Extracellular vesicle biomarkers in pancreatic ductal adenocarcinoma: from bulk detection to single-extracellular vesicle profiling and endoscopic liquid biopsy

Pancreatic ductal adenocarcinoma (PDAC) remains a deadly malignancy owing to its late presentation and the limited sensitivity of current serum biomarkers and imaging for early detection. Extracellular vesicles (EVs), which carry proteins, nucleic acids, and lipids that reflect tumor-stromal interactions, have emerged as promising biomarkers for early diagnosis, disease monitoring, and treatment response. Several EV-derived proteins, microRNAs, long noncoding RNAs, and DNA alterations linked to early carcinogenesis and therapeutic resistance have been identified. However, variability in pre-analytical handling and analytical platforms has hindered reproducibility. Global standardization efforts, such as European Liquid Biopsy Society (ELBS), The Blood Profiling Atlas in Cancer (BloodPAC), International Liquid Biopsy Standardization Alliance (ILSA), and Minimal information for studies of extracellular vesicles (MISEV), are currently helping to unify methodologies for EV isolation and molecular profiling. Advances in analytical technologies have shifted the field from bulk EV measurements to high-resolution single-vesicle approaches. Techniques, such as nanoflow cytometry, super-resolution imaging, Raman spectroscopy, and surface-enhanced Raman scattering, enable the detection of rare, mutation-bearing, or functionally distinct EV subpopulations, which may enhance diagnostic precision. In gastroenterology, a major opportunity lies in integrating single-EV analytics with the endoscopic sampling of tumor-proximal fluids. EVs obtained from pancreatic juice, bile, duodenal fluid, or portal venous blood via endoscopic retrograde cholangiopancreatography (ERCP) or endoscopic ultrasound (EUS) provide spatially enriched molecular information beyond peripheral blood. Combining endoscopic access with particle-level EV characterization may allow real-time, mechanism-informed assessment of tumor biology and premalignant lesions, offering a promising strategy for early detection and risk stratification in PDAC. Together, these developments have positioned EV-based liquid biopsy as a rapidly maturing field with strong translational potential.