Background Glioblastoma (GBM) is the most aggressive primary brain tumor with a survival rate of 14–16 months. Several ongoing clinical trials employ vaccine, immune checkpoint inhibitors and/or adoptive T cell therapy to treat GBM. However, the impact of irradiation as part of the standard of care on the immune microenvironment and response to immunotherapy is ill-defined.

Methods In the present study, we employed clinically relevant fractionated irradiation (FIR) at a dose of 4x 2 Gy to investigate the response of experimental orthotopic glioma expressing the tumor-associated antigen glycoprotein (gp)100 in immunocompetent syngeneic mice to a gp100 peptide vaccine as well as to adoptively transferred T cells. We employed single cell RNA sequencing to decipher the post-irradiation changes in the GBM immune microenvironment and elucidate the impact FIR on the efficacy of T cell-mediated immunotherapies.

Results FIR boosted the response of orthotopically-implanted GL261 tumors to a gp100 vaccine. Furthermore, FIR enhanced the infiltration of GL261-gp100 gliomas by adoptively-transferred gp100 T cell receptor-transgenic T cells, which also displayed a more activated and less exhausted phenotype post-irradiation. Single cell transcriptomic studies of tumor-infiltrating CD45⁺ leukocytes revealed that FIR promotes the expression of leukocyte-endothelial adhesion molecules in T cells, and transcripts encoding proinflammatory M1-like genes in tumor-associated microglia and macrophages.

Conclusions Our preclinical findings support the use of FIR with to increase response to T cell-based immunotherapies.