HAVCR2

Gene Summary

Gene:HAVCR2; hepatitis A virus cellular receptor 2
Aliases: TIM3, CD366, KIM-3, TIMD3, Tim-3, TIMD-3, HAVcr-2
Location:5q33.3
Summary:The protein encoded by this gene belongs to the immunoglobulin superfamily, and TIM family of proteins. CD4-positive T helper lymphocytes can be divided into types 1 (Th1) and 2 (Th2) on the basis of their cytokine secretion patterns. Th1 cells are involved in cell-mediated immunity to intracellular pathogens and delayed-type hypersensitivity reactions, whereas, Th2 cells are involved in the control of extracellular helminthic infections and the promotion of atopic and allergic diseases. This protein is a Th1-specific cell surface protein that regulates macrophage activation, and inhibits Th1-mediated auto- and alloimmune responses, and promotes immunological tolerance. [provided by RefSeq, Sep 2011]
Databases:VEGA, OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:hepatitis A virus cellular receptor 2
Source:NCBIAccessed: 16 March, 2017

Ontology:

What does this gene/protein do?
HAVCR2 is implicated in:
- integral to membrane
Data from Gene Ontology via CGAP

Cancer Overview

Research Indicators

Publications Per Year (1992-2017)
Graph generated 16 March 2017 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Acute Myeloid Leukaemia
  • TNF
  • Galectins
  • fms-Like Tyrosine Kinase 3
  • Up-Regulation
  • CD8-Positive T-Lymphocytes
  • Membrane Proteins
  • T-Lymphocytes
  • Liver Cancer
  • Receptors, Virus
  • Genetic Predisposition
  • Stomach Cancer
  • Cancer Gene Expression Regulation
  • Asian Continental Ancestry Group
  • Genotype
  • Hepatocellular Carcinoma
  • Biomarkers, Tumor
  • Liver Cirrhosis
  • Lymphocyte Activation
  • Risk Factors
  • Transfection
  • VEGFA
  • CD4-Positive T-Lymphocytes
  • Recombinant Proteins
  • Case-Control Studies
  • Single Nucleotide Polymorphism
  • Interferon-gamma
  • Th1 Cells
  • Programmed Cell Death 1 Receptor
  • Cancer RNA
  • Chromosome 5
  • Transforming Growth Factor beta
  • Hepatitis B, Chronic
  • Genetic Association Studies
  • Flow Cytometry
  • Pancreatic Cancer
  • Hepatitis A Virus Cellular Receptor 2
  • Cell Proliferation
  • Melanoma
  • Young Adult
  • Haplotypes
Tag cloud generated 16 March, 2017 using data from PubMed, MeSH and CancerIndex

Specific Cancers (5)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: HAVCR2 (cancer-related)

Li Z, Li N, Li F, et al.
Immune checkpoint proteins PD-1 and TIM-3 are both highly expressed in liver tissues and correlate with their gene polymorphisms in patients with HBV-related hepatocellular carcinoma.
Medicine (Baltimore). 2016; 95(52):e5749 [PubMed] Free Access to Full Article Related Publications
Immune checkpoint proteins programmed death-1 (PD-1) and T-cell immunoglobulin domain and mucin domain containing molecule-3 (TIM-3) expression and their gene polymorphisms have separately been shown to be associated with hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC). This study simultaneously examined PD-1 and TIM-3 expression in liver tissues and PD1 and TIM3 polymorphisms and analyzed their correlations in 171 patients with HBV-related HCC and 34 patients with HBV-related cirrhosis.PD-1 and TIM-3 expression in liver tissues were examined by immunohistochemistry and the genotypes of PD1 rs10204525 and TIM3 rs10053538 polymorphisms were determined using genomic DNA extracted from peripheral blood as template.Both PD-1 and TIM-3 expressions in liver infiltrating lymphocytes of HCC tumor tissues were significantly higher than those in tumor adjacent tissues or cirrhotic tissues. The elevated PD-1 and TIM-3 expressions were significantly associated with higher tumor grades. The levels between PD-1 and TIM-3 expression in tumor tissues and tumor adjacent tissues had a significant positive intercorrelation. The expressions of PD-1 and TIM-3 in tumor tissues, tumor adjacent tissues, and cirrhotic tissues were significantly associated with PD1 and TIM3 polymorphisms, with genotype AA of PD1 rs10204525 and genotypes GT+TT of TIM3 rs10053538 being associated with significantly increased PD-1 and TIM-3 expression, respectively.These findings support the potential to improve the efficiency of immune checkpoint-targeted therapy and reduce resistance to the therapy by blocking both PD-1 and TIM-3 and suggest the potential to apply the genotype determination of PD1 rs10204525 and TIM3 rs10053538 as biomarkers of immune checkpoint-directed therapies.

Wang Z, Yin N, Zhang Z, et al.
Upregulation of T-cell Immunoglobulin and Mucin-Domain Containing-3 (Tim-3) in Monocytes/Macrophages Associates with Gastric Cancer Progression.
Immunol Invest. 2017; 46(2):134-148 [PubMed] Related Publications
T-cell immunoglobulin and mucin-domain containing-3 (Tim-3) is an important immune regulatory molecule in cancer immune system. However, expression and function of Tim-3 in monocytes/macrophages in cancer progression mainly remain unclear. In this study, we analyzed Tim-3 levels in peripheral blood mononuclear cells (PBMCs) from 62 gastric cancer patients and 45 healthy controls using flow cytometry and then associated Tim-3 levels with clinical pathological data from patients. We found Tim-3 level was significantly upregulated in monocytes from gastric cancer patients compared with those from healthy controls, and that upregulated Tim-3 levels associated with depth of tumor invasion and tumor lymph node metastasis and advanced clinical stages of gastric cancer patients. Furthermore, tumor-bearing mouse experiments revealed that Tim-3 level on monocytes/macrophages associated with xenograft formation and growth. In addition, culture of monocytes from healthy controls with gastric cancer cell-conditioned medium upregulated Tim-3 expression, but IL-10, TNF-α, IFN-γ, or GM-CSF treatment or T-bet, Eomes, and T-bet/Eomes double gene knockout did not affect Tim-3 levels in blood monocytes/macrophages from human or mouse, respectively. Gal-9/Tim-3 signal was able to significantly stimulate monocyte to secrete IL-6, IL-8, and IL-10, but not IL-1β, IL-12p70, or TNF-α in presence of LPS. In conclusion, our study demonstrated that Tim-3 expressed by monocyte/macrophages might be an important mechanism in gastric cancer progression.

Feng ZM, Guo SM
Tim-3 facilitates osteosarcoma proliferation and metastasis through the NF-κB pathway and epithelial-mesenchymal transition.
Genet Mol Res. 2016; 15(3) [PubMed] Related Publications
The aim of this study was to investigate the expression of T-cell immunoglobulin mucin domain molecule-3 (Tim-3) in osteosarcoma tissues, and analyze its effect on cell proliferation and metastasis in an osteosarcoma cell line. Tim-3 mRNA and protein expression in osteosarcoma tissue was detected by reverse transcriptase-polymerase chain reaction and immunohistochemistry, respectively. Additionally, the cell viability, apoptosis rate, and invasive ability of the osteosarcoma cell line MG-63 were tested using the methyl thiazolyl tetrazolium assay, Annexin V-propidium iodide flow cytometry, and a Transwell assay, respectively, following Tim-3 interference using small interfering RNA (siRNA). We also analyzed the expression of Snail, E-cadherin, vimentin, and nuclear factor (NF)-kB in the cells by western blot. We observed that Tim-3 mRNA and protein was significantly overexpressed in osteosarcoma tissues, compared to the adjacent normal tissue (P < 0.01). Moreover, MG-63 cells transfected with the Tim-3 siRNA presented lower cell viability, a greater number of apoptotic cells, and decreased invasive ability (P < 0.01), compared to control cells. Additionally, we observed a decrease in Snail and vimentin expression, an increase in the E-cadherin level, and an increase in NF-kB p65 phosphorylation (P < 0.01) in Tim-3 siRNA-transfected MG-63 cells. Based on these results, we concluded that Tim-3 is highly expressed in osteosarcoma tissue. Moreover, we speculated that interfering in Tim-3 expression could significantly suppress osteosarcoma cell (MG-63) proliferation and metastasis via the NF-kB/Snail signaling pathway and epithelial-mesenchymal transition.

Fooladinezhad H, Khanahmad H, Ganjalikhani-Hakemi M, Doosti A
Negative regulation of TIM-3 expression in AML cell line (HL-60) using miR-330-5p.
Br J Biomed Sci. 2016; 73(3):129-133 [PubMed] Related Publications
BACKGROUND: Uncontrolled proliferation and accumulation of leukaemic stem cells (LSCs) in bone marrow leads to acute myeloma leukaemia (AML). T cell immunoglobulin and mucine domain (TIM)-3 is a specific surface marker for LSCs and is highly expressed on LSCs compared with normal bone marrow cells, haematopoietic stem cells. Studies have indicated that microRNAs can affect AML progression through targeting different genes expressions like TIM-3. So, based on bioinformatics assessments, we predicted that miR-330-5p may highly inhibit TIM-3 expression. The purpose of the present study was to prove the silencing effect of miR-330-5p on TIM-3 gene expression in AML cell line (HL-60) in vitro.
METHODS: HL-60 cells were cultured in RPMI 1640 supplied with 10% FBS. TIM-3 expression was induced in the cells using phorbol myristate acetate (PMA). The cells were transfected with miR-330-5p and then, the gene and protein expression of TIM-3 were measured using q-RT-PCR and flow-cytometry methods, respectively.
RESULTS: The results of our bioinformatics surveys revealed that miR-330-5p has high predicted ability to silence TIM-3 gene expression. Accordingly, our experiments confirmed that miR-330-5p is able to strongly silence TIM-3 expression (98.15% silencing) in HL-60 cell line (p = 0.0001).
CONCLUSION: According to our results, miR-330-5p has a strong inhibitory effect on TIM-3 expression in AML cell line. Thus, the bioinformatics prediction of Mirwalk and Target Scan softwares for silencing effect of miR-330-5p on TIM-3 is confirmed.

Antonia SJ, Vansteenkiste JF, Moon E
Immunotherapy: Beyond Anti-PD-1 and Anti-PD-L1 Therapies.
Am Soc Clin Oncol Educ Book. 2016; 35:e450-8 [PubMed] Related Publications
Advanced-stage non-small cell lung cancer (NSCLC) and small cell lung cancer are cancers in which chemotherapy produces a survival benefit, although it is small. We now know that anti-PD-1/PD-L1 has substantial clinical activity in both of these diseases, with an overall response rate (ORR) of 15%-20%. These responses are frequently rapid and durable, increase median overall survival (OS) compared with chemotherapy, and produce long-term survivors. Despite these very significant results, many patients do not benefit from anti-PD-1/PD-L1. This is because of the potential for malignancies to co-opt myriad immunosuppressive mechanisms other than aberrant expression of PD-L1. Conceptually, these can be divided into three categories. First, for some patients there is likely a failure to generate sufficient functional tumor antigen-specific T cells. Second, for others, tumor antigen-specific T cells may be generated but fail to enter into the tumor parenchyma. Finally, there are a large number of immunosuppressive mechanisms that have the potential to be operational within the tumor microenvironment: surface membrane immune checkpoint proteins PD-1, CTLA-4, LAG3, TIM3, BTLA, and adenosine A2AR; soluble factors and metabolic alterations interleukin (IL)-10, transforming growth factor (TGF)-β, adenosine, IDO, and arginase; and inhibitory cells, cancer-associated fibroblasts (CAFs), regulatory T cells, myeloid-derived suppressor cells (MDSCs), and tumor-associated macrophages. In this article, we discuss three strategies to generate more tumor-reactive T cells for patients: anti-CTLA-4, therapeutic tumor vaccination, and adoptive cellular therapy, with T cells redirected to tumor antigens using T-cell receptor (TCR) or chimeric antigen receptor (CAR) gene modification. We also review some of the various strategies in development to thwart tumor microenvironment immunosuppressive mechanisms. Strategies to drive more T cells into tumors remain a significant challenge.

Poh SL, Linn YC
Immune checkpoint inhibitors enhance cytotoxicity of cytokine-induced killer cells against human myeloid leukaemic blasts.
Cancer Immunol Immunother. 2016; 65(5):525-36 [PubMed] Related Publications
We studied whether blockade of inhibitory receptors on cytokine-induced killer (CIK) cells by immune checkpoint inhibitors could increase its anti-tumour potency against haematological malignancies. CIK cultures were generated from seven normal donors and nine patients with acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL) or multiple myeloma (MM). The inhibitory receptors B and T lymphocyte attenuator, CD200 receptor, lymphocyte activation gene-3 (LAG-3) and T cell immunoglobulin and mucin-domain-containing-3 (TIM-3) were present at variable percentages in most CIK cultures, while cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death-1 (PD-1) and killer cell immunoglobulin-like receptors (KIR2DL1/2/3) were expressed at low level in most cultures. Without blockade, myeloid leukaemia cells were susceptible to autologous and allogeneic CIK-mediated cytotoxicity. Blockade of KIR, LAG-3, PD-1 and TIM-3 but not CTLA-4 resulted in remarkable increase in killing against these targets, even in those with poor baseline cytotoxicity. ALL and MM targets were resistant to CIK-mediated cytotoxicity, and blockade of receptors did not increase cytotoxicity to a meaningful extent. Combination of inhibitors against two receptors did not further increase cytotoxicity. Interestingly, potentiation of CIK killing by blocking antibodies was not predicted by expression of receptors on CIK and their respective ligands on the targets. Compared to un-activated T and NK cells, blockade potentiated the cytotoxicity of CIK cells to a greater degree and at a lower E:T ratio, but without significant increase in cytotoxicity against normal white cell. Our findings provide the basis for clinical trial combining autologous CIK cells with checkpoint inhibitors for patients with AML.

Kikushige Y, Miyamoto T
Identification of TIM-3 as a Leukemic Stem Cell Surface Molecule in Primary Acute Myeloid Leukemia.
Oncology. 2015; 89 Suppl 1:28-32 [PubMed] Related Publications
Acute myeloid leukemia (AML) originates from self-renewing leukemic stem cells (LSCs), an ultimate therapeutic target in AML. Eradication of LSCs should be a critical and efficient therapeutic approach for the cure of AML. T-cell immunoglobulin mucin-3 (TIM-3) is expressed in most types of AML LSCs, but not in normal hematopoietic stem cells (HSCs); therefore, TIM-3 would be one of the promising therapeutic targets to specifically kill AML LSCs, sparing normal HSCs. In xenograft models reconstituted with human AML LSCs or human normal HSCs, an anti-human TIM-3 mouse antibody with cytotoxic activities exerts a potent anti-leukemic effect by targeting AML LSCs but does not affect normal human hematopoiesis in vivo. Here, we would like to introduce the recent studies on TIM-3 in normal and malignant hematopoiesis.

Xu L, Huang Y, Tan L, et al.
Increased Tim-3 expression in peripheral NK cells predicts a poorer prognosis and Tim-3 blockade improves NK cell-mediated cytotoxicity in human lung adenocarcinoma.
Int Immunopharmacol. 2015; 29(2):635-41 [PubMed] Related Publications
T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) has been shown to play an important role in mediating NK-cell function in human diseases. However, the relationship between Tim-3 expression in natural killer (NK) cells and human lung adenocarcinoma remains unclear. We therefore investigated the expression of Tim-3 in NK cells and explored the effect of Tim-3 blockade on NK cell-mediated activity in human lung adenocarcinoma. Upregulated expression of Tim-3 on CD3-CD56+ cells (P<0.05) and CD3-CD56(dim) cells (P<0.05) of patients with lung adenocarcinoma was detected by flow cytometry. Moreover, Tim-3 expression in CD3-CD56+ NK cells was higher in patients with lung adenocarcinoma with lymph node metastasis (LNM) (P<0.05) or with tumor stage T3-T4 (P<0.05). Tim-3 expression in CD56(dim) NK-cell subset was higher in patients with tumor size ≥3cm (P<0.05), or LNM (P<0.05) or with tumor stage T3-T4 (P<0.05). Further analysis showed that higher expressions of Tim-3 on both CD3-CD56+ NK cells and CD56(dim) NK-cell subset were independently correlated with shorter overall survival of patients with lung adenocarcinoma (log-rank test, P=0.0418, 0.0406, respectively). Importantly, blockade of Tim-3 signaling with anti-Tim-3 antibodies resulted in the increased cytotoxicity and IFN-γ production of peripheral NK cells from patients with lung adenocarcinoma. Our data indicate that Tim-3 expression in NK cells can function as a prognostic biomarker in human lung adenocarcinoma and support that Tim-3 could be a new target for an immunotherapeutic strategy.

Gonçalves Silva I, Gibbs BF, Bardelli M, et al.
Differential expression and biochemical activity of the immune receptor Tim-3 in healthy and malignant human myeloid cells.
Oncotarget. 2015; 6(32):33823-33 [PubMed] Free Access to Full Article Related Publications
The T cell immunoglobulin and mucin domain 3 (Tim-3) is a plasma membrane-associated receptor which is involved in a variety of biological responses in human immune cells. It is highly expressed in most acute myeloid leukaemia (AML) cells and therefore may serve as a possible target for AML therapy. However, its biochemical activities in primary human AML cells remain unclear. We therefore analysed the total expression and surface presence of the Tim-3 receptor in primary human AML blasts and healthy primary human leukocytes isolated from human blood. We found that Tim-3 expression was significantly higher in primary AML cells compared to primary healthy leukocytes. Tim-3 receptor molecules were distributed largely on the surface of primary AML cells, whereas in healthy leukocytes Tim-3 protein was mainly expressed intracellularly. In primary human AML blasts, both Tim-3 agonistic antibody and galectin-9 (a Tim-3 natural ligand) significantly upregulated mTOR pathway activity. This was in line with increased accumulation of hypoxia-inducible factor 1 alpha (HIF-1α) and secretion of VEGF and TNF-α. Similar results were obtained in primary human healthy leukocytes. Importantly, in both types of primary cells, Tim-3-mediated effects were compared with those induced by lipopolysaccharide (LPS) and stem cell factor (SCF). Tim-3 induced comparatively moderate responses in both AML cells and healthy leukocytes. However, Tim-3, like LPS, mediated the release of both TNF-α and VEGF, while SCF induced mostly VEGF secretion and did not upregulate TNF-α release.

Zhou E, Huang Q, Wang J, et al.
Up-regulation of Tim-3 is associated with poor prognosis of patients with colon cancer.
Int J Clin Exp Pathol. 2015; 8(7):8018-27 [PubMed] Free Access to Full Article Related Publications
Tim-3 (T cell immunoglobulin and mucin domain 3), belonging to the member of the novel Tim family, has been confirmed that it plays a critical negative role in regulating the immune responses against viral infection and carcinoma. Recently, it has also been reported that the over-expression of Tim-3 is associated with poor prognosis in solid tumors. However, the role of Tim-3 in colorectal cancer remains largely unknown. In the current study, we aim to investigate the expression of Tim-3 in colorectal carcinoma and discuss the relationship between Tim-3 expression and colon cancer prognosis, thus speculating the possible role of Tim-3 in colon cancer progression. Colon cancer tissues and paired normal tissue were obtained from 201 patients with colon cancer for preparation of tissue microarray. Tim-3 expression was evaluated by immunohistochemical staining. The Tim-3 expression level was evaluated by q-RT-PCR, western blot and immunocytochemistry in four colon cancer cell lines (HT-29, HCT116, LoVo, SW620). Tim-3 was expressed in 92.5% tumor tissue samples and 86.5% corresponding normal tissue samples. Expression of Tim-3 was significantly higher in tumor tissues than in normal tissues (P < 0.0001). Tim-3 expression in colon cancer tissues is in correlation with colon cancer lymphatic metastasis and TNM (P < 0.0001). Multivariate analysis demonstrated that Tim-3 expression could be a potential independent prognostic factor for colon cancer patients (P < 0.0001). Kaplan-Meier survival analysis result showed that patients with higher Tim-3 expression had a significantly shorter survival time than those with lower Tim-3 expression patients. Our results indicated that Tim-3 might participate in the tumorgenesis of colon cancer and Tim-3 expression might be a potential independent prognostic factor for patients with colorectal cancer.

Taipale K, Liikanen I, Juhila J, et al.
Chronic Activation of Innate Immunity Correlates With Poor Prognosis in Cancer Patients Treated With Oncolytic Adenovirus.
Mol Ther. 2016; 24(1):175-83 [PubMed] Free Access to Full Article Related Publications
Despite many clinical trials conducted with oncolytic viruses, the exact tumor-level mechanisms affecting therapeutic efficacy have not been established. Currently there are no biomarkers available that would predict the clinical outcome to any oncolytic virus. To assess the baseline immunological phenotype and find potential prognostic biomarkers, we monitored mRNA expression levels in 31 tumor biopsy or fluid samples from 27 patients treated with oncolytic adenovirus. Additionally, protein expression was studied from 19 biopsies using immunohistochemical staining. We found highly significant changes in several signaling pathways and genes associated with immune responses, such as B-cell receptor signaling (P < 0.001), granulocyte macrophage colony-stimulating factor (GM-CSF) signaling (P < 0.001), and leukocyte extravasation signaling (P < 0.001), in patients surviving a shorter time than their controls. In immunohistochemical analysis, markers CD4 and CD163 were significantly elevated (P = 0.020 and P = 0.016 respectively), in patients with shorter than expected survival. Interestingly, T-cell exhaustion marker TIM-3 was also found to be significantly upregulated (P = 0.006) in patients with poor prognosis. Collectively, these data suggest that activation of several functions of the innate immunity before treatment is associated with inferior survival in patients treated with oncolytic adenovirus. Conversely, lack of chronic innate inflammation at baseline may predict improved treatment outcome, as suggested by good overall prognosis.

Cai C, Xu YF, Wu ZJ, et al.
Tim-3 expression represents dysfunctional tumor infiltrating T cells in renal cell carcinoma.
World J Urol. 2016; 34(4):561-7 [PubMed] Related Publications
PURPOSE: Renal cell carcinoma (RCC) is the most common cancer of kidney. Evidences have shown that RCC is sensitive to various immunotherapies. Tim-3 plays a role in suppressing Th1-mediated immune responses. However, no study has yet examined the effect of Tim-3 on tumor infiltrating lymphocytes (TILs) in RCC.
METHODS: We investigated the expression and function of Tim-3 on TIL CD4+ T cells and TIL CD8+ T cells from 30 RCC patients.
RESULTS: Levels of Tim-3 were significantly increased on both TIL CD4+ T cells and TIL CD8+ T cells and were associated with higher stages of the cancer. Also, GATA-3 and interferon gamma (IFN-γ) were down-regulated, whereas T-bet was up-regulated in TIL Tim-3+ T cells, indicating that Tim-3 expression defined a population of dysfunctional TIL Th1/Tc1 cells. Mechanism analyses showed that TIL Tim-3-expressing CD8+ T cells exhibited impaired Stat5 and p38 signaling pathway. Blocking the Tim-3 pathway restored cell proliferation and increased IFN-γ production in TIL CD4+ and CD8+ T cells of RCC.
CONCLUSIONS: These results suggest that Tim-3 may be used as a novel target for increasing immune responses in RCC tumor microenvironment.

Cheng L, Ruan Z
Tim-3 and Tim-4 as the potential targets for antitumor therapy.
Hum Vaccin Immunother. 2015; 11(10):2458-62 [PubMed] Free Access to Full Article Related Publications
Both Tim-3 and Tim-4 belong to the T-cell immunoglobulin and mucin domain (Tim) gene family, which plays a critical role in immunoregulation. Tim-3 has been suggested as a negative regulator of anti-tumor immunity due to its function on inducing T cells exhaustion in cancer. In addition to its expression on exhausted T cells, Tim-3 also has been reported to up-regulate on nature killer (NK) cells and promote NK cells functionally exhausted in cancer. While Tim-3 selectively expression on most types of leukemia stem cells, it promotes the progression of acute myeloid leukemia. Recently, data from experimental models of tumor discovered that Tim-3 and Tim-4 up-regulation on tumor associated dendritic cells and macrophages attenuated the anti-tumor effects of cancer vaccines and chemotherapy. Moreover, co-blockage of Tim-3 and PD-1, Tim-3 and CD137, Tim-3 and carcinoembryonic antigen cell adhesion molecule 1 (CEACAM1) could enhance cell-mediated immunity in advanced tumor, and combined treatment with anti-Tim-3 and anti-Tim-4 mAbs further increase the efficacy of cancer vaccines. The therapeutic manipulation of TIM-3 and TIM-4 may provide a novel strategy to improve the clinical efficacy of cancer immunotherapy.

Heon EK, Wulan H, Macdonald LP, et al.
IL-15 induces strong but short-lived tumor-infiltrating CD8 T cell responses through the regulation of Tim-3 in breast cancer.
Biochem Biophys Res Commun. 2015; 464(1):360-6 [PubMed] Related Publications
IL-15 has pivotal roles in the control of CD8(+) memory T cells and has been investigated as a therapeutic option in cancer therapy. Although IL-15 and IL-2 share many functions together, including the stimulation of CD8 T cell proliferation and IFN-γ production, the different in vivo roles of IL-15 and IL-2 have been increasingly recognized. Here, we explored the different effects of IL-15 and IL-2 on tumor-infiltrating (TI) T cells from resected breast tumors. We found that neither IL-2 nor IL-15 induced intratumoral CD8 T cell proliferation by itself, but after CD3/CD28-stimulation, IL-15 induced significantly higher proliferation than IL-2 during early time points, at day 2, day 3 and day 6. However, the IL-15-induced proliferation leveled off at day 9 and day 12, whereas IL-2 induced lower but progressive proliferation at each time point. Furthermore, IL-15 caused an early and robust increase of IFN-γ in the supernatant of TI cell cultures, which diminished at later time points, while the IL-2-induced IFN-γ production remained constant over time. In addition, the IL-15-costimulated CD8 T cells presented higher frequencies of apoptotic cells. The diminishing IL-15-induced response was possibly due to regulatory and/or exhaustion mechanisms. We did not observe increased IL-10 or PD-1 upregulation, but we have found an increase of Tim-3 upregulation on IL-15-, but not IL-2-stimulated cells. Blocking Tim-3 function using anti-Tim-3 antibodies resulted in increased IL-15-induced proliferation and IFN-γ production for a prolonged period of time, whereas adding Tim-3 ligand galectin 9 led to reduced proliferation and IFN-γ production. Our results suggest that IL-15 in combination of Tim-3 blocking antibodies could potentially act as an IL-2 alternative in tumor CD8 T cell expansion in vitro, a crucial step in adoptive T cell therapy.

Webb JR, Milne K, Nelson BH
PD-1 and CD103 Are Widely Coexpressed on Prognostically Favorable Intraepithelial CD8 T Cells in Human Ovarian Cancer.
Cancer Immunol Res. 2015; 3(8):926-35 [PubMed] Related Publications
αE(CD103)β7 is a TGFβ-regulated integrin that mediates retention of lymphocytes in peripheral tissues by binding to E-cadherin expressed on epithelial cells. We recently reported that αE(CD103)β7 specifically demarcates intraepithelial CD8(+) tumor-infiltrating lymphocytes (CD8 TIL) in ovarian cancer and that CD103(+) TIL have a surface profile consistent with an active effector phenotype (HLA-DR(+), Ki67(+), and CD127(lo)). These findings led us to hypothesize that, over time, CD103-mediated retention of CD8 TIL within the tumor epithelium might result in chronic stimulation by tumor antigen, which in turn might lead to an exhausted phenotype. To investigate this possibility, we evaluated PD-1 expression in a large cohort of ovarian tumors (N = 489) with known CD103(+) TIL content. PD-1(+) cells were present in 38.5% of high-grade serous carcinomas (HGSC), but were less prevalent in other histologic subtypes. PD-1(+) TIL were strongly associated with increased disease-specific survival in HGSC (HR, 0.4864; P = 0.0007). Multicolor immunohistochemistry and flow cytometry revealed a high degree of PD-1 and CD103 coexpression, specifically within the CD8 TIL compartment. PD-1(+)CD103(+) CD8 TIL were quiescent when assessed directly ex vivo yet were capable of robust cytokine production after pharmacologic stimulation. Moreover, they showed negligible expression of additional exhaustion-associated markers, including TIM-3, CTLA-4, and LAG-3. Thus, as hypothesized, CD103(+) CD8 TIL express PD-1 and appear quiescent in the tumor microenvironment. However, these cells retain functional competence and demonstrate strong prognostic significance. We speculate that, after standard treatment, PD-1(+)CD103(+) CD8 TIL might regain functional antitumor activity, an effect that potentially could be augmented by immune modulation.

Sehgal K, Das R, Zhang L, et al.
Clinical and pharmacodynamic analysis of pomalidomide dosing strategies in myeloma: impact of immune activation and cereblon targets.
Blood. 2015; 125(26):4042-51 [PubMed] Free Access to Full Article Related Publications
In preclinical studies, pomalidomide mediated both direct antitumor effects and immune activation by binding cereblon. However, the impact of drug-induced immune activation and cereblon/ikaros in antitumor effects of pomalidomide in vivo is unknown. Here we evaluated the clinical and pharmacodynamic effects of continuous or intermittent dosing strategies of pomalidomide/dexamethasone in lenalidomide-refractory myeloma in a randomized trial. Intermittent dosing led to greater tumor reduction at the cost of more frequent adverse events. Both cohorts experienced similar event-free and overall survival. Both regimens led to a distinct pattern but similar degree of mid-cycle immune activation, manifested as increased expression of cytokines and lytic genes in T and natural killer (NK) cells. Pomalidomide induced poly-functional T-cell activation, with increased proportion of coinhibitory receptor BTLA(+) T cells and Tim-3(+) NK cells. Baseline levels of ikaros and aiolos protein in tumor cells did not correlate with response or survival. Pomalidomide led to rapid decline in Ikaros in T and NK cells in vivo, and therapy-induced activation of CD8(+) T cells correlated with clinical response. These data demonstrate that pomalidomide leads to strong and rapid immunomodulatory effects involving both innate and adaptive immunity, even in heavily pretreated multiple myeloma, which correlates with clinical antitumor effects. This trial was registered at www.clinicaltrials.gov as #NCT01319422.

Djenidi F, Adam J, Goubar A, et al.
CD8+CD103+ tumor-infiltrating lymphocytes are tumor-specific tissue-resident memory T cells and a prognostic factor for survival in lung cancer patients.
J Immunol. 2015; 194(7):3475-86 [PubMed] Related Publications
We had previously demonstrated the role of CD103 integrin on lung tumor-infiltrating lymphocyte (TIL) clones in promoting specific TCR-mediated epithelial tumor cell cytotoxicity. However, the contribution of CD103 on intratumoral T cell distribution and functions and the prognosis significance of TIL subpopulations in non-small cell lung carcinoma (NSCLC) have thus far not been systematically addressed. In this study, we show that an enhanced CD103(+) TIL subset correlates with improved early stage NSCLC patient survival and increased intraepithelial lymphocyte infiltration. Moreover, our results indicate that CD8(+)CD103(+) TIL, freshly isolated from NSCLC specimens, display transcriptomic and phenotypic signatures characteristic of tissue-resident memory T cells and frequently express PD-1 and Tim-3 checkpoint receptors. This TIL subset also displays increased activation-induced cell death and mediates specific cytolytic activity toward autologous tumor cells upon blockade of the PD-1-PD-L1 interaction. These findings emphasize the role of CD8(+)CD103(+) tissue-resident memory T cells in promoting intratumoral CTL responses and support the rationale for using anti-PD-1 blocking Ab to reverse tumor-induced T cell exhaustion in NSCLC patients.

Yang ZZ, Grote DM, Ziesmer SC, et al.
PD-1 expression defines two distinct T-cell sub-populations in follicular lymphoma that differentially impact patient survival.
Blood Cancer J. 2015; 5:e281 [PubMed] Free Access to Full Article Related Publications
To determine the biological and clinical relevance of programmed death 1 (PD-1) in follicular lymphoma (FL), we characterized PD-1(+) T-cell subsets and assessed their biological function as well as potential clinical impact. We found that PD-1 is expressed on intratumoral CD4(+) T cells with both bright and dim intensity, representing two different sub-populations of cells. By immunohistochemistry, we found that CD4(+)PD-1(high) T cells predominantly reside in the lymph node follicles, while PD-1(low) T cells are mainly located in an interfollicular pattern. Intratumoral CD4(+)PD-1(high) T cells have a TFH cell phenotype, express CXCR5, secrete IL-21 and are BCL-6 positive with no TIM-3 expression. In contrast, CD4(+)PD-1(low) T cells have an exhausted phenotype, express TIM-3 and do not express BCL-6 and CXCR5. Functionally, CD4(+)PD-1(high) T cells actively supported B-cell growth, while CD4(+)PD-1(low) T cells displayed a reduced cytokine production and cell-signal transduction. Clinically, we observed that the numbers of CD4(+) or CD8(+)PD-1(low) T cells significantly correlate with a reduced overall survival in FL patients (P=0.007 and 0.04 respectively; n=32). In contrast, the number of CD4(+)PD-1(high) T cells was not associated with patient outcome. Taken together, these results indicated that PD-1 expression defines two sub-populations with distinct functions that differentially impact patient outcome in FL.

Yan W, Liu X, Ma H, et al.
Tim-3 fosters HCC development by enhancing TGF-β-mediated alternative activation of macrophages.
Gut. 2015; 64(10):1593-604 [PubMed] Related Publications
OBJECTIVE: Tumour-associated macrophages (TAMs) and their alternative activation contribute greatly to the development of hepatocellular carcinoma (HCC). Tim-3 is highly expressed on macrophages and regulates macrophage functions in several conditions. However, whether Tim-3 is involved in the activation and the function of TAMs has not been reported.
DESIGN: Tim-3 expression in HCC samples was evaluated by flow cytometry, immunohistochemistry and confocal analysis. We analysed the effects of Tim-3 knockdown on macrophages in growth of H22 tumour homografts in BALB/c mice. Tim-3 interference was performed by neutralising antibody, small interfering RNA or short hairpin RNA-expressing lentivirus. Cytokine production was evaluated by reverse transcription PCR, ELISA or Cytometric Bead Array. The effects of Tim-3 interference in macrophages were examined with regard to alternative activation of macrophages and proliferation and migration of Hepa1-6 cells. Cell growth curve, colony formation and transwell assays were involved to estimate cell proliferation and migration.
RESULTS: Tim-3 expression was significantly increased in both peripheral blood monocytes and TAMs in patients with HCC. The Tim-3 expression in monocytes/TAMs strongly correlated with higher tumour grades and the poor survival of patients with HCC. Consistently, HCC conditioned medium or transforming growth factor-β fostered Tim-3 expression and the alternative activation of macrophages. Moreover, Tim-3 interference in macrophages significantly inhibited the alternative activation of macrophages and suppressed HCC cell growth both in vitro and in vivo. Blocking interleukin 6 reversed the Tim-3-mediated effects on HCC cell growth in vitro.
CONCLUSIONS: Tim-3 displays critical roles in microenvironment-induced activation and protumoral effects of TAMs in HCC. Interference of Tim-3 might be great potential in HCC therapy.

Mrizak D, Martin N, Barjon C, et al.
Effect of nasopharyngeal carcinoma-derived exosomes on human regulatory T cells.
J Natl Cancer Inst. 2015; 107(1):363 [PubMed] Related Publications
BACKGROUND: Regulatory T cells (Treg) and tumor-exosomes are thought to play a role in preventing the rejection of malignant cells in patients bearing nasopharyngeal carcinoma (NPC).
METHODS: Treg recruitment by exosomes derived from NPC cell lines (C15/C17-Exo), exosomes isolated from NPC patients' plasma (Patient-Exo), and CCL20 were tested in vitro using Boyden chamber assays and in vivo using a xenograft SCID mouse model (n = 5), both in the presence and absence of anti-CCL20 monoclonal antibodies (mAb). Impact of these NPC exosomes (NPC-Exo) on Treg phenotype and function was determined using adapted assays (FACS, Q-PCR, ELISA, and MLR). Experiments were performed in comparison with exosomes derived from plasma of healthy donors (HD-Exo). The Student's t test was used for group comparisons. All statistical tests were two-sided.
RESULTS: CCL20 allowed the intratumoral recruitment of human Treg. NPC-Exo also facilitated Treg recruitment (3.30 ± 0.34 fold increase, P < .001), which was statistically significantly inhibited (P < .001) by an anti-CCL20 blocking mAb. NPC-Exo also recruited conventional CD4(+)CD25(-) T cells and mediated their conversion into inhibitory CD4(+)CD25(high) cells. Moreover, NPC-Exo enhanced (P = .0048) the expansion of human Treg, inducing the generation of Tim3(Low) Treg with increased expression of CD25 and FOXP3. Finally, NPC-Exo induced an overexpression of cell markers associated with Treg phenotype, properties and recruitment capacity. For example, GZMB mean fold change was 21.45 ± 1.75 (P < .001). These results were consistent with a stronger suppression of responder cells' proliferation and the secretion of immunosuppressive cytokines (IL10, TGFB1).
CONCLUSION: Interactions between NPC-Exo and Treg represent a newly defined mechanism that may be involved in regulating peripheral tolerance by tumors and in supporting immune evasion in human NPC.

Markwick LJ, Riva A, Ryan JM, et al.
Blockade of PD1 and TIM3 restores innate and adaptive immunity in patients with acute alcoholic hepatitis.
Gastroenterology. 2015; 148(3):590-602.e10 [PubMed] Related Publications
BACKGROUND & AIMS: Susceptibility to bacterial infection is a feature of alcohol-related liver disease. Programmed cell death 1 (PD1), the T-cell immunoglobulin and mucin domain-containing protein 3 (TIM3, also known as hepatitis A virus cellular receptor 2), and their respective ligands-CD274 (also known as PD ligand 1 [PDL1]) and galectin-9-are inhibitory receptors that regulate the balance between protective immunity and host immune-mediated damage. However, their sustained hyperexpression promotes immune exhaustion and paralysis. We investigated the role of these immune inhibitory receptors in driving immune impairments in patients with alcoholic liver disease.
METHODS: In a prospective study, we collected blood samples from 20 patients with acute alcoholic hepatitis (AAH), 16 patients with stable advanced alcohol-related cirrhosis, and 12 healthy individuals (controls). Whole blood or peripheral blood mononuclear cells were assessed for expression of PD1, PDL1, TIM3, galectin-9, and Toll-like receptors on subsets of innate and adaptive immune effector cells. We measured antibacterial immune responses to lipopolysaccharide (endotoxin) using ELISpot assays, and used flow cytometry to quantify cytokine production, phagocytosis, and oxidative burst in the presence or absence of blocking antibodies against PD1 or TIM3.
RESULTS: Antibacterial innate and adaptive immune responses were greatly reduced in patients with AAH, compared with controls, and patients with alcohol-related cirrhosis had less severe dysfunctions in innate immune effector cells and preserved functional T-cell responses. Fewer T cells from patients with AAH produced interferon gamma in response to lipopolysaccharide, compared with controls. In addition, patients with AAH had greater numbers of interleukin 10-producing T cells, and reduced levels of neutrophil phagocytosis and oxidative burst in response to Escherichia coli stimulation, compared with controls. T cells from patients with AAH, but not alcohol-related cirrhosis, expressed higher levels of PD1 and PDL1, or TIM3 and galectin-9, than T cells from controls. Antibodies against PD1 and TIM3 restored T-cell production of interferon gamma, reduced the numbers of interleukin 10-producing T cells, and increased neutrophil antimicrobial activities. Circulating levels of endotoxin in plasma from patients with AAH caused over expression of immune inhibitory receptors on T cells via Toll-like receptor 4 binding to CD14(+) monocytes.
CONCLUSIONS: Antibacterial immune responses are impaired in patients with AAH. Lymphocytes from these patients express high levels of immune inhibitory receptors, produce lower levels of interferon gamma, and have increased IL10 production due to chronic endotoxin exposure. These effects can be reversed by blocking PD1 and TIM3, which increase the antimicrobial activities of T cells and neutrophils.

Fend L, Gatard-Scheikl T, Kintz J, et al.
Intravenous injection of MVA virus targets CD8+ lymphocytes to tumors to control tumor growth upon combinatorial treatment with a TLR9 agonist.
Cancer Immunol Res. 2014; 2(12):1163-74 [PubMed] Related Publications
Effector T-cell access to tumor tissue is a limiting step for clinical efficacy of antigen-specific T cell-based immunotherapies. Ectopic mouse tumor models, in which a subcutaneously (s.c.) implanted tumor is treated with s.c. or intramuscular therapeutic immunization, may not be optimal for targeting effector T cells to an organ-borne tumor. We used an orthotopic renal carcinoma model to evaluate the impact of injection routes on therapeutic efficacy of a Modified Vaccinia virus Ankara viral vector expressing the human mucin 1 tumor-associated xeno-antigen (MVA-MUC1). We show that intravenous (i.v.) administration of MVA-MUC1 displayed enhanced efficacy when compared with s.c. injection. Therapeutic efficacy of MVA-MUC1 was further enhanced by i.v. injection of a TLR9 agonist. In all cases, infiltration of tumor-bearing kidney by CD8(+) lymphocytes was associated with control of tumor growth. Biodistribution experiments indicate that, following i.v. injection, MVA-encoded antigens are quickly expressed in visceral organs and, in particular, in splenic antigen-presenting cells, compared with those following s.c. injection. This appears to result in a faster generation of MUC1-specific CD8(+) T cells. Lymphocytes infiltrating tumor-bearing kidneys are characterized by an effector memory phenotype and express PD-1 and Tim3 immune checkpoint molecules. Therapeutic efficacy was associated with a modification of the tumor microenvironment toward a Th1-type immune response and recruitment of activated lymphocytes. This study supports the clinical evaluation of MVA-based immunotherapies via the i.v. route.

Yang J, Zhu L, Cai Y, et al.
Role of downregulation of galectin-9 in the tumorigenesis of gastric cancer.
Int J Oncol. 2014; 45(3):1313-20 [PubMed] Related Publications
Galectin-9 (Gal-9), a member of the β-galactoside-binding galectin family, plays a role in immune response, apoptosis, cell proliferation and cell death. Recent studies have shown that abnormal expression of Gal-9 is involved in certain primary cancers. The present study is the first investigation of the role of Gal-9 gene expression in clinically diagnosed primary gastric cancer tissues. Gal-9 mRNA expression was assessed in 44 clinically diagnosed frozen primary cancer tissue samples using quantitative PCR (qPCR). Analysis of the qPCR data revealed a significant reduction (>2-fold decreased) of Gal-9 gene expression in gastric cancer tissues in 77% (34/44) of patients. In patients with gastric cancer, although no statistically significant difference was found between adjacent (<2 cm away from the cancer tissue) and normal tissues (>5 cm away from the cancer tissue), a >2-fold reduction in Gal-9 expression was observed in the adjacent tissues of 34% of the patients. Compared to matched normal or adjacent tissues, the gene expression of Gal-9 was significantly decreased in tumor tissues (p<0.001). The correlation of Gal-9 expression and clinicopathological features in gastric cancer was analyzed according to the TNM classification system using AJCC stage grouping. In patients with gastric cancer, clinical staging, tumor pathological stage (pT stage), tumor cell differentiation, lymph node metastasis and survival rate were found to be associated with Gal-9 expression. However, no significant association was found between Gal-9 expression and distant metastasis (p>0.05). No significant difference was found between patients of different genders, levels of cell differentiation, distant metastasis status or different survival time of patients. Compared to normal tissues, >2-fold reduction of Tim-3 expression in gastric cancer tissues occurred in 59% of patients, but no correlation was found between Gal-9 and Tim-3 in gastric cancer. These results strongly suggest that Gal-9 is involved in tumorigenesis of gastric cancer.

Jajosky AN, Coad JE, Vos JA, et al.
RepSox slows decay of CD34+ acute myeloid leukemia cells and decreases T cell immunoglobulin mucin-3 expression.
Stem Cells Transl Med. 2014; 3(7):836-48 [PubMed] Free Access to Full Article Related Publications
Despite initial response to therapy, most acute myeloid leukemia (AML) patients relapse. To eliminate relapse-causing leukemic stem/progenitor cells (LPCs), patient-specific immune therapies may be required. In vitro cellular engineering may require increasing the "stemness" or immunogenicity of tumor cells and activating or restoring cancer-impaired immune-effector and antigen-presenting cells. Leukapheresis samples provide the cells needed to engineer therapies: LPCs to be targeted, normal hematopoietic stem cells to be spared, and cancer-impaired immune cells to be repaired and activated. This study sought to advance development of LPC-targeted therapies by exploring nongenetic ways to slow the decay and to increase the immunogenicity of primary CD34(+) AML cells. CD34(+) AML cells generally displayed more colony-forming and aldehyde dehydrogenase activity than CD34(-) AML cells. Along with exposure to bone marrow stromal cells and low (1%-5%) oxygen, culture with RepSox (a reprogramming tool and inhibitor of transforming growth factor-β receptor 1) consistently slowed decline of CD34(+) AML and myelodysplastic syndrome (MDS) cells. RepSox-treated AML cells displayed higher CD34, CXCL12, and MYC mRNA levels than dimethyl sulfoxide-treated controls. RepSox also accelerated loss of T cell immunoglobulin mucin-3 (Tim-3), an immune checkpoint receptor that impairs antitumor immunity, from the surface of AML and MDS cells. Our results suggest RepSox may reduce Tim-3 expression by inhibiting transforming growth factor-β signaling and slow decay of CD34(+) AML cells by increasing CXCL12 and MYC, two factors that inhibit AML cell differentiation. By prolonging survival of CD34(+) AML cells and reducing Tim-3, RepSox may promote in vitro immune cell activation and advance development of LPC-targeted therapies.

Gros A, Robbins PF, Yao X, et al.
PD-1 identifies the patient-specific CD8⁺ tumor-reactive repertoire infiltrating human tumors.
J Clin Invest. 2014; 124(5):2246-59 [PubMed] Free Access to Full Article Related Publications
Adoptive transfer of tumor-infiltrating lymphocytes (TILs) can mediate regression of metastatic melanoma; however, TILs are a heterogeneous population, and there are no effective markers to specifically identify and select the repertoire of tumor-reactive and mutation-specific CD8⁺ lymphocytes. The lack of biomarkers limits the ability to study these cells and develop strategies to enhance clinical efficacy and extend this therapy to other malignancies. Here, we evaluated unique phenotypic traits of CD8⁺ TILs and TCR β chain (TCRβ) clonotypic frequency in melanoma tumors to identify patient-specific repertoires of tumor-reactive CD8⁺ lymphocytes. In all 6 tumors studied, expression of the inhibitory receptors programmed cell death 1 (PD-1; also known as CD279), lymphocyte-activation gene 3 (LAG-3; also known as CD223), and T cell immunoglobulin and mucin domain 3 (TIM-3) on CD8⁺ TILs identified the autologous tumor-reactive repertoire, including mutated neoantigen-specific CD8⁺ lymphocytes, whereas only a fraction of the tumor-reactive population expressed the costimulatory receptor 4-1BB (also known as CD137). TCRβ deep sequencing revealed oligoclonal expansion of specific TCRβ clonotypes in CD8⁺PD-1⁺ compared with CD8⁺PD-1- TIL populations. Furthermore, the most highly expanded TCRβ clonotypes in the CD8⁺ and the CD8⁺PD-1⁺ populations recognized the autologous tumor and included clonotypes targeting mutated antigens. Thus, in addition to the well-documented negative regulatory role of PD-1 in T cells, our findings demonstrate that PD-1 expression on CD8⁺ TILs also accurately identifies the repertoire of clonally expanded tumor-reactive cells and reveal a dual importance of PD-1 expression in the tumor microenvironment.

Yang ZZ, Grote DM, Xiu B, et al.
TGF-β upregulates CD70 expression and induces exhaustion of effector memory T cells in B-cell non-Hodgkin's lymphoma.
Leukemia. 2014; 28(9):1872-84 [PubMed] Free Access to Full Article Related Publications
Transforming growth factor beta (TGF-β) has an important role in mediating T-cell suppression in B-cell non-Hodgkin lymphoma (NHL). However, the underlying mechanism responsible for TGF-β-mediated inhibition of effector memory T (Tm) cells is largely unknown. As reported here, we show that exhaustion is a major mechanism by which TGF-β inhibits Tm cells, and TGF-β mediated exhaustion is associated with upregulation of CD70. We found that TGF-β upregulates CD70 expression on effector Tm cells while it preferentially induces Foxp3 expression in naive T cells. CD70 induction by TGF-β is Smad3-dependent and involves IL-2/Stat5 signaling. CD70+ T cells account for TGF-β-induced exhaustion of effector Tm cells. Both TGF-β-induced and preexisting intratumoral CD70+ effector Tm cells from B-cell NHL have an exhausted phenotype and express higher levels of PD-1 and TIM-3 compared with CD70- T cells. Signaling transduction, proliferation and cytokine production are profoundly decreased in these cells, and they are highly susceptible to apoptosis. Clinically, intratumoral CD70-expressing T cells are prevalent in follicular B-cell lymphoma (FL) biopsy specimens, and increased numbers of intratumoral CD70+ T cells correlate with an inferior patient outcome. These findings confirm TGF-β-mediated effector Tm cell exhaustion as an important mechanism of immune suppression in B-cell NHL.

Jiang J, Jin MS, Kong F, et al.
Decreased galectin-9 and increased Tim-3 expression are related to poor prognosis in gastric cancer.
PLoS One. 2013; 8(12):e81799 [PubMed] Free Access to Full Article Related Publications
INTRODUCTION: Galectin-9 (Gal-9) induces adhesion and aggregation of certain cell types and inhibits the metastasis of tumor cells. T-cell immunoglobulin-and mucin domain-3-containing molecule 3 (TIM-3) plays a pivotal role in immune regulation. The aim of this study is to investigate Gal-9 and TIM-3 alterations in gastric cancer and their prognostic values.
METHODS: Gal-9 and Tim-3 expression was evaluated using a tissue microarray immunohistochemistry method in 305 gastric cancers, of which 84 had paired adjacent normal samples. Cell lines SGC-7901, BGC-823, MGC-803, MKN45 and GES-1 were also stained. Correlations were analyzed between expression levels of Gal-9 and Tim-3 protein and tumor parameters or clinical outcomes.
RESULTS: Gal-9 and Tim-3 stained positive on tumor cells in 86.2% (263/305), and 60.0% (183/305) patients with gastric cancer, respectively. Gal-9 expression was significantly higher in cancer than in normal mucosa (P<0.001). Reduced Gal-9 expression was associated with lymph-vascular invasion, lymph node metastasis, distant metastasis and worse TNM staging (P = 0.034, P = 0.009, P = 0.002 and P = 0.043, respectively). In contrast, Tim-3 expression was significantly lower in cancer than in control mucosa (P<0.001). Patients with lymph-vascular invasion had higher expression levels of Tim-3 (P<0.001). Moreover, multivariate analysis shows that both high Gal-9 expression and low Tim-3 expression were significantly associated with long overall survival (P = 0.002, P = 0.010, respectively); the combination of Gal-9 and Tim-3 expression was an independent prognostic predictor for patients with gastric cancer (RR: 0.43; 95%CI: 0.20-0.93). H.pylori infection status was not associated with Gal-9 and Tim-3 expression (P = 0.102, P = 0.565).
CONCLUSION: The results suggest that expression of Gal-9 and Tim-3 in tumor cells may be a potential, independent prognostic factor for patients with gastric cancer. Gal-9 and TIM-3 may play an important part in the gastric carcinogenesis.

Roth CG, Garner K, Eyck ST, et al.
TIM3 expression by leukemic and non-leukemic myeloblasts.
Cytometry B Clin Cytom. 2013; 84(3):167-72 [PubMed] Related Publications
BACKGROUND: T-cell immunoglobulin mucin-3 (TIM3) has recently been described as an acute myeloid leukemia (AML) stem cell antigen expressed on leukemic myeloblasts, but not on normal hematopoietic stem cells. TIM3 is also expressed by monocytes, natural killer cells, and several T cell subsets; however, normal myeloblasts have not been well-characterized or compared to AML. A specific flow cytometric marker capable of separating leukemic myeloblasts from non-neoplastic myeloblasts would be diagnostically useful, especially in the post-chemotherapy setting.
METHODS: TIM3 myeloblast expression was assessed in 69 bone marrow and/or peripheral blood specimens, including 27 AML and 42 non-neoplastic cases (20 with a recent history of chemotherapy). TIM3 median fluorescence intensity (MFI) was evaluated within myeloblast, monocyte, T cell, and natural killer cell populations.
RESULTS: The median percentage of myeloblasts positive for TIM3 was lower in non-neoplastic specimens without a history of recent chemotherapy (50.3%) as compared to AML (71.4%), but not significantly different as compared to non-leukemic myeloblasts in the post-chemotherapy setting (72.4%). Mean myeloblast TIM3 MFI was higher in AML myeloblasts and non-leukemic myeloblasts in the post-chemotherapy setting as compared to non-neoplastic myeloblasts in cases lacking a history of chemotherapy. Mean monocyte, natural killer cell, and T-cell TIM3 MFI remained relatively constant in varied clinical settings.
CONCLUSIONS: We confirm that leukemic myeloblasts overexpress TIM3 as compared to non-neoplastic controls; however, high levels of expression may also be seen among non-leukemic myeloblasts in the post-chemotherapy setting. This overlap limits the diagnostic utility of TIM3 as a specific marker of neoplasia.

Song H, Ma S, Cha Z, et al.
T-cell immunoglobulin- and mucin-domain-containing molecule 3 genetic variants and HIV+ non-Hodgkin lymphomas.
Inflammation. 2013; 36(4):793-9 [PubMed] Related Publications
T cell immunoglobulin- and mucin-domain-containing molecule 3 (TIM-3) has been established as a negative regulatory molecule and plays a critical role in inflammatory diseases such as rheumatoid arthritis, hepatitis B and C, and human immunodeficiency virus (HIV)-related inflammation. Recent studies have shown that chronic inflammation may greatly affect the pathogenesis of non-Hodgkin lymphomas (NHL). The aim of this study was to investigate whether polymorphisms in the TIM-3 gene were associated with susceptibility to non-NHL and HIV-related NHL. Three polymorphisms in TIM-3 gene (-1516G/T, -574G/T, and +4259T/G) were identified by polymerase chain reaction-restriction fragment length polymorphism in 434 NHL patients, 62 HIV-related NHL cases, and 512 healthy controls. Results showed that the prevalence of -574GT genotype and +4259TG genotype were significantly increased in the NHL cases than in controls (odds ratio (OR) = 2.72, 95% confidence interval (CI) = 1.50-4.92, p = 0.0006 and OR = 2.59, 95% CI = 1.49-4.49, p = 0.0005, respectively). The -1516G/T polymorphism did not reveal significant difference between patients and healthy controls. When analyzing the TIM-3 polymorphisms in HIV-related NHL patients, data showed that HIV+ NHL patients had higher prevalence of -574GT or +4259TG genotypes than those cases without HIV infection (OR = 3.48, 95% CI = 1.67-7.28, p = 0.0005 and OR = 2.92, 95% CI = 1.42-6.01, p = 0.0026, respectively). These results suggested polymorphisms in TIM-3 gene could be new risk factors for NHL as well as HIV-related NHL and suggested a possible role of the inflammatory factor in these diseases.

Bai J, Li X, Tong D, et al.
T-cell immunoglobulin- and mucin-domain-containing molecule 3 gene polymorphisms and prognosis of non-small-cell lung cancer.
Tumour Biol. 2013; 34(2):805-9 [PubMed] Related Publications
Lung cancer is the leading cause of death worldwide. Non-small-cell lung cancer (NSCLC) accounts for most of these cases. T-cell immunoglobulin- and mucin-domain-containing molecule 3 (TIM-3) has been established as a negative regulatory molecule and plays a critical role in immune tolerance. Studies have shown that polymorphisms in TIM-3 gene can be associated with various diseases. The aim of this study was to investigate whether polymorphisms in the TIM-3 gene were associated with susceptibility to NSCLC. Three polymorphisms in TIM-3 gene (-1516G/T, -574G/T, and +4259T/G) were identified by polymerase chain reaction-restriction fragment length polymorphism in 432 NSCLC patients and 466 healthy controls. Results showed that frequencies of TIM-3 +4259TG genotype for cases and controls were 10.9 and 4.1 %, respectively; subjects carrying the +4259TG genotype had a 2.81-fold increased risk of NSCLC compared to the wild-type genotype (P < 0.0001). The TIM-3 -1516G/T and -574G/T polymorphisms did not show any correlation with NSCLC. In addition, when analyzing the survival time of NSCLC patients with TIM-3 +4259T/G polymorphism, cases with +4259TG genotype had significantly shorter survival time compared to the wild-type patients (15.2 months vs. 26.7 months, P = 0.007). These results suggested polymorphism in TIM-3 gene is associated with increased susceptibility to NSCLC and could be used as prognostic factor for this malignancy.

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