ANTICANCER RESEARCH 34: 4741-4746 (2014)
Serum EpCAM Expression in Pancreatic Cancer FLORIAN GEBAUER1,2,3*, LEA STRUCK1*, MICHAEL TACHEZY1, YOGESH VASHIST1, DANIEL WICKLEIN2,3, UDO SCHUMACHER2,3, JAKOB R. IZBICKI1 and MAXIMILIAN BOCKHORN1 1Department
of General, Visceral and Thoracic Surgery, 2Institute of Anatomy and Experimental Morphology, and Cancer Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
3University
Abstract. Background: Aim of the study was to assess the diagnostic and prognostic impact of serum EpCAM levels in patients with pancreatic adenocarcinoma (PDAC). Materials and Methods: For quantitative measurement of preoperative serum EpCAM levels, a sandwich enzyme immunoassay kit was used (ELISA). Sixty-six patients with PDAC were included in the study and for comparison 43 patients with chronic pancreatitis and 104 healthy blood donors without any clinical evidence of cancer were analyzed as well. Results: Serum EpCAM levels differed significantly between the groups. The average value for patients with PDAC was 0.240±0.833 ng/ml, in patients with CP 0.192±0.590 ng/ml and 0.626±1.164 ng/ml in normal blood donor sera. With a cut-off level of 0.422 ng/ml EpCAM, the calculated sensitivity of detecting PDAC was 66.7% with corresponding specificity of 77.5%. A correlation with clinico-pathological data (pT, pN, M, R-status, grading, UICC stage) was not found and in addition there was no difference in overall survival between patients with high- and low-preoperative serum EpCAM levels. Conclusion: EpCAM can be detected in the serum in patients with PDAC though the sensitivity for detecting PDAC is low and a correlation with clinical parameters was not found. Pancreatic cancer is the third most common malignancy of the gastrointestinal tract and is characterized by its rapid progression and dismal prognosis (2). Currently, the only potentially curative therapy option is pancreaticoduodenectomy, but only 20% of patients have resectable
*These Authors contributed equally to this study. Correspondence to: Florian Gebauer, MD, Department of General, Visceral and Thoracic Surgery, University Medical Centre HamburgEppendorf, Martinistr. 52, 20246 Hamburg, Germany. Tel: +49 40741050152, Fax: +49 40741054995, e-mail:
[email protected] Key Words: EpCAM, pancreatic cancer, diagnosis, prognosis.
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disease at diagnosis (22). Due to local extension or metastases at the time of diagnosis, the overall 5-year survival rate is only 5-6% (11). Therefore, there is an urgent need for the identification and establishment of clinically relevant markers for earlier diagnosis and evaluation of prognosis for patients with pancreatic cancer. Epithelial cell adhesion molecule (EpCAM, also referred to as 17-1A, ESA and EGP40) is a 40-kDA epithelial transmembrane glycoprotein and a calcium-independent homophilic cell adhesion molecule (15). The EpCAM protein consists of two epidermal growth factor-like (EGFlike) repeats, a transmembrane domain, and a short cytoplasmatic tail (5). It is expressed at low levels in the majority of healthy epithelial tissues and on primary tumors at elevated levels in cancer of various origins, including colon and rectum, liver and oesophagus (7, 10, 13). Despite its overexpression on cancer cells, EpCAM seem to have a dual role in tumor progression, with tumorpromoting and anti-tumorgenic effects (34). Due to its function as a cell adhesion molecule, EpCAM is able to mediate homophilic adhesive interactions and is therefore likely to prevent cell scattering and to play a role in inhibition of invasion (6, 16). The influence of EpCAM expression in different carcinomas on the clinical prognosis has been shown to be controversial. While some reports show high EpCAM expression to be a factor of good prognosis, other studies have shown it to be a factor for poor prognosis (23, 24, 26, 27, 29, 35). These diverse results propose that EpCAM may have a different role in each type of cancer and its function might be influenced by various factors. For pancreatic cancer, the influence of EpCAM expression on cancer cell activity or prognosis after resection has not been solved. In this study, we investigated the potential role of EpCAM as a diagnostic molecule in pancreatic cancer by determining serum EpCAM levels in patients with pancreatic cancer, patients with chronic pancreatitis and in healthy blood donors. Furthermore, correlations of these data with clinicopathological features of the tumor and prognosis are discussed.
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ANTICANCER RESEARCH 34: 4741-4746 (2014) Materials and Methods Patients and samples. The study was performed on 66 patients (26 females and 40 males) with ductal adenocarcinoma of the pancreas (PADC) who underwent potentially curative surgery without any preoperative therapy at the Department of General, Visceral and Thoracic surgery, University Medical Center of Hamburg, Germany, between 2003 and 2009. The median age of the patients was 62 years and the median follow-up was 18.1 months. The study was approved by the Ethics Committee of the Chamber of Physicians in Hamburg, Germany (PVN1045). Written consent for using the samples for research purposes was obtained from all patients prior to surgery or blood drawing. Information on sex, age, stage of disease and histopathological factors was extracted from medical records, follow-up data were obtained from a combination of clinical and pathological record reviews, from outpatient clinic medical records, or communication with patients and their attending physicians, or from the cancer registry. Overall survival (OS) was calculated from the date of operation to the date of death or last follow-up. Patients who did not survive the first 30 days after surgery were excluded from the survival analysis. Tumor stage was classified according to the TNM classification of the International Union Against Cancer (UICC seventh edition) (25). As comparators, serum samples from 43 patients with chronic pancreatitis (CP) (13 females and 30 males) and samples from 104 healthy blood donors without any clinical evidence of cancer were analyzed. Serum EpCAM quantification. Blood samples of patients with PDAC and CP were obtained from central venous catheter from each patient directly before surgery. After collection of blood samples, serum was separated from the blood by centrifugation at 3400 × g for 10 min and these serum samples were kept frozen at –80˚C. Samples from healthy blood donors were obtained from a peripheral venous catheter during voluntary blood donation. For quantitative measurement of serum EpCAM levels, a monoclonal sandwich enzyme immunoassay kit was used (EpCAM/TROP I DuoSet, R&D Systems, Minneapolis, USA). The EpCAM-coated antibody was used at a concentration of 4 μg/ml in 96-well plates (Nunc immunoplate F96 maxisorp; Thermo Scientific, Schwerte, Germany), capture antibody was applied at a concentration of 0.2 μg/ml. Recombinant human EpCAM (R&D Systems) was used as standard. Before running the assay, all patient samples were thawed at room temperature and then diluted 1:5 using Dulbecco’s phosphate buffered saline (PBS) (GIBCO by Invitrogen, Grand Island, NY, USA) based on the instructions in the kit protocol. Incubation was carried out at 26˚C, determination of the H2SO4 stopping reaction was performed with a ELISA plate reader (DynaTech MR 5000; Denkendorf, Germany). All samples were analyzed in duplicates, and the mean value was used for data analysis. In the case of variance >5% within the duplicates, repetitive measurements were made. In order to ensure that the immunoassay was suitable for measuring clinical serum samples, reproducibility and linearity were examined. The assay showed excellent linearity with use of serial dilutions and showed
