BACKGROUND: Strategies to modulate the tumor microenvironment (TME) have opened new therapeutic avenues with dramatic yet heterogeneous intertumoral efficacy in multiple cancers, including glioblastomas (GBMs). Therefore, investigating molecular actors of TME may help understand the interactions between tumor cells and TME. Immune checkpoint proteins such as a Cluster of Differentiation 80 (CD80) and CD86 are expressed on the surface of tumor cells and infiltrative tumor lymphocytes. However, their expression and prognostic value in GBM microenvironment are still unclear. METHODS: In this study, we investigated, in a retrospective local discovery cohort and a validation TCGA dataset, expression of CD80 and CD86 at mRNA level and their prognostic significance in response to standard of care. Furthermore, CD80 and CD86 at the protein level were investigated in the discovery cohort. RESULTS: Both CD80 and CD86 are expressed heterogeneously in the TME at mRNA and protein levels. In a univariate analysis, the mRNA expression of CD80 and CD86 was not significantly correlated with OS in both local OncoNeuroTek dataset and TCGA datasets. CD80 and CD86 mRNA high expression was significantly associated with shorter progression free survival (PFS) (p < 0.05). These findings were validated using the TCGA cohort; higher CD80 and CD86 expressions were correlated with shorter PFS (p < 0.05). In multivariate analysis, CD86 mRNA expression was an independent prognostic factor for PFS in the TCGA dataset only (p < 0.05). CONCLUSION: CD86 could be used as a potential biomarker for the prognosis of GBM patients treated with immunotherapy; however, additional studies are needed to validate these findings.