Subtype-specific NK cell-TAM interactions drive a novel prognostic signature in HNSCC.

Background: The immune microenvironment of head and neck squamous cell carcinoma (HNSCC) is highly complex, and the mechanisms underlying interactions between natural killer (NK) cells and tumor-associated macrophages (TAMs) remain unclear. This study investigates the cellular heterogeneity, interaction patterns, and prognostic significance of NK-TAM crosstalk through multi-omics analyses. Methods: A total of 58 HNSCC tissue samples were analyzed. NK and TAM subsets were identified using immunohistochemistry (CD16, CD64, CD163), single-cell RNA sequencing (GSE139324), and public databases (TCGA-HNSC, GSE65858). CellChat was used to infer ligand-receptor interactions, while spatial proximity was assessed via the CSOmap algorithm and validated by immunofluorescence. A prognostic model was constructed using LASSO Cox regression and validated in an immunotherapy cohort (PRJEB23709, phs000452.v2.p1). Results: High CD16/CD64 expression correlated with favorable prognosis, while CD163 indicated poor outcomes (P < 0.05). NK cells were divided into IL32+NK (antiviral, T cell-activating), NFKBIA+NK (ribosome-related), and STMN1+NK (DNA repair-related) subsets. TAMs included APOE+TAM (M2-like), IL1B+/CXCL10+TAM (M1-like), and HSP+TAM (stress-responsive). IL32+NK interacted most strongly with APOE+TAM and CXCL10+TAM via SPP1, MIF, and ITGB2 pathways. Spatial mapping and immunofluorescence confirmed proximity and a positive correlation between IL32 and CXCL10 (R = 0.641, P < 0.001), and a negative correlation with APOE (R=-0.686, P < 0.001). A 23-gene NK-TAM interaction-related signature (CINT) effectively stratified patient risk in both training and validation cohorts (P < 0.05) and predicted survival benefit in immunotherapy-treated patients. Conclusion: This study uncovers subtype-specific NK-TAM interactions in HNSCC and introduces CINT as a robust prognostic and immunotherapy response model, offering a new strategy for immune microenvironment-targeted therapy.