Aging strongly promotes inflammation responses, which may predispose individuals after cancer therapies to lethal system toxicities and pathology that can be partially prevented by TNF blockade.

Protective cell-mediated immune responses in cancer are critically dependent on T-helper type 1 (T(H)1) cytokines such as interferon-gamma (IFN-gamma). We have previously shown that the combination of CD40 stimulation and interleukin-2 (IL-2) leads to synergistic antitumor responses in several models of advanced metastatic disease. We now report that after this treatment and other immunotherapy regimens, the CD4+ T-cell population, in contrast to CD8+ T cells, did not significantly increase but rather exhibited a substantial level of apoptosis that was dependent on IFN-gamma. Mice immunized with tumor cells and treated with an immunotherapy regimen that was initially protective were later unable to mount effective memory responses compared with immunized mice not receiving immunotherapy. Immunotherapy given to tumor-bearing Ifngr-/- mice resulted in restoration of secondary responses. Thus, although immunotherapeutic regimens inducing strong IFN-gamma responses can lead to successful early antitumor efficacy, they may also impair the development of durable antitumor responses.

Natural killer (NK) cells are powerful effector cells that can be directed to eliminate tumor cells through tumor-targeted monoclonal antibodies (mAbs). Some tumor-targeted mAbs have been successfully applied in the clinic and are included in the standard of care for certain malignancies. Strategies to augment the antitumor response by NK cells have led to an increased understanding of how to improve their effector responses. Next-generation reagents, such as molecularly modified mAbs and mAb-cytokine fusion proteins (immunocytokines, ICs) designed to augment NK-mediated killing, are showing promise in preclinical and some clinical settings. Continued research into the antitumor effects induced by NK cells and tumor-targeted mAbs suggests that additional intrinsic and extrinsic factors may influence the antitumor response. Therefore more research is needed that focuses on evaluating which NK cell and tumor criteria are best predictive of a clinical response and which combination immunotherapy regimens to pursue for distinct clinical settings.

Memory T cells exhibit tremendous antigen specificity within the immune system and accumulate with age. Our studies reveal an antigen-independent expansion of memory, but not naive, CD8 ؉ T cells after several immunotherapeutic regimens for cancer resulting in a distinctive phenotype. Signaling through T-cell receptors (TCRs) or CD3 in both mouse and human memory CD8 ؉ T cells markedly up-regulated programmed death-1 (PD-1) and CD25 (IL-2 receptor ␣ chain), and led IntroductionMemory T cells represent an arm of the adaptive immune system that are long-lived, and capable of rapid antigen-specific responses. Memory T cells have been shown to have functional advantages more than naive T cells, as they develop more rapidly into cytolytic effector cells and produce greater amounts of cytokines after antigenic stimulation. 1 Although T cells classically require T-cell receptor (TCR) engagement and proper costimulation for complete activation and proliferation, memory T cells have also been observed to proliferate in response to various cytokines during viral infections. [2][3][4][5][6] These "bystander cells" proliferate and gain effector functions in response to the cytokine milieu produced during the course of viral and bacterial infections in mice and humans. [7][8][9][10] Cytokines alone can induce this as a single dose of recombinant type-I interferon (IFN) resulting in a transient increase in the proliferation of CD8 ϩ , CD62L ϩ CD44 high memory T cells, which was independent of coligation of the TCRs. 11 Such proliferation was not induced by the direct effects of type-I IFNs on CD8 ϩ T cells, but was because of type-I IFN-driven production of secondary cytokines such as 12 Effector and memory CD8 ϩ T cells express elevated levels of the receptors for IL-12 and IL-18, and secrete IFN-␥ in response to stimulation with both cytokines, 13 which suggests that other cytokine pathways can also induce their expansion. Similar to the secondary cytokine-driven proliferation observed after type-I IFN stimulation, IL-2, and toll-like receptor (TLR) agonists, that is, CpG and Poly:IC have also been described as having the capacity to induce bystander proliferation of CD8 ϩ CD44 high T cells. 12,14,15 The extent of antigen-specific proliferation versus bystander expansion has been the subject of considerable debate and may be contingent on the pathogen model and tissue examined. 9,13,16 Cancer therapies that target the stimulation of the immune system via agonist antibodies, cytokine-based modalities, or TLR agonists have been shown to result in potent CD8 ϩ T cell-mediated antitumor effects. 17,18 We have previously shown that a combination immunotherapy consisting of an agonist CD40 antibody and IL-2 results in synergistic antitumor effects. 19 Treatment of mice with other cytokine or TLR agonist combinations, such as CpGs and IL-15 or IL-2 and IL-12, also resulted in marked antitumor effects. 18 In all of these models, the antitumor effects were associated with rapid, extensive CD8 ϩ T-cell expansion. The antitumor effec...

Many strategies for cancer treatment use combinations of immunotherapeutic agents for enhanced anti-tumor responses. However, these approaches are often complicated by a need to overcome tumor-induced immune suppression in the tumor microenvironment. In this regard, T regulatory (Treg) cells and myeloid-derived suppressor cells (MDSC) have been identified as functional suppressor cells within tumors (1, 2). The most effective immunotherapeutic regimens are likely to consist of agents that restructure, within the tumor microenvironment, the composition of tumor-infiltrating leukocytes away from these inhibitory elements in favor of effector cells, such as NK cells and CD8 ϩ T cells.Chemokine expression can regulate the polarization of immune responses (3). For example, CXCR3 and CCR5 are preferentially expressed on Th1 T cells and M1 macrophages and their respective ligands are associated with enhanced cellmediated immune responses (3-5) and favorable prognosis in human RCC (5, 6). Another chemokine, monocyte chemoattractant protein (MCP)-1 activates macrophages for enhanced anti-tumor activities (7), however MCP-1 expression is also associated with the recruitment of mononuclear cells capable of producing tumor promoting factors (8, 9), as well as MDSC that contribute to tumor progression through the inhibition of effector cell functions (8, 10).We reported previously that IL-2 and agonistic antibody to CD40 (␣CD40) synergize for the regression of metastatic tumors in mice (11). Although we identified CD8 ϩ T cells and host IFN␥ expression as critical components of this therapeutic approach (11), the specific mechanisms underlying the IL-2/␣CD40 synergistic anti-tumor responses within the microenvironment remain unclear. We demonstrate in a murine model of metastatic renal cancer that ␣CD40 may be limited by its dependency upon MCP-1 and an inability to remove Tregs and MDSC specifically from within the tumor microenvironment, allowing for eventual tumor progression. In contrast, synergistic anti-tumor responses and protection achieved by IL-2/␣CD40 are associated with the expression of Th1 chemokines that are associated with favorable prognosis in RCC (5, 6), an augmentation of effector leukocytes and concomitant removal of suppressive cells specifically within the tumor microenvironment. Results CCR2 Expression Is Required for ␣CD40, but Not IL-2/␣CD40 MediatedAnti-Tumor Responses. Our previous study showed that IL-2/ ␣CD40 exhibited strong synergy for treatment of established metastatic tumors in mice, as compared to IL-2 or ␣CD40 as single agents (11). Furthermore, we found that ␣CD40 treatment of Renca-bearing mice induced significant reduction in tumors in association with high levels of systemic MCP-1 levels, suggesting a possible role for MCP-1 in leukocyte recruitment into tumors and CD40-dependent anti-tumor effects (12). To determine the relative contribution of MCP-1 to the ␣CD40-and IL-2/␣CD40-mediated anti-tumor responses, we compared tumor outcomes in treated WT and mice deficient in CCR2, the rec...