Differential Diagnosis of Benign/Malignant Biliary Stenoses by DNA Analysis Using Bile

Abstract

Diagnosis of benign/malignant common bile duct (CBD) stenoses is often difficult. No currently available diagnostic modality ensures absolute accuracy in differentiating malignant from nonmalignant CBD stenoses. Cell-free nucleic acids in bile might serve as potential diagnostic biomarkers. We investigated whether cell-free DNA in bile could distinguish malignant from nonmalignant CBD stenoses. Bile samples were collected from 28 patients undergoing therapeutic endoscopic retrograde cholangiopancreatography at university hospitals for CBD stenosis of malignant (biliary cancer, n=9; pancreatic cancer, n=9) or nonmalignant (n=10) disease. KRAS mutations and DNA methylation were analyzed by quantitative pyrosequencing. Receiver operating characteristic area under the curve (AUC) was used to determine diagnostic accuracy. KRAS mutations were detected in 67% of biliary cancers, 56% of pancreatic cancers, and 10% of the nonmalignant disease. No significant differences were found in DNA methylation levels of CDKN2A, MINT25, SOX17, and miR-34b/c between malignant and nonmalignant diseases. BARHL2 methylation levels were significantly higher in bile samples from patients with malignant CBD stenoses than in those with nonmalignant CBD stenoses. Moreover, bile BARHL2 methylation levels distinguished malignant from nonmalignant CBD stenoses with 88% sensitivity and 100% specificity (AUC=0.95556). Thus, the analysis of bile BARHL2 methylation levels may be useful for the diagnosis of pancreatobiliary malignancies in clinical settings.