CCR5 Marks an Immunosuppressive Macrophage-Associated Microenvironment and Predicts Poor Prognosis in Glioma: An Integrative Multi-Omics Analysis

Background: Glioma develops within a strongly immunosuppressive tumor microenvironment (TME), which contributes to poor therapeutic response and unfavorable prognosis. Although C-C Chemokine Receptor Type 5 (CCR5) has previously been associated with glioma progression, prognosis, and tumor microenvironment remodeling, its cell-type-specific expression pattern and spatial context within the heterogeneous glioma microenvironment remain incompletely characterized. Methods: A comprehensive multi-omics analysis was conducted by integrating bulk transcriptomic profiles from The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) cohorts with single-cell RNA sequencing and spatial transcriptomics datasets. The expression pattern, prognostic value, and immune-related functions of CCR5 were systematically evaluated. To estimate immune infiltration, Tumor Immune Estimation Resource (TIMER), Cell-type Identification By Estimating Relative Subsets Of RNA Transcripts (CIBERSORT), and Single-sample Gene Set Enrichment Analysis (ssGSEA) algorithms computational approaches were applied. A prognostic nomogram incorporating CCR5 and clinical variables was constructed and validated. Experimental validation was performed using immunohistochemistry, Western blotting, and dual immunofluorescence staining. Results: CCR5 expression was markedly elevated in glioma and associated with more aggressive clinicopathological characteristics, including advanced WHO grade, Isocitrate Dehydrogenase (IDH) wild-type status, and 1p/19q non-codeletion. Elevated CCR5 expression predicted significantly worse overall survival across the TCGA and CGGA cohorts, while remaining independently associated with patient prognosis following multivariate analysis. Functional annotation indicated that CCR5 was predominantly associated with immune-related biological processes and signaling pathways, particularly cytokine–cytokine receptor interactions. Immune infiltration analyses demonstrated that CCR5 expression was closely associated with macrophage-related immune features, regulatory T cells, and multiple inhibitory immune checkpoint molecules. Single-cell transcriptomic analysis showed that detectable CCR5 transcripts were preferentially distributed in macrophage-associated populations. Spatial transcriptomic analysis further revealed sparse and heterogeneous CCR5 distribution, with context-dependent spatial relationships between CCR5-detectable regions and macrophage-associated transcriptional features. Experimental validation confirmed increased CCR5 protein expression in high-grade glioma tissues, while dual immunofluorescence demonstrated CCR5 colocalization with CD163-positive macrophages in glioma tissues. Conclusions: Our integrative analyses associate elevated CCR5 expression with unfavorable prognosis and a macrophage-enriched immunosuppressive microenvironment in glioma. By resolving CCR5 expression across bulk, single-cell, and spatial transcriptomic levels, this study provides additional cellular and spatial context for its previously reported clinical and immunological relevance. These findings support further investigation of the potential role of CCR5 in macrophage-associated immune remodeling in glioma.