Single-cell and spatial transcriptomics uncover a THBS2⁺ CAF–KRT17⁺ malignant cell axis drives progression and immunotherapy resistance in pancreatic cancer

Background Pancreatic ductal adenocarcinoma (PDAC) features a fibrotic stroma dominated by cancer-associated fibroblasts (CAFs), yet the functional heterogeneity of CAFs and their crosstalk with malignant cells in metastasis and immune evasion remain unclear. Methods We performed an integrated analysis of single‑cell RNA‑seq data and spatial transcriptomics of PDAC. Pseudotime trajectory reconstruction, regulon activity inference, and ligand–receptor interaction mapping were applied to dissect fibroblast heterogeneity and stromal–epithelial crosstalk. Bulk transcriptomic data from multiple independent cohorts were used to evaluate prognostic and immunotherapeutic implications. Results We identified nine CAF subsets, including a THBS2⁺ ECM‑remodeling population enriched in liver metastases. Spatial transcriptomics localized these THBS2⁺ CAFs to dense stromal regions surrounding malignant ducts, where they established an immune‑excluded microenvironment correlated with T‑cell exclusion and immunotherapy resistance. Pseudotime analysis revealed PRRX2 as a master regulator driving this CAF subset. Epithelial sub-clustering uncovered KRT17⁺ malignant cells (a transcriptionally distinct epithelial subpopulation associated with EMT and metastasis) that co-enriched with THBS2⁺ CAFs in metastatic sites. Ligand–receptor analysis showed that THBS2⁺ CAFs are computationally predicted to signal to KRT17⁺ cells via the THBS2-SDC4 axis, while malignant cells reciprocally activate CAFs through MDK–LRP1 and SPP1–integrin pathways. Co‑infiltration of both populations strongly predicted poor prognosis and resistance to anti‑PD‑L1 therapy across multiple cohorts. Conclusions The THBS2⁺ CAF–KRT17⁺ malignant cell axis drives metastatic progression and immune exclusion in PDAC via THBS2–SDC4 signaling, representing a promising prognostic biomarker and therapeutic target.