Distinct tumor immune microenvironmental landscapes drive divergent immunotherapy responses in glioblastoma

BACKGROUND: Immunotherapies have improved outcomes in many cancers but remain largely ineffective in glioblastoma (GBM). We investigated whether immunotherapy could be rationally tailored to GBM by functionally subtyping the tumor immune microenvironment (TIME) and associated vascular landscapes. METHODS: Single-cell and single-nucleus RNA sequencing, multiplex immunohistochemistry, and flow cytometry were used to define TIME subtypes in human and murine GBMs. Therapeutic responses to anti-angiogenic and immunomodulatory therapies, including CD40 agonist, αPD-L1, and PI3Kγ/δ inhibition, were assessed in orthotopic syngeneic GBM mouse models. RESULTS: Three distinct functional TIME subtypes with unique vascular-immune landscapes were identified in human and murine GBM. TIME-low tumors were immune-low/deserted with a leaky vasculature. TIME-med GBM exhibited intermediate immune-infiltration, prominent angiogenesis, and active immune responses. TIME-high tumors showed dense infiltration of immunosuppressive myeloid cells and dysfunctional T cells. Representative mouse models demonstrated subtype-specific sensitivities to anti-angiogenic immunomodulating therapies. TIME-low GBMs exhibited transient T-cell activation but relapsed due to myeloid-driven immunosuppression and mesenchymal transition. TIME-med tumors displayed the most favorable responses across anti-angiogenic immunomodulating therapies. TIME-high GBMs were largely resistant, although therapeutic efficacy improved with myeloid-targeted PI3Kγ inhibition. In contrast, CD40 agonist therapy worsened survival by enhancing angiogenesis, amplifying immunosuppression, impairing T cell function, reducing NK-cell recruitment, and promoting tumor propagation. CONCLUSIONS: GBM comprises 3 functional TIME subtypes with divergent vascular-immune landscapes that require subtype-specific therapeutic strategies. TIME-med tumors are most amenable to immunotherapies. TIME-low tumors derive transient effects with anti-angiogenic immunomodulating therapies, and TIME-high are resistant or even experience worse outcome without targeted reversal of myeloid immunosuppression.