SPP1⁺ macrophage-derived WNT5A contributes to MMP11⁺ myCAF-mediated matrix remodeling and immunotherapy resistance in lung adenocarcinoma

Lung adenocarcinoma (LUAD) is the leading subtype of non-small cell lung cancer. However, the stromal-immune circuits that coordinate matrix remodeling and immunotherapy resistance in LUAD remain incompletely understood. This study aimed to characterize the stromal-immune interactions associated with LUAD progression and poor response to PD-1 blockade. Bulk transcriptomic datasets, single-cell RNA sequencing, and spatial transcriptomic data were integrated with clinical immunotherapy cohorts. Weighted gene co‑expression network analysis, survival meta‑analysis, and tissue enrichment scoring identified high‑risk cell subpopulations. Cell‑cell communication, virtual gene knockout, and spatial deconvolution were used to predict ligand‑receptor interactions and colocalization. Multiplex immunohistochemistry of clinical specimens validated spatial relationships. In vitro, MMP11-overexpressing cancer-associated fibroblasts (CAFs) and SPP1-overexpressing macrophages (Macs) were generated, and a macrophage-CAF co‑culture system was constructed. Migration, contractility, extracellular matrix deposition, and YAP/INHBA pathway activation was measured. In vivo, a murine tumor model was established and evaluated using in vivo imaging, flow cytometry, atomic force microscopy assay, HE, masson staining, and multiplexed IHC staining. MMP11⁺ myofibroblastic CAF (MMP11⁺ myCAF) was identified as a high-risk CAF subset enriched in primary tumor and metastatic lesions, characterized by elevated expression of matrix-remodeling and contractile genes, including MMP11 , ACTA2 , and TAGLN . Increased abundance of MMP11⁺ myCAF was associated with advanced tumor stage, metastasis, poor overall survival, and reduced benefit from immune checkpoint blockade (ICB). Spatial transcriptomic deconvolution and multiplex immunohistochemistry revealed that MMP11⁺ myCAFs spatially co-localized with SPP1⁺ Mac, a pro-tumor myeloid population enriched in malignant and metastatic LUAD tissues. Ligand-receptor and downstream target analyses identified WNT5A as a key macrophage-derived mediator linking SPP1⁺ Mac to MMP11⁺ myCAF activation. Mechanistically, SPP1⁺ Mac exhibited activation of NFKB2/RELB-associated transcriptional programs, and SPP1 overexpression increased Nfkb2, Relb, and WNT5A protein levels in macrophages. WNT5A stimulation promoted YAP/INHBA-associated signaling in CAFs, enhancing CAF migration, contractility and extracellular matrix deposition. In animal models, activation of WNT5A signaling impaired the therapeutic efficacy of PD-1 blockade, reduced intratumoral CD8⁺ T-cell infiltration, and increased MMP11⁺ CAF-associated collagen deposition and tissue stiffness. SPP1⁺ Mac-derived WNT5A may contribute to MMP11⁺ myCAF-mediated matrix remodeling and immunotherapy resistance in LUAD, highlighting a potential therapeutic strategy to overcome ICB resistance.