Human Gammadelta T-cells Research Articles

Crucial role of calcium signaling in TCR-dependent functions of Vgamma2Vdelta2 T cell (89.23)

Abstract Vgamma2Vdelta2 T cells in human peripheral blood recognize phosphoantigen and play important roles in host defense and immunoregulation. However, the molecular mechanisms responsible for phophogantigen-mediated activation of human gammadelta T-cells remain unclear. We have previously reported that phosphoantigen isopentyl pyrophosphate (IPP) activated MEK/Erk and PI-3K/Akt pathways rapidly. Specific inhibition of Erk and Akt significantly impaired the functional response to IPP. In the present study, we found that IL2 can also activate MEK/Erk and PI-3K/Akt pathways without inducing functional responses. When given together with IPP, the effect of IL2 was to enhance cytokines production, degranulation and cytotoxicity. These data implicate the activation of MEK/Erk and PI-3K/Akt pathways as necessary but not sufficient to induce functional responses in Vgamma2Vdelta2 T cells. Further, we found that FK506 or cyclosporine A (CsA) inhibitors of calcineurin, completely blocked IPP-induced cytokines production, and partly inhibited TCR-dependent degranulation and cytotoxicity functions of Vgamma2Vdelta2 T cells. IPP but not IL2 induced the calcium-dependent nuclear translocation of NFAT. Our results indicate that calcium signaling pathway plays a crucial role in TCR-dependent functions in Vgamma2Vdelta2 T cells that can be distinguished from the effect of IL2.

Highly Active Microbial Phosphoantigen Induces Rapid yet Sustained MEK/Erk- and PI-3K/Akt-Mediated Signal Transduction in Anti-Tumor Human γδ T-Cells

BackgroundThe unique responsiveness of Vγ9Vδ2 T-cells, the major γδ subset of human peripheral blood, to non-peptidic prenyl pyrophosphate antigens constitutes the basis of current γδ T-cell-based cancer immunotherapy strategies. However, the molecular mechanisms responsible for phosphoantigen-mediated activation of human γδ T-cells remain unclear. In particular, previous reports have described a very slow kinetics of activation of T-cell receptor (TCR)-associated signal transduction pathways by isopentenyl pyrophosphate and bromohydrin pyrophosphate, seemingly incompatible with direct binding of these antigens to the Vγ9Vδ2 TCR. Here we have studied the most potent natural phosphoantigen yet identified, (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), produced by Eubacteria and Protozoa, and examined its γδ T-cell activation and anti-tumor properties.Methodology/Principal FindingsWe have performed a comparative study between HMB-PP and the anti-CD3ε monoclonal antibody OKT3, used as a reference inducer of bona fide TCR signaling, and followed multiple cellular and molecular γδ T-cell activation events. We show that HMB-PP activates MEK/Erk and PI-3K/Akt pathways as rapidly as OKT3, and induces an almost identical transcriptional profile in Vγ9+ T-cells. Moreover, MEK/Erk and PI-3K/Akt activities are indispensable for the cellular effects of HMB-PP, including γδ T-cell activation, proliferation and anti-tumor cytotoxicity, which are also abolished upon antibody blockade of the Vγ9+ TCR Surprisingly, HMB-PP treatment does not induce down-modulation of surface TCR levels, and thereby sustains γδ T-cell activation upon re-stimulation. This ultimately translates in potent human γδ T-cell anti-tumor function both in vitro and in vivo upon transplantation of human leukemia cells into lymphopenic mice,Conclusions/SignificanceThe development of efficient cancer immunotherapy strategies critically depends on our capacity to maximize anti-tumor effector T-cell responses. By characterizing the intracellular mechanisms of HMB-PP-mediated activation of the highly cytotoxic Vγ9+ T-cell subset, our data strongly support the usage of this microbial antigen in novel cancer clinical trials.

Expression of intercellular adhesion molecule (ICAM)‐1 or ICAM‐2 is critical in determining sensitivity of pancreatic cancer cells to cytolysis by human γδ‐T cells: Implications in the design of γδ‐T‐cell‐based immunotherapies for pancreatic cancer

gammadelta-T cells can recognize and kill malignant cells, particularly those of epithelial origin, through mechanisms which do not require the recognition of tumor-specific antigens (innate immune response). This natural ability of gammadelta-T cells to kill tumor cells in a tumor antigen-independent manner provides a strong rationale for developing clinical trials designed to exploit the innate antitumor properties of gammadelta-T cells. In vitro studies were carried out to asses the sensitivity of pancreatic cancer cells (MIA PaCa2, BxPC-3, PANC-1) to killing by ex vivo expanded human gammadelta-T cells. The capacity of gammadelta-T cells to bind to as well as to kill pancreatic cancer cells correlated with the degree of surface expression of key intercellular adhesion molecules (ICAM) present on pancreatic cancer cells. Moreover, pancreatic cancer cells expressing neither ICAM-1 nor ICAM-2 were bound poorly by gammadelta-T cells and were found to be resistant to gammadelta-T-cell killing. However, upon transfection of resistant cells with ICAM-1 or ICAM-2, gammadelta-T cells were then able to bind to and subsequently kill these cells. In vitro, the expression of ICAM-1 or ICAM-2 on human pancreatic cancer cells is critically important in determining the extent to which these cells are sensitive to killing by human gammadelta-T cells. Accordingly, in ongoing and future clinical studies using gammadelta-T cells for the treatment of a variety of epithelial-derived solid tumors-including pancreatic cancer-interventions intended to modulate ICAM expression on tumor cells may become important adjuncts to gammadelta-T-cell-based immunotherapies.

Metabolite antigens and pathway incompatibility.

Vgamma9Vdelta2 cells, which constitute a small portion of peripheral blood T-cells (approximately 5%), are known to be the biggest subset of human gammadelta T-cells ( approximately 70%) in circulating blood. The Vgamma9Vdelta2 T-cells expressing Vgamma9Vdelta2 T-cell receptors have an ability to recognize non-peptide antigens directly or indirectly, for example, phosphorylated metabolites referred to as phosphoantigens, synthesized aminobisphosphonates used for therapeutic purpose such as pamidronate, and alkylamines. These chemical compounds recognized by Vgamma9Vdelta2 T-cells are produced by many prokaryotic and eukaryotic organisms. Previous works show that the phosphoantigens recognized by Vgamma9Vdelta2 T-cells are found as the intermediates in the pathogens' pathway producing IPP. In this paper, we show that the other compounds recognized by Vgamma9Vdelta2 T-cells are found on the pathogens' biosynthetic pathways leading to production of shared compounds with human. In addition, many compounds having high structural similarity with alkylamine antigens are also found only in pathogen's biosynthetic pathways or produced only by non-human enzymes.

Innate Anti-breast Cancer Immunity of Apoptosis-resistant Human γδ-T cells

We previously identified a CD2-initiated signaling pathway which inhibits activation-induced cell death in mitogen-stimulated human gammadelta-T cells permitting the large-scale expansion of these cells. Here we report the innate anti-tumor activity of expanded human gammadelta-T cells against human breast cancer cells. Apoptosis-resistant human gammadelta-T cells which were expanded in vitro from cultured human peripheral blood mononuclear cells displayed lytic activity against breast cancer cell lines MDA-MB-231, MCF-7 and T-47D, but failed to kill normal human skin fibroblasts and normal human liver cells. Monoclonal antibodies (mAb) directed against the gammadelta-T cell receptor (TCR) or mAb directed against either the Vgamma9 or the Vdelta2 TCR chains were able to block gammadelta-T cell-mediated lysis of MDA-MB-231 cells. In addition, mAb against intercellular adhesion molecules-1 (ICAM-1/CD54) or CD18 (beta subunit of ICAM-1 counter-receptor) also blocked gammadelta-T cell-mediated killing of MDA-MB-231 cells. Ex vivo expanded human gammadelta-T cells are thus able to innately recognize and kill human breast cancer cells in a gammadelta-TCR-dependent manner; ICAM-1 and CD18 also appear to be involved in the interactions between sensitive breast cancer cells and cytolytic gammadelta-T cells. As apoptosis-resistant human gammadelta-T cells can now readily be expanded to large numbers (clinical scale), these findings must be considered in the context of developing adoptive immunotherapy strategies to exploit gammadelta-T cell innate immune responses for the primary or adjuvant treatment of breast cancer.

EX VIVO EXPANDED HUMAN Vλ9Vδ2+ λδ-T CELLS MEDIATE INNATE ANTITUMOR ACTIVITY AGAINST HUMAN PROSTATE CANCER CELLS IN VITRO

We have previously identified a CD2 mediated, interleukin-12 dependent signaling pathway that inhibits activation induced cell death in mitogen stimulated human gammadelta-T cells, permitting the large-scale expansion of these cells. Herein we report the innate antitumor activity of expanded human Vgamma9Vdelta2+ gammadelta-T cells against human prostate cancer cells. Apoptosis resistant human gammadelta-T cells were expanded in vitro from cultured human peripheral blood mononuclear cells and then enriched to high purity by immunomagnetic separation. In vitro cytotoxicity of expanded gammadelta-T cells was measured against human prostate cancer cell lines using standard cytotoxicity assays. gammadelta-T cells derived from various donors consistently showed lytic activity against the prostate cancer cell lines DU-145 and PC-3 but not LNCaP. mAbs against Vgamma9 or Vdelta2 T-cell receptor chains as well as mAb against intercellular adhesion molecule-1 (ICAM-1) or CD18, the beta subunit of ICAM-1 counter receptors, blocked gammadelta-T cell mediated killing of prostate cancer cells. gammadelta-T cells lysed prostate cancer cell lines largely through the perforin/granzyme pathway. Ex vivo, expanded human Vgamma9Vdelta2+ gammadelta-T cells are able innately to recognize and kill certain human prostate tumor cell lines in vitro. The recognition and killing of prostate cancer cells occurs in a gammadelta-T-cell receptor dependent manner and it also appears to involve interactions between ICAM-1 and CD18. Because apoptosis resistant human Vgamma9Vdelta2+ gammadelta-T cells can readily be expanded to large numbers (clinical scale), these findings must be considered in the context of developing adoptive immunotherapy strategies to exploit gammadelta-T cell innate immune responses to prostate cancer.

Contribution of Complementarity-Determining Region 3 of the T-Cell Receptor Vδ2 Chain to the Recognition of Aminobisphosphonates by Human γδ T-Cells.

Human gammadelta T-lymphocytes expressing Vgamma2Vdelta2 T-cell receptors (TCRs) can be stimulated by aminobisphosphonates and can kill certain tumor cells. Although germline-encoded lysine residues on the Jgamma1.2 segment have been demonstrated to be essential for the recognition of nonpeptide antigens by human gammadelta T-cells, the role of the junctional sequences of the TCR delta chain in the recognition of aminobisphosphonates by Vgamma2Vdelta2+ T-cells remains unknown. We examined the structure of complementarity-determining region 3 (CDR3) of Vdelta2 chains expressed by aminobisphosphonate-stimulated human gammadelta T-cells. CDR3 size-spectratyping analysis of Vdelta2 chains revealed that risedronate did not induce a CDR3 size distribution pattern of Vdelta2 cells that was distinct from that of Vdelta2 cells cultured without risedronate. The clonality of risedronate-expanded Vdelta2 T-cells was also determined by sequencing analysis, with the result that no particular consensus sequences were observed. However, most Vdelta2 T-cells freshly isolated from peripheral blood carried a distinctive junctional motif consisting of a hydrophobic amino acid residue (valine, leucine, or isoleucine [Val/Leu/Ile]) at conserved position 97, and this feature was not altered by risedronate stimulation. A significant proportion of Vdelta1 T-cells from the same individual had Leu at position 97, but Vdelta1 T-cells did not expand in response to risedronate. Moreover, Vdelta2 T-cells from the nonresponder against risedronate also carried a Val/Leu/Ile amino acid residue at position 97. These results suggest that the presence of a hydrophobic amino acid residue at position 97 in CDR3 of the TCR delta chain is not sufficient to account for the recognition of aminobisphosphonate by human gammadelta T-cells.

Human gammadelta-T cells in adoptive immunotherapy of malignant and infectious diseases.

Human gammadelta-T cells are capable of mediating both innate antitumor and antiviral activity, functions that theoretically might be exploitable in the treatment of a variety of malignant or infectious diseases. Nonetheless, experimental therapies incorporating the adoptive transter of human gammadelta-T cells have remained unfeasible to date owing largely to the difficulty of isolating or expanding sufficient numbers of gammadelta-T cells. It is in this context that recent discoveries from our laboratory are presented. By identifying specific signaling pathways that selectively inhibit activation-induced apoptosis in apoptosis-prone gammadelta-T cells, we have been able to expand large numbers of viable and functional human gammadelta-T cells, an undertaking until now notpossible. As important, these apoptosis-resistant gammadelta-Tcells appear to retain major histocompatibility complex-unrestricted (innate) antitumor activity against a wide variety of human tumor cells both in vitro and in vivo. Moreover, apoptosis-resistant gammadelta-T cells also display potent innate antiviral activity in vitro against human immunodeficiency virus-1. Both the biologic and practical implications of these findings are considered and discussed particularly as they relate to the development of future adoptive immunotherapy strategies.

Down-regulation of IL-2 receptor alpha (CD25) characterizes human gammadelta-T cells rendered resistant to apoptosis after CD2 engagement in the presence of IL-12.

We recently identified a CD2-mediated, IL-12-dependent signaling pathway that inhibits apoptosis in mitogen-stimulated human gammadelta-T cells. Here we show that gammadelta-T cells which acquire resistance to mitogen-induced apoptosis upregulate IL-12 receptor beta 1 subunit (IL-12Rbeta1); in contrast, gammadelta-T cells which remain sensitive to mitogen-induced apoptosis fail to express IL-12Rbeta1. Next we show that gammadelta-T cells which are rendered resistant to mitogen-induced apoptosis attenuate their expression of the IL-2 receptor alpha chain (IL-2Ralpha/CD25), this in part accounting for their acquired resistance to IL-2-induced death. In contrast, apoptosis-sensitive gammadelta-T cells are shown to persist in their expression of IL-2Ralpha/CD25, thus remaining sensitive to IL-2-induced death. Moreover, we show that apoptosis-resistant, but not apoptosis-sensitive, gammadelta-T cells display an enhanced responsiveness to IL-15, a finding in keeping with the known function of IL-15 as a growth and survival factor. Finally, we present evidence to suggest that this differential responsiveness to IL-15 occurs in part by the increased expression of the IL-15Ralpha chain on apoptosis-resistant gammadelta-T cells, compared to apoptosis-sensitive gammadelta-T cells. The biological and clinical implications of these findings are discussed.

Recognition of non-peptide antigens and the role of human gamma delta T-cells in innate and adaptive immunity

Recognition of non-peptide antigens and the role of human gamma delta T-cells in innate and adaptive immunity

Human gamma delta T-cells in the epithelium of the gut and in the inflamed synovial tissue preferentially express the V gamma 8 T-cell receptor chain.

Annals of the New York Academy of SciencesVolume 756, Issue 1 p. 406-409 Human γδ T-Cells in the Epithelium of the Gut and in the Inflamed Synovial Tissue Preferentially Express the Vγ8 T-Cell Receptor Chain KALLE SÖDERSTRÖM, KALLE SÖDERSTRÖM Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this authorANDERS BUCHT, ANDERS BUCHT Kabi Pharmacia Uppsala, SwedenSearch for more papers by this authorEVA HALAPI, EVA HALAPI Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this authorCARINA LUNDQVIST, CARINA LUNDQVIST Department of Immunology University of Umeå Umeå, SwedenSearch for more papers by this authorALVAR GRÖNBERG, ALVAR GRÖNBERG Kabi Pharmacia Uppsala, SwedenSearch for more papers by this authorETHEL NILSSON, ETHEL NILSSON Department of Rheumatology Karolinska Hospital Stockholm, SwedenSearch for more papers by this authorDANIELA L. M. ORSINI, DANIELA L. M. ORSINI Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorYVONNE van de WAL, YVONNE van de WAL Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorFRITS KONING, FRITS KONING Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorMARIE-LOUISE HAMMARSTRÖM, MARIE-LOUISE HAMMARSTRÖM Department of Immunology University of Umeå Umeå, SwedenSearch for more papers by this authorAND ROLF KIESSLING, AND ROLF KIESSLING Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this author KALLE SÖDERSTRÖM, KALLE SÖDERSTRÖM Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this authorANDERS BUCHT, ANDERS BUCHT Kabi Pharmacia Uppsala, SwedenSearch for more papers by this authorEVA HALAPI, EVA HALAPI Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this authorCARINA LUNDQVIST, CARINA LUNDQVIST Department of Immunology University of Umeå Umeå, SwedenSearch for more papers by this authorALVAR GRÖNBERG, ALVAR GRÖNBERG Kabi Pharmacia Uppsala, SwedenSearch for more papers by this authorETHEL NILSSON, ETHEL NILSSON Department of Rheumatology Karolinska Hospital Stockholm, SwedenSearch for more papers by this authorDANIELA L. M. ORSINI, DANIELA L. M. ORSINI Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorYVONNE van de WAL, YVONNE van de WAL Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorFRITS KONING, FRITS KONING Department of Immunohaematology and Blood Bank University Hospital Leiden, the NetherlandsSearch for more papers by this authorMARIE-LOUISE HAMMARSTRÖM, MARIE-LOUISE HAMMARSTRÖM Department of Immunology University of Umeå Umeå, SwedenSearch for more papers by this authorAND ROLF KIESSLING, AND ROLF KIESSLING Microbiology and Tumor Biology Center Karolinska Institute, Box 280 S-171 77, Stockholm, SwedenSearch for more papers by this author First published: July 1995 https://doi.org/10.1111/j.1749-6632.1995.tb44547.xCitations: 4AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume756, Issue1T‐Cell Receptor Use in Human Autoimmune DiseasesJuly 1995Pages 406-409 RelatedInformation