Single-cell spatial transcriptional profiling of pancreatic ductal adenocarcinoma uncovers key immune-modulating and pro-metastatic mechanisms

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy characterized by profound tumor microenvironment (TME) heterogeneity and therapeutic resistance. However, the cellular dynamics, spatial architecture, and molecular networks driving progression from primary tumors (PTs) to hepatic metastases (HMs) remain incompletely understood. Here, we performed high-definition single-cell spatial transcriptomics using Xenium In Situ (5K) on formalin-fixed paraffin-embedded (FFPE) matched PDAC PTs and HMs, generating a single-cell spatial atlas encompassing more than 900,000 resolved cells. Integrated analysis with single-cell RNA-seq revealed distinct cellular composition, spatial organization, and cell–cell communication patterns between PTs and HMs. Hypoxia and genomic instability were identified as core drivers of tumor cell plasticity toward a basal-like phenotype, while cancer-associated fibroblasts (CAFs) further shaped malignant cell fate. Compared with stroma-rich PTs, HMs displayed spatial immune-stromal remodeling and formed an immunogenic invasive border with actionable therapeutic vulnerabilities. We further identified functionally specialized macrophage subsets that spatially orchestrate pro-metastatic programs: TREM2 + macrophages drive tumor vascular remodeling, whereas MMP12 + macrophages promote CAF-mediated extracellular matrix (ECM) reorganization to facilitate metastasis. Moreover, CXCL9/10 + cells recruit and activate CD8 + T cells, and ectopic expression of Cxcl9/10 significantly sensitizes PDAC to immune checkpoint blockade in immunocompetent mouse models. Collectively, our study delineates the spatially coordinated tumor–stromal–immune ecosystem that sustains PDAC progression and metastasis, unveils novel immune-modulating and pro-metastatic mechanisms, and provides targetable vulnerabilities to improve immunotherapy for metastatic PDAC.