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.