Single-cell spatial analysis of pediatric high-grade glioma reveals a novel population of SPP1+/GPNMB+ myeloid cells with immunosuppressive and tumor-promoting capabilities

Abstract

BackgroundPediatric-type diffuse high-grade gliomas (pHGG) are a leading cause of pediatric cancer-related mortality. Although immunotherapy offers a promising treatment avenue, clinical responses in pHGG patients remain limited. A detailed understanding of the tumor immune microenvironment (TIME) is essential for advancing immunotherapeutic strategies. MethodsWe performed single-cell spatial analysis integrating cyclical immunofluorescence imaging and Spatial Molecular Imaging to interrogate the proteomic and transcriptomic landscape of pHGG. A tissue microarray comprising 32 diagnostic patient-derived pHGG samples was utilized to map the spatial distribution of immune and tumor cells. ResultsOur analyses reveal that the pHGG TIME is predominantly composed of myeloid cells, including brain-resident microglia and monocyte-derived macrophages, with only few T cells. A significant subset of these myeloid cells express mesenchymal-like genes and are positive for SPP1 and GPNMB. Spatial mapping further demonstrated that SPP1+/GPNMB+ myeloid cells localize in close proximity to mesenchymal-like tumor cells, and negatively correlate with the location and presence of CD8+ T cells. These cells also express genes related to immunosuppression and epithelial-to-mesenchymal transition, indicating their potential role in establishing an immunosuppressive niche. ConclusionsOur findings reveal a distinct immune landscape in pHGG characterized by SPP1+/GPNMB+ myeloid cells which may contribute to the exclusion of CD8+ T cells. This spatially resolved insight identifies these myeloid cells as promising therapeutic targets and provides a rationale for developing novel immunotherapeutic strategies to improve outcomes in pediatric high-grade gliomas.