The glioblastoma (GBM) microenvironment is enriched in immunosuppressive factors that potently interfere with the function of cytotoxic T lymphocytes. Cancer cells can directly affect the immune system, but the mechanisms driving these interactions are not completely clear. Here, we demonstrate that the polyamine metabolite spermidine (SPD) was elevated in the GBM tumor microenvironment. Exogenous administration of SPD drove tumor aggressiveness in an immune-dependent manner in preclinical mouse models via reduction of CD8 ⁺ T cell frequency and reduced cytotoxic function. Knockdown of ornithine decarboxylase, the rate-limiting enzyme in SPD synthesis, did not affect cancer cell growth in vitro but did result in extended survival. Furthermore, patients with GBM with a more favorable outcome had a significant reduction in SPD compared with patients with a poor prognosis. Our results demonstrate that SPD functions as a cancer cell–derived metabolite that drives tumor progression by reducing CD8 ⁺ T cell numbers and function.

Matrix-assisted laser desorption/ionization mass spectrometry imaging allows for the study of metabolic activity in the tumor microenvironment of brain cancers. The detectable metabolites within these tumors are contingent upon the choice of matrix, deposition technique, and polarity setting. In this study, we compared the performance of three different matrices, two deposition techniques, and the use of positive and negative polarity in two different brain cancer types and across two species. Optimal combinations were confirmed by a comparative analysis of lipid and small-molecule abundance by using liquid chromatography–mass spectrometry and RNA sequencing to assess differential metabolites and enzymes between normal and tumor regions. Our findings indicate that in the tumor-bearing brain, the recrystallized α-cyano-4-hydroxycinnamic acid matrix with positive polarity offered superior performance for both detected metabolites and consistency with other techniques. Beyond these implications for brain cancer, our work establishes a workflow to identify optimal matrices for spatial metabolomics studies.

High-grade gliomas are aggressive primary brain cancers with poor response to standard regimens, driven by immense heterogeneity. In isocitrate dehydrogenase ( IDH ) wild-type high-grade glioma (glioblastoma, GBM), increased intra-tumoral heterogeneity is associated with more aggressive disease. Recently, spatial technologies have emerged to dissect this complex heterogeneity within the tumor ecosystem by preserving cellular organization in situ . Here, we construct a high-resolution molecular landscape of GBM and IDH -mutant high-grade glioma patient samples to investigate the cellular subtypes and spatial communities that compose high-grade glioma using digital spatial profiling and spatial molecular imaging. This uncovered striking diversity of the tumor and immune microenvironment, that is embodied by the heterogeneity of the inferred copy-number alterations in the tumor. Reconstructing the tumor architecture revealed two distinct niches, one composed of tumor cell states that most closely resemble normal glial cells, associated with microglia; and the other niche populated by monocytes and mesenchymal tumor cells. We further reveal that communication between tumor and immune cells is underpinned by tumor-specific ligands, such as TGFβ signaling in astrocyte-like tumor cells. This primary study reveals high levels of intra-tumoral heterogeneity in high-grade gliomas, associated with a diverse immune landscape within spatially localized regions.