Introduction

With a 5-year survival rate of about 10%, pancreatic ductal adenocarcinoma (PDAC) is one of the most deadly cancers in the world. Even among diseases of similar stages, patients' responses to chemotherapy vary greatly, as evidenced by the stark differences in short and median overall survival. Liquid biopsy has emerged as a promising tool for non-invasive tumor profiling, but its application in bile has not been extensively studied. Given the direct contact of bile with pancreatic and biliary malignancies, bile could represent an important substrate for tumor biomarkers and may provide diagnostic and prognostic insights. This study aimed to evaluate the feasibility of circulating tumor DNA (ctDNA) extraction from the bile.

Aims & Methods

A prospective observational study was conducted on patients with PDAC-associated obstructive jaundice undergoing endoscopic or percutaneous biliary drainage. Bile samples were collected during the procedure, alongside plasma and tissue biopsies obtained via EUS-guided fine-needle biopsy (EUS-FNB) or intraductal biopsies. DNA extraction and next-generation sequencing (NGS) were performed to analyze genetic alterations. Clinical data, including tumor stage, treatment response, and survival outcomes, were recorded. Statistical analyses were conducted using Stata 15, with significance set at p<0.05. The primary endpoint was to compare cfDNA extracted from bile with that from plasma and tumor tissue. Secondary outcomes included assessing the prognostic value of bile cfDNA and its correlation with disease progression and treatment response.

Results

A total of 46 patients with an histological diagnosis of PDAC were included in the preliminary analysis. The mean tumor size was 35 mm (SD 13,1), and 39% of patients had metastatic disease at diagnosis. Biliary drainage was performed by endoscopic retrograde cholangiopancreatography (ERCP) in 60.9% of cases, EUS-guided drainage in 32%, and percutaneous drainage in 7,1%.
Quantitative analysis revealed significantly higher cfDNA concentrations in bile (mean 49 ng/uL) compared to plasma (mean 3,3 ng/uL, p=0.001) Figure 1. cfDNA levels in bile correlated with disease stage, showing higher concentrations in metastatic patients (mean 81 ng/uL) than in locally advanced (22 ng/uL) or resectable cases (55ng/uL). On the contrary, no significant correlation was found between plasma cfDNA levels and tumor stage (p=0.94). At three-month follow-up, 25.9% of patients showed disease progression, while 48.2% had stable disease, and 11.1% achieved complete remission. Survival analyses suggested a non-significant correlation between bile cfDNA concentrations and patient’s prognosis (Spearman coefficient -0.23, p=0.2).

Conclusion

These preliminary findings demonstrate the feasibility and a potential role of bile liquid biopsy in PDAC patients. Bile showed significantly higher concentrations of cfDNA than plasma cfDNA and representing a promising substrate for future tumor analysis and for a deeper comprehension of patient’s prognosis. Further analyses, including genetic sequencing and larger cohort validation, are warranted to establish the clinical utility of bile-derived biomarkers in guiding personalized therapeutic strategies for PDAC patients.