ABSTRACT
INTRODUCTION: Regulatory T cell (Treg) therapy has been demonstrated to facilitate long-term allograft survival in preclinical models of transplantation and may permit reduction of immunosuppression and its associated complications in the clinical setting. Phase 1 clinical trials have shown Treg therapy to be safe and feasible in clinical practice. Here we describe a protocol for the TWO study, a phase 2b randomised control trial of Treg therapy in living donor kidney transplant recipients that will confirm safety and explore efficacy of this novel treatment strategy. METHODS AND ANALYSIS: 60 patients will be randomised on a 1:1 basis to Treg therapy (TR001) or standard clinical care (control). Patients in the TR001 arm will receive an infusion of autologous polyclonal ex vivo expanded Tregs 5 days after transplantation instead of standard monoclonal antibody induction. Maintenance immunosuppression will be reduced over the course of the post-transplant period to low-dose tacrolimus monotherapy. Control participants will receive a standard basiliximab-based immunosuppression regimen with long-term tacrolimus and mycophenolate mofetil immunosuppression. The primary endpoint is biopsy proven acute rejection over 18 months; secondary endpoints include immunosuppression burden, chronic graft dysfunction and drug-related complications. ETHICS AND DISSEMINATION: Ethical approval has been provided by the National Health Service Health Research Authority South Central-Oxford A Research Ethics Committee (reference 18/SC/0054). The study also received authorisation from the UK Medicines and Healthcare products Regulatory Agency and is being run in accordance with the principles of Good Clinical Practice, in collaboration with the registered trials unit Oxford Clinical Trials Research Unit. Results from the TWO study will be published in peer-reviewed scientific/medical journals and presented at scientific/clinical symposia and congresses. TRIAL REGISTRATION NUMBER: ISRCTN: 11038572; Pre-results.
Subject(s)
Kidney Transplantation , T-Lymphocytes, Regulatory , Graft Rejection/prevention & control , Humans , Immunosuppression Therapy , Immunosuppressive Agents/adverse effects , Kidney Transplantation/methods , Living Donors , Randomized Controlled Trials as Topic , State Medicine , Tacrolimus/therapeutic useABSTRACT
Short-term outcomes in kidney transplantation are marred by progressive transplant failure and mortality secondary to immunosuppression toxicity. Immune modulation with autologous polyclonal regulatory T cell (Treg) therapy may facilitate immunosuppression reduction promoting better long-term clinical outcomes. In a Phase I clinical trial, 12 kidney transplant recipients received 1-10 × 106 Treg per kg at Day +5 posttransplantation in lieu of induction immunosuppression (Treg Therapy cohort). Nineteen patients received standard immunosuppression (Reference cohort). Primary outcomes were rejection-free and patient survival. Patient and transplant survival was 100%; acute rejection-free survival was 100% in the Treg Therapy versus 78.9% in the reference cohort at 48 months posttransplant. Treg therapy revealed no excess safety concerns. Four patients in the Treg Therapy cohort had mycophenolate mofetil withdrawn successfully and remain on tacrolimus monotherapy. Treg infusion resulted in a long-lasting dose-dependent increase in peripheral blood Tregs together with an increase in marginal zone B cell numbers. We identified a pretransplantation immune phenotype suggesting a high risk of unsuccessful ex-vivo Treg expansion. Autologous Treg therapy is feasible, safe, and is potentially associated with a lower rejection rate than standard immunosuppression. Treg therapy may provide an exciting opportunity to minimize immunosuppression therapy and improve long-term outcomes.
Subject(s)
Kidney Transplantation , Feasibility Studies , Graft Rejection/etiology , Graft Rejection/prevention & control , Humans , Immunosuppressive Agents/therapeutic use , Living Donors , Monitoring, Immunologic , T-Lymphocytes, RegulatoryABSTRACT
BACKGROUND: Use of cell-based medicinal products (CBMPs) represents a state-of-the-art approach for reducing general immunosuppression in organ transplantation. We tested multiple regulatory CBMPs in kidney transplant trials to establish the safety of regulatory CBMPs when combined with reduced immunosuppressive treatment. METHODS: The ONE Study consisted of seven investigator-led, single-arm trials done internationally at eight hospitals in France, Germany, Italy, the UK, and the USA (60 week follow-up). Included patients were living-donor kidney transplant recipients aged 18 years and older. The reference group trial (RGT) was a standard-of-care group given basiliximab, tapered steroids, mycophenolate mofetil, and tacrolimus. Six non-randomised phase 1/2A cell therapy group (CTG) trials were pooled and analysed, in which patients received one of six CBMPs containing regulatory T cells, dendritic cells, or macrophages; patient selection and immunosuppression mirrored the RGT, except basiliximab induction was substituted with CBMPs and mycophenolate mofetil tapering was allowed. None of the trials were randomised and none of the individuals involved were masked. The primary endpoint was biopsy-confirmed acute rejection (BCAR) within 60 weeks after transplantation; adverse event coding was centralised. The RTG and CTG trials are registered with ClinicalTrials.gov, NCT01656135, NCT02252055, NCT02085629, NCT02244801, NCT02371434, NCT02129881, and NCT02091232. FINDINGS: The seven trials took place between Dec 11, 2012, and Nov 14, 2018. Of 782 patients assessed for eligibility, 130 (17%) patients were enrolled and 104 were treated and included in the analysis. The 66 patients who were treated in the RGT were 73% male and had a median age of 47 years. The 38 patients who were treated across six CTG trials were 71% male and had a median age of 45 years. Standard-of-care immunosuppression in the recipients in the RGT resulted in a 12% BCAR rate (expected range 3·2-18·0). The overall BCAR rate for the six parallel CTG trials was 16%. 15 (40%) patients given CBMPs were successfully weaned from mycophenolate mofetil and maintained on tacrolimus monotherapy. Combined adverse event data and BCAR episodes from all six CTG trials revealed no safety concerns when compared with the RGT. Fewer episodes of infections were registered in CTG trials versus the RGT. INTERPRETATION: Regulatory cell therapy is achievable and safe in living-donor kidney transplant recipients, and is associated with fewer infectious complications, but similar rejection rates in the first year. Therefore, immune cell therapy is a potentially useful therapeutic approach in recipients of kidney transplant to minimise the burden of general immunosuppression. FUNDING: The 7th EU Framework Programme.
Subject(s)
Cell- and Tissue-Based Therapy/methods , Graft Rejection/prevention & control , Immunosuppression Therapy/methods , Immunosuppressive Agents/therapeutic use , Kidney Transplantation , Cell- and Tissue-Based Therapy/adverse effects , Dendritic Cells/immunology , Graft Rejection/immunology , Humans , Immunosuppression Therapy/adverse effects , Macrophages/immunology , T-Lymphocytes, Regulatory/immunologyABSTRACT
INTRODUCTION: To gain insights into the needs, attitudes, perceptions, and preferences of people living with obesity using an online bulletin board (OBB) study. METHODS: The OBB is a moderated asynchronous online qualitative market research method that allows interactive discussion among participants. Participants were recruited via physician referral followed by screening questions to ensure eligibility and willingness to participate. The discussions in the OBB were moderated and allowed anonymized open answers and responses. Analysis was performed using various qualitative analytical tools. RESULTS: This OBB study included 23 participants (n = 11, UK; n = 12, USA). Participants expressed negative emotions associated with obesity. Obesity impacted various aspects of their life, and the feeling of loneliness caused food indulgence, especially during the evenings. Their appearance was their primary cause of anxiety, whilst health considerations were secondary. The participants felt trapped in a cycle where food was (ab)used to overcome problems associated with being obese. Participants were pessimistic about weight management measures as a result of unsuccessful past attempts, with little/no support from healthcare providers, friends, and family for weight management. They preferred medications that would allow them to maintain their current lifestyle yet cause visible weight reduction. Along with medications, they expressed a strong preference for an online support group with similar peers for motivation, support, and sustained outcomes. CONCLUSIONS: As losing excess weight is a challenge for most overweight individuals, the qualitative insights from this OBB can inform the planning and successful execution of various weight management and drug development programs. FUNDING: Novartis Pharma AG, Basel Switzerland.
Subject(s)
Obesity , Quality of Life , Self Concept , Adult , Body Dissatisfaction/psychology , Body Mass Index , Female , Health Knowledge, Attitudes, Practice , Humans , Male , Middle Aged , Obesity/psychology , Obesity/therapy , Qualitative Research , Weight Reduction Programs/methodsABSTRACT
The concept of regulatory T cell (Treg)-based immunotherapy has enormous potential for facilitating tolerance in autoimmunity and transplantation. Clinical translation of Treg cell therapy requires production processes that satisfy the rigors of Good Manufacturing Practice (GMP) standards. In this regard, we report our findings on the implementation of a robust GMP compliant process for the ex vivo expansion of clinical grade Tregs, demonstrating the feasibility of this developed process for the manufacture of a final product for clinical application. This Treg isolation procedure ensured the selection of a pure Treg population that underwent a 300-fold expansion after 36 days of culture, while maintaining a purity of more than 75% CD4+CD25+FOXP3+ cells and a suppressive function of above 80%. Furthermore, we report the successful cryopreservation of the final product, demonstrating the maintenance of phenotype and function. The process outlined in this manuscript has been implemented in the ONE study, a multicenter phase I/IIa clinical trial in which cellular therapy is investigated in renal transplantation.
ABSTRACT
The measurement of immunological reactivity to donor antigens in transplant recipients is likely to be crucial for the successful reduction or withdrawal of immunosuppression. The mixed leukocyte reaction (MLR), limiting dilution assays, and trans-vivo delayed-type hypersensitivity (DTH) assay have all been applied to this question, but these methods have limited predictive ability and/or significant practical limitations that reduce their usefulness. Imaging flow cytometry is a technique that combines the multiparametric quantitative powers of flow cytometry with the imaging capabilities of fluorescent microscopy. We recently made use of an imaging flow cytometry approach to define the proportion of recipient T cells capable of forming mature immune synapses with donor antigen-presenting cells (APCs). Using a well-characterized mouse heart transplant model, we have shown that the frequency of in vitro immune synapses among T-APC membrane contact events strongly predicted allograft outcome in rejection, tolerance, and a situation where transplant survival depends on induced regulatory T cells. The frequency of T-APC contacts increased with T cells from mice during acute rejection and decreased with T cells from mice rendered unresponsive to alloantigen. The addition of regulatory T cells to the in vitro system reduced prolonged T-APC contacts. Critically, this effect was also seen with human polyclonally expanded, naturally occurring regulatory T cells, which are known to control the rejection of human tissues in humanized mouse models. Further development of this approach may allow for a deeper characterization of the alloreactive T-cell compartment in transplant recipients. In the future, further development and evaluation of this method using human cells may form the basis for assays used to select patients for immunosuppression minimization, and it can be used to measure the impact of tolerogenic therapies in the clinic.
Subject(s)
Flow Cytometry/methods , Isoantigens/immunology , T-Lymphocytes/immunology , Animals , Antigens/immunology , CD4-Positive T-Lymphocytes/immunology , Dendritic Cells/immunology , Graft Survival/immunology , Heart Transplantation/methods , Immune Tolerance , Immunosuppression Therapy/methods , Mice, Inbred CBA , T-Lymphocytes, Regulatory/immunology , Transplantation, HomologousABSTRACT
Distinctive molecular characteristics of functionally diverse lymphocyte populations may represent novel pharmacological targets for immunotherapy. The intrinsic apoptosis pathway is differently regulated among conventional and regulatory T cells (Tregs). Targeted pharmacological modulation of this pathway with a small molecule Bcl-2/Bcl-xL inhibitor (ABT-737) caused a selective depletion of effector T cells and a relative enrichment of Tregs in vivo. Treatment with ABT-737 resulted in a tolerogenic milieu, which was exploited to alleviate graft-versus-host disease, to prevent allograft rejection in a stringent fully MHC-mismatched skin transplantation model and to induce immunological tolerance in combination with bone marrow transplantation. This concept has the potential to find various applications for immunotherapy, since it allows pharmacologic exploitation of the immunomodulatory properties of Tregs without the need for cell manipulation ex vivo.
ABSTRACT
Solid organ transplantation is the treatment of choice for patients with end-stage organ failure. To prevent rejection of the transplanted organ continuous treatment with immunosuppressive medication is needed. Immunosuppression may be harmful to the transplant recipient, increasing the risk of cancer, infections and cardiovascular disease. To improve transplant and patient survival, there is a need for an immune-modulatory regimen that is not only potent in preventing rejection of the transplanted organ, but has less side effects compared to current immunosuppressive regimens. Increasingly, transplantation research focusses on regulatory T cell (Treg) therapy to achieve this aim, in which Treg are used as a strategy to allow reduction of immunosuppression. Currently, the first clinical trials are underway investigating the safety and feasibility of Treg therapy in renal transplantation. This review gives an overview of the rationale of using Treg therapy in transplantation, previous experience with Treg therapy in humans, and the expected safety, potential efficacy and cost-effectiveness of Treg therapy in solid organ transplantation.
Subject(s)
Graft Rejection/prevention & control , Immunotherapy, Adoptive/methods , Organ Transplantation/methods , T-Lymphocytes, Regulatory/transplantation , Animals , Follow-Up Studies , Humans , Immune Tolerance/physiology , Immunosuppressive Agents/therapeutic use , Organ Transplantation/adverse effects , Patient Safety/statistics & numerical data , T-Lymphocytes, Regulatory/immunology , Time Factors , Transplantation Immunology/physiology , Treatment OutcomeABSTRACT
Improper activation of the immune system contributes to a variety of clinical conditions, including autoimmune and allergic diseases as well as solid organ and bone marrow transplantation. One approach to counteract this activation is through adoptive therapy with regulatory T cells (Tregs). Efforts to manufacture these cells have led to good maunfacturing practice-compliant protocols, and Treg products are entering early clinical trials. Here, we report the stance of the European Union Cooperation in Science and Technology Action BM1305, "Action to Focus and Accelerate Cell-based Tolerance-inducing Therapies-A FACTT," which identifies hurdles hindering Treg clinical applications in Europe and provides possible solutions.
Subject(s)
Immunotherapy , T-Lymphocytes, Regulatory/immunology , Clinical Trials as Topic , Drug Approval , Humans , Immunotherapy/economics , Treatment OutcomeABSTRACT
CD4(+) CD25(+) Foxp3(+) regulatory T (Treg) cells mediate immunological self-tolerance and suppress immune responses. Retinoic acid (RA), a natural metabolite of vitamin A, has been reported to enhance the differentiation of Treg cells in the presence of TGF-ß. In this study, we show that the co-culture of naive T cells from C57BL/6 mice with allogeneic antigen-presenting cells (APCs) from BALB/c mice in the presence of TGF-ß, RA, and IL-2 resulted in a striking enrichment of Foxp3(+) T cells. These RA in vitro-induced regulatory T (RA-iTreg) cells did not secrete Th1-, Th2-, or Th17-related cytokines, showed a nonbiased homing potential, and expressed several cell surface molecules related to Treg-cell suppressive potential. Accordingly, these RA-iTreg cells suppressed T-cell proliferation and inhibited cytokine production by T cells in in vitro assays. Moreover, following adoptive transfer, RA-iTreg cells maintained Foxp3 expression and their suppressive capacity. Finally, RA-iTreg cells showed alloantigen-specific immunosuppressive capacity in a skin allograft model in immunodeficient mice. Altogether, these data indicate that functional and stable allogeneic-specific Treg cells may be generated using TGF-ß, RA, and IL-2. Thus, RA-iTreg cells may have a potential use in the development of more effective cellular therapies in clinical transplantation.
Subject(s)
Graft Rejection/prevention & control , Skin Transplantation , Skin/immunology , T-Lymphocytes, Regulatory/immunology , Tretinoin/pharmacology , Adoptive Transfer , Allografts , Animals , Coculture Techniques , Dendritic Cells/cytology , Dendritic Cells/drug effects , Dendritic Cells/immunology , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/immunology , Gene Expression , Graft Survival , Interleukin-2/pharmacology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Ovalbumin/administration & dosage , Skin/cytology , Spleen/cytology , Spleen/immunology , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/drug effects , T-Lymphocytes, Regulatory/transplantation , Transforming Growth Factor beta/pharmacologyABSTRACT
Organ transplantation results in the activation of both innate and adaptive immune responses to the foreign antigens. While these responses can be limited with the use of systemic immunosuppressants, the induction of regulatory cell populations may be a novel strategy for the maintenance of specific immunological unresponsiveness that can reduce the severity of the detrimental side effects of current therapies. Our group has extensively researched different regulatory T-cell induction protocols for use as cellular therapy in transplantation. In this review, we address the cellular and molecular mechanisms behind regulatory T-cell suppression and their stability following induction protocols. We further discuss the use of different hematopoietically derived regulatory cell populations, including regulatory B cells, regulatory macrophages, tolerogenic dendritic cells, and myeloid-derived suppressor cells, for the induction of transplantation tolerance in light of new clinical trials developing therapies with some of these populations.
Subject(s)
Graft Rejection/prevention & control , Hematopoietic Stem Cell Transplantation , Organ Transplantation , T-Lymphocytes, Regulatory/immunology , Transplantation Tolerance , Adoptive Transfer , Animals , Graft Rejection/immunology , Graft Survival , Hematopoietic Stem Cell Transplantation/adverse effects , Humans , Immunosuppressive Agents/therapeutic use , Mice , Organ Transplantation/adverse effects , T-Lymphocytes, Regulatory/drug effects , T-Lymphocytes, Regulatory/transplantation , Transplantation Tolerance/drug effects , Treatment OutcomeABSTRACT
OBJECTIVE: Left ventricular assist device placement is an increasingly common treatment for cardiac failure, resulting in cardiac unloading and potentially reversing the remodelling changes seen in heart failure. A popular animal model for human ventricular unloading is the rodent heterotopic non-working heart transplant; the volume loading status of this preparation is important to interpreting the resulting reverse remodelling yet has not been previously investigated. This study was designed to assess the variability of left ventricular volume loading in the rodent transplant model. METHODS: Heterotopic abdominal heart transplant was performed on syngeneic rats; high resolution cine magnetic resonance imaging was subsequently performed on the heterotopic transplanted hearts in anesthetised rats, after variable post-transplant recovery times, in order to assess ventricular loading status. RESULTS: Highly variable left ventricular volume loading status was demonstrated, with some hearts exhibiting considerable ventricular filling and ejection. CONCLUSIONS: These observations call into question the assumption that studies using this model are consistently examining fully unloaded ventricles, and indicate the desirability of in vivo imaging of such hearts to quantify the degree of ventricular loading.
ABSTRACT
Cellular therapy is emerging as a promising alternative to conventional immunosuppression in the fields of hematopoietic stem cell (HSC) transplantation, autoimmune disease, and solid organ transplantation. Determining the persistence of cell-based therapies in vivo is crucial to understanding their regulatory function and requires the combination of an extremely sensitive detection technique and a stable, long-lifetime cell labeling agent. This paper reports the first application of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) to perform single cell detection of T cell populations relevant to cellular immunotherapy. Purified human CD4(+) T cells were labeled with commercially available Gd-based magnetic resonance imaging (MRI) contrast agents, Omniscan and Dotarem, which enabled passive loading of up to 10(8) Gd atoms per cell. In mixed preparations of labeled and unlabeled cells, LA-ICP-MS was capable of enumerating labeled cells at close to the predicted ratio. More importantly, LA-ICP-MS single cell analysis demonstrated that the cells retained a sufficient label to remain detectable for up to 10 days post-labeling both in vitro and in vivo in an immunodeficient mouse model.
Subject(s)
CD4-Positive T-Lymphocytes/cytology , Cell Tracking/methods , Gadolinium/pharmacokinetics , Laser Therapy/methods , Mass Spectrometry/methods , Animals , CD4-Positive T-Lymphocytes/physiology , Contrast Media , Humans , Magnetic Resonance Imaging , Mice , Mice, Inbred BALB C , Tissue DistributionABSTRACT
Induction of mixed hematopoietic chimerism results in donor-specific immunological tolerance by apoptosis-mediated deletion of donor-reactive lymphocytes. A broad clinical application of this approach is currently hampered by limited predictability and toxicity of the available conditioning protocols. We developed a new therapeutic approach to induce mixed chimerism and tolerance by a direct pharmacological modulation of the intrinsic apoptosis pathway in peripheral T cells. The proapoptotic small-molecule Bcl-2 inhibitor ABT-737 promoted mixed chimerism induction and reversed the antitolerogenic effect of calcineurin inhibitors by boosting the critical role of the proapoptotic Bcl-2 factor Bim. A short conditioning protocol with ABT-737 in combination with costimulation blockade and low-dose cyclosporine A resulted in a complete deletion of peripheral donor-reactive lymphocytes and was sufficient to induce mixed chimerism and robust systemic tolerance across full major histocompatibility complex barriers, without myelosuppression and by using moderate doses of bone marrow cells. Thus, immunological tolerance can be achieved by direct modulation of the intrinsic apoptosis pathway in peripheral lymphocytes-a new approach to translate immunological tolerance into clinically applicable protocols.
Subject(s)
Apoptosis/drug effects , Biphenyl Compounds/pharmacology , Graft Survival/drug effects , Hematopoiesis/drug effects , Molecular Targeted Therapy/methods , Nitrophenols/pharmacology , Sulfonamides/pharmacology , Transplantation Chimera , Animals , Apoptosis/immunology , Biphenyl Compounds/therapeutic use , Cells, Cultured , Graft Survival/physiology , Hematopoiesis/physiology , Immunosuppression Therapy , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred CBA , Mice, Knockout , Nitrophenols/therapeutic use , Piperazines/pharmacology , Piperazines/therapeutic use , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Sulfonamides/therapeutic use , Transplantation Chimera/immunology , Transplantation Chimera/physiology , Transplantation Conditioning/methods , Transplantation Tolerance/drug effects , Up-Regulation/drug effects , Up-Regulation/immunologyABSTRACT
Immune regulation is fundamental to any immune response to ensure that it is appropriate for the perceived threat to the host. Following cell and organ transplantation, it is essential to control both the innate immune response triggered by the injured tissue and the adaptive immune response stimulated by mismatched donor and recipient histocompatibility antigens to enable the transplant to survive and function. Here, we discuss the leukocyte populations that can promote immune tolerance after cell or solid-organ transplantation. Such populations include regulatory T cells, B cells and macrophages, as well as myeloid-derived suppressor cells, dendritic cells and mesenchymal stromal cells. We consider the potential of these regulatory immune cells to develop and function in transplant recipients and their potential use as cellular therapies to promote long-term graft function.
Subject(s)
Immune System/cytology , Organ Transplantation/methods , Transplantation Immunology , Animals , Graft vs Host Disease/immunology , Graft vs Host Disease/prevention & control , HLA Antigens/immunology , Humans , Immune Tolerance/immunology , Immunosuppressive Agents/pharmacologyABSTRACT
Decreasing the incidence of chronic rejection and reducing the need for life-long immunosuppression remain important goals in clinical transplantation. In this article, we will review how regulatory T cells (Treg) came to be recognized as an attractive way to prevent or treat allograft rejection, the ways in which Treg can be manipulated or expanded in vivo, and the potential of in vitro expanded/generated Treg for cellular therapy. We will describe the first regulatory T cell therapies that have been or are in the process of being conducted in the clinic as well as the safety concerns of such therapies and how outcomes may be measured.
Subject(s)
Cell- and Tissue-Based Therapy , Graft Rejection/immunology , Organ Transplantation , T-Lymphocytes, Regulatory/immunology , Transplantation Tolerance , Animals , Cell- and Tissue-Based Therapy/adverse effects , Cell- and Tissue-Based Therapy/trends , Chemotherapy, Adjuvant , Clinical Trials as Topic , Europe , Graft Rejection/diagnosis , Graft Rejection/prevention & control , Humans , Immunomodulation , T-Lymphocytes, Regulatory/transplantation , Treatment OutcomeABSTRACT
Regulatory T cells (T(regs)) manipulated ex vivo have potential as cellular therapeutics in autoimmunity and transplantation. Although it is possible to expand naturally occurring T(regs), an attractive alternative possibility, particularly suited to solid organ and bone marrow transplantation, is the stimulation of total T cell populations with defined allogeneic antigen-presenting cells (APCs) under conditions that lead to the generation or expansion of donor-reactive, adaptive T(regs). Here we demonstrate that stimulation of mouse CD4(+) T cells by immature allogeneic dendritic cells combined with pharmacological inhibition of phosphodiesterase 3 (PDE) resulted in a functional enrichment of Foxp3(+) T cells. Without further manipulation or selection, the resultant population delayed skin allograft rejection mediated by polyclonal CD4(+) effectors or donor-reactive CD8(+) T cell receptor transgenic T cells and inhibited both effector cell proliferation and T cell priming for interferon-γ production. Notably, PDE inhibition also enhanced the enrichment of human Foxp3(+) CD4(+) T cells driven by allogeneic APCs. These cells inhibited T cell proliferation in a standard in vitro mixed lymphocyte assay and, moreover, attenuated the development of vasculopathy mediated by autologous peripheral blood mononuclear cells in a functionally relevant humanized mouse transplant model. These data establish a method for the ex vivo generation of graft-reactive, functional mouse and human T(regs) that uses a clinically approved agent, making pharmacological PDE inhibition a potential strategy for T(reg)-based therapies.
Subject(s)
Cyclic Nucleotide Phosphodiesterases, Type 3/drug effects , Graft Rejection/prevention & control , T-Lymphocytes, Regulatory/cytology , Animals , Forkhead Transcription Factors/genetics , Methylation , Mice , Quinolones/pharmacology , T-Lymphocytes, Regulatory/drug effectsABSTRACT
Exposure to alloantigen in vivo or in vitro induces alloantigen reactive regulatory T cells that can control transplant rejection. The mechanisms that underpin the activity of alloantigen reactive regulatory T cells in vivo are common with those of regulatory T cells that prevent autoimmunity. The identification and characterization of regulatory T cells that control rejection and contribute to the induction of immunologic unresponsiveness to alloantigens in vivo has opened up exciting opportunities for new therapies in transplantation. Findings from laboratory studies are informing the design of clinical protocols using regulatory T cells as a cellular therapy.
Subject(s)
Graft Rejection/therapy , Immune Tolerance , Immunotherapy, Adoptive , Isoantigens/immunology , T-Lymphocytes, Regulatory/immunology , Animals , Autoimmunity , Clinical Protocols , Graft Rejection/immunology , Humans , Organ Transplantation , T-Lymphocytes, Regulatory/transplantationABSTRACT
Naturally occurring FOXP3(+) CD4(+) Treg have a crucial role in self-tolerance. The ability to generate similar populations against alloantigens offers the possibility of preventing transplant rejection without indefinite global immunosuppression. Exposure of mice to donor alloantigens combined with anti-CD4 antibody induces operational tolerance to cardiac allografts, and generates Treg that prevent skin and islet allograft rejection in adoptive transfer models. If protocols that generate Treg in vivo are to be developed in the clinical setting it will be important to know the origin of the Treg population and the mechanisms responsible for their generation. In this study, we demonstrate that graft-protective Treg arise in vivo both from naturally occurring FOXP3(+) CD4(+) Treg and from non-regulatory FOXP3(-) CD4(+) cells. Importantly, tolerance induction also inhibits CD4(+) effector cell priming and T cells from tolerant mice have impaired effector function in vitro. Thus, adaptive tolerance induction shapes the immune response to alloantigen by converting potential effector cells into graft-protective Treg and by expanding alloreactive naturally occurring Treg. In relation to clinical tolerance induction, the data indicate that while the generation of alloreactive Treg may be critical for long-term allograft survival without chronic immunosuppression, successful protocols will also require strategies that target potential effector cells.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , T-Lymphocytes, Regulatory/immunology , Transplantation Tolerance/immunology , Adoptive Transfer , Animals , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/metabolism , Cell Differentiation , Forkhead Transcription Factors/metabolism , Graft Rejection/immunology , Graft Rejection/prevention & control , Heart Transplantation/immunology , Islets of Langerhans Transplantation/immunology , Isoantigens , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , Mice, Inbred DBA , Mice, Knockout , Mice, Transgenic , Skin Transplantation/immunology , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/metabolism , Th1 Cells/cytology , Th1 Cells/immunology , Tissue Donors , Transplantation, HomologousABSTRACT
IFN-γ was originally characterized as a proinflammatory cytokine with T helper type 1 inducing activity, but it is now clear that it also has important immunoregulatory functions. Regulatory T cells play an important role in models of autoimmunity, GVHD, and transplantation, and offer potential as a cellular therapy. In rodent models, in vivo-generated CD25(+)CD4(+) T cells can prevent allograft rejection, but therapeutic exploitation of Treg will more likely depend on protocols that allow the generation or selection of Treg ex vivo. The experiments described in this chapter will show that alloantigen-reactive Treg can be generated/expanded ex vivo using IFN-γ, a cytokine more usually associated with allograft rejection. Although IFN-γ production has hitherto been generally regarded as nonpermissive for allograft survival, we believe this paradoxical "good-bad" role for IFN-γ may reflect an important physiological negative feedback loop.