Studies and clinical trials of immunotherapy for GBM pointed out the immunosuppressive influence of the GBM microenvironment as a significant hurdle, however, GBM infiltrating immune cells have been found to express immune checkpoint molecules. Blocking these immunosuppressive mechanisms while generating a strong antitumor response is an intuitive strategy for cancer immunotherapy. A variety of tools are now available to test this strategy empirically and move these agents into clinical trials. Our group recently published CGS-9343B results demonstrating that PD-1 blockade, in combination with stereotactic radiation therapy resulted in a durable, long-term survival in GL261 bearing mice. Antibodies against co-stimulatory molecules, such as 4-1BB and Cytotoxic T-Lymphocyte Antigen 4 have the potential to enhance immune responses and produce anti-tumor immunity. 4-1BB is expressed on activated T cells and engagement of 4-1BB with its ligand drives proliferation of CD8 + T cells, increased pro-inflammatory cytokine production and plays an essential role in the formation of long-lived memory cytotoxic T cells. CTLA-4 signaling impairs the BMS-199264 hydrochloride capacity of T cells to proliferate and to produce pro-inflammatory cytokines. Blockade of CTLA-4 removes these suppressive signals and allows antigen-specific T cells to expand and perform their effector functions. Ipilimumab, a human monoclonal antibody that blocks CTLA- 4, has been approved by the FDA for first line treatment of advanced melanoma. In phase III trials, ipilimumab improved survival in patients with metastatic melanoma and produced a durable anti-tumor memory response. Ipilimumab has also been shown to induce regression of melanoma brain metastases and may be potentiated by radiation therapy. However, some patients treated with ipilimumab suffered from severe immune-related adverse events, which was consistent with the proposed mechanism of CTLA-4 blockade. An approach to overcome this burden is to combine CTLA-4 blockade with 4-1BB activation: both individual antibodies cause inflammation to selective organs, however, a combination of the two antibodies increased cancer immunity while reducing inflammation and autoimmune effects. To bolster the anti-tumor immunity created by the monoclonal antibodies anti-CTLA-4 and anti-4-1BB, our group investigated the effects of radiation on glioma treatment as well.