Function Of TIGIT Research Articles

TIGIT: A promising target to overcome the barrier of immunotherapy in hematological malignancies.

Immune evasion through up-regulating checkpoint inhibitory receptors on T cells plays an essential role in tumor initiation and progression. Therefore, immunotherapy, including immune checkpoint inhibitor targeting programmed cell death protein 1 (PD-1) and chimeric antigen receptor T cell (CAR-T) therapy, has become a promising strategy for hematological malignancies. T cell immunoreceptor with immunoglobulin and ITIM domain (TIGIT) is a novel checkpoint inhibitory receptor expressed on immune cells, including cytotoxic T cells, regulatory T cells, and NK cells. TIGIT participates in immune regulation via binding to its ligand CD155. Blockage of TIGIT has provided evidence of considerable efficacy in solid tumors in preclinical research and clinical trials, especially when combined with PD-1 inhibition. However, the mechanism and function of TIGIT in hematological malignancies have not been comprehensively studied. In this review, we focus on the role of TIGIT in hematological malignancies and discuss therapeutic strategies targeting TIGIT, which may provide a promising immunotherapy target for hematological malignancies.

Fcγ receptor-mediated cross-linking codefines the immunostimulatory activity of anti-human CD96 antibodies.

New strategies that augment T cell responses are required to broaden the therapeutic arsenal against cancer. CD96, TIGIT, and CD226 are receptors that bind to a communal ligand, CD155, and transduce either inhibitory or activating signals. The function of TIGIT and CD226 is established, whereas the role of CD96 remains ambiguous. Using a panel of engineered antibodies, we discovered that the T cell stimulatory activity of anti-CD96 antibodies requires antibody cross-linking and is potentiated by Fcγ receptors. Thus, soluble "Fc silent" anti-CD96 antibodies failed to stimulate human T cells, whereas the same antibodies were stimulatory after coating onto plastic surfaces. Remarkably, the activity of soluble anti-CD96 antibodies was reinstated by engineering the Fc domain to a human IgG1 isotype, and it was dependent on antibody trans-cross-linking by FcγRI. In contrast, neither human IgG2 nor variants with increased Fcγ receptor IIB binding possessed stimulatory activity. Anti-CD96 antibodies acted directly on T cells and augmented gene expression networks associated with T cell activation, leading to proliferation, cytokine secretion, and resistance to Treg suppression. Furthermore, CD96 expression correlated with survival in HPV+ head and neck squamous cell carcinoma, and its cross-linking activated tumor-infiltrating T cells, thus highlighting the potential of anti-CD96 antibodies in cancer immunotherapy.

Beyond PD-1: Investigating the Therapeutic Potential of TIGIT Blockade in DLBCL

Background: The PD-1/PD-L1 axis is a dominant cancer immune escape pathway, and PD-1 blockade therapy has greatly benefited patients with select solid tumors and lymphomas. Unfortunately, anti-PD-1 monotherapy has limited efficacy against relapsed/refractory (r/r) diffuse large B cell lymphoma (DLBCL) - a disease where new therapies are needed. Because numerous inhibitory checkpoint receptors have been implicated in driving tumor-specific T cell dysfunction, we hypothesized that combinatorial checkpoint blockade therapy (CBT) would enhance the activity of PD-1-based therapy in r/r DLBCL. T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) is a recently identified co-inhibitory receptor expressed on dysfunctional tumor-infiltrating T cells. PD-1 and TIGIT co-blockade therapy has demonstrated impressive activity in pre-clinical solid tumor and myeloma models. However, the degree to which TIGIT is involved in mediating T cell suppression in DLBCL is not fully known. Methods: TIGIT expression on lymphoma-infiltrating T cells (LITs) from 18 fresh lymphoma samples was analyzed by flow cytometry. Multiplex IHC on tissue microarrays (TMAs) was also performed to investigate TIGIT expression in DLBCL samples. The syngeneic murine A20 B cell lymphoma model was employed to study: 1) the kinetics of TIGIT and other co-receptor expression on LITs, 2) the association of TIGIT expression with effector function among LITs, and 3) the effectiveness of anti-TIGIT mono- and combination CBT in mice with established A20 lymphomas. A20 lymphoma tumors were established in groups of Balb/c mice by subcutaneous (SQ) injections of 5 x 106 cells. Expression of TIGIT and other co-receptors in A20 LITs was examined by flow cytometry at various time points. Function of TIGIT+ LITs was assessed by examining cytokine production following ex vivo stimulation with PMA and ionomycin. To test the efficacy of TIGIT blockade, mice received intraperitoneal injections of anti-TIGIT, anti-PD-1, anti-4-1BB, or combinations of these antibodies. Treatments began once tumors reached a diameter of 10 mm and were continued every 3 days for 5 doses. Tumor growth was monitored and compared to that in A20-bearing mice treated with isotype control antibodies. In some experiments, mice that achieved complete tumor rejection following single or dual CBT were re-challenged with A20 cells to investigate immunological memory responses. Results: Across a variety of human lymphomas, flow cytometric analysis revealed that TIGIT was broadly upregulated on LITs, including regulatory T cells and conventional CD4+and CD8+ T cells (Figure A and B). TIGIT expression on LITs in DLBCL was particularly high. Nearly all TIGIT+ LITs were also PD-1+, suggesting that these receptors co-orchestrate a T cell dysfunctional state in the lymphoma environment. Multiplex immunofluorescence staining of DLBCL samples demonstrated that TIGIT was most highly expressed on CD8+ T cells and that TIGIT+ T cells tended to be localized near, and in some cases, surrounding CD20+ lymphoma cells. Consistent with observations in human lymphomas, LITs isolated from murine A20 lymphoma commonly co-expressed TIGIT and PD-1, and the degree of expression correlated directly with tumor volume. This correlation was also present for other co-receptors, including 4-1BB, TIM3, and CTLA-4. Ex vivo restimulation of A20 LITs revealed that TIGIT+ T cells produced lower levels of effector cytokines, such as TNF-α, compared with TIGIT- T cells. In mice with established A20 lymphomas, both TIGIT and PD-1 mono-blockade led to modest delays in tumor outgrowth compared with mice treated with isotype control antibodies. Strikingly, however, combined PD-1 and TIGIT blockade resulted in complete rejection of A20 lymphomas in most mice and led to significantly prolonged survival compared to mice treated with single agent CBT (Figure C). Combination TIGIT and 4-1BB CBT was also remarkably effective in driving rejection of A20 lymphomas, and led to remarkable memory responses. Conclusions: TIGIT promotes immune tolerance in the DLBCL environment. While TIGIT monotherapy has anti-lymphoma activity, combinatorial CBT incorporating anti-TIGIT antibodies drives extremely potent rejection of established lymphomas in mice. These results provide rationale for further study of TIGIT blockade as a therapeutic strategy in r/r lymphomas, including DLBCL. Disclosures Sunseri: iTeos Therapeutics: Research Funding. Wald:iTeos Therapeutics: Employment. Preillon:iTeos Therapeutics: Employment. Smith:Portola Pharmaceuticals: Research Funding. Driessens:iTeos Therapeutics: Employment. Kline:Merck: Honoraria; Merck: Research Funding.

Expression of the Inhibitory Receptor TIGIT Is Up-Regulated Specifically on NK Cells With CD226 Activating Receptor From HIV-Infected Individuals.

Natural killer (NK) cells are important for maintenance of innate immune system stability and serve as a first line of defense against tumors and virus infections; they can act either directly or indirectly and are regulated via co-operation between inhibitory and stimulatory surface receptors. The recently reported inhibitory receptor, TIGIT, can be expressed on the NK cell surface; however, the expression level and function of TIGIT on NK cells during HIV infection is unknown. In this study, for the first time, we investigated the expression and function of TIGIT in NK cells from HIV-infected individuals. Our data demonstrate that the level of TIGIT is higher on NK cells from patients infected with human immunodeficiency virus (HIV) compared with HIV-negative healthy controls. TIGIT expression is inversely correlated with CD4+ T cell counts and positively correlated with plasma viral loads. Additionally, levels of the TIGIT ligand, CD155, were higher on CD4+ T cells from HIV-infected individuals compared with those from healthy controls; however, there was no difference in the level of the activating receptor, CD226, which recognizes the same ligands as TIGIT. Furthermore, TIGIT was found to specifically up-regulated on CD226+ NK cells in HIV-infected individuals, and either rIL-10, or rIL-12 + rIL-15, could induce TIGIT expression on these cells. In addition, high TIGIT expression inhibited the production of interferon-gamma (IFN-γ) by NK cells, while TIGIT inhibition restored IFN-γ production. Overall, these results highlight the important role of TIGIT in NK cell function and suggest a potential new avenue for the development of therapeutic strategies toward a functional cure for HIV.

Abstract 4720: TIGIT blockade enhances cytolytic function in antigen-specific CTLs in a manner non-redundant to PD1 blockade

Abstract Background Many mechanisms contribute to the dysfunction of CTLs in the tumor microenvironment, rendering these cells unable to kill. TIGIT is a co-inhibitory receptor on T cells that is notably expressed at high levels by antigen-specific PD1-expressing CTLs in tumors. The ligands for TIGIT, CD155 and CD112, are in turn expressed by tumor cells and tumor-infiltrating immune stroma. TIGIT-mediated suppression of T cell function results from T cell intrinsic competition with the co-stimulatory molecule CD226 for engagement of CD155 and CD112, or enhancement of cell intrinsic repressive function of TIGIT downstream of the T cell receptor (TCR). To dissect the release of TIGIT-mediated suppression of polyclonal antigen-specific CTLs, we 1) examined the expression of TIGIT, CD226, and their shared ligands CD112 and CD155 in primary human tumor disaggregates and 2) studied the functional consequences of TIGIT blockade in an assay in which human polyclonal antigen-specific CD8+ CTLs were expanded and co-cultured with human tumor cell lines that endogenously express their cognate antigen. Methods Primary human tumor disaggregates were examined for expression of TIGIT, CD226, CD155 and CD112 by flow cytometry. To further examine the repressive function of TIGIT, CTLs from donor human PBMCs were expanded in the presence of a specific major histocompatibility class I (MHC I) peptide antigen and subsequently screened for enrichment of activation and exhaustion markers to confirm expansion. CTLs were then co-cultured with CD155+ and CD112+ tumor cell lines endogenously expressing the MHC I-binding peptide of interest, in the presence and absence of TIGIT or PD1 blocking antibodies. To further investigate the mechanism of cytolysis, inhibitors of TCR signaling, cytoskeletal rearrangement, and cytokine receptor signaling were investigated. Summary of Results TIGIT blockade induced a rapid, potent, and reproducible cytolytic response in co-culture experiments. The maximal effect was similar to that observed with PD1 inhibition, and moreover the combination of PD1 and TIGIT blockade was greater than either monotherapy, implying non-redundancy of mechanism. Inhibitors of cytoskeletal function and signal transduction downstream of the TCR appeared to block TIGIT mediated cytotoxicity. Conclusion TIGIT has a clearly defined role in the repression of cytotoxic function in antigen-specific CTLs, in a manner non-redundant with PD1. The enhancement in cytotoxicity subsequent to TIGIT blockade is dependent on cytoskeletal function as well as additional signal transduction downstream of TCR engagement. Strategies to enhance these signal transduction events may be of further interest to potentiate TIGIT blocking therapy. Citation Format: Deepali Malhotra, Gordon Moody, Michael Overstreet, John Mumm. TIGIT blockade enhances cytolytic function in antigen-specific CTLs in a manner non-redundant to PD1 blockade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4720.

Expression and Function of Tigit in B-Cell Non-Hodgkin Lymphoma

Background:T cell Ig and ITIM domain (TIGIT) is an immune checkpoint molecule and a novel member of CD28 family. TIGIT is expressed on NK cells, effector and memory T cells, and Treg cells. Upon ligation with CD155, TIGIT delivers an inhibitory signal and negatively regulates anti-tumor responses. While important in normal T-cell biology, the expression and function of TIGIT in the blood and tumor microenvironment of patients with B-cell non-Hodgkin lymphoma (NHL) is completely unknown. Goal: To phenotypically and functionally characterize TIGIT+T cell subsets in B-cell NHL and compare expression of TIGIT on intratumoral T cells to normal controls. Results: In peripheral blood, TIGIT expression was not detected on resting T cells. In contrast, when we analyzed biopsy specimens from B-cell NHL patients, we observed that TIGIT is variably but highly expressed on a subset of intratumoral T cells. A median of 41.1% (range: 24.9-56.1, n=5) of CD4+ or 22.2% (range: 13.4-33.3, n=5) of CD8+ T cells from tumor specimens expressed TIGIT on the cell surface. In normal tonsil tissue, expression level of TIGIT was modest or negligible. A median of 12.0% (range: 9.1-15.8, n=5) of CD4+ or 3.9% (range: 1.3-5.7, n=5) of CD8+ T cells express TIGIT on the cell surface, which is significantly lower than that in B-cell NHL (p=0.005 and p=0.003 for CD4+ and CD8+, respectively). We observed that both CD45RA+ and CD45RA- T cells express TIGIT with the greatest expression on the CD45RA- population in B-cell NHL specimens. Furthermore, we found that TIGIT+ T cells coexpressed other immune checkpoint molecules including PD-1 and TIM-3. Functionally, TIGIT+ T cells displayed reduced cytokine production, as the number of IFN-γ- and IL-2-producing cells was lower in the TIGIT+ population than in TIGIT-T cells. Unlike TIM-3 that is coexpressed with PD-1 and whose expression is upregulated by IL-12, TIGIT expression is not upregulated by IL-12, suggesting that a different mechanism is involved in TIGIT upregulation in B-cell NHL. Conclusion: Taken together, these results indicated that TIGIT is highly expressed on intratumoral T cells in B-cell NHL and is expressed on a population of T-cells with suppressed immune function. TIGIT, while co-expressed with PD-1 and TIM-3, is not upregulated by cytokines such as IL-12, suggesting that it is regulated by an alternate pathway. Inhibition of TIGIT signaling may be an additional mechanism to prevent T-cell suppression and exhaustion in B-cell NHL. Disclosures No relevant conflicts of interest to declare.

Identification of TIGIT as an HBZ-induced gene by genome-wide analyses: its association with evasion of host defense

Human T-cell leukemia virus type 1 (HTLV-1) is a causative virus of adult T-cell leukemia (ATL), and inflammatory diseases. HTLV-1 bZIP factor (HBZ) is encoded on the minus strand of HTLV-1, and expressed in all ATL cases. We performed RNA-seq and ChIP-seq using HBZ transduced T cells, and found that expressions of several Treg-related genes, including Foxp3, CD25, CCR4, CCR5, PD-1, NRP1, IKZF family genes, and T-cell immunoglobulin and ITIM domain receptor (Tigit), were upregulated by HBZ. We focused on TIGIT in this study. TIGIT expression was also upregulated in HTLV-1 infected cells in HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients and CD4+ T cells of ATL patients. HBZ induced the histone modification of the promoter of TIGIT and enhanced its expression. HBZ also enhanced acetylation of H3K18, which is the specific target of p3. HBZ augments transcription of Tigit gene in the presence of PMA/Ionomycin stimulation. Although the detailed mechanism of HBZ mediated activation of Tigit transcription is still unclear, p3 and activated pathway by PMA/Ionomycin is likely involved in its regulation. Furthermore, we analyzed the function of Tigit in HBZ-expressing T cells and CD4+ T cells from HBZ-transgenic mice (HBZ-Tg), and found that stimulation of Tigit with its ligand, PVR, enhanced expression of inhibitory cytokines, such as IL-10, in HBZ-Tg. These results suggest that the enhanced Tigit expression by HBZ may play a key role to modulate the microenvironment where anti-tumor immune response is attenuated by increased expression of IL-10. This environment seems to be associated with the evasion of HTLV-1 infected cells from anti-tumor host immune response and the pathogenesis of HTLV-1 associated inflammatory diseases and ATL.