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

This repository contains the raw data files of the NanoString CosMx spatial transcriptomic experiment used to generate the figures within the publication (doi: https://doi.org/10.1101/2025.03.18.643953).

The bulk RNA-sequencing data of patients is access-restricted and access can be requested at the European Genome-phenome Archive under the accession number EGAS00001008002. Published DMG single cell RNA-sequencing data were downloaded under accession code GSE184357. Published DMG Visium data were downloaded under accession code GSE194329. Published bulk RNA-sequencing data was downloaded from https://pedcbioportal.org/.

Pediatric-type diffuse high-grade gliomas (pHGG) are the leading cause of pediatric cancer-related deaths. Immunotherapy recently emerged as a promising novel treatment opportunity, but so far, clinical responses remain limited to only a small proportion of pHGG patients. Therefore, more insights into the pHGG’s tumor immune microenvironment, the target of immunotherapy, are urgently needed. As emerging evidence in other cancers points to the spatial architecture of the tumor immune microenvironment as a major determinant of therapeutic efficacy, we employed single-cell spatial analysis at the proteomic and transcriptomic level by combining cyclical immunofluorescence imaging and Spatial Molecular Imaging. We analyzed 32 patient-derived pHGG samples assembled into a tissue microarray and mapped their single-cell spatial landscapes. Our findings revealed that the tumor immune microenvironment is dominated by myeloid cells, including brain-resident microglia and monocyte-derived macrophages, with limited T cells. A large proportion of these myeloid cells express mesenchymal-like genes and were found to be positive for SPP1 and GPNMB. Spatial analysis uncovered that these cells are positioned close to mesenchymal-like tumor cells, and negatively correlated with the location of CD8⁺ T cells. SPP1⁺/GPNMB⁺ cells express genes related to immunosuppression and epithelial-to-mesenchymal transition, suggesting that these cells are an important component of the immunosuppressive tumor microenvironment of pHGG, and may serve as a potential novel immunotherapeutic target.