SPP1⁺ tumor-associated macrophages promote cervical cancer progression through a CD44-dependent mechanism involving NF-κB activation

Tumor-associated macrophages (TAMs) contribute to cervical cancer progression, but the macrophage states that mediate tumor–macrophage crosstalk remain incompletely defined. This study aimed to characterize SPP1⁺ macrophages and evaluate their clinical and functional relevance in cervical cancer. TCGA-CESC and GSE44001 cohorts were used to develop and evaluate a five-gene prognostic signature using genes associated with CIBERSORT-inferred M2 macrophage abundance. Public and in-house single-cell RNA-seq datasets spanning normal cervix, cervical lesions, and cervical cancer were integrated to define macrophage subsets. Clinical specimens were assessed by immunohistochemistry and multiplex immunofluorescence. Cell–cell communication analysis, pathway scoring, macrophage phenotyping, functional assays, CD44 neutralization, and xenograft experiments were performed to investigate SPP1-associated tumor progression. The five-gene signature stratified patients with cervical cancer into distinct prognostic groups and was associated with increased macrophage infiltration. Single-cell analysis identified an SPP1 + macrophage state enriched in cervical cancer. In clinical specimens, SPP1 was predominantly localized to CD68⁺ macrophage-enriched stromal regions adjacent to CD44⁺CK5⁺ tumor epithelial cells. Cell–cell communication analysis suggested enhanced SPP1-CD44 signaling in cervical cancer, and CD44-high epithelial cells showed increased NF-κB pathway activity. In THP-1-derived macrophages, SPP1 overexpression increased the CD206 + fraction and IL10 expression while reducing CD86 expression. Functionally, SPP1-overexpressing macrophages enhanced cervical cancer cell viability and migration. CD44 neutralization attenuated migration and NF-κB p65 phosphorylation induced by conditioned medium from SPP1-overexpressing macrophages. SPP1-overexpressing macrophages also accelerated xenograft growth. These findings support that macrophage-derived SPP1 promotes cervical cancer progression through tumor-cell CD44-dependent NF-κB activation.