Interleukin-7 (IL-7), a cytokine produced by stromal cells, is required for thymic development and peripheral homeostasis of most major subsets of T cells. We IntroductionRegulatory T (Treg) cells have a critical suppressive function for maintenance of self-tolerance and prevention of autoimmunity, 1,2 and their deficiency can predispose to gastritis, thyroiditis, diabetes and graft-versus-host disease. Initially, Treg cells were identified as a small percentage, approximately 10% to 15%, of mouse CD4 ϩ T cells that expressed CD25, the ␣ chain of IL-2R. 3,4 Treg cells also are reported to express CD45RB, 5 CD62L, 6 cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), 7-9 glucocorticoidinduced tumor necrosis factor (TNF)-like receptor (GITR), [10][11][12] CD103 (␣E␤7 integrin) 13 and forkhead box transcription factor 3 (Foxp3), an intracellular transcriptional regulator. [14][15][16] In vitro, Treg cells can block proliferative responses of both CD4 ϩ and CD8 ϩ CD25 Ϫ cells by a mechanism that remains to be clearly defined but appears to be based on cell contact and to be independent of cytokine production. [17][18][19][20][21] A very recent paper suggests Tregs suppress target cells by interleukin-2 (IL-2) deprivation and subsequent apoptosis. 22 In vivo, Treg cells suppress activation and expansion of self-reactive T cells that have escaped thymic clonal deletion. 1,3,[23][24][25] In addition to cell contact, the suppressive cytokines IL-10 and transforming growth factor (TGF)␤ have been implicated in vivo as mediators of inhibition. [26][27][28][29][30][31][32] Several studies suggest that CD4 ϩ CD25 ϩ T cells mature in the thymus as a distinct T-cell population. 5,7,10,15,29,33 High-affinity IL-2 receptors are constitutively expressed on Treg cells, and IL-2 has been implicated in the development, maintenance, and function of these cells. 34 It has been reported that IL-2 may be required for peripheral expansion and homeostasis, [35][36][37][38] and IL-2 appears to be required for Treg cell function in the periphery. 17,[39][40][41][42] Mice deficient in IL-2, IL-2R␣, or IL-2R␤ lack regulatory T cells 43,44 and develop severe autoimmune disease. 42,[45][46][47][48][49] IL-7 is a cytokine that is produced by stromal cells in lymphoid tissues and is required for development and homeostasis of most subsets of T cells. [50][51][52] The IL-7 receptor is composed of IL-7R␣ and the common cytokine receptor ␥ chain, ␥ c 53,54 . A related stromal factor, thymic stromal lymphopoietin (TSLP), also shares IL-7R␣ but additionally has a distinctive receptor subunit, TSLPR. 55,56 Naive and memory CD4 ϩ T cells require IL-7 for homeostatic survival. 57,58 In vitro it has been shown that IL-7 can, albeit less well than IL-2 or IL-4, promote the proliferation and suppressor function of CD4 ϩ CD25 ϩ cells stimulated with anti-CD3. 40 The Tr1 cell, another type of suppressor cell that acts by secreting suppressive cytokines, has been shown to respond to IL-7 in vitro. 59 It has recently been reported that Treg cells develop, s...

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...

The use of interleukin-18 (IL-18) together with IL-12 induced high levels of IFN-; in tumor-bearing mice and regression of liver tumors that was abolished in IFN-;(À/À) mice. Natural killer (NK) and NKT cells were the major producers of IFN-; in the livers of mice treated with IL-18 and/or IL-12. Liver NK cells were significantly increased by treatment with IL-18/ IL-12, whereas the degree of liver NKT cell TCR detection was diminished by this treatment. Reduction of NK cells with antiasGM1 decreased the antitumor activity of IL-18/IL-12 therapy and revealed NK cells to be an important component for tumor regression in the liver. In contrast, the antitumor effects of both IL-18 and IL-12 were further increased in CD1d (À/À) mice, which lack NKT cells. Our data, therefore, show that the antitumor activity induced in mice by IL-18/IL-12 is NK and IFN-; dependent and is able to overcome an endogenous immunosuppressive effect of NKT cells in the liver microenvironment. These results suggest that immunotherapeutic approaches that enhance NK cell function while eliminating or altering NKT cells could be effective in the treatment of cancer in the liver. (Cancer Res 2006; 66(22): 11005-12)

Drosophila peripheral nerves, structured similarly to their mammalian counterparts, comprise a layer of motor and sensory axons wrapped by an inner peripheral glia (analogous to the mammalian Schwann cell) and an outer perineurial glia (analogous to the mammalian perineurium). Growth and proliferation within mammalian peripheral nerves are increased by Ras pathway activation: loss-of-function mutations in Nf1, which encodes the Ras inhibitor neurofibromin, cause the human genetic disorder neurofibromatosis, which is characterized by formation of neurofibromas (tumors of peripheral nerves). However, the signaling pathways that control nerve growth downstream of Ras remain incompletely characterized. Here we show that expression specifically within the Drosophila peripheral glia of the constitutively active Ras V12 increases perineurial glial thickness. Using chromosomal loss-of-function mutations and transgenes encoding dominant-negative and constitutively active proteins, we show that this nonautonomous effect of Ras V12 is mediated by the Ras effector phosphatidylinositol 3-kinase (PI3K) and its downstream kinase Akt. We also show that the nonautonomous, growthpromoting effects of activated PI3K are suppressed by coexpression within the peripheral glia of FOXO ϩ (forkhead box O) a transcription factor inhibited by Akt-dependent phosphorylation. We suggest that Ras-PI3K-Akt activity in the peripheral glia promotes growth of the perineurial glia by inhibiting FOXO. In mammalian peripheral nerves, the Schwann cell releases several growth factors that affect the proliferative properties of neighbors. Some of these factors are oversecreted in Nf1 mutants. Our results raise the possibility that neurofibroma formation in individuals with neurofibromatosis might result in part from a Ras-PI3K-Akt-dependent inhibition of FOXO within Schwann cells.

Type I IFNs (IFN-α/β) are pleitropic cytokines widely used in the treatment of certain malignancies, hepatitis B and C, and multiple sclerosis. IFN resistance is a challenging clinical problem to overcome. Hence, understanding the molecular mechanism by which IFN immunotherapy ceases to be effective is of translational importance. In this study, we report that continuous IFN-α stimulation of the human Jurkat variant H123 led to resistance to type I IFN-induced apoptosis due to a loss of signal transducers and activators of transcription 2 (STAT2) expression. The apoptotic effects of IFN-α were hampered as STAT2-deficient cells were defective in activating the mitochondrial-dependent death pathway and ISGF3-mediated gene activation. Reconstitution of STAT2 restored the apoptotic effects of IFN-α as measured by the loss of mitochondrial membrane potential, cytochrome c release from mitochondria, caspase activation, and ultimately cell death. Nuclear localization of STAT2 was a critical event as retention of tyrosine-phosphorylated STAT2 in the cytosol was not sufficient to activate apoptosis. Furthermore, silencing STAT2 gene expression in Saos2 and A375S.2 tumor cell lines significantly reduced the apoptotic capacity of IFN-α. Altogether, we show that STAT2 is a critical mediator in the activation of type I IFN-induced apoptosis. More importantly, defects in the expression or nuclear localization of STAT2 could lessen the efficacy of type I IFN immunotherapy. Mol Cancer Res; 8(1); 80-92. ©2010 AACR.