Ultra-low-pass whole-genome sequencing (ULP-WGS) of bile cell-free DNA (cfDNA) may improve the early detection of biliary tract malignancies, distinguish cancer types, and support prognostic assessment, according to an exploratory study involving 95 patients with biliary strictures. The approach demonstrated potential for identifying malignancy before confirmation through routine diagnostic procedures.
Distinguishing malignant from benign biliary strictures remains a significant clinical challenge. Although targeted mutation analysis using the Bilemut assay enables sensitive detection of malignancy, the technique cannot reliably differentiate cholangiocarcinoma (CCA) from pancreatic ductal adenocarcinoma (PDAC). Additional approaches are therefore needed to improve early diagnosis, determine tumour origin, and identify patients at increased risk of poor outcomes.
The investigation evaluated bile cfDNA using ULP-WGS, a genomic technique designed to assess tumour-derived genetic material with low DNA requirements. Bile samples were collected from 95 patients undergoing endoscopic retrograde cholangiopancreatography (ERCP) for biliary strictures suspected of malignancy. Initial cytology or biopsy findings classified 26 cases as malignant and 69 as indeterminate.
Tumour fraction and copy-number alterations (CNAs) were assessed using genomic analysis, while tumour mutational burden (TMB) was evaluated using the Bilemut panel. Diagnostic performance, tumour classification, and overall survival were subsequently examined. Bile-based ULP-WGS demonstrated greater sensitivity for tumour detection than plasma-based analysis in matched samples. Among patients with an initial malignant diagnosis, tumour-derived DNA was detected in 73% of cases.
Among patients initially classified as indeterminate but subsequently confirmed to have malignancy, tumour DNA was detected in 51% of cases. Detection preceded the final diagnosis by a median of 38 days. In seven patients, tumour-derived DNA was identified more than 100 days before routine diagnostic confirmation, indicating the potential for earlier identification of malignancy. Specificity reached 96% among patients whose biliary strictures were ultimately determined to be benign, supporting the potential utility of the approach in diagnostic assessment.
Copy-number alteration profiling identified recurrent chromosomal changes consistent with previously reported tissue-based findings. A classifier incorporating 50 genomic regions distinguished CCA from PDAC with an accuracy of 86.4%, demonstrating potential for determining tumour origin. Neither tumour fraction nor TMB independently showed an association with overall survival. However, combining both measures identified a subgroup with poorer prognosis, suggesting that integrated genomic assessment may provide additional prognostic information.
The findings indicate that bile-based ULP-WGS performed during the first ERCP could complement existing diagnostic methods for suspected biliary malignancies. The technique may facilitate earlier detection, improve differentiation between CCA and PDAC, and support prognostic stratification when combined with mutation-based analysis. Further validation in larger patient populations is required to establish diagnostic accuracy, clinical utility, and cost-effectiveness.
References
- Rojo C, Rullan M, Sogbe M, Herranz JM, Elizalde M, Oyón D, et al. Ultra-low-pass whole-genome sequencing of bile cfDNA in patients with biliary strictures. eGastroenterology. 2026;4(3):e100425.