Abstract BACKGROUND: The benefits of immunotherapy are often hampered by immunosuppressive networks in the tumor microenvironment. Accumulating evidence indicates that local ionizing radiation (IR) can synergize with immunotherapy and improve tumor response.CD137/4-1BB is a co-stimulatory molecule expressed on activated T-cells that provides a survival signal to promote the expansion and differentiation of antigen-specific T-cells. Pre-clinical studies have shown the ability of agonistic anti-CD137 antibodies to enhance anti-tumor immunity. 4T1 is a poorly immunogenic experimental mouse model of metastatic breast cancer. 4T1 cells injected s.c. form a tumor and rapidly spread systemically giving rise to lung metastases that kill the animals. We have previously shown that IR as single modality can delay growth of the irradiated 4T1 s.c. tumors but not extend mice survival.METHODS: BALB/c mice were inoculated s.c. with 4T1 cells on day 0. Treatment with anti-CD137 mAb (200 mg, BMS-469492 from Bristol Myers Squibb) was started on day 7 or 15 and repeated every 3 days for two or three times. For radiotherapy mice were randomized into 4 groups receiving (1) no treatment, (2) IR alone, (3) anti-CD137 mAb and (4) IR+ anti-CD137 mAb. IR was delivered exclusively to the tumor in 2 fractions of 12 Gy at 24 hours interval starting on day 13. Mice were followed for tumor growth and survival.RESULTS: Administration of anti-CD137 mAb to 4T1 tumor-bearing mice starting at day 7 resulted in significant tumor growth delay which correlated with an 8-fold increase in the ratio of intratumural effector CD8 T cells (Teff) to regulatory T cells (Treg) at day 15. In contrast, when administration of anti-CD137 mAb was delayed until day 15 no anti-tumor effect was observed.The median survival of mice (39 days) was not increased by treatment with anti-CD137 mAb or IR used as single modalities (39 days). On the other hand, the combination of IR and anti-CD137 mAb increased survival to 49 days, although no complete tumor remission was observed.CONCLUSIONS:Overall, our studies indicate that with tumor progression mice became resistant to immunotherapy with single modality anti-CD137 mAb. However, a therapeutic effect was seen when anti-CD137 mAb was combined with local radiotherapy. We are currently investigating the mechanisms of resistance to anti-CD137 mAb treatment that develop with 4T1 tumor progression. Accumulation of Treg and myeloid-derived suppressor cells (MDSC) within the tumors likely contribute to resistance to immunotherapy by creating a highly immunosuppressive microenvironment. Identification of these mechanisms will provide new therapeutic targets to improve the response to treatment. Importantly, data support the combination of local radiotherapy with immunotherapy in advanced breast cancer. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4134.