Isolation of circulating tumor cells from pancreatic cancer by automated filtration

// Nora Brychta 1, * , Michael Drosch 1, 6,* , Christiane Driemel 2 , Johannes C. Fischer 2 , Rui P. Neves 2 , Irene Esposito 3 , Wolfram Knoefel 2 , Birte Mohlendick 2 , Claudia Hille 1, 5 , Antje Stresemann 1 , Thomas Krahn 1 , Matthias U. Kassack 4 , Nikolas H. Stoecklein 2 and Oliver von Ahsen 1 1 Bayer AG, Biomarker Research, 13353 Berlin, Germany 2 Department of General, Visceral and Pediatric Surgery, Medical Faculty, University Hospital of the Heinrich-Heine-University Duesseldorf, 40225 Duesseldorf, Germany 3 Institute of Pathology, Heinrich-Heine-University of Duesseldorf, 40225 Duesseldorf, Germany 4 Institute of Pharmaceutical & Medicinal Chemistry, University of Duesseldorf, 40225 Duesseldorf, Germany 5 Current/Present address: University Medical Center Hamburg-Eppendorf, Department of Tumor Biology, 20246 Hamburg, Germany 6 Current/Present address: JPT Peptide Technologies GmbH, 12489 Berlin, Germany * These authors have contributed equally to this work Correspondence to: Oliver von Ahsen, email: oliver.vonahsen@bayer.com Nikolas H. Stoecklein, email: nikolas.stoecklein@uni-duesseldorf.de Keywords: pancreatic cancer, circulating tumor cells, KRAS , diagnostic leukapheresis, EMT Received: March 24, 2017      Accepted: August 07, 2017      Published: September 16, 2017 ABSTRACT It is now widely recognized that the isolation of circulating tumor cells based on cell surface markers might be hindered by variability in their protein expression. Especially in pancreatic cancer, isolation based only on EpCAM expression has produced very diverse results. Methods that are independent of surface markers and therefore independent of phenotypical changes in the circulating cells might increase CTC recovery also in pancreatic cancer. We compared an EpCAM-dependent (IsoFlux) and a size-dependent (automated Siemens Healthineers filtration device) isolation method for the enrichment of pancreatic cancer CTCs. The recovery rate of the filtration based approach is dramatically superior to the EpCAM-dependent approach especially for cells with low EpCAM-expression (filtration: 52%, EpCAM-dependent: 1%). As storage and shipment of clinical samples is important for centralized analyses, we also evaluated the use of frozen diagnostic leukapheresis (DLA) as source for isolating CTCs and subsequent genetic analysis such as KRAS mutation detection analysis. Using frozen DLA samples of pancreatic cancer patients we detected CTCs in 42% of the samples by automated filtration.