Integrated Bile Metabolomics and Serum Biomarkers for Differentiating Cholangiocarcinoma from Cholelithiasis

Background: We investigated bile metabolites and serum variables for differentiating cholangiocarcinoma (CCA) from cholelithiasis. Methods: Bile from 31 patients with CCA and 48 with cholelithiasis was profiled by high-resolution LC-MS. Candidate metabolites and serum variables were integrated in a seven-variable classifier, and public single-cell data provided tissue-level biological context. Results: Of 2269 annotated metabolite features, 532 met the exploratory screening criteria. The selected signature comprised pyruvic acid, lipoxin A4, CEA, APOA1, 3-chloro-L-tyrosine, ADP-ribose, and glutaric acid; pyruvic acid had the largest contribution in the SHAP analysis. Random forest achieved an AUC of 0.894 in the classifier comparison, a training cross-validation AUC of 0.820, and an AUC of 0.914 in the 16-patient test subset. Because selection used all 79 patients before evaluation, these internal performance estimates may be optimistic. Conclusions: Integrated bile and serum profiling identified a promising seven-variable signature for differentiating CCA from cholelithiasis in this cohort. Added diagnostic value over routine serum markers and performance in other benign biliary conditions have not been established.