scholarly journals Graded RhoA GTPase Expression in Treg Cells Distinguishes Tumor Immunity From Autoimmunity

2021 ◽  
Vol 12 ◽  
Author(s):  
Khalid W. Kalim ◽  
Jun-Qi Yang ◽  
Vishnu Modur ◽  
Phuong Nguyen ◽  
Yuan Li ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Treg Cell ◽  
Tumor Immunity ◽  
Th17 Cell ◽  
Treg Cells ◽  
Effector T Cells ◽  
Conditional Knockout ◽  
Cell Plasticity ◽  
Cell Homeostasis

RhoA of the Rho GTPase family is prenylated at its C-terminus. Prenylation of RhoA has been shown to control T helper 17 (Th17) cell-mediated colitis. By characterizing T cell-specific RhoA conditional knockout mice, we have recently shown that RhoA is required for Th2 and Th17 cell differentiation and Th2/Th17 cell-mediated allergic airway inflammation. It remains unclear whether RhoA plays a cell-intrinsic role in regulatory T (Treg) cells that suppress effector T cells such as Th2/Th17 cells to maintain immune tolerance and to promote tumor immune evasion. Here we have generated Treg cell-specific RhoA-deficient mice. We found that homozygous RhoA deletion in Treg cells led to early, fatal systemic inflammatory disorders. The autoimmune responses came from an increase in activated CD4+ and CD8+ T cells and in effector T cells including Th17, Th1 and Th2 cells. The immune activation was due to impaired Treg cell homeostasis and increased Treg cell plasticity. Interestingly, heterozygous RhoA deletion in Treg cells did not affect Treg cell homeostasis nor cause systemic autoimmunity but induced Treg cell plasticity and an increase in effector T cells. Importantly, heterozygous RhoA deletion significantly inhibited tumor growth, which was associated with tumor-infiltrating Treg cell plasticity and increased tumor-infiltrating effector T cells. Collectively, our findings suggest that graded RhoA expression in Treg cells distinguishes tumor immunity from autoimmunity and that rational targeting of RhoA in Treg cells may trigger anti-tumor T cell immunity without causing autoimmune responses.

scholarly journals Targeting of Cdc42 GTPase in regulatory T cells unleashes anti-tumor T cell immunity

2021 ◽  
Author(s):  
Khalid W Kalim ◽  
Jun-Qi Yang ◽  
Mark Wunderlich ◽  
Vishnu Modur ◽  
Phuong Nguyen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Treg Cell ◽  
Cell Function ◽  
Treg Cells ◽  
T Cell Response ◽  
T Cell Immunity ◽  
Cell Plasticity ◽  
Effector T Cell ◽  
Cell Immunity

Regulatory T (Treg) cells play an important role in maintaining immune tolerance through inhibiting effector T cell function. In the tumor microenvironment, Treg cells are utilized by tumor cells to counteract effector T cell-mediated tumor killing. Targeting Treg cells may thus unleash the anti-tumor activity of effector T cells. While systemic depletion of Treg cells can cause excessive effector T cell responses and subsequent autoimmune diseases, controlled targeting of Treg cells may benefit cancer patients. Here we show that Treg cell-specific heterozygous deletion or pharmacological targeting of Cdc42 GTPase does not affect Treg cell numbers but induces Treg cell plasticity, leading to anti-tumor T cell immunity without detectable autoimmune reactions. Cdc42 targeting potentiates an immune checkpoint blocker anti-PD-1 antibody-mediated T cell response against mouse and human tumors. Mechanistically, Cdc42 targeting induces Treg cell plasticity and unleashes anti-tumor T cell immunity through carbonic anhydrase I-mediated pH changes. Thus, rational targeting of Cdc42 in Treg cells holds therapeutic promises in cancer immunotherapy.

scholarly journals P038 Microbial regulation of T-cell fate towards regulatory profiles during T-cell transfer induced colitis

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S150-S151
Author(s):  
A Machicote ◽  
P Pelczar ◽  
M Nawrocki ◽  
A Fazio ◽  
B Liu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antibiotic Treatment ◽  
Cd4 T Cells ◽  
Th17 Cells ◽  
Treg Cells ◽  
Effector T Cells ◽  
Cell Transfer ◽  
Cell Plasticity ◽  
T Cell Transfer

Abstract Background During Inflammatory Bowel Diseases (IBD), CD4+ effector T cells are main mediators of the tissue damage. Among them, Th17 cells strongly contribute to the inflammatory response. Interestingly, our lab previously showed that Th17 cells can convert into regulatory T cells, thereby controlling inflammation. However, the forces controlling the plasticity of T cells during IBD remain largely unknown. Our aim is to understand, how CD4+ T-cell plasticity can be modulated from a pro-inflammatory towards an anti-inflammatory profile during IBD. It is currently known that both Th17 and Foxp3 Treg cells can recognize microbiota-derived antigens and that changes in the microbiota are commonly observed in IBD. We hypothesize that the microbiota is a key candidate to modulate T-cell plasticity. Methods T-cell transfer mouse IBD model was performed by transferring CD4+ naïve T-cells into Rag1 -/- receptors. To study Th17 T-cell plasticity, we used as donors IL-17A Fate-mapping mice. These mice comprise of an IL-17ACRE/R26fl/fl YFP construct where the cells that previously expressed IL-17A turn into YFP+ cells. Ciprofloxacin and metronidazole, two antibiotics commonly given to IBD patients, were used to treat the mice. Stool microbiota composition was analysed longitudinally by 16S rRNA amplicon sequencing. Colitis development was evaluated by colonoscopy. CD4+ T cells were isolated from the colon and reporter expression was analysed by Flow Cytometry. Results After T-cell transfer colitis induction, ciprofloxacin and metronidazole treatment alleviated gut inflammation, as determined by weight loss (p=0.005), colitis score (p=0.004) and colon length (p=0.01). Antibiotic treatment increased the relative abundance of bacteria previously associated with beneficial IBD outcomes (e.g. Bifidobacterium). Interestingly, colonic CD4+ T-cells showed an increased conversion from Th17 towards a Foxp3+ Treg profile (Foxp3ExTh17) (p=0.03). Conclusion The conversion of CD4+ effector T cells into Treg cells is a promising approach to counteract inflammation in IBD patients. We showed that antibiotic treatment not only correlates with a relative expansion of beneficial bacteria, but perhaps most interesting, with an increased conversion of effector Th17 cells towards Foxp3+ Treg cells in the colon. Altogether, these data support the manipulation of the microbiota as a tool to revert intestinal inflammation in IBD patients.

517 Regulatory T cell functional identity is sustained by a glucose:lactate axis that is exploited in the tumor microenvironment

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A553-A553
Author(s):  
McLane Watson ◽  
Paolo Vignali ◽  
Steven Mullet ◽  
Abigail Overacre-Delgoffe ◽  
Ronal Peralta ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Microenvironment ◽  
Treg Cell ◽  
High Glucose ◽  
Cell Function ◽  
Treg Cells ◽  
Effector T Cells ◽  
Flux Analysis ◽  
Major Barrier ◽  
Suppressive Function

BackgroundRegulatory T (Treg) cells are vital for preventing autoimmunity but are a major barrier to robust cancer immunity as the tumor microenvironment (TME) recruits and promotes their function. The deregulated cellular metabolism of tumor cells leads to a metabolite-depleted, hypoxic, and acidic TME. While the TME impairs the effector function of highly glycolytic tumor infiltrating CD8 T cells, Treg cell suppressive function is maintained. Further, studies of in vitro induced and ex vivo Treg cells reveal a distinct metabolic profile compared to effector T cells. Thus, it may be that the altered metabolic landscape of the TME and the increased activity of intratumoral Treg cells are linked.MethodsFlow cytometry, isotopic flux analysis, Foxp3 driven Cre-lox, glucose tracers, Seahorse extracellular flux analysis, RNA sequencing.ResultsHere we show Treg cells display heterogeneity in terms of their glucose metabolism and can engage an alternative metabolic pathway to maintain their high suppressive function and proliferation within the TME and other tissues. Tissue derived Treg cells (both at the steady state and under inflammatory conditions) show broad heterogeneity in their ability to take up glucose. However, glucose uptake correlates with poorer suppressive function and long-term functional stability, and culture of Treg cells in high glucose conditions decreased suppressive function. Treg cells under low glucose conditions upregulate genes associated with the uptake and metabolism of the glycolytic end-product lactic acid. Treg cells withstand high lactate conditions, and lactate treatment prevents the destabilizing effects of high glucose culture. Treg cells utilize lactate within the TCA cycle and generate phosphoenolpyruvate (PEP), a critical intermediate that can fuel intratumoral Treg cell proliferation in vivo. Using mice with a Treg cell-restricted deletion of lactate transporter Slc16a1 (MCT1) we show MCT1 is dispensable for peripheral Treg cell function but required intratumorally, resulting in slowed tumor growth and prolonged survival.ConclusionsThese data support a model in which Treg cells are metabolically flexible such that they can utilize ‘alternative’ metabolites present in the TME to maintain their suppressive identity. Further, our studies support the notion that tumors avoid immune destruction not only by depriving effector T cells of essential nutrients, but also by metabolically supporting regulatory T cells.

scholarly journals PTCy ameliorates GVHD by restoring regulatory and effector T-cell homeostasis in recipients with PD-1 blockade

2019 ◽  
Vol 3 (23) ◽  
pp. 4081-4094 ◽  
Author(s):  
Shuntaro Ikegawa ◽  
Yusuke Meguri ◽  
Takumi Kondo ◽  
Hiroyuki Sugiura ◽  
Yasuhisa Sando ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Effector T Cells ◽  
T Cell Subsets ◽  
T Cell Homeostasis ◽  
Cell Homeostasis ◽  
Effector T Cell ◽  
Key Points

Key Points PD-1 blockade exacerbated GVHD by altering the homeostasis of Tregs and effector T cells after HSCT. PTCy ameliorated GVHD after PD-1 blockade by restoring the homeostatic balance of T-cell subsets.

scholarly journals Treatment of advanced tumors with agonistic anti-GITR mAb and its effects on tumor-infiltrating Foxp3+CD25+CD4+ regulatory T cells

2005 ◽  
Vol 202 (7) ◽  
pp. 885-891 ◽  
Author(s):  
Kuibeom Ko ◽  
Sayuri Yamazaki ◽  
Kyoko Nakamura ◽  
Tomohisa Nishioka ◽  
Keiji Hirota ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Immunity ◽  
Cd8 T Cells ◽  
Effector T Cells ◽  
Tumor Rejection ◽  
Cell Stimulation ◽  
T Cell Stimulation ◽  
Ifn Γ ◽  
Advanced Tumors

T cell stimulation via glucocorticoid-induced tumor necrosis factor receptor family–related protein (GITR) can evoke effective tumor immunity. A single administration of agonistic anti-GITR monoclonal antibody (mAb) to tumor-bearing mice intravenously or directly into tumors provoked potent tumor-specific immunity and eradicated established tumors without eliciting overt autoimmune disease. A large number of CD4+ and CD8+ T cells, including interferon (IFN)-γ–secreting cells, infiltrated regressing tumors. Tumor-specific IFN-γ–secreting CD4+ and CD8+ T cells also increased in the spleen. The treatment led to tumor rejection in IFN-γ–intact mice but not IFN-γ–deficient mice. Furthermore, coadministration of anti-GITR and anti–CTLA-4 mAbs had a synergistic effect, leading to eradication of more advanced tumors. In contrast, coadministration of anti-CD25 and anti-GITR mAbs was less effective than anti-GITR treatment alone, because anti-CD25 depleted both CD25+-activated effector T cells and CD25+CD4+ naturally occurring regulatory T (T reg) cells. Importantly, CD4+ T cells expressing the T reg–specific transcription factor Foxp3 predominantly infiltrated growing tumors in control mice, indicating that tumor-infiltrating natural Foxp3+CD25+CD4+ T reg cells may hamper the development of effective tumor immunity. Taken together, T cell stimulation through GITR attenuates T reg–mediated suppression or enhances tumor-killing by CD4+ and CD8+ effector T cells, including those secreting IFN-γ, or both. Agonistic anti-GITR mAb is therefore instrumental in treating advanced cancers.

Regulation of Hematopoietic Colony Forming Cells by CD4+CD25+ T Cells: A Role for T Regulatory Cells in Hematopoiesis?.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3181-3181
Author(s):  
Maite Urbieta ◽  
Isabel Barao ◽  
Monica Jones ◽  
William J. Murphy ◽  
Robert B. Levy
Keyword(s):  
T Cells ◽  
Growth Factors ◽  
T Cell ◽  
Treg Cell ◽  
Mhc Class Ii ◽  
Progenitor Cell ◽  
Cell Activity ◽  
Class Ii ◽  
Treg Cells

Abstract CD4+CD25+ T cells (Treg) comprise a small population within the normal peripheral CD4 T cell compartment. Their primary physiological role appears to be the regulation of autoimmune responses, however, in recent years it has been established that they can modulate anti-tumor as well as transplantation responses. Treg cells have been found to exert their affects on multiple types of immunologically relevant cells including CD4, CD8 and NK populations. Although model dependent, cytokines including TGFβ and IL-10 have been identified as mediators of this population’s regulatory activity and ex-vivo, the inhibition effected is generally contact dependent. Based upon the expanding application of Treg cells in stem cell transplants for the control of GVHD, rejection (HVG) and GVL responses, we hypothesized that following T cell receptor engagement and activation in recipients, CD4+CD25+ cells may modulate hematopoietic responses via production of effector cytokines. To address this question, various populations of CD4+CD25+ T cells were initially co-cultured with unfractionated syngeneic bone marrow cells (BMC) for 24–48 hours in medium supplemented with growth factors to maintain progenitor cell (i.e. CFU) function. Following co-culture, cells were collected and replated in triplicate in methylcellulose containing medium together with hematopoietic growth factors and five-seven days later, colonies were counted. CD4+CD25+ T cells were purified from BALB/c or B6–CD8−/− mice which were then activated for 3–8 days with anti-CD3/CD28 beads (a gift of Dr. B. Blazar, U. Minn.) These cells inhibited syngeneic CFU-IL3 colony ($25 cells) formation at ratios as low as 2:1 and 0.5:1 CD4+CD25+: BMC. Notably, Tregs from B6-CD8−/− mice exhibited comparable inhibition of allogeneic (BALB/c) CFU-IL3. Non-activated CD4+CD25+ T cells co-cultured with BMC did not exhibit this inhibitory activity nor did CD4+CD25− cells which contaminated (<10%) CD4+CD25+ populations. Activated Treg cells were also found to inhibit the production of CFU-HPP, a multi-potential marrow progenitor cell population. Contact dependency was found to be required for this effect as separation of activated CD4+CD25+ T cells from BMC “targets” in trans-well cultures abrogated inhibition. Prior depletion of CD25+ cells in vivo resulted in increases in CFU-GM 7–9 days after syngeneic BMT in mice suggesting that Tregs can inhibit hematopoietic reconstitution in vivo. To examine a potential contribution of TGFβ in this model, neutralizing anti-TGFβ mab was added during CD4+CD25+ T cell + BMC co-culture. The inhibition of CFU activity was abrogated in the presence of this antibody. To begin investigating the role of MHC class II molecules in this Treg cell activity, c-kit+ enriched (>85%) BMC from B6-MHC class II KO and B6-wt mice were co-cultured with B6 Treg cells from CD8−/− mice. In contrast to B6-wt c-kit enriched populations, CFU inhibition was not detected against the MHC class II deficient c-kit enriched BMC population. Antibody experiments are in progress to determine if cognate interaction is required between c-kit enriched cells and CD4+CD25+ T cells. In summary, this is the first report demonstrating that CD4+CD25+ T cells can alter hematopoietic progenitor cell activity. We hypothesize that membrane bound TGFβ may participate in effecting such regulation via direct Treg cell interactions with progenitor cell populations.

Inhibiting IL-2 Signaling and the Regulatory T-Cell Pathway Using Computationally Designed Novel Peptides

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3875-3875
Author(s):  
Tammy Price-Troska ◽  
David Diller ◽  
Alexander Bayden ◽  
Mark Jarosinski ◽  
Joseph Audies ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immune Function ◽  
Treg Cell ◽  
Immune Cell ◽  
Conflicts Of Interest ◽  
Interleukin 2 ◽  
Computational Design ◽  
Treg Cells ◽  
Cell Numbers

Abstract Regulatory T-cells (TREG) are the gateway to immune function and typically regulate immune cell activation. Cytokines, including interleukin-2 (IL-2), induce T-cell differentiation and promote a regulatory phenotype. Once activated via the IL-2 receptor (IL-2R), a cascade of events in T-cells initiate signal transducer and activator of transcription 5 (STAT5) and Forkhead box P3 (FOXP3) activation which appear to function as important regulators of this immunologic pathway and promote the development and function of TREG cells. In non-Hodgkin lymphoma (NHL), we have found that intratumoral TREG cells are increased in number and suppress immune function. In previous work, we have found that TREG cells inhibit T-cell proliferation, suppress cytokine production and limit effector cell cytotoxicity. We have also shown that increased serum levels of soluble sIL-2Rα is a prognostic factor in NHL and that sIL-2Rα can bind to IL-2 and promote its signaling thereby increasing TREG cell numbers. In this study, we developed a strategy to inhibit the binding of IL-2 to sIL-2Rα with the goal of suppressing the induction of FOXP3 and decreasing TREG cell numbers. To do this, we developed peptides designed to disrupt the interaction between IL2 and sILRα. In collaboration with CMDBioscienceSM, we developed and analyzed 22 peptide compounds derived by structure-based computational design. Initially, we screened each peptide at increasing concentrations using an ELISA assay to test the inhibition of IL-2/IL-2Rα binding by the solubilized peptide. Candidate peptides were then further tested using upregulation of pSTAT5 and FOXP3 in T-cells measured by flow cytometry as a measure of inhibition of IL-2 signaling. The peptides were developed according to different design hypotheses and grouped into different families; the screening ELISA results indicated 4 promising peptides that inhibited IL2/IL2Rα binding (up to 100% inhibition; max peptide concentration of 100uM). These peptides were then used to determine their effect on STAT5 and FOXP3 expression. A lead candidate peptide consistently reduced the expression of FOXP3 and STAT5 expression compared to cells not exposed to peptide. Use of the peptide to disrupt IL-2 signaling inhibited the development of cells with a TREG phenotype. We conclude that structure-based peptide design can be used to identify novel peptide inhibitors that block IL-2/IL-2Rα signaling and inhibit STAT5 and FOXP3 upregulation. These peptides could be used as new therapeutic agents to limit immune suppression by TREG cells and promote a more effective anti-tumor immune response in NHL. Disclosures No relevant conflicts of interest to declare.

Human CD4+ CD25+ Regulatory T Cells Effectively Control Xenogeneic-GvHD Induced by Autologous T Cells in Rag2−/− γc−/− Immune-Deficient Mice.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 301-301
Author(s):  
Tuna Mutis ◽  
Rozemarijn S. van Rijn ◽  
Elles R. Simonetti ◽  
Tineke Aarts ◽  
Maarten Emmelot ◽  
...  
Keyword(s):  
T Cells ◽  
Treg Cell ◽  
Graft Versus Host Disease ◽  
Peripheral Blood ◽  
Treg Cells ◽  
Effector T Cells ◽  
Graft Versus Host ◽  
Cell Engraftment ◽  
High Doses ◽  
The Impact

Abstract The curative Graft-versus-Leukemia (GvL) effect of allogeneic stem cell transplantation (SCT) and Donor Lymphocyte Infusions (DLI) is frequently complicated by Graft-versus-Host Disease (GvHD). To date, it is not possible to prevent GvHD without sacrificing the GvL effect. Recently, in a number of murine transplantation studies, administration of naturally occurring CD4+CD25+ regulatory T (Treg) cells in recipients of allogeneic bone marrow effectively prevented GvHD without abrogating GvL. If human (hu)CD4+CD25+ Treg cells also possess such properties, they may become new cellular immunotherapeutics for the prevention of GvHD. Therefore, we have started to investigate the impact of huTreg cells on GvHD in a recently developed, highly relevant xenogeneic(x)-GvHD model in immunodeficient Rag2−/− γc−/− mice. This model represents several features of human allo-GvHD, such as the involvement of both CD4 and CD8 T cells, the association of GvHD with a “cytokine storm” of several Th1/Th2 and inflammatory cytokines and the similarity of skin histopathology to the human allo-GvHD(1). As in this model the x-GvHD is induced by the i.v. injection of huPBMC and the severity of x-GvHD correlates with the number of T cells in the administered PBMC, we explored the impact of Treg cells on x-GvHD either by depletion of Treg cells from huPBMC at different administration doses of effector T cells (4-15 x106 CD25− T cells) and or co-injection of autologous Treg cells at high doses of effector T cells (12-15 x106 T cells). PBMC were isolated from the buffycoats of healthy blood bank donors. Part of the PBMC was used as effector cells, the remaining cells were fractionated into CD25+ and CD25− subsets, which contain Treg cells and conventional T cells, respectively. Different groups of mice were injected with low to high doses of Treg-cell-depleted-PBMC or with high doses PBMC supplemented with 4-6 x106 Treg cell-enriched CD25+ cells. Control mice received equivalent numbers of unmodified PBMC only. The development of x-GvHD was monitored weekly by determination of body weight, clinical scores (ruffled fur, alopecia, mobility) and survival. Peripheral blood obtained from orbital vein was analyzed for human T cell engraftment and expansion. In three independent experiments, depletion of Treg cells significantly exacerbated the x-GvHD signs and lethality. In striking contrast, the development of x-GvHD was significantly inhibited by the co-injection of Treg cell enriched cell fractions. In two independent experiments Treg cells completely protected mice from lethal x-GvHD. Phenotypical analyses of peripheral blood revealed that addition of Treg cells did not disturb huT cell engraftment, but inhibited the expansion of huT cells between 3-5 weeks of administration. These results demonstrate the effective control of x-GvHD in Rag2−/− γc−/− mice by huTreg cells. Studies are underway to reveal the mechanism of GvHD inhibition and the impact of huTreg cells on GvL. (1) R.S. van Rijn, E.R. Simonetti, M.C.H. Hogenes, G. Storm, A. Hagenbeek, H. Spits, K. Weijer, A. C. M. Martens, and S.B. Ebeling. A new in vivo model for graft-versus-host disease by intravenous transfer of human peripheral blood mononuclear cells in RAG2−/− γc−/− double mutant mice.

CD28 Costimulation of Effector T Cells During An Anti-Tumor Attack Sustains Its Repression by Regulatory T Cells Which Can Be Overcome by Preventing Lck Activation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3569-3569
Author(s):  
David M Kofler ◽  
Markus Chmielewski ◽  
Tobias Riet ◽  
Andreas Hombach ◽  
Michael Hallek ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Chimeric Antigen Receptor ◽  
Cell Activation ◽  
Antigen Receptor ◽  
Treg Cells ◽  
Effector T Cells ◽  
Cd28 Costimulation ◽  
Signaling Domain

Abstract Abstract 3569 Poster Board III-506 Background Massive infiltrations of the tumor tissue with CD4+CD25highFoxP3+ regulatory T (Treg) cells render a productive T cell anti-tumor response ineffective resulting in unrestricted tumor growth despite the presence of tumor-specific, cytolytic T cells. Methods Using a human/CD1-deficient mouse model we investigated the impact of human regulatory T cells on redirected effector T cells expressing a tumor-specific chimeric antigen receptor. The chimeric antigen receptor consists of an antibody derived binding domain for antigens in the extracellular part and of the TCR/CD3zeta or the combined CD3zeta-CD28 signaling domain in the intracellular part. Upon antigen binding the chimeric antigen receptor transmits an activation signal via the CD3zeta or CD3zeta-CD28 domain to drive T cell activation, resulting in cytokine secretion, T cell proliferation, and cytolytic activity. Results We revealed that effector T cells redirected by a tumor-specific chimeric antigen receptor are more effectively repressed by Treg cells when they are activated through a combined CD3zeta-CD28 signal compared to a CD3zeta signal without CD28 costimulation. Mutations in the CD28 signaling domain of the chimeric antigen receptors resulted in abolished IL-2 secretion by prevention of CD28 mediated lck activation. Abolished IL-2 induction in redirected effector T cells expressing the modified CD3zetaCD28delta antigen receptor increased their in vivo efficacy in an anti-tumor response by reduced sustaining of Treg cell suppression. Conclusions While data indicate the dichotomy of CD28 costimulation in inducing full effector T cell activation and sustaining Treg repression, our findings provide a strategy to improve the efficacy of the T cell anti-tumor attack in the presence of Treg cells for use in adoptive immunotherapy of cancer. Disclosures: No relevant conflicts of interest to declare.

Regulatory T Cells Enhance the Allogeneic Activity of Endothelial-Specific Cytotoxic T Lymphocytes – Protective Effect of Defibrotide

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4693-4693
Author(s):  
Guenther Eissner ◽  
Isabel Hartmann ◽  
Altug Kesikli ◽  
Silvia Haffner ◽  
Tanja Sax ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
T Cell ◽  
T Lymphocytes ◽  
Treg Cell ◽  
Interleukin 2 ◽  
Treg Cells ◽  
Protective Function ◽  
Phenotypic Quality

Abstract Abstract 4693 BACKGROUND: Damage to the vascular endothelium is the primary event of transplant related complications and often precedes loss of organ function. Depending on the amount of co-stimulatory signals, endothelial cells can either act as stimulating or inhibiting antigen presenting cells (APC). On the other hand, numerous data indicate that CD4+CD25+FoxP3+ T cells (Treg cells) can attenuate alloresponses of conventional T lymphocytes against classical APC and thus qualify for clinical use in various transplant settings. However, it is unknown whether Treg cells also influence T cell – endothelial cell interactions. Defibrotide (DF) is a polydisperse mixture of single-stranded deoxyribonucleotides with anti-thrombotic and anti-inflammatory activity, known to modulate the antigenicity of vascular endothelial cells. METHODS: CD8+ T lymphocytes (CTL) were isolated by magnetic microbead separation of peripheral blood mononuclear cells (PBMC) from healthy human blood donors and stimulated with mito-inactivated cells of a human microvascular endothelial cell line (CDC/EU.HMEC-1, further referred to as EC) and other primary and transformed micro- and macrovascular ECs for 7 days in the presence of interleukin 2 (IL-2). Treg cells from the CTL donor were prepared by CD4 (untouched) and a double CD25 microbead separation as well as a CD127bright depletion, followed by anti-CD3/CD28 expansion in the presence of IL-2 and a phenotypic quality control. Treg cells were added to the CTL-EC co-culture (1:1:1) prior to 51Cr release or flow cytometric cytotoxicity assays. Additionally, Treg cells were also tested for their capacity to influence CTL lysis of Epstein-Barr-Virus-transformed B-LCL, which as classical APC were HLA-matched to the HMEC. Furthermore, EC targets were incubated in the presence or absence of DF (25μM) for 24 hrs to assess the drug's protective function on the allogenicity of EC. RESULTS: EC-stimulated CTL showed a specific MHC class I-restricted target lysis. Addition of Treg cells prior to the cytotoxicity assay and during the afferent immune phase surprisingly increased EC lysis by CTL. In contrast, Treg cells alone did not show any lytic activity against EC. As a control, conventional CD4+CD25- T cells did not influence CTL activity either. Treg cell-mediated enhancement was endothelial-specific, since B-LCL lysis was not influenced. Further subpopulation analysis revealed the existence of CD8+/CD28-/CD57+ CTL, requiring cell-to-cell contact with Treg cells for their increased activity towards EC. Importantly, DF could almost fully protect EC against lysis by allogeneic CD28- CTL and the Treg cell-mediated enhancement. Of note, DF exclusively protected EC and did influence T cell function nor viability, suggesting a strong tropism for the endothelial cell type. CONCLUSION: There is no doubt about the potential therapeutic efficacy of Treg cells to ameliorate outcome of allogeneic transplants, but the endothelium might require additional protective interventions to prevent specific alloreactivity, such as DF. Disclosures: Eissner: Gentium, Sp.A.: Consultancy. Iacobelli:Gentium SpA: Employment.

Sign in / Sign up

Export Citation Format

Share Document