No abstract

The mechanisms by which tumor microenvironments modulate nucleic acid–mediated innate immunity remain unknown. Here we identify the receptor TIM-3 as key in circumventing the stimulatory effects of nucleic acids in tumor immunity. Tumor-associated dendritic cells (DCs) in mouse tumors and patients with cancer had high expression of TIM-3. DC-derived TIM-3 suppressed innate immune responses through the recognition of nucleic acids by Toll-like receptors and cytosolic sensors via a galectin-9-independent mechanism. In contrast, TIM-3 interacted with the alarmin HMGB1 to interfere with the recruitment of nucleic acids into DC endosomes and attenuated the therapeutic efficacy of DNA vaccination and chemotherapy by diminishing the immunogenicity of nucleic acids released from dying tumor cells. Our findings define a mechanism whereby tumor microenvironments suppress antitumor immunity mediated by nucleic acids.

CD4+ T helper 1 (TH1) cells are important mediators of inflammation and are regulated by numerous pathways, including the negative immune receptor Tim-3. We found that Tim-3 is constitutively expressed on cells of the innate immune system in both mice and humans, and that it can synergize with Toll-like receptors. Moreover, an antibody agonist of Tim-3 acted as an adjuvant during induced immune responses, and Tim-3 ligation induced distinct signaling events in T cells and dendritic cells; the latter finding could explain the apparent divergent functions of Tim-3 in these cell types. Thus, by virtue of differential expression on innate versus adaptive immune cells, Tim-3 can either promote or terminate TH1 immunity and may be able to influence a range of inflammatory conditions.

Tumor-associated immune suppression can lead to defective T cell-mediated antitumor immunity. Here, we identified a unique phenotype of exhausted T cells in mice with advanced acute myelogenous leukemia (AML). This phenotype is characterized by the coexpression of Tim-3 and PD-1 on CD8 ؉ T cells in the liver, the major first site of AML metastases. PD-1 and Tim-3 coexpression increased during AML progression. PD-1 ؉ Tim-3 ؉ CD8 ؉ T cells were deficient in their ability to produce IFN-␥, TNF-␣, and IL-2 in response to PD-1 ligand (PDL1) and Tim-3 ligand (galectin-9) expressing AML cells. PD-1 knockout (KO), which were partially resistant to AML challenge, up-regulated Tim-3 during AML progression and such Tim-3 ؉ PD-1-KO CD8 ؉ T cells had reduced cytokine production. Galectin-9 KO mice were more resistant to AML, which was associated with reduced T-regulatory cell accumulation and a modest induction of PD-1 and Tim-3 expression on CD8 ؉ T cells. Whereas blocking the PD-1/ PDL1 or Tim-3/galectin-9 pathway alone was insufficient to rescue mice from AML lethality, an additive effect was seen in reducing-albeit not eliminating-both tumor burden and lethality when both pathways were blocked. Therefore, combined PD-1/PDL1 and Tim-3/galectin-9 blockade may be beneficial in preventing CD8 ؉ T-cell exhaustion in patients with hematologic malignancies such as advanced AML. (Blood. 2011;117(17):4501-4510) Introduction T-cell exhaustion, a state of T-cell dysfunction characterized by diminished cytokine production, impaired killing, and hypoproliferation, was first characterized in the settings of chronic lymphocytic choriomeningitis virus (LCMV) infection. 1,2-5 Since its discovery, the process of T-cell exhaustion has been of intense interest and has been the subject of study in viral infections such as hepatitis C virus 2,6 and HIV, 3,7 as well as in tumor models. 8,9,10,11 Cell-surface antigen determinants such as program death-1 (PD-1), CTLA-4, and, in some instances, CD28 (eg, hepatitis C viral infection) can be used to identify antigen-specific T cells that are at an exhaustion stage. 4 T-cell immunoglobulin and mucin domain-containing protein 3 (Tim-3) is a type I membrane glycoprotein and its expression can be found on terminally differentiated Th1 cells and innate immune cells. [12][13][14] is its only confirmed Tim-3 ligand to date, 15,16 although it is known that Tim-3 can also bind to certain carbohydrate moieties. 17 Ligation of Tim-3 on T cells and gal-9 inhibits Th1 responses and plays an important role in infection, autoimmunity, peripheral tolerance, and inflammation. 14,[18][19][20][21] In addition to its negative regulatory role in dampening the immune system, a recent report showed a synergistic effect of Tim-3 signaling and lipopolysaccharide in producing proinflammatory cytokines by naive dendritic cells (DCs) and monocytes, 22 indicating a dual role of the Tim-3 signaling pathway at a different phase of immune responses.Studies have demonstrated a strong correlation between PD-1 and Tim-3 coexpressi...

No abstract

Galectin-3 is a β-galactoside-binding protein implicated in diverse biological processes. We found that galectin-3 induced human monocyte migration in vitro in a dose-dependent manner, and it was chemotactic at high concentrations (1.0 μM) but chemokinetic at low concentrations (10–100 nM). Galectin-3-induced monocyte migration was inhibited by its specific mAb and was blocked by lactose and a C-terminal domain fragment of the protein, indicating that both the N-terminal and C-terminal domains of galectin-3 are involved in this activity. Pertussis toxin (PTX) almost completely blocked monocyte migration induced by high concentrations of galectin-3. Galectin-3 caused a Ca2+ influx in monocytes at high, but not low, concentrations, and both lactose and PTX inhibited this response. There was no cross-desensitization between galectin-3 and any of the monocyte-reactive chemokines examined, including monocyte chemotactic protein-1, macrophage inflammatory protein-1α, and stromal cell-derived factor-1α. Cultured human macrophages and alveolar macrophages also migrated toward galectin-3, but not monocyte chemotactic protein-1. Finally, galectin-3 was found to cause monocyte accumulation in vivo in mouse air pouches. These results indicate that galectin-3 is a novel chemoattractant for monocytes and macrophages and suggest that the effect is mediated at least in part through a PTX-sensitive (G protein-coupled) pathway.

NK-cell function is regulated by the integration of signals received from activating and inhibitory receptors. Here we show that a novel immune receptor, T-cell Ig and mucin-containing domain-3 (Tim-3), is expressed on resting human NK cells and is up-regulated on activation. The NK92 NK-cell line engineered to overexpress Tim-3 showed a marked increase in IFN-␥ production in the presence of soluble rhGal-9 or Raji tumor cells engineered to express Gal-9. The Tim-3 ؉ population of low-dose IL-12/IL-18-activated primary NK cells significantly increased IFN-␥ production in response to soluble rhGal-9, Gal-9 presented by cell lines, and primary acute myelogenous leukemia (AML) targets that endogenously express Gal-9. This effect is highly specific as Tim-3 Ab blockade significantly decreased IFN-␥ production, and Tim-3 cross-linking induced ERK activation and degradation of IB␣. Exposure to Gal-9-expressing target cells had little effect on CD107a degranulation. Reconstituted NK cells obtained from patients after hematopoietic cell transplantation had diminished expression of Tim-3 compared with paired donors. This observation correlates with the known IFN-␥ defect seen early posttransplantation. In conclusion, we show that Tim-3 functions as a human NK-cell coreceptor to enhance IFN-␥ production, which has important implications for control of infectious disease and cancer. (Blood. 2012;119(13): 3064-3072) IntroductionHuman NK cells are lymphocytes that develop from hematopoietic progenitor cells in the BM and secondary lymphoid tissues. 1 Peripheral blood (PB) NK cells are phenotypically defined as expressing the surface receptor CD56 (neural cell adhesion molecule [NCAM]) and lacking expression of CD3. 2 They mediate their function through the exocytosis of lytic granules that contain perforin and granzymes, 3 the expression of death receptor ligands, 4 the expression of FcR␥III, which mediates Ab-dependent cellmediated cytotoxicity, 5 and the secretion of cytokines and chemokines. 6 As a result, NK cells take part in both the innate and adaptive immune systems and play important roles in the control of viral infections, pregnancy, tumor immunosurveillance, and hematopoietic cell transplantation (HCT). 7,8 The ability of NK cells to differentiate normal healthy cells (self) from virally infected or malignantly transformed cells (nonself) is regulated by a sophisticated repertoire of cell-surface receptors that control their activation, proliferation, and effector functions. [9][10][11] Recently, a novel receptor, T-cell Ig and mucincontaining domain-3 (Tim-3), has been described to have various roles in immune regulation and is highly expressed on NK cells in mice and humans. [12][13][14][15][16][17] Tim-3 is a type I membrane glycoprotein that was first identified as a cell marker of terminally differentiated CD4 ϩ Th1 cells. 18 Galectin-9 (Gal-9), a 40-kDa S-type ␤-galactoside binding lectin, is a known ligand for Tim-3 and is highly expressed in tissues of the immune system, such as the BM, lymph nodes, thymu...

Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC)is the third most frequent virus-associated human malignancy. How this tumor escapes immune recognition despite the expression of several viral antigens has remained poorly understood. Our previous in vitro studies have shown that NPC cells release exosomes containing high amounts of galectin-9, a ligand of the membrane receptor Tim-3, which is able to induce apoptosis in mature Th1 lymphocytes.Here, we sought to determine whether galectin-9-carrying exosomes were produced in NPC patients and whether such exosomes might play a role in the immune evasion of NPC cells. We report that galectin-9-containing exosomes are selectively detected in plasma samples from NPC patients and mice xenografted with NPC tumors. The incorporation into exosomes protects galectin-9 against proteolytic cleavage but retains its Tim-3-binding capacity. Importantly, NPC exosomes induce massive apoptosis in EBVspecific CD4 ؉ cells used as a model of target T cells. This effect is inhibited by both anti-Tim-3 and antigalectin-9 blocking antibodies. These results indicate that blocking galectin-9/Tim-3 interaction in vivo might alleviate the Th1-suppressive effect of NPC exosomes and sustain antitumoral T-cell responses and thereby improve clinical efficacy of immunotherapeutic approaches against NPC. (Blood. 2009;113:1957-1966 IntroductionNasopharyngeal carcinoma (NPC) is an Epstein-Barr virus (EBV)-associated malignancy and the third most frequent virus-associated human malignancy after hepatocarcinomas and cervix carcinomas. Each year, approximately 80 000 new cases are diagnosed worldwide. The geographic distribution of NPC is not uniform. It is relatively rare in European and North American countries. Very high incidence foci are found in South China, especially in Guandong and Guangxi provinces (25-40 per 100 000 per year). 1 Areas of intermediate incidence (approximately 3-8 per 100 000 per year) include a large number of developing or emerging countries, especially in North and Central Africa (Tunisia, Algeria, Morocco, Somalia, and Kenya) and in Southeast Asia (Philippines, Vietnam, Indonesia).NPC is an epithelial malignancy with a complex etiology involving viral, environmental, and hereditary factors. Except for some very rare cases of atypical highly differentiated NPC occurring in Western countries, the intact EBV genome is always contained in the nuclei of all malignant cells. 2,3 Many of the approximately 80 EBV genes are silent, but several viral RNAs and proteins are consistently expressed in NPC and contribute to the malignant phenotype. 4 NPC oncogenesis also requires a variable assortment of cellular genetic or epigenetic alterations. 5 Another important biologic feature of NPC is the presence of a massive lymphoid infiltrate in the primary tumor. This infiltrate contains mostly T lymphocytes and a minority of B cells, monocytes, dendritic cells, and eosinophils. The abundant production by malignant NPC cells of inflammatory cytokines, including interle...