A major factor limiting the efficacy of cancer immunotherapy is the development of mechanisms allowing tumors to resist or escape immune rejection. We are studying these mechanisms in vivo in preclinical models in order to define strategies to boost immunotherapy by overcoming tumoral immune resistance. We focus on the role of regulatory T lymphocytes, myeloid suppressor cells, indoleamine 2,3-dioxygenase, and other potential immunosuppressive molecules. Natural regulatory T cells. Our current results show that depletion of natural regulatory T cells (by anti-CD25 mAb treatment) induces rejection of P815 mastocytoma in 70-to-80% of mice. These observations suggest that natural regulatory T cells inhibit the spontaneous tumor-specific immune response, a hypothesis consistent with the infiltration of cells displaying the phenotype of natural T regulatory T cells in P815. We are characterizing the specificity, phenotype and function of Treg infiltrating the tumor at various time points (using FACS, confocal microscopy, RT-PCR, in vitro coculture) in order to test whether distinct subpopulations , i.e. natural or induced, infiltrate a growing P815 tumor. Indoleamine 2, 3-dioxygenase (IDO) In addition to tumor cells, antigen-presenting-cells have been shown to produce IDO, in particular upon engagement with CTLA-4 receptor on regulatory T cells. We are evaluating the impact of IDO expressed by tumor cells versus immune cells, using tumor cells expressing various amounts of IDO (available at ICP) and WT or IDO-KO mice (kindly provided by A. Mellor). We have evidence that regulatory T cell populations may induce IDO production by DC in vitro and in vivo Myeloid suppressor cells The comparison of cell populations which infiltrate progressing versus regressing P815 tumors led us to identify a population of CD11c+ CD11b+ Gr1+ cells which infiltrates regressing tumors. By contrast, a population of CD11c- CD11b+ Gr1+ cells is detected in progressing P815 tumors. The latter cells, which have been observed in other cancer models and also in some cancer patients, are thought to bear immunosuppressive properties and have been called Myeloid Suppressor Cells (MSC). We will characterize the phenotype and function of both subpopulations, and in particular analyse their production of cytokines known to influence the outcome of tumor growth. [A major factor limiting the efficacy of cancer immunotherapy is the development of mechanisms allowing tumors to resist or escape immune rejection. We are studying these mechanisms in vivo in preclinical models in order to define strategies to boost immunotherapy by overcoming tumoral immune resistance. We focus on the role of regulatory T lymphocytes, myeloid suppressor cells, indoleamine 2,3-dioxygenase, and other potential immunosuppressive molecules. Natural regulatory T cells. Our current results show that depletion of natural regulatory T cells (by anti-CD25 mAb treatment) induces rejection of P815 mastocytoma in 70-to-80% of mice. These observations suggest that natural regulatory T cells inhibit the spontaneous tumor-specific immune response, a hypothesis consistent with the infiltration of cells displaying the phenotype of natural T regulatory T cells in P815. We are characterizing the specificity, phenotype and function of Treg infiltrating the tumor at various time points (using FACS, confocal microscopy, RT-PCR, in vitro coculture) in order to test whether distinct subpopulations , i.e. natural or induced, infiltrate a growing P815 tumor. Indoleamine 2, 3-dioxygenase (IDO). In addition to tumor cells, antigen-presenting-cells have been shown to produce IDO, in particular upon engagement with CTLA-4 receptor on regulatory T cells. We are evaluating the impact of IDO expressed by tumor cells versus immune cells, using tumor cells expressing various amounts of IDO (available at ICP) and WT or IDO-KO mice (kindly provided by A. Mellor). We have evidence that regulatory T cell populations may induce IDO production by DC in vitro and in vivo Myeloid suppressor cells The comparison of cell populations which infiltrate progressing versus regressing P815 tumors led us to identify a population of CD11c+ CD11b+ Gr1+ cells which infiltrates regressing tumors. By contrast, a population of CD11c- CD11b+ Gr1+ cells is detected in progressing P815 tumors. The latter cells, which have been observed in other cancer models and also in some cancer patients, are thought to bear immunosuppressive properties and have been called Myeloid Suppressor Cells (MSC). We will characterize the phenotype and function of both subpopulations, and in particular analyse their production of cytokines known to influence the outcome of tumor growth.]