scholarly journals GABAA receptor α4-subunit knockout enhances lung inflammation and airway reactivity in a murine asthma model

2017 ◽  
Vol 313 (2) ◽  
pp. L406-L415 ◽  
Author(s):  
Gene T. Yocum ◽  
Damian L. Turner ◽  
Jennifer Danielsson ◽  
Matthew B. Barajas ◽  
Yi Zhang ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Immune Cells ◽  
Cell Activation ◽  
Cell Function ◽  
Immune Cell ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Asthma Model

Emerging evidence indicates that hypnotic anesthetics affect immune function. Many anesthetics potentiate γ-aminobutyric acid A receptor (GABAAR) activation, and these receptors are expressed on multiple subtypes of immune cells, providing a potential mechanistic link. Like immune cells, airway smooth muscle (ASM) cells also express GABAARs, particularly isoforms containing α4-subunits, and activation of these receptors leads to ASM relaxation. We sought to determine if GABAAR signaling modulates the ASM contractile and inflammatory phenotype of a murine allergic asthma model utilizing GABAAR α4-subunit global knockout (KO; Gabra40/0) mice. Wild-type (WT) and Gabra4 KO mice were sensitized with house dust mite (HDM) antigen or exposed to PBS intranasally 5 days/wk for 3 wk. Ex vivo tracheal rings from HDM-sensitized WT and Gabra4 KO mice exhibited similar magnitudes of acetylcholine-induced contractile force and isoproterenol-induced relaxation ( P = not significant; n = 4). In contrast, in vivo airway resistance (flexiVent) was significantly increased in Gabra4 KO mice ( P < 0.05, n = 8). Moreover, the Gabra4 KO mice demonstrated increased eosinophilic lung infiltration ( P < 0.05; n = 4) and increased markers of lung T-cell activation/memory (CD62L low, CD44 high; P < 0.01, n = 4). In vitro, Gabra4 KO CD4+ cells produced increased cytokines and exhibited increased proliferation after stimulation of the T-cell receptor as compared with WT CD4+ cells. These data suggest that the GABAAR α4-subunit plays a role in immune cell function during allergic lung sensitization. Thus GABAAR α4-subunit-specific agonists have the therapeutic potential to treat asthma via two mechanisms: direct ASM relaxation and inhibition of airway inflammation.

scholarly journals IMMU-46. EXAMINATION OF THE EFFECTS OF DEXAMETHASONE ON THE EFFICACY OF IMMUNOTHERAPY IN GLIOMA USING GENE-MEDIATED CYTOTOXIC IMMUNOTHERAPY

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi129-vi129
Author(s):  
Marilin Koch ◽  
Mykola Zdioruk ◽  
M Oskar Nowicki ◽  
Estuardo Aguilar ◽  
Laura Aguilar ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cell Function ◽  
Immune Cell ◽  
Symptomatic Treatment ◽  
Tumor Cell Death ◽  
The Impact

Abstract RATIONALE Dexamethasone is frequently used in symptomatic treatment of glioma patients, although it is known to cause immune suppression. Checkpoint inhibitor immunotherapies have not yet been successful in glioma treatments. Gene-mediated cytotoxic immunotherapy (GMCI) is an immunotherapeutic approach that uses aglatimagene besadenovec with an anti-herpetic prodrug to induce immunogenic tumor cell death and immune cell attraction to the tumor site with potent CD8 T cell activation. GMCI is currently in clinical trials for solid tumors including glioblastoma, where it showed encouraging survival results in a Phase 2 study that did not limit the use of dexamethasone. However, the effects of dexamethasone on its efficacy have not been explored. METHODS We investigated the effects of dexamethasone on GMCI in vitro using cytotoxicity and T-cell-killing assays in glioblastoma cell lines. The impact of dexamethasone in vivo was assessed in an orthotopic syngeneic murine glioblastoma model. RESULTS Cyotoxicity assays showed that Dexamethasone has a slight impact on GMCI in vitro. In contrast, we observed a highly significant effect in T-cell-functional assays in which killing was greatly impaired. Immune cell response assays revealed a reduced T-cell proliferation after co-culture with supernatant from dexamethasone or combination treated glioblastoma cells in contrast to GMCI alone. In a murine model, the combination of GMCI and dexamethasone resulted in a significant reduction in median symptom-free survival (29d) in comparison to GMCI alone (39.5d) (P = 0.0184). CONCLUSION Our data suggest that high doses of dexamethasone may negatively impact the efficacy of immunotherapy for glioma, which may be a consequence of impaired T cell function. These results support the idea that there is a need in identifying possible alternatives to dexamethasone to maximize the effectiveness of immunostimulatory therapies such as GMCI.

scholarly journals 886 Targeting VSIG4, a novel macrophage checkpoint, repolarizes suppressive macrophages which induces an inflammatory response in primary cell in vitro assays and fresh human tumor cultures

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A928-A928
Author(s):  
Steve Sazinsky ◽  
Phuong Nguyen ◽  
Mohammad Zafari ◽  
Ryan Phennicie ◽  
Joe Wahle ◽  
...  
Keyword(s):  
Lung Cancer ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Immune Cell ◽  
Ex Vivo ◽  
Syngeneic Mouse ◽  
Pathogen Clearance

BackgroundVSIG4 (V-set immunoglobulin-domain-containing 4) is a B7 family related protein with known roles as a complement receptor involved in pathogen clearance as well as a negative regulator of T cell activation by an undetermined mechanism.1–3 VSIG4 is expressed in tumor associated macrophages (TAMs) with exquisite specificity. In cancer, increased expression of VSIG4 has been associated with worse survival in multiple indications, including non-small cell lung cancer, multiple myeloma, ovarian cancer, and glioma, suggesting an important role in tumor immune evasion.3–6 Based upon computational analysis of transcript data across thousands of primary cancer and normal tissue samples, we hypothesized that VSIG4 has an important regulatory role in promoting M2-like immune suppressive macrophages in the tumor microenvironment, and that targeting VSIG4 via a monoclonal antibody could relieve VSIG4-mediated macrophage suppression by repolarizing TAMs to an inflammatory phenotype capable of coordinating an anti-tumor immune response.MethodsThe ability of anti-VSIG4 antibodies to repolarize M2-like macrophages and induce T cell activation was assessed in vitro and ex vivo, by measuring production of inflammatory mediators. In vitro assays were performed primarily with M-CSF plus IL-10 driven monocyte-derived M2c macrophages from healthy donors. Ex vivo assays were performed with fresh, patient-derived tumor samples in culture. To determine whether targeting VSIG4 can lead to an anti-tumor effect in vivo, syngeneic mouse models were dosed with anti-mouse VSIG4 antibodies and characterized for changes in tumor volume and immune cell populations.ResultsIn in vitro and ex vivo assays anti-VSIG4 antibodies repolarize M2 macrophages and induce an immune response culminating in T cell activation. Targeting VSIG4 upregulates pro-inflammatory cytokines in M2c macrophages, as well as upregulates pro-inflammatory myeloid-derived cytokines and T cell-derived cytokines in M2c macrophages co-cultured with autologous T cells in the presence of staphylococcal enterotoxin B (SEB) activation. To assess targeting VSIG4 in a relevant translational model, fresh, patient-derived tumor samples were treated ex vivo with anti-VSIG4. Across multiple tumor types, anti-VSIG4 treatment resulted in a significant upregulation of cytokines involved in TAM repolarization and T cell activation, and chemokines involved in immune cell recruitment, at levels greater than observed by treatment with anti-PD-1 or a clinical macrophage repolarizing agent (anti-ILT-4). In vivo, tumor growth inhibition is observed in syngeneic mouse models dosed with anti-mouse-VSIG4 alone and in combination with anti-PD-1.ConclusionsTaken together, these data suggest that VSIG4 represents a promising new target capable of stimulating an anti-cancer response via multiple key immune mechanisms.Referencesvan Lookeren Campagne M, Verschoor A. Pathogen clearance and immune adherence “revisited”: immuno-regulatory roles for CRIg. Semin Immunol 2018;37:4–11.Xu S, Sun Z, Li L, Liu J, He J, Song D, Shan G, Liu H, Wu X. Induction of T cells suppression by dendritic cells transfected with VSIG4 recombinant adenovirus. Immunol Lett 2010;128(1):46–50.Liao Y, Guo S, Chen Y, Cao D, Xu H, Yang C, Fei L, Ni B, Ruan Z. VSIG4 expression on macrophages facilitates lung cancer development. Lab Invest 2014;94(7):706–715.Roh J, Jeon Y, Lee A, Lee S, Kim Y, Sung C, Park C, Hong J, Yoon D, Suh C, Huh J, Choi I, Park C. The immune checkpoint molecule V-set Ig domain-containing 4 is an independent prognostic factor for multiple myeloma. Oncotarget 2017;8(35):58122–58132.Xu T, Jiang Y, Yan Y, Wang H, Lu C, Xu H, Li W, Fu D, Lu Y, Chen J. VSIG4 is highly expressed and correlated with poor prognosis of high-grade glioma patients. Am J Transl Res 2015;7(6):1172–1180.Byun J, Jeong D, Choi I, Lee D, Kang M, Jung K, Jeon Y, Kim Y, Jung E, Lee K, Sung M, Kim K. The significance of VSIG4 expression in ovarian cancer. Int J Gynecol Cancer 2017;27(5):872–878.Ethics ApprovalAll legal and ethical requirements were met with regards to the humane treatment of animals described in the study. The animal study was conducted in compliance with CRL IACUC under IACUC No. I033.

scholarly journals CD14 Expressing Precursors Give Rise to Highly Functional Conventional Dendritic Cells for Use as Dendritic Cell Vaccine

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3818
Author(s):  
Maud Plantinga ◽  
Denise A. M. H. van den Beemt ◽  
Ester Dünnebach ◽  
Stefan Nierkens
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
Cord Blood ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ex Vivo ◽  
Dendritic Cell Vaccine ◽  
Cell Vaccine ◽  
Activated T Cells

Induction of long-lasting immunity by dendritic cells (DCs) makes them attractive candidates for anti-tumor vaccination. Although DC vaccinations are generally considered safe, clinical responses remain inconsistent in clinical trials. This initiated studies to identify subsets of DCs with superior capabilities to induce effective and memory anti-tumor responses. The use of primary DCs has been suggested to overcome the functional limitations of ex vivo monocyte-derived DCs (moDC). The ontogeny of primary DCs has recently been revised by the introduction of DC3, which phenotypically resembles conventional (c)DC2 as well as moDC. Previously, we developed a protocol to generate cDC2s from cord blood (CB)-derived stem cells via a CD115-expressing precursor. Here, we performed index sorting and single-cell RNA-sequencing to define the heterogeneity of in vitro developed DC precursors and identified CD14+CD115+ expressing cells that develop into CD1c++DCs and the remainder cells brought about CD123+DCs, as well as assessed their potency. The maturation status and T-cell activation potential were assessed using flow cytometry. CD123+DCs were specifically prone to take up antigens but only modestly activated T-cells. In contrast, CD1c++ are highly mature and specialized in both naïve as well as antigen-experienced T-cell activation. These findings show in vitro functional diversity between cord blood stem cell-derived CD123+DC and CD1c++DCs and may advance the efficiency of DC-based vaccines.

scholarly journals TRPM2 Is Not Required for T-Cell Activation and Differentiation

2022 ◽  
Vol 12 ◽  
Author(s):  
Niels C. Lory ◽  
Mikolaj Nawrocki ◽  
Martina Corazza ◽  
Joanna Schmid ◽  
Valéa Schumacher ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Transient Receptor Potential ◽  
Cell Function ◽  
Intestinal Inflammation ◽  
Activation Markers

Antigen recognition by the T-cell receptor induces a cytosolic Ca2+ signal that is crucial for T-cell function. The Ca2+ channel TRPM2 (transient receptor potential cation channel subfamily M member 2) has been shown to facilitate influx of extracellular Ca2+ through the plasma membrane of T cells. Therefore, it was suggested that TRPM2 is involved in T-cell activation and differentiation. However, these results are largely derived from in vitro studies using T-cell lines and non-physiologic means of TRPM2 activation. Thus, the relevance of TRPM2-mediated Ca2+ signaling in T cells remains unclear. Here, we use TRPM2-deficient mice to investigate the function of TRPM2 in T-cell activation and differentiation. In response to TCR stimulation in vitro, Trpm2-/- and WT CD4+ and CD8+ T cells similarly upregulated the early activation markers NUR77, IRF4, and CD69. We also observed regular proliferation of Trpm2-/- CD8+ T cells and unimpaired differentiation of CD4+ T cells into Th1, Th17, and Treg cells under specific polarizing conditions. In vivo, Trpm2-/- and WT CD8+ T cells showed equal specific responses to Listeria monocytogenes after infection of WT and Trpm2-/- mice and after transfer of WT and Trpm2-/- CD8+ T cells into infected recipients. CD4+ T-cell responses were investigated in the model of anti-CD3 mAb-induced intestinal inflammation, which allows analysis of Th1, Th17, Treg, and Tr1-cell differentiation. Here again, we detected similar responses of WT and Trpm2-/- CD4+ T cells. In conclusion, our results argue against a major function of TRPM2 in T-cell activation and differentiation.

scholarly journals Signaling function of PRC2 is essential for TCR-driven T cell responses

2018 ◽  
Vol 215 (4) ◽  
pp. 1101-1113 ◽  
Author(s):  
Marc-Werner Dobenecker ◽  
Joon Seok Park ◽  
Jonas Marcello ◽  
Michael T. McCabe ◽  
Richard Gregory ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Therapeutic Potential ◽  
Cell Types ◽  
Polycomb Repressive Complex 2 ◽  
Histone Lysine Methyltransferases

Differentiation and activation of T cells require the activity of numerous histone lysine methyltransferases (HMT) that control the transcriptional T cell output. One of the most potent regulators of T cell differentiation is the HMT Ezh2. Ezh2 is a key enzymatic component of polycomb repressive complex 2 (PRC2), which silences gene expression by histone H3 di/tri-methylation at lysine 27. Surprisingly, in many cell types, including T cells, Ezh2 is localized in both the nucleus and the cytosol. Here we show the presence of a nuclear-like PRC2 complex in T cell cytosol and demonstrate a role of cytosolic PRC2 in T cell antigen receptor (TCR)–mediated signaling. We show that short-term suppression of PRC2 precludes TCR-driven T cell activation in vitro. We also demonstrate that pharmacological inhibition of PRC2 in vivo greatly attenuates the severe T cell–driven autoimmunity caused by regulatory T cell depletion. Our data reveal cytoplasmic PRC2 is one of the most potent regulators of T cell activation and point toward the therapeutic potential of PRC2 inhibitors for the treatment of T cell–driven autoimmune diseases.

T-cell redirected killing and cure of pancreatic and gastric cancer xenografts by targeting Trop-2.

2015 ◽  
Vol 33 (3_suppl) ◽  
pp. 262-262
Author(s):  
David M. Goldenberg ◽  
Edmund A. Rossi ◽  
Diane L Rossi ◽  
Thomas M. Cardillo ◽  
Chien-Hsing Chang
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ex Vivo ◽  
Control Group ◽  
Calcium Signal ◽  
Target Cells ◽  
Normal Tissues

262 Background: Trop-2 [also called tumor-associated calcium signal transducer 2 (TACSTD2), EGP-1 (epithelial glycoprotein-1), GA733-1, or M1S1]is a 35 kDa transmembrane glycoprotein that is overexpressed relative to normal tissues in a variety of human cancers, including pancreatic and gastric carcinomas, where increased expression correlates with poor prognosis. Trop-2 appears to be more tumor-specific than the related molecule, EpCAM (Trop-1). MT110, the EpCAM antibody x CD3 bispecific T-cell engager (BiTE), is currently undergoing a Phase I study in various solid tumors, including lung, gastric, colorectal, breast, prostate, and ovarian cancers. We produced a similar T-cell redirecting bispecific tandem scFv, E1-3, using the variable domains of hRS7 (humanized anti-Trop-2 mAb) and Okt-3 (anti-CD3 mAb). Methods: T-cell activation, cytokine induction and cytotoxicity were evaluated ex vivo using PBMCs or purified T cells with human pancreatic (Capan-1 and BxPC3) and gastric (NCI-N87) cancer cell lines as target cells. In vivo activity was assayed with NCI-N87 xenografts that were inoculated s.c. in a mixture with twice the number of human PBMCs and matrigel. Results: In the presence of target cells and PBMCs, E1-3 potently induced T-cell activation, proliferation, and dose-dependent cytokine production of IL-2 (>2 ng/mL), IL-6 (>1 ng/mL), IL-10 (>7 ng/mL), TNF-α (>1 ng/mL) and IFN-γ (>50 ng/mL). In vitro, E1-3 mediated a highly potent T-cell lysis of BxPC3 [IC50=0.09(±0.04) pM], Capan-1 [IC50=1.2(±1.1) pM] and NCI-N87 [IC50=1.2(±1.2) pM] target cells. In vivo, two 50-µg doses of E1-3 given three days apart cured all of the mice (N=8) bearing NCI-N87 xenografts (P=0.0005; Log-Rank). Tumors in the control group (PBMCs only) reached the endpoint (TV>1 cm3) with a median of 39.5 days. All mice remained tumor-free in the E1-3 group at 78 days. Conclusions: Trop-2 is an attractive target for T-cell-mediated killing of pancreatic, gastric and other epithelial cancers.

scholarly journals A high-affinity human TCR-like antibody detects celiac disease gluten peptide–MHC complexes and inhibits T cell activation

2021 ◽  
Vol 6 (62) ◽  
pp. eabg4925
Author(s):  
Rahel Frick ◽  
Lene S. Høydahl ◽  
Jan Petersen ◽  
M. Fleur du Pré ◽  
Shraddha Kumari ◽  
...  
Keyword(s):  
Celiac Disease ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Therapeutic Potential ◽  
Human Leukocyte ◽  
Fab Fragment ◽  
Biopsy Material ◽  
Human T Cell

Antibodies specific for peptides bound to human leukocyte antigen (HLA) molecules are valuable tools for studies of antigen presentation and may have therapeutic potential. Here, we generated human T cell receptor (TCR)–like antibodies toward the immunodominant signature gluten epitope DQ2.5-glia-α2 in celiac disease (CeD). Phage display selection combined with secondary targeted engineering was used to obtain highly specific antibodies with picomolar affinity. The crystal structure of a Fab fragment of the lead antibody 3.C11 in complex with HLA-DQ2.5:DQ2.5-glia-α2 revealed a binding geometry and interaction mode highly similar to prototypic TCRs specific for the same complex. Assessment of CeD biopsy material confirmed disease specificity and reinforced the notion that abundant plasma cells present antigen in the inflamed CeD gut. Furthermore, 3.C11 specifically inhibited activation and proliferation of gluten-specific CD4+ T cells in vitro and in HLA-DQ2.5 humanized mice, suggesting a potential for targeted intervention without compromising systemic immunity.

scholarly journals Cepharanthine Blocks Presentation of Thyroid and Islet Peptides in a Novel Humanized Autoimmune Diabetes and Thyroiditis Mouse Model

2021 ◽  
Vol 12 ◽  
Author(s):  
Cheuk Wun Li ◽  
Roman Osman ◽  
Francesca Menconi ◽  
Larissa C. Faustino ◽  
Kookjoo Kim ◽  
...  
Keyword(s):  
T Cell ◽  
Mouse Model ◽  
Small Molecules ◽  
T Cell Activation ◽  
Cell Activation ◽  
Therapeutic Potential ◽  
Autoimmune Diabetes ◽  
Autoimmune Response ◽  
Hla Dr3

Autoimmune polyglandular syndrome type 3 variant (APS3v) refers to an autoimmune condition in which both type 1 diabetes (T1D) and autoimmune thyroiditis (AITD) develop in the same individual. HLA-DR3 confers the strongest susceptibility to APS3v. Previously we reported a unique amino acid signature pocket that predisposes to APS3v. We found that this pocket is flexible and can trigger APS3v by presenting both thyroid (Tg.1571, TPO.758) and islet (GAD.492) peptides to induce autoimmune response. We hypothesized that blocking the specific APS3v-HLA-DR3 pocket from presenting thyroid/islet antigens can block the autoimmune response in APS3v. To test this hypothesis we performed a virtual screen of small molecules blocking APS3v-HLA-DR3, and identified 11 small molecules hits that were predicted to block APS3v-HLA-DR3. Using the baculovirus-produced recombinant APS3v-HLA-DR3 protein we tested the 11 small molecules in an in vitro binding assay. We validated 4 small molecule hits, S9, S5, S53 and S15, that could block the APS3v-HLA-DR3 pocket in vitro. We then developed a novel humanized APS3v mouse model induced by co-immunizing a peptide mix of Tg.1571, TPO.758 and GAD.492. The immunized mice developed strong T-cell and antibody responses to the thyroid/islet peptides, as well as mouse thyroglobulin. In addition, the mice showed significantly lower free T4 levels compared to controls. Using the APS3v mouse model, we showed that one of the 4 small molecules, Cepharanthine (S53), blocked T-cell activation by thyroid/islet peptides ex vivo and in vivo. These findings suggested Cepharanthine may have a therapeutic potential in APS3v patients carrying the specific APS3v-HLA-DR3 pocket.

scholarly journals Nanoengineering of Immune Cell Function

2009 ◽  
Vol 1209 ◽  
Author(s):  
Keyue Shen ◽  
Michael C Milone ◽  
Michael L. Dustin ◽  
Lance Cameron Kam
Keyword(s):  
T Cells ◽  
Immune System ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cell Function ◽  
Immune Cell ◽  
Cell Communication ◽  
Cell Therapies ◽  
Nano Scale

AbstractT lymphocytes are a key regulatory component of the adaptive immune system. Understanding how the micro- and nano-scale details of the extracellular environment influence T cell activation may have wide impact on the use of T cells for therapeutic purposes. In this article, we examine how the micro- and nano-scale presentation of ligands to cell surface receptors, including microscale organization and nanoscale mobility, influences the activation of T cells. We extend these studies to include the role of cell-generated forces, and the rigidity of the microenvironment, on T cell activation. These approaches enable delivery of defined signals to T cells, a step toward understanding the cell-cell communication in the immune system, and developing micro/nano- and material- engineered systems for tailoring immune responses for adoptive T cell therapies.

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