CD44 identified as a diagnostic biomarker for highly malignant CA19-9 negative pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer with limited diagnostic biomarkers. Carbohydrate antigen 19-9 (CA19-9) is a widely used clinical biomarker and is generally considered to correlate with PDAC malignancy. However, the relationship between CA19-9 expression levels and tumor aggressiveness remains underexplored. In this study, we report a biphasic relationship between CA19-9 expression levels and PDAC malignancy, where both negative (<5 U/mL) and high (>37 U/mL) CA19-9 levels are associated with increased tumor aggressiveness. We defined CA19-9 negative PDAC as tumors that lack CA19-9 expression intracellulary, on the cell membrane, and in secreted form. In PDAC cell lines and patient-derived organoids, CA19-9 negativity, confirmed by immunofluorescence, flow cytometry and ELISA, correlated with more aggressive behaviors. In PDAC patients, tumors from those with serum CA19-9 levels below 5 U/mL exhibited stronger metabolically activity, more immunosuppressive tumor microenvironment, and worse survival than CA19-9 positive tumors, with over 90 % showing absent CA19-9 expression by immunohistochemistry (IHC). Glycoproteomics profiling identified CD44 as a highly expressed biomarker in CA19-9 negative PDAC. Elevated CD44 expression effectively distinguished CA19-9 negative PDAC from both CA19-9 positive PDAC and CA19-9 negative benign pancreatic diseases, suggesting its potential as a diagnostic tool. Furthermore, we developed a radionuclide-labeled CD44 antibody ⁸⁹Zr-1M2E3, which specifically recognized CA19-9 negative PDAC tumors in preclinical models using PET-CT imaging. These findings highlight CD44 as a promising biomarker and therapeutic target for diagnosing and treating CA19-9 negative PDAC.