CD134 and CD137 primed CD8 T cells mount powerful effector responses upon recall, but even without recall these dual costimulated T cells respond to signal 3 cytokines like IL-12. We searched for alternative signal 3 receptor pathways and found the IL-1 family member IL-36R. While IL-36 alone did not stimulate effector CD8 T cells, in combination with IL-12, or more surprisingly IL-2, induced striking and rapid TCR-independent IFN-γ synthesis. To understand how signal 3 responses functioned in dual costimulated T cells we showed that IL-2 induced IL-36R gene expression in a JAK/STAT dependent manner. These data help delineate a sequential stimulation process where IL-2 conditioning must precede IL-36 for IFN-γ synthesis. Importantly, this responsive state was transient, and functioned only in effector T cells capable of aerobic glycolysis. Specifically, as the effector T cells metabolized glucose and consumed O2, they also retained potential to respond through IL-36R. This suggests that T cells use innate receptor pathways like the IL-36R-axis when programmed for aerobic glycolysis. To explore a function for IL-36R in vivo we showed that dual costimulation therapy reduced B16 melanoma tumor growth while increasing IL-36R gene expression. In sum, cytokine therapy to eliminate tumors may target effector T cells, even outside of TCR specificity, as long as the effectors are in the correct metabolic state.

The ability of immune-based cancer therapies to elicit beneficial CD8+ CTL is limited by tolerance pathways that inactivate tumor-specific CD4 helper T cells. A strategy to bypass this problem is to engage tumor-unrelated CD4 helper T cells. Thus, CD4 T cells, regardless of their specificity per se, can boost CD8+ CTL priming so long as the cognate epitopes are linked via presentation on the same dendritic cell. Here, we assessed the therapeutic impact of engaging tumor-unrelated CD4 T cells during dual costimulation with CD134 plus CD137 that not only provide help via the above-mentioned classical linked pathway, but also provide non-linked help that facilitates CTL function in T cells not directly responding to cognate antigen. We found that engagement of tumor-unrelated CD4 helper T cells dramatically boosted the ability of dual costimulation to control the growth of established B16 melanomas. Surprisingly, this effect depended upon a CD134-dependent component that was extrinsic to the tumor-unrelated CD4 T cells, suggesting that the dual-costimulated helper cells are themselves helped by a CD134+ cell(s). Nevertheless, the delivery of therapeutic help tracked with an increased frequency of tumor-infiltrating granzyme B+ effector CD8 T cells and a reciprocal decrease in Foxp3+CD4+ cell frequency. Notably, the tumor-unrelated CD4 helper T cells also infiltrated the tumors, and their deletion several days following initial T cell priming negated their therapeutic impact. Taken together, dual costimulation programs tumor-unrelated CD4 T cells to deliver therapeutic help during both the priming and effector stages of the anti-tumor response.

No abstract

Host anti-tumor immunity can be hindered by various mechanisms present within the tumor microenvironment, including the actions of myeloid-derived suppressor cells (MDSCs). We investigated the role of the CCR2/MCP-1 pathway in MDSC-associated tumor progression in murine lung cancer models. Phenotypic profiling revealed maximal expression of CCR2 by tumor-resident MDSCs, and MCP-1 by transplanted TC1 tumor cells, respectively. Use of CCR2-knockout (CCR2-KO) mice showed dependence of tumor growth on CCR2 signaling. Tumors in CCR2-KO mice had fewer CCR2low MDSCs, CD4 T cells and Tregs than WT mice, and increased infiltration by CD8 T cells producing IFN-γ and granzyme-B. Effects were MDSC specific, since WT and CCR2-KO conventional T (Tcon) cells had comparable proliferation and production of inflammatory cytokines, and suppressive functions of WT and CCR2-KO Foxp3+ Treg cells were also similar. We used a thioglycolate-induced peritonitis model to demonstrate a role for CCR2/MCP-1 in trafficking of CCR2+ cells to an inflammatory site, and showed the ability of a CCR2 antagonist to inhibit such trafficking. Use of this CCR2 antagonist promoted anti-tumor immunity and limited tumor growth. In summary, tumor cells are the prime source of MCP-1 that promotes MDSC recruitment, and our genetic and pharmacologic data demonstrate that CCR2 targeting may be an important component of cancer immunotherapy.