Single-dose radiotherapy is more effective than fractionation when combined with anti-PD-1 immunotherapy in glioblastoma.

Glioblastoma (GBM), the most common adult primary brain tumor, has an average survival of only 15-18 months. Recently, the combination of immune checkpoint blockers paired with radiotherapy has shown promise in preclinical murine GBM models. Human clinical trials have largely failed. One reason for this may be the discrepancy between radiation protocols utilized in preclinical models versus clinical practice. For translational relevance, defining correct and comparable radiation dosages and schedules to achieve optimal synergy with immunotherapeutic drugs, is essential. We used the GL261-based syngeneic mouse GBM model to compare the effects of two radiation regimens on tumor cell growth and survival. We assessed the in vivo effects of a single dose of 10 Gy (10Gyx1) or five consecutive doses of 2 Gy (2Gyx5) on the tumor immune microenvironment over time and compared their efficacy when combined with anti-PD-1 in vivo. Our data show that the 10Gyx1 regimen is more effective than 2Gyx5 at inhibiting tumor cell proliferation and growth in vitro and in vivo. Both regimens preserved the antigen-presenting ability of both dendritic cells and local microglia, but 10Gyx1 led to the highest lymphocyte infiltration. The combination of radiation with the checkpoint blocker anti-PD-1 was advantageous for both radiation regimens with animals treated with the 10Gyx1 regimen surviving the longest. Our study highlights how radiation regimen choices may impact the translation of preclinical findings, and in particular, the effects of radiation and immunotherapy in GBM. This work and literature data on the effects of positive hypofractionation in human GBM patients suggest that applying fewer, higher-dose radiation fractions may benefit GBM patients and lead to tumoricidal effects without sacrificing favorable anti-tumor immune responders.