Simple SummaryCanine oligodendrogliomas are universally fatal primary brain tumors. Glioma-associated microglia/macrophages (GAMs) have been shown to contribute to immunosuppression and tumor progression in human glioblastoma (GBM). While a robust GAM infiltrate has been observed in canine oligodendrogliomas, their corresponding molecular signature has not previously been explored. The results of this study show that GAMs variably infiltrate canine oligodendrogliomas. We observed marked differences in GAM density within individual tumors and across tumors. We further observed elevations in several GAM-derived pro-tumorigenic molecules, suggesting that GAMs likely contribute to canine oligodendroglioma pathogenesis. However, similar to GAM density, the tumor tissue expression of the majority of molecules assayed demonstrated significant variability. This is in contrast to our previous work on canine astrocytomas, which more consistently demonstrated robust increases in several GAM-derived pro-tumorigenic molecules. This study raises the possibility that the immune microenvironment across oligodendrogliomas and astrocytomas has key differences which may be relevant to future therapeutic targeting.AbstractThe goal of this study was to define the glioma-associated microglia/macrophage (GAM) response and associated molecular landscape in canine oligodendrogliomas. Here, we quantified the intratumoral GAM density of low- and high-grade oligodendrogliomas compared to that of a normal brain, as well as the intratumoral concentration of several known GAM-derived pro-tumorigenic molecules in high-grade oligodendrogliomas compared to that in a normal brain. Our analysis demonstrated marked intra- and intertumoral heterogeneity of GAM infiltration. Correspondingly, we observed significant variability in the intratumoral concentrations of several GAM-associated molecules, unlike what we previously observed in high-grade astrocytomas. However, high-grade oligodendroglioma tumor homogenates (n = 6) exhibited an increase in the pro-tumorigenic molecules hepatocyte growth factor receptor (HGFR) and vascular endothelial growth factor (VEGF), as we observed in high-grade astrocytomas. Moreover, neoplastic oligodendrocytes displayed robust expression of GAL-3, a chimeric galectin implicated in driving immunosuppression in human glioblastoma. While this work identifies shared putative therapeutic targets across canine glioma subtypes (HGFR, GAL-3), it highlights several key differences in the immune landscape. Therefore, a continued effort to develop a comprehensive understanding of the immune microenvironment within each subtype is necessary to inform therapeutic strategies going forward.