Colorectal cancer (CRC) screening relies primarily on stool analysis to identify occult blood. However, its sensitivity for detecting precancerous lesions is limited, requiring the development of new tools to improve CRC screening. Carcinogenesis involves significant alterations in mucosal epithelium glycocalyx that decisively contribute to disease progression. Building on this knowledge, we examined patient series comprehending premalignant lesions, colorectal tumors, and healthy controls for the T-antigen—a short-chain O-glycosylation of proteins considered a surrogate marker of malignancy in multiple solid cancers. We found the T-antigen in the secretions of dysplastic lesions as well as in cancer. In CRC, T-antigen expression was associated with the presence of distant metastases. In parallel, we analyzed a broad number of stools from individuals who underwent colonoscopy, which showed high T expressions in high-grade dysplasia and carcinomas. Employing mass spectrometry-based lectin-affinity enrichment, we identified a total of 262 proteins, 67% of which potentially exhibited altered glycosylation patterns associated with cancer and advanced pre-cancerous lesions. Also, we found that the stool (glyco)proteome of pre-cancerous lesions is enriched for protein species involved in key biological processes linked to humoral and innate immune responses. This study offers a thorough analysis of the stool glycoproteome, laying the groundwork for harnessing glycosylation alterations to improve non-invasive cancer detection.

Early detection of colorectal cancer (CRC) significantly improves overall prognosis and increases 5-year survival rates up to 90%. Current non-invasive screening methods for CRC, such as the Faecal Immunohistochemical Test (FIT), have some drawbacks, namely, low sensitivity and a high false-positive rate. The Sialyl-Tn (STn) antigen, frequently expressed in pre-malignant lesions and adenocarcinomas, has been shown to be detected by the novel monoclonal antibody L2A5. In this study, we explored the potential of L2A5 as a non-invasive CRC screening method in an attempt to overcome current limitations. The subjects were categorised into four groups based on colonoscopy findings: no lesion (NL), low-grade dysplasia (LGD), high-grade dysplasia (HGD), and colorectal cancer (CRC). Slot blot analysis using the L2A5 antibody was performed on stool samples from 95 colonoscopy patients. Our findings showed a differential STn expression between the different clinical groups, evidencing excellent discrimination between NL and CRC (AUC, 0.8252; 95% CI: 0.6983–0.9521; sensitivity, 70%). Moreover, moderate discrimination between the NL+LGD and HGD+CRC groups was discerned (AUC, 0.7766; 95% CI: 0.6792–0.8740; sensitivity, 58%). These findings support the application of L2A5 as a tool for detecting STn, allowing for the identification of advanced lesions in non-invasive CRC screening.