For this purpose, a novel tissue microarray (TMA) based analysis platform was developed. TMAs were constructed containing 8 different tissue cylinders from 144 primary NSCLCs. From 62 of these patients additional nodal metastases were sampled. EGFR gene copy number and EGFR expression was analyzed by FISH and immunohistochemistry according to the suggested guidelines. 13 (9.0%) of the 144 evaluated tumors showed EGFR amplification and 37 (25.7%) tumors high polysomy in at least one tumor area. In 7 (53.8%) of 13 amplified cases the analysis of different tumor areas revealed subclones without EGFR gene copy gain next to subclones with amplification. All of the 36 evaluable tumors with high polysomy showed heterogeneity of EGFR gene copy number with areas negative for gene copy gain within the individual tumors. Heterogeneity of EGFR gene copy gain in lung cancer challenges the concept of using small biopsies for the analysis of EGFR FISH status. EGFR gene copy number is highly heterogeneous within individual NSCLCs and this finding might well be a reason for the controversial clinical data existing regarding responsiveness to anti-EGFR therapy., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Introduction: The GNAS1 T393C single nucleotide polymorphism (T393C-SNP) correlates with Gαs mRNA stability and protein expression and augmented apoptosis. Genetic germ line variations as stable and reproducible markers potentially serve as prognostic marker in oncology. The aim of this study was to evaluate the potential prognostic value of T393C-SNP in complete resected non-small cell lung cancer (NSCLC)., Patients and Methods: In total 163 Caucasian patients, who had been surgically treated for NSCLC between 1998 and 2010, were included in this study. Genotyping of peripheral blood cells was performed by polymerase chain reaction and digestion using the restriction enzyme FokI. The T393C-SNP was correlated with clinic-pathological parameters and survival. Chi-square test, Kaplan-Meier estimator and cox regression hazard model were used to assess the prognostic value of the T393C-SNP., Results: C-allele carriers had a higher recurrence rate (p=0.018) and a shorter disease-free survival compared to homozygous T-allele carriers (12.26 months vs. 44.65 months, p=0.009). The overall survival in homozygous C allele carriers was shorter (19.10 months vs. 53.95 months, p=0.019). Multivariate Cox regression identified the CC genotype as a negative independent prognostic factor for recurrence (hazard ratio 2.36, p=0.007) and survival (hazard ratio 2.51, p=0.008)., Conclusion: Determination of T393C-SNP preoperatively potentially allows allocation of NSCLC patients into different risk profiles and may influence the therapeutic strategy., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background: The activated leukocyte cell adhesion molecule (ALCAM, CD166) has been reported to be involved in tumorigenesis of colorectal cancer (CRC) and to function as a cancer stem cell marker. Controversial data exist regarding the prognostic power of ALCAM expression in CRC. Here, we evaluate the expression of ALCAM in a cohort of CRC patients and its usage as a prognostic marker for survival., Materials and Methods: Tissue specimens from 299 patients with CRC treated between 1993 and 2006 were analyzed via ALCAM immunohistochemistry (clone MOG/07) using a tissue microarray. Results were correlated with clinical, histopathological, and patient survival data (Chi-square test, Kaplan-Meier analysis, and log-rank test, respectively). Multivariate analysis also was performed (Cox regression)., Results: ALCAM is expressed in most primary (76%) and secondary (62%) CRC lesions (P = 0.014). Immunohistochemistry revealed an inverse association with tumor grading (P = 0.002) but not with any other clinical or histopathological data. Kaplan-Meier survival analysis revealed a significant overall survival benefit in the group of ALCAM-positive patients (P = 0.019). Multivariate analysis showed that ALCAM is an independent positive prognostic marker for overall survival (P = 0.023)., Conclusions: ALCAM expression is a positive prognostic marker for overall survival of CRC patients, and its detection might help to optimize the existing prognostic staging system. Elevated expression in higher differentiated tumors might indicate a potential role in the early steps of tumorigenesis, and its loss might be associated with reduced cellular adhesion, resulting in a higher metastatic potential of the tumor. Further studies must be conducted investigating these hypotheses., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background and Aim: Gastric carcinoma is the second most frequent cause of cancer-related death worldwide. As PTEN is a potential modifier of tumour response to trastuzumab, a recently approved therapy in metastatic HER2 positive gastric cancer, the existence of PTEN deletions in primary gastric cancer was investigated., Methods: 230 primary gastric cancers were analysed in a tissue microarray format by dual labelling fluorescence in situ hybridisation for PTEN deletion. HER2 analysis was also performed. To study PTEN deletion heterogeneity, all available large tissue sections from primary cancer and corresponding metastases were analysed in seven patients., Results: Eight of 180 interpretable primary gastric cancer spots showed PTEN deletions (4.4%), including seven hemizygous and one homozygous deletion. PTEN deletion was correlated with nodal (8 of 122 cases (6.6%); p=0.041) and distant metastases (4 of 19 (21.1%); p<0.001). Large section validation showed a homogeneous distribution of PTEN deletion. HER2 positivity was seen in one PTEN deleted case., Conclusion: Genomic PTEN deletion is a rare event in gastric adenocarcinoma but correlates with metastatic disease. The homogeneous distribution pattern indicates that this alteration occurs early in tumour development.

Purpose: VEGFR-2 gene displays several functional germline polymorphisms with impact on VEGFR-2 mediated angiogenesis. Our purpose was to evaluate VEGFR-2 polymorphisms as prognostic markers for tumor recurrence and overall survival (OS) in non-small-cell lung cancer (NSCLC)., Methods: In total, 209 Caucasian patients who had been surgically treated for NSCLC between 1996 and 2010 were included in this study. Genotyping of peripheral blood cells was performed by TaqMan® genotyping assays or polymerase chain reaction for five VEGFR-2 polymorphisms. Chi- square test, Kaplan-Meier estimator, and Cox regression hazard model were used to assess the prognostic value of VEGFR-2 polymorphisms., Results: VEGFR-2+4422 (AC)10-14 polymorphism was identified as a positive prognostic marker for time to metastasis (11/12 vs. 11/11 (AC) repeats: hazard ratio (HR), 0.28; 95% confidence interval (CI), 0.11-0.75; p=0.012) and OS (11/12 vs. 11/11 (AC) repeats: HR, 0.41; 95% CI, 0.21-0.82; p=0.012) in squamous cell carcinoma. For adenocarcinoma, VEGFR-2-906 C>T (C/T vs. CC: HR, 0.19; 95% CI, 0.43-0.82; p=0.027) and VEGFR-2-271 G>A (G/A vs. G/G: HR, 0.25; 95% CI, 0.07-0.86; p=0.027) predicted longer time to local recurrence and VEGFR-2-906 C>T was a predictor for better OS (T/T vs. C/C: HR, 0.28; 95% CI, 0.09-0.84; p=0.024)., Conclusions: VEGFR2 germline polymorphisms predict tumor recurrence and OS in NSCLC.

Background: L1 cell adhesion molecule (CD171) has been detected in different malignant tumors and is associated with unfavorable outcome. It thus represents a target for tumor diagnosis and therapy. In this study, we assessed L1 expression in more than 8000 normal human tissues and different types of tumors, both malignant and non-malignant, and neural and non-neural., Materials and Methods: Tissue micro-arrays, including a multi-tumor-array of 128 different tumor types, up to 50 samples of each type (approximately 5500 different samples), arrays with approximately 3000 different prostate and 600 mesenchymal tumor samples, and a normal human tissue-array were analyzed by immunohistochemistry with a monoclonal antibody using immunoperoxidase staining., Results: L1 expression was detected in tumors of neural and neural crest origin and other types of non-neural tumors, but not in those of epithelial origin. In normal human tissues, L1 was detected in skin basal cells and small blood vessels, most notably in the mature placenta and peripheral nerves., Conclusion: This first comprehensive study of the importance of L1 expression in human demonstrates strong L1 overexpression in tumors of neuroectodermal and neural crest origin and an expression in only very few normal human tissues. L1 thus is a potentially important therapeutic target, particularly with respect to malignant melanoma, gastrointestinal stromal tumor, neuroblastoma, and certain subtypes of non-neural tumors., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The high mortality rate of lung cancer patients is mainly due to the late stage at which lung cancer is diagnosed. For effective cancer prevention programs and early diagnosis, better blood-based markers are needed. Hence, blood-based microarray profiling of microRNA (miR) expression was performed in preoperative serum of 21 non-small cell lung cancer (NSCLC) patients and 11 healthy individuals by microfluid biochips containing 1158 different miRs. Two out of the 30 most dysregulated miRs were further validated in serum of 97 NSCLC patients, 20 patients with benign lung diseases and 30 healthy individuals by TaqMan MicroRNA Assays. Microarray profiling showed that miR-361-3p and miR-625* were significantly down-regulated in serum of lung cancer patients. Their further evaluation by quantitative RT-PCR showed that the levels of miR-361-3p and miR-625* were lower in NSCLC than in benign disease (p = 0.0001) and healthy individuals (p = 0.0001, p = 0.0005, respectively). Moreover, the levels of miR-625* were significantly lower in patients with large cell lung cancer (LCLC, p = 0.014) and smoking patients (p = 0.030) than in patients with adenocarcinoma and non-smoking patients, respectively. A rise in the levels of both miRs was observed in the postoperative samples compared with the preoperative levels (p = 0.0001). Functional analyses showed that Smad2 and TGFß1 are not dysregulated by miR-361-3p and miR-625* in the lung cell line A549, respectively. Our present pilot study suggests that miR-361-3p and miR-625* might have a protective influence on the development of NSCLC, and the quantitative assessment of these miRs in blood serum might have diagnostic potential to detect NSCLC, in particular in smokers.

Background: Activated leukocyte cell adhesion molecule (ALCAM, CD166) is a cell membrane protein that is aberrantly expressed in different tumors, including pancreatic neuroendocrine tumors (PNET). The aim of this study was to determine the expression of ALCAM in PNET to learn more about the prevalence and clinical significance of ALCAM expression in PNET., Methods: Primary tumors (n = 38) and corresponding lymph node (n = 5) and liver metastases (n = 9) of patients with PNET, treated at the University Medical Center Hamburg-Eppendorf between 1993 and 2006, were analyzed via ALCAM immunohistochemistry in a tissue microarray format. The results were correlated with clinical and histopathologic data, including the WHO classification of PNET., Results: The majority of primary (74%) and secondary (50%) lesions of PNET showed strong ALCAM expression. Immunohistochemistry of primary tumors revealed an association between high ALCAM expression and the hormone production status of the tumor (P = 0.037), and an inverse correlation with metastasis status (P = 0.041) and tumor size (cut off level 2 cm, P = 0.013). Elevated ALCAM expression was a significant positive prognostic factor for recurrence-free (P = 0.002) and disease-specific survival (P = 0.009) in Kaplan-Meier survival analysis (log rank test)., Conclusions: ALCAM is abundantly expressed in PNET and decreased expression is significantly associated with poor prognosis. ALCAM may be a potential marker for risk prediction in patients diagnosed with PNET. Further studies with larger patient collectives are required to validate the results of this study and investigate the functional role of ALCAM in PNET., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: L1 cell adhesion molecule (CD171) is expressed in many malignant tumors and its expression correlates with unfavourable outcome. It thus represents a target for tumor diagnosis and therapy. An earlier study conducted by our group identified L1 expression levels in primary gastrointestinal stromal tumors (GIST) as a prognostic marker. The aim of the current study was to compare L1 serum levels of GIST patients with those of healthy controls and to determine whether levels of soluble L1 in sera could serve as a prognostic marker., Methods: Using a sensitive enzyme-linked immunosorbent assay (ELISA), soluble L1 was measured in sera of 93 GIST patients und 151 healthy controls. Soluble L1 levels were then correlated with clinicopathological data., Results: Median levels of soluble L1 were significantly higher (p < 0.001; Mann-Whitney U test) in sera of GIST patients compared to healthy individuals. Median soluble L1 levels were particularly elevated in patients with recurrence and relapse (p < 0.05; Mann Whitney U test)., Conclusion: These results suggest that high soluble L1 levels predict poor prognosis and may thus be a promising tumor marker that can contribute to individualise therapy.

Background: Current available preoperative diagnostic workup is insufficient to differentiate between benign and malignant pancreatic neuroendocrine tumors (PNET). The aim of the present study was to evaluate the potential prognostic role of the promoter GTn repeat polymorphism (GTn) of the heme oxygenase-1 gene in PNET., Methods: Tumor, metastasis, corresponding healthy tissue, and peripheral blood leukocyte DNA of 46 patients who underwent surgical resection for PNET were analyzed for GTn by PCR, capillary electrophoresis, and DNA-sequencing. The GTn was correlated to clinicopathologic parameters and clinical outcome., Results: GTn was classified into short (<25) and long (≥ 25) alleles and three (SS, SL, and LL) genotypes were defined. There was no difference in GTn genotype among tumor, healthy tissue, metastasis, and peripheral blood leukocyte DNA. The SS and SL genotype displayed significantly more poor differentiated tumors (P = 0.001) and higher tumor recurrence rate (P = 0.0001) compared with LL patients. The LL genotype patients presented predominantly benign PNET (P < 0.001). The LL genotype had the longest disease-free (P < 0.001) and overall survival (P = 0.006). Besides the WHO classification the GTn was identified as a strong predictor of tumor recurrence (hazard ratio 3.1, 95% confidence interval 1.3-7.3) in PNET., Conclusion: GTn differentiates between benign and malignant PNET and is a strong predictor of tumor recurrence., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: A previous study identified midkine (MK) expression in primary gastrointestinal stromal tumor (GIST) as a prognostic marker. The aim of the current study was to compare serum midkine (S-MK) concentrations of GIST patients with those of healthy controls and to determine if MK can serve as a prognostic serum marker for these patients., Materials and Methods: S-MK concentrations were measured by enzyme-linked immunosorbent assay in GIST patients (n = 96) and healthy controls (n = 148). S-MK levels were then correlated with clinicopathological data and the administration of imatinib therapy. In addition, MK expression was evaluated in 39 surgically resected GIST and in 17 leiomyoma specimens on a tissue microarray., Results: S-MK concentrations in GIST patients were significantly higher than in healthy controls: median (25th and 75th percentiles) S-MK concentration was 235 (139 and 376) pg/ml in the GIST patients and 99 (33 and 198) pg/ml in the controls (P < 0.001; Mann-Whitney U test). Significantly higher median S-MK concentrations were found in GIST with recurrence compared with those without (295 vs 230; P = 0.009). GIST patients with S-MK levels higher than 400 pg/ml showed a significantly worse recurrence-free survival (P = 0.026; log-rank test). Patients receiving imatinib therapy had decreased median S-MK concentrations compared with those who were not treated with imatinib (331 vs 201; P < 0.001)., Conclusions: S-MK concentration is a potential marker for evaluating the progression and prognosis of GIST, especially during imatinib therapy. Further studies could focus on the role of midkine in the tumorigenesis of GIST and responsiveness toward imatinib therapy.

Insulin-like growth factor-1 receptor (IGF-1R) and human epidermal growth factor receptor-2 (HER2) receptor expression has been found to be a key regulator of tumorigenesis. The purpose of our study was to establish the prognostic significance of IGF-1R in esophageal cancer and to determine the effect of IGF-1R and HER2 targeting with alpha-IR3 and Herceptin antibodies on the proliferation of esophageal cancer cells in vitro. IGF-1R expression and clinicopathological correlations were analyzed with a tissue microarray containing 234 esophageal cancer specimens (133 adenocarcinomas and 101 squamous cell carcinomas). Proliferation changes associated with Herceptin and alpha-IR3 blockage were evaluated with the unique human esophageal cancer cell lines Pt1590 and LN1590. IGF-1R and HER2 expression levels, activation and phosphorylation status of downstream signaling proteins involved in the activation pathways were analyzed by Western blotting. IGF-1R overexpression was detected in 121 (52%) of the 234 esophageal tumors examined. In the subgroup of 87 HER2-positive tumors, 93.1% showed concordant overexpression for IGF-1R. IGF-1R was identified as a variable associated with reduced overall survival for adenocarcinoma (p = 0.05), but not for squamous cell carcinoma. The combination of Herceptin and alpha-IR3 was more effective in inhibiting in vitro proliferation than treatment with either agent alone (p < 0.01). This was associated with a decrease in HER2 and IGF-1R protein levels and suppression of Akt- and MAP kinase phosphorylation. IGF-1R expression can be used as a novel prognostic marker for adenocarcinomas of the esophagus. Cotreatment with IGF-1R and HER2 antibodies might become a valuable and effective treatment option in esophageal adenocarcinoma.

Background: Intraductal papillary-mucinous neoplasm (IPMN) of the pancreas is an increasingly diagnosed entity since its definition by the World Health Organization in 1996. It has a broad clinical spectrum ranging from benign to malignant tumors. Optimum treatment is controversial and a better understanding of the development of IPMN of the pancreas and identification of potential prognostic factors will help to address this. Angiogenesis plays an elementary role in the development of malignant tumors and may well also be important in the development of IPMN of the pancreas. Therefore we investigated endothelial cell marker CD31 (PECAM-1) and angiogenesis associated marker CD105 (endoglin) by immunohistochemistry., Methods: Thirty-two cases of surgically resected IPMN were chosen retrospectively and clinical data were obtained. Specimens were stained for proliferation marker (Ki-67), CD31 and CD105 by immunohistochemistry. A CD105/CD31 Angiogenesis ratio (AR) was established to determine the proliferating fraction of endothelial cells., Results: The AR is significantly elevated in invasive IPMN of the pancreas (Mann-Whitney-U Test, p<0.05) and is associated with the Ki-67-labelling-index, demonstrating synergy between tumor-growth and neovascularisation. Invasive IPMN of the pancreas is associated with significantly lower recurrence-free and overall survival., Conclusions: Neovascularisation plays an important role in the tumorigenesis of invasive IPMN of the pancreas, and therefore angiogenesis-associated molecules like CD105 and CD31 might be useful tools as prognostic markers. Furthermore, the results indicate a potential role for adjuvant anti-angiogenic therapies in selected patients with recurring and/or invasive IPMN of the pancreas.

Background: The Y-box-binding protein-1 (YB-1) is a member of a family of DNA-binding proteins and an oncogenic transcription factor that is highly expressed in cancers of the breast, lung and prostate. To date, no data are available on its role in neuroblastoma. The aim of the present study was to evaluate the YB-1 expression in neuroblastoma., Materials and Methods: A tumour tissue microarray (TMA) was constructed from 36 neuroblastoma samples which were analysed by immunohistochemistry for YB-1 expression., Results: Expression of YB-1 was detected in 35 of 37 (94.6%) neuroblastoma cases examined. Nevertheless, no correlation of YB-1 expression with survival, risk factors or stage of the disease was observed., Conclusion: As the majority of neuroblastomas express YB-1, this protein may play an important role in tumour pathogenesis. The results of this study suggest that YB-1 may serve as a novel immune marker for neuroblastoma and may be potentially useful as a therapeutic target.

Background: Esophageal adenocarcinoma is currently the most rapidly increasing cancer in Western populations. L1 (CD171), a neural cell adhesion molecule, has an essential function in tumor progression and has been shown to be expressed in the proliferating cells of the intestinal crypts in mice. The aim of the current study was to determine L1 expression in esophageal cancer and to evaluate whether L1 could serve as a potential marker and therapeutic target for this tumor type., Materials and Methods: L1 expression was assessed on a tissue microarray with 257 surgically resected esophageal cancer samples by immunohistochemistry with a monoclonal antibody (Clone UJ127). L1 expression was correlated with clinicopathological data., Results: L1 was detected in 22 (9%) of 257 esophageal cases, whereas 235 (91%) were L1 negative. Nineteen (86%) of the 22 L1-positive cases were adenocarcinoma. Cross table analysis showed a significant association between L1 expression and adenocarcinoma subtype (p<0.001), but not squamous cell carcinoma., Conclusion: L1 expression in a subgroup of esophageal cancer is specifically prevalent in adenocarcinoma. Data suggest L1 as a potential target for biological therapy in L1-positive esophageal adenocarcinoma patients.