Abstract
Aim: Previous studies suggest that circulating tumor cells (CTC) are present at very low frequencies in blood of pancreatic cancer (PC) patients. However, no technique has proven efficient for their detection, in part due to the lack of accurate tumor markers. Here, we evaluated the potential utility of two marker candidates - Mucin 16 (MUC16) and Tetraspanin 1 (TSPAN1) - identified through a detailed review of the literature.Methods: To evaluate the pattern of expression of both markers in pancreatic tumor cells vs. normal blood, we used cell lines derived from pancreatic cancer patients and blood from healthy adults.Results: Antibodies against both MUC16 and TSPAN1 showed expression in three pancreatic cancer (PC) cell lines while they were absent in blood cells. To evaluate the efficiency of isolating tumor cells from blood, PC cell lines were spiked at different frequencies in blood, sequentially stained with biotin-conjugated TSPAN1 and MUC16 antibodies and a streptavidin ferrofluids, followed by immunomagnetic enrichment. The recovery of spiked TSPAN1+ tumor cells was high with limited contamination by leukocytes. In contrast, no PC cells were isolated when the biotin MUC16 reagent was used because the biotin-conjugated clone did not recognize PC cells.Conclusion: The combination of MUC16, TSPAN1, and epithelial cell adhesion molecule (EpCAM) antibodies will likely increase the efficiency of capturing circulating tumor cell in blood of pancreatic ductal adenocarcinoma. To further develop a protocol for isolation of circulating tumor cell in blood of PC patients, high amounts of antibodies (5-10 mg) against EpCAM, MUC16, and TSPAN1 will be needed.
Highlights
Pancreatic ductal adenocarcinoma (PDAC) represents ~3% of all newly-diagnosed cancer patients[1] (10.5 cases per 100,000/year in the EU)
We developed a PDAC-oriented approach for diagnosis and monitoring of PDAC patients, based on detection and positive selection of blood cells expressing the epithelial cell adhesion molecule (EpCAM) compared to other protein biomarkers (MUC16 and Tetraspanin 1 (TSPAN1)) that might increase the efficiency of capturing circulating PDAC tumor cells in blood
peripheral blood (PB) samples were drawn by venipuncture into 10 mL CellSave collection tubes (Menarini Silicon Biosystems, Huntingdon Valley, PA) or vacutainer tubes containing EDTA as anticoagulant [Becton/ Dickinson (BD), Franklin Lakes NJ].The study and blood collection were approved by the local ethics committees of the University Hospital of Salamanca (Salamanca, Spain) and the University of Twente (Enschede, The Netherlands), respectively, and the research complied with all applicable laws and institutional guidelines
Summary
Pancreatic ductal adenocarcinoma (PDAC) represents ~3% of all newly-diagnosed cancer patients[1] (10.5 cases per 100,000/year in the EU) It represents the fourth cause of death in the western world due to its very poor prognosis, with a five-year survival rate of 5%[2,3,4,5], mainly caused by delayed diagnosis and resistance to conventional therapy. The potential utility of serum biomarkers such as carbohydrate antigen 19-9 (CA19-9) for the diagnostic screening of PDAC has been extensively evaluated. These markers (e.g., CA19-9) lack sensitivity and specificity for early PDAC detection with a significant percentage of both false negative and false positive results. Thereby, robust (sensitive and specific), cost-effective markrs are still required for early (minimally-invasive) diagnosis of PDAC that would lead to early treatment and improved PDAC patient outcome
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