Stroma-associated extracellular vesicles snRNAs and piRNAs as non-invasive diagnostic and prognostic biomarkers in colon and pancreatic cancer

Background: Colon cancer (CC) and pancreatic ductal adenocarcinoma (PDAC) are among the most lethal gastrointestinal (GI) malignancies, largely due to limitations in early detection and risk stratification. While most circulating biomarkers currently under clinical evaluation are tumor cell–derived, the tumor stroma, and particularly cancer-associated fibroblasts (CAFs), represents an underexplored source of clinically informative signals. Methods: Based on our prior characterization of CAF-derived extracellular vesicles (EVs) cargo, we evaluated two small nuclear RNAs (snRNAs) and one PIWI-interacting RNA (piRNA) as EV-associated biomarkers in peripheral blood samples obtained prior to surgical intervention. EV RNA was isolated from serum samples of patients with colon pathology (including benign, premalignant, and malignant lesions), PDAC patients, and healthy donors. Small non-coding RNA (sncRNA) expression was quantified by RT–qPCR. Multiple machine learning models were applied to assess diagnostic and prognostic performance. The prognostic relevance of predicted target genes regulated by these sncRNAs was further evaluated in a large, publicly available CC cohort from The Cancer Genome Atlas. Results: The sncRNA panel discriminated CC patients from healthy individuals with high diagnostic accuracy (AUC = 0.866) and enabled discrimination of benign and premalignant colonic lesions. In PDAC, the same signature achieved robust diagnostic performance (AUC = 0.782), and combined analysis of CC and PDAC supported its ability to detect GI malignancy. In stage I–III CC patients, pre-operative EV sncRNA levels were associated with disease relapse, allowing stratification into low- and high-risk groups (AUC = 0.725; log-rank p = 0.009). Analysis of 43 predicted sncRNA target genes in a TCGA cohort of 597 CC patients revealed strong prognostic value for overall survival (AUC = 0.813; log-rank p < 0.001). Conclusions: CAF-associated EV snRNAs and piRNAs can be detected in peripheral blood and provide clinically relevant diagnostic and pre-operative prognostic information in CC and PDAC. By capturing stromal-derived signals distinct from tumor cell–intrinsic biomarkers, these molecules offer a complementary liquid biopsy strategy with potential utility for early detection, pre-surgical risk stratification, and personalized patient management. Together, our findings support the tumor stroma as a systemic source of accessible biomarkers and highlight CAF-associated EV sncRNAs as promising tools for improving clinical decision-making in GI cancers.