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Mouse TIGIT inhibits NK-cell cytotoxicity upon interaction with PVR.

ABSTRACT: The activity of natural killer (NK) cells is controlled by a balance of signals derived from inhibitory and activating receptors. TIGIT is a novel inhibitory receptor, recently shown in humans to interact with two ligands: PVR and Nectin2 and to inhibit human NK-cell cytotoxicity. Whether mouse TIGIT (mTIGIT) inhibits mouse NK-cell cytotoxicity is unknown. Here we show that mTIGIT is expressed by mouse NK cells and interacts with mouse PVR. Using mouse and human Ig fusion proteins we show that while the human TIGIT (hTIGIT) cross-reacts with mouse PVR (mPVR), the binding of mTIGIT is restricted to mPVR. We further demonstrate using surface plasmon resonance (SPR) and staining with Ig fusion proteins that mTIGIT binds to mPVR with higher affinity than the co-stimulatory PVR-binding receptor mouse DNAM1 (mDNAM1). Functionally, we show that triggering of mTIGIT leads to the inhibition of NK-cell cytotoxicity, that IFN-γ secretion is enhanced when mTIGIT is blocked and that the TIGIT-mediated inhibition is dominant over the signals delivered by the PVR-binding co-stimulatory receptors. Additionally, we identify the inhibitory motif responsible for mTIGIT inhibition. In conclusion, we show that TIGIT is a powerful inhibitory receptor for mouse NK cells.

SUBMITTER: Stanietsky N

PROVIDER: S-EPMC3863769 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Mouse TIGIT inhibits NK-cell cytotoxicity upon interaction with PVR.

Stanietsky Noa N Rovis Tihana Lenac TL Glasner Ariella A Seidel Einat E Tsukerman Pinchas P Yamin Rachel R Enk Jonatan J Jonjic Stipan S Mandelboim Ofer O

European journal of immunology 20130704 8

The activity of natural killer (NK) cells is controlled by a balance of signals derived from inhibitory and activating receptors. TIGIT is a novel inhibitory receptor, recently shown in humans to interact with two ligands: PVR and Nectin2 and to inhibit human NK-cell cytotoxicity. Whether mouse TIGIT (mTIGIT) inhibits mouse NK-cell cytotoxicity is unknown. Here we show that mTIGIT is expressed by mouse NK cells and interacts with mouse PVR. Using mouse and human Ig fusion proteins we show that w ...[more]

PMID: 23677581

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Project description:BackgroundInhibitors targeting immune checkpoint were proved effective in cancer immunotherapy, such as PD-1/PD-L1 blockade. The novel immune checkpoint TIGIT/PVR plays critical roles in suppressing the anti-tumor effects of CD8+ T and NK cells, and dual blockade of TIGIT/PVR and PD-1/PD-L1 by antibody can elicit synergistic effects in tumor models and clinical trials. However, small molecules for TIGIT/PVR blockade have not been investigated.MethodsThe expression of PVR in tumors were analyzed by using TCGA, Oncomine and GEO database, and in cancer cell lines examined by flow cytometry. Natural product compounds were docked to PVR for virtual screening by using the software Molecular Operating Environment (MOE). Candidate compounds were further tested by biolayer interferometry-based binding assay, microscale thermophoresis assay and cell based blocking assay. The in vitro activity of the candidate compound was determined by MTT, peripheral blood mononuclear cells (PBMCs) activation assay and coculture assay. The anti-tumor effects and mechanism were also investigated by using MC38 tumor-bearing mice model and immune cell depletion tumor model.ResultsPVR was over-expressed in many tumor tissues and cancer cell lines, making it a promising therapeutic target. Through virtual screening, binding, and blocking assay, liothyronine was discovered to bind PVR and block the interaction of TIGIT/PVR. Liothyronine could enhance the function of CD4+ and CD8+ T cells in PBMCs. Besides, in the Jurkat-hTIGIT and CHOK1-hPVR coculture assay, liothyronine could reverse the IL-2 secretion inhibition resulted by TIGIT/PVR ligation. Although had no influence on the proliferation of tumor cells in vitro, liothyronine could significantly inhibit tumor growth when administrated in vivo, by enhancing CD8+ T cell infiltration and immune responses in the tumor bearing mice. The immune cell depletion model showed that the anti-tumor effects of liothyronine depends on CD4+ T cells, CD8+ T cells and NK cells.ConclusionsA small molecule liothyronine was discovered to serve as a potential candidate for cancer immunotherapy by blocking the immune checkpoint TIGIT/PVR. Video abstract.

Project description:The latent human immunodeficiency virus (HIV) reservoir presents the biggest obstacle to curing HIV chronic infection. Consequently, finding novel strategies to control the HIV reservoir is critical. Natural killer (NK) cells are essential for antiviral immunity. However, the influence of NK cell subsets and their associated inhibitory or activating receptors on their cytotoxicity toward the HIV reservoir has not been fully studied. We investigated the relationship between the percentage of NK cells or NK cell subsets and the HIV reservoir. Our results indicated that the percentage of CD56-CD16+ NK cells was positively associated with HIV reservoir size (i.e., HIV DNA, HIV msRNA, or HIV usRNA). Additionally, we observed that the percentage of IFN-γ+ NK cells was inversely related to the HIV reservoir. Furthermore, the expression of TIGIT on NK cells, particularly CD56-CD16+ and CD56dim NK cell subsets, positively correlated with the HIV reservoir. Notably, individuals with higher percentage of TIGIT+ NK and lower percentage of CD226+ NK cells exhibited larger HIV reservoir. Mechanistically, we discovered that TIGIT could inhibit the PI3K-Akt-mTOR-mTORC1 (s6k) signaling pathway to decrease the production of IFN-γ in NK cells. Importantly, inhibiting TIGIT in NK cells enhanced their ability to eliminate reactivated latently infected CD4+ T cells. Our experiments underscored the crucial role of NK cells in controlling the HIV reservoir and suggested that TIGIT serves as a promising target for enhancing the NK cell-mediated clearance of the HIV reservoir.ImportanceAs a major barrier to human immunodeficiency virus (HIV) cure, HIV reservoir persist in viremia-suppressed infected individuals. NK cells are important antiviral cells, and their impact on reservoir has rarely been reported. We analyzed the relationship between the size of reservoir and NK cell subsets, inhibitory receptor TIGIT expression. Our analysis found that the percentage of CD56-CD16+ NK cells was positively associated with HIV reservoir size. Furthermore, TIGIT expression on NK cells and CD56-CD16+ NK cells or CD56dim NK cells has a positive correlation with the HIV reservoir. TIGIT can inhibit the PI3K-Akt-mTOR-mTORC1 (s6k) signaling pathway to decrease the production of IFN-γ on NK cells. Blocking TIGIT in NK cells can enhance their ability to eliminate reactivated latently infected CD4+ T cells. Our study indicated that NK cells are critical to the control of the reservoir size, and TIGIT may be a target for enhancing the NK cell-mediated elimination of the reservoir.

Project description:Treatments targeting TIGIT have gained a lot of attention due to strong preclinical and early clinical results, particularly with anti-PD-(L)1 therapeutics. However, this combination has failed to meet progression-free survival endpoints in phase III trials. Most of our understanding of TIGIT comes from studies of T cell function. Yet, this inhibitory receptor is often upregulated to the same, or higher, extent on NK cells in cancers. Studies in murine models have demonstrated that TIGIT inhibits NK cells and promotes exhaustion, with its effects on tumor control also being dependent on NK cells. However, there are limited studies assessing the role of TIGIT on the function of human NK cells (hNK), particularly in lung cancer. Most studies used NK cell lines or tested TIGIT blockade to reactivate exhausted cells obtained from cancer patients. For therapeutic advancement, a better understanding of TIGIT in the context of activated hNK cells is crucial, which is different than exhausted NK cells, and critical in the context of adoptive NK cell therapeutics that may be combined with TIGIT blockade. In this study, the effect of TIGIT blockade on the anti-tumor activities of human ex vivo-expanded NK cells was evaluated in vitro in the context of lung cancer. TIGIT expression was higher on activated and/or expanded NK cells compared to resting NK cells. More TIGIT+ NK cells expressed major activating receptors and exerted anti-tumor response as compared to TIGIT- cells, indicating that NK cells with greater anti-tumor function express more TIGIT. However, long-term TIGIT engagement upon exposure to PVR+ tumors downregulated the cytotoxic function of expanded NK cells while the inclusion of TIGIT blockade increased cytotoxicity, restored the effector functions against PVR-positive targets, and upregulated immune inflammation-related gene sets. These combined results indicate that TIGIT blockade can preserve the activation state of NK cells during exposure to PVR+ tumors. These results support the notion that a functional NK cell compartment is critical for anti-tumor response and anti-TIGIT/adoptive NK cell combinations have the potential to improve outcomes.

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Mouse TIGIT inhibits NK-cell cytotoxicity upon interaction with PVR.

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Mouse TIGIT inhibits NK-cell cytotoxicity upon interaction with PVR.

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