Unravelling the diagnostic dilemma of pancreatic head lesions: A circulating microRNA panel to diagnose distal cholangiocarcinoma

Introduction: Accurate diagnosis of distal cholangiocarcinoma (CCA) and pancreatic ductal adenocarcinoma (PDAC) is a challenge with clinical consequences. Both are lethal malignancies with distinct therapeutic options. This study aimed to identify a circulating microRNA (miRNA) signature to diagnose and discriminate distal CCA from PDAC. Methods: In the discovery phase, microarray profiling of 752 miRNAs was performed on plasma samples of seven patients with distal CCA and seven age- and sex-matched healthy controls. Significant candidate miRNAs were selected for validation based on predefined selection criteria. In the validation phase, these miRNAs were analyzed by RT-qPCR in an independent cohort of healthy controls (n=32), benign periampullary disease (n=20), distal CCA (n=24), and age- sex- and stage-matched PDAC (n=24). Data were normalized to a combination of two reference genes. The optimal diagnostic combination of miRNAs was determined by logistic regression. Sensitivity and specificity were evaluated by ROC curves and AUC values. Results: In the discovery phase, 19 miRNAs were significantly deregulated in patients with distal CCA compared to healthy controls. In the validation phase, 12 candidate miRNAs were selected for validation by RT-qPCR based on pre-defined selection criteria. A two-miRNA panel of downregulated miR-16 and upregulated miR-877 was able accurately distinguish distal CCA from benign disease as well as from PDAC. Conclusion: This is the first study to identify a combined panel of plasma miRNAs which shows promising diagnostic capability to serve as distal CCA signature with the potential to discriminate distal CCA from PDAC. For the discovery cohort of this study, a microarray was performed on 7 healthy individuals and 7 patients with distal CCA.Plasma samples were collected at diagnosis and RNA was isolated using the small RNA isolation kit (Exiqon). Technical duplicated were included in the subsequent analysis. The plasma miRNA expression levels of healthy individuals were compared to distal CCA.