Dissecting the immunosuppressive microenvironment of LAMA3 + malignant cells and SPP1 + macrophages in esophageal squamous cell carcinoma using single-cell and spatial transcriptomics

Esophageal squamous cell carcinoma (ESCC) exhibits pronounced tumor microenvironment (TME) heterogeneity, which contributes to tumor invasion, immune evasion, and resistance to therapy. However, the cellular and spatial mechanisms driving this heterogeneity remain incompletely understood. We integrated single-cell RNA sequencing and spatial transcriptomics to characterize the cellular composition, spatial organization, and tumor–immune interactions within ESCC tissues. Ligand–receptor communication was inferred using NicheNet modeling, and functional roles of key genes were evaluated through in vitro assays. We identified a LAMA3⁺ epithelial subpopulation enriched at the invasive tumor front, characterized by strong epithelial–mesenchymal transition (EMT) signatures and poor prognostic relevance. Tumor-enriched SPP1⁺ macrophages with M2-like phenotypes were also identified and associated with adverse clinical outcomes. Spatial analyses revealed close colocalization of LAMA3⁺ epithelial cells and SPP1⁺ macrophages, while NicheNet modeling indicated reciprocal ligand–receptor interactions that directly mediated the extracellular matrix, and promoted desmoplastic niche formation. This cooperative network restricted lymphocyte infiltration despite a high tumor mutational burden, leading to an immune-excluded microenvironment. Clinically, co-enrichment of LAMA3 and SPP1 predicted impaired responses to anti–PD-L1 therapy. Functional assays further confirmed that LAMA3 promotes tumor cell proliferation, migration, and invasion. Our study identifies a synergistic LAMA3–SPP1 axis that promotes desmoplastic remodeling and immune evasion in ESCC, suggesting a potential therapeutic target to improve immunotherapy outcomes.