Micrornas in ascites as potential biomarkers for peritoneal carcinomatosis and peritonitis

Abstract

Introduction: Peritoneal carcinomatosis (PCA) has a prognostic role in patients with gastrointestinal cancers. Despite the low sensitivity, cytology remains the gold standard in differential diagnosis of PCA to peritonitis (for example spontaneous bacterial peritonitis, SBP) or uncomplicated ascites due to portal hypertension (no SBP/PCA). MicroRNAs (miRNAs) are considered as promising biomarkers and are commonly dysregulated in cancer. Aims & Methods: In this proof-of-principle study, we systematically evaluated preanalytical factors and potential of miRNAs as ascites biomarkers. We prospectively examined samples from patients with ascites with benign and malignant conditions including: PCA (n¼15), SBP (n¼15) and portal hypertension (no SBP/PCA, n¼15). Various extraction kits were used to compare the total RNA extraction. Furthermore, we systematically evaluated the influence of storage, stability and sample processing (uncentifuged, pelleted etc.) on miRNA expression in ascites. MiRNA expression profiling using TaqMan Low Density Array (TLDA) and quantitative RT-PCR (TaqMan/SYBRgreen) were used to evaluate the expression. Results: Systematic analysis of miRNAs stability confirms that miRNAs in ascites are well preserved from degradation with good short- (0h, 12h, 24h, and 48h) and long-term stability (-30C,80C for 2 years). Several miRNAs that were selected for the proof-of-principle analysis (miR-21 and miR-16) were reproducibly detectable in ascites samples. MiRNA expression profiling in patients with PCA compared to those with uncomplicated portal hypertension revealed miR-21, miR-186, miR-222 and miR-483-5p to be up-regulated and miR-26b to be down-regulated. MiRNA expression validation analysis confirmed higher expression of miR-21 (mean delta CTSD; 11.111.2 vs. 8.463.46 vs. 9.652.55 for no SBP/PCA, PCA and SBP, respectively, ANOVA, p¼0.026; posttest no SBP/PCA vs. PCA p50.05