Page 90 - Demo

Chapter 588Materials and MethodsStudy population and blood samples Between October 2004 and September 2013, a total of 82 patients with histologically confirmed locally advanced or metastatic EC or GC were enrolled in a randomized phase II study that was performed in the Amsterdam UMC, location VUmc, and the Noordwest Ziekenhuisgroep in Alkmaar, both in the Netherlands. The institutional Medical Ethical board of the VUmc and Noordwest Ziekenhuisgroep approved the trial, which was in accordance with the Declaration of Helsinki and Good Clinical Practice. Written informed consent was obtained from all patients prior to inclusion into the study. The primary endpoint of this study was to determine whether supplementation of folic acid and vitamin B12 could increase the response rate (RR) of patients with advanced esophagogastric cancer treated with the combination of gemcitabine and cisplatin. The addition of folic acid and vitamin B12 did not improve RR, PFS or OS in this study [28]. Secondary endpoints included survival and pharmacokinetic analysis. Seventyeight patients were randomized to treatment with up to six cycles of cisplatin and gemcitabine in a three-weekly cycle with or without vitamin B12 and folic acid suppletion. Blood samples were drawn prior to onset of the study and collected in BECTON DICKINSON Vacutainer Heparin Tubes in which 10 %u00b5g/ml tetrahydrouridine (THU) was spiked in to avoid conversion of gemcitabine into 2%u2019,2%u2019-difluorodeoxyuridine (dFdU). This was done for pharmacokinetic analyses as previously reported. Follow-up samples were drawn before the third (n=35) and the sixth (n=12) cycle of chemotherapy. After centrifugation the plasma was taken off and stored at -25 %u00b0C until analysis. The plasma samples collected during the course of this study were used for ci-miRNA analysis. miRNA selectionSeven miRNAs were selected for measurement based on the literature (Table 1). PubMed search criteria included micro RNA and gastric or oesophageal cancer (MESH, all fields, and similar articles). miRNAs were selected if evidence was presented in at least two independent publications. miR-375, miR-200c-3p, miR21-5p and miR-148a-3p were selected for measurement in patients with EAC and ESCC. miR-200c-3p, miR-141-3p, miR-146a-5p and miR-218-5p were selected for measurement in patients with GC. These miRNAs were all previously detected in blood samples [29].