Publications by authors named "Giovanna Lombardi"

149 Publications

Potential Application of T-Follicular Regulatory Cell Therapy in Transplantation.

Front Immunol 2020 2;11:612848. Epub 2021 Feb 2.

Centre for Nephrology, Urology and Transplantation, School of Immunology and Microbial Sciences, King's College London, London, United Kingdom.

Regulatory T cells (Tregs) constitute a small proportion of circulating CD4 T cells that function to maintain homeostasis and prevent autoimmunity. In light of their powerful immunosuppressive and tolerance-promoting properties, Tregs have become an interesting potential candidate for therapeutic use in conditions such as solid organ transplant or to treat autoimmune and inflammatory conditions. Clinical studies have demonstrated the safety of polyclonally expanded Tregs in graft-versus-host disease, type 1 diabetes, and more recently in renal and liver transplantation. However, Tregs are heterogenous. Recent insights indicate that only a small proportion of Tregs, called T follicular regulatory cells (Tfr) regulate interactions between B cells and T follicular helper (Tfh) cells within the germinal center. Tfr have been mainly described in mouse models due to the challenges of sampling secondary lymphoid organs in humans. However, emerging human studies, characterize Tfr as being CD4CD25FOXP3CXCR5 cells with different levels of PD-1 and ICOS expression depending on their localization, in the blood or the germinal center. The exact role they play in transplantation remains to be elucidated. However, given the potential ability of these cells to modulate antibody responses to allo-antigens, there is great interest in exploring translational applications in situations where B cell responses need to be regulated. Here, we review the current knowledge of Tfr and the role they play focusing on human diseases and transplantation. We also discuss the potential future applications of Tfr therapy in transplantation and examine the evidence for a role of Tfr in antibody production, acute and chronic rejection and tertiary lymphoid organs. Furthermore, the potential impact of immunosuppression on Tfr will be explored. Based on preclinical research, we will analyse the rationale of Tfr therapy in solid organ transplantation and summarize the different challenges to be overcome before Tfr therapy can be implemented into clinical practice.
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http://dx.doi.org/10.3389/fimmu.2020.612848DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7884443PMC
February 2021

Therapy for the Treatment of Sickle Cell Disease: Taking Advantage of the Fetal Immune System.

Front Cell Dev Biol 2020 22;8:624477. Epub 2021 Jan 22.

School of Immunology & Microbial Sciences, King's College London, London, United Kingdom.

Sickle Cell Disease (SCD) is an autosomal recessive disorder resulting from a β-globin gene missense mutation and is among the most prevalent severe monogenic disorders worldwide. Haematopoietic stem cell transplantation remains the only curative option for the disease, as most management options focus solely on symptom control. Progress in prenatal diagnosis and fetal therapeutic intervention raises the possibility of treatment. SCD can be diagnosed prenatally in high-risk patients using chorionic villus sampling. Among the possible prenatal treatments, stem cell transplantation (IUSCT) shows the most promise. IUSCT is a non-myeloablative, non-immunosuppressive alternative conferring various unique advantages and may also offer safer postnatal management. Fetal immunologic immaturity could allow engraftment of allogeneic cells before fetal immune system maturation, donor-specific tolerance and lifelong chimerism. In this review, we will discuss SCD, screening and current treatments. We will present the therapeutic rationale for IUSCT, examine the early experimental work and initial human experience, as well as consider primary barriers of clinically implementing IUSCT and the promising approaches to address them.
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http://dx.doi.org/10.3389/fcell.2020.624477DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7862553PMC
January 2021

Spatiotemporal tracking of polyclonal human regulatory T cells (Tregs) reveals a role for innate immune cells in Treg transplant recruitment.

Mol Ther Methods Clin Dev 2021 Mar 10;20:324-336. Epub 2020 Dec 10.

MRC Centre for Transplantation, Peter Gorer Department of Immunobiology, School of Immunology and Microbial Science, King's College London, Guy's Hospital, London SE1 9RT, UK.

Regulatory T cells (Tregs) are emerging as a new cell-based therapy in solid organ transplantation. Adoptive transfer of Tregs has been shown preclinically to protect from graft rejection, and the safety of Treg therapy has been demonstrated in clinical trials. Despite these successes, the distribution and persistence of adoptively transferred Tregs remained elusive, which hampers clinical translation. Here we isolated human Tregs using a GMP-compatible protocol and lentivirally transduced them with the human sodium iodide symporter to render them traceable by radionuclide imaging. Engineered human Tregs were characterized for phenotype, survival, suppressive capacity, and reporter function. To study their trafficking behavior, they were subsequently administered to humanized mice with human skin transplants. Traceable Tregs were quantified in skin grafts by non-invasive nano-single-photon emission computed tomography (nanoSPECT)/computed tomography (CT) for up to 40 days, and the results were validated . Using this approach, we demonstrated that Treg trafficking to skin grafts was regulated by the presence of recipient Gr-1 innate immune cells. We demonstrated the utility of radionuclide reporter gene-afforded quantitative Treg tracking, addressing a fundamental need in Treg therapy development and offering a clinically compatible methodology for future Treg therapy imaging in humans.
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http://dx.doi.org/10.1016/j.omtm.2020.12.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811063PMC
March 2021

Purification and Immunophenotypic Characterization of Human CD24CD38 and CD24CD27 Regulatory B Cells in Transplantation.

Methods Mol Biol 2021 ;2270:451-467

MRC Centre for Transplantation, School of Immunology & Mucosal Biology, King's College London, London, UK.

Transplantation is still the treatment of choice for organ failure; however, allograft induces inflammatory immune responses that require immunosuppressive treatment. The role of regulatory B cells (Bregs) in downregulating inflammation has been reported to be significant in several diseases including transplant rejection. Many reports have analyzed different B-cell subpopulations, including Bregs, in tolerant, stable, and rejecting transplant recipients as well as the influence of immunosuppressant on the frequencies and functions of the different B-cell subsets. In this chapter, the key techniques required to investigate human Breg frequencies and functions in transplant patients are discussed.
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http://dx.doi.org/10.1007/978-1-0716-1237-8_24DOI Listing
January 2021

The Theoretical Basis of In Utero Hematopoietic Stem Cell Transplantation and Its Use in the Treatment of Blood Disorders.

Stem Cells Dev 2021 Jan;30(2):49-58

Immunoregulation Laboratory, School of Immunology, Microbial Sciences, and Faculty of Life Sciences and Medicine, King's College London, United Kingdom.

Since its conception, prenatal therapy has been successful in correction of mainly anatomical defects, although the range of application has been limited. Research into minimally invasive fetal surgery techniques and prenatal molecular diagnostics has facilitated the development of in utero stem cell transplantation (IUT)-a method of delivering healthy stem cells to the early gestation fetus with the hope of engraftment, proliferation, and migration to the appropriate hematopoietic compartment. An area of application that shows promise is the treatment of hematopoietic disorders like hemoglobinopathies. The therapeutic rationale of IUT with hematopoietic stem cells (HSCs) is based on the proposed advantages the fetal environment offers based on its unique physiology. These advantages include the immature immune system facilitating the development of donor-specific tolerance, the natural migration of endogenous hematopoietic cells providing space for homing and engraftment of donor cells, and the fetal environment providing HSCs with the same opportunity to survive and proliferate regardless of their origin (donor or host). Maternal immune tolerance to the fetus and placenta also implies that the maternal environment could be accepting of donor cells. In theory, the fetus is a perfect recipient for stem cell transplant. Clinically, however, IUT is yet to see widespread success calling into question these assumptions of fetal physiology. This review aims to discuss and evaluate research surrounding these key assumptions and the clinical success of IUT in the treatment of thalassemia.
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http://dx.doi.org/10.1089/scd.2020.0181DOI Listing
January 2021

Feasibility, long-term safety, and immune monitoring of regulatory T cell therapy in living donor kidney transplant recipients.

Am J Transplant 2020 Nov 10. Epub 2020 Nov 10.

Peter Gorer Department of Immunobiology, School of Immunology and Microbial Science, Kings College London, London, UK.

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 × 10 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.
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http://dx.doi.org/10.1111/ajt.16395DOI Listing
November 2020

The Future of Regulatory T Cell Therapy: Promises and Challenges of Implementing CAR Technology.

Front Immunol 2020 24;11:1608. Epub 2020 Jul 24.

Peter Gorer Department of Immunobiology, MRC Centre for Transplantation, School of Immunology and Microbial Science, King's College London (KCL), Guy's Hospital, London, United Kingdom.

Cell therapy with polyclonal regulatory T cells (Tregs) has been translated into the clinic and is currently being tested in transplant recipients and patients suffering from autoimmune diseases. Moreover, building on animal models, it has been widely reported that antigen-specific Tregs are functionally superior to polyclonal Tregs. Among various options to confer target specificity to Tregs, genetic engineering is a particularly timely one as has been demonstrated in the treatment of hematological malignancies where it is in routine clinical use. Genetic engineering can be exploited to express chimeric antigen receptors (CAR) in Tregs, and this has been successfully demonstrated to be robust in preclinical studies across various animal disease models. However, there are several caveats and a number of strategies should be considered to further improve on targeting, efficacy and to understand the distribution and fate of CAR-Tregs. Here, we review the differing approaches to confer antigen specificity to Tregs with emphasis on CAR-Tregs. This includes an overview and discussion of the various approaches to improve CAR-Treg specificity and therapeutic efficacy as well as addressing potential safety concerns. We also discuss different imaging approaches to understand the biodistribution of administered Tregs. Preclinical research as well as suitability of methodologies for clinical translation are discussed.
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http://dx.doi.org/10.3389/fimmu.2020.01608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7393941PMC
July 2020

An autocrine Vitamin D-driven Th1 shutdown program can be exploited for COVID-19.

bioRxiv 2020 Jul 19. Epub 2020 Jul 19.

Pro-inflammatory immune responses are necessary for effective pathogen clearance, but cause severe tissue damage if not shut down in a timely manner . Excessive complement and IFN-γ-associated responses are known drivers of immunopathogenesis and are among the most highly induced immune programs in hyper-inflammatory SARS-CoV2 lung infection . The molecular mechanisms that govern orderly shutdown and retraction of these responses remain poorly understood. Here, we show that complement triggers contraction of IFN-γ producing CD4 T helper (Th) 1 cell responses by inducing expression of the vitamin D (VitD) receptor (VDR) and CYP27B1, the enzyme that activates VitD, permitting T cells to both activate and respond to VitD. VitD then initiates the transition from pro-inflammatory IFN-γ Th1 cells to suppressive IL-10 Th1 cells. This process is primed by dynamic changes in the epigenetic landscape of CD4 T cells, generating superenhancers and recruiting c-JUN and BACH2, a key immunoregulatory transcription factor . Accordingly, cells in psoriatic skin treated with VitD increased BACH2 expression, and BACH2 haplo-insufficient CD4 T cells were defective in IL-10 production. As proof-of-concept, we show that CD4 T cells in the bronchoalveolar lavage fluid (BALF) of patients with COVID-19 are Th1-skewed and that VDR is among the top regulators of genes induced by SARS-CoV2. Importantly, genes normally down-regulated by VitD were de-repressed in CD4 BALF T cells of COVID-19, indicating that the VitD-driven shutdown program is impaired in this setting. The active metabolite of VitD, alfacalcidol, and cortico-steroids were among the top predicted pharmaceuticals that could normalize SARS-CoV2 induced genes. These data indicate that adjunct therapy with VitD in the context of other immunomodulatory drugs may be a beneficial strategy to dampen hyperinflammation in severe COVID-19.
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http://dx.doi.org/10.1101/2020.07.18.210161DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7388488PMC
July 2020

Regulatory T Cell Extracellular Vesicles Modify T-Effector Cell Cytokine Production and Protect Against Human Skin Allograft Damage.

Front Cell Dev Biol 2020 20;8:317. Epub 2020 May 20.

Immunoregulation Laboratory, MRC Centre for Transplantation, School of Immunology & Microbial Sciences, King's College London, Guy's Hospital, London, United Kingdom.

Regulatory T cells (Tregs) are a subpopulation of CD4 T cells with a fundamental role in maintaining immune homeostasis and inhibiting unwanted immune responses using several different mechanisms. Recently, the intercellular transfer of molecules between Tregs and their target cells has been shown via trogocytosis and the release of small extracellular vesicles (sEVs). In this study, CD4CD25CD127 human Tregs were found to produce sEVs capable of inhibiting the proliferation of effector T cells (Teffs) in a dose dependent manner. These vesicles also modified the cytokine profile of Teffs leading to an increase in the production of IL-4 and IL-10 whilst simultaneously decreasing the levels of IL-6, IL-2, and IFNγ. MicroRNAs found enriched in the Treg EVs were indirectly linked to the changes in the cytokine profile observed. In a humanized mouse skin transplant model, human Treg derived EVs inhibited alloimmune-mediated skin tissue damage by limiting immune cell infiltration. Taken together, Treg sEVs may represent an exciting cell-free therapy to promote transplant survival.
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http://dx.doi.org/10.3389/fcell.2020.00317DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7251034PMC
May 2020

B lymphocytes contribute to indirect pathway T cell sensitization via acquisition of extracellular vesicles.

Am J Transplant 2020 Jun 2. Epub 2020 Jun 2.

MRC Centre for Transplantation, Peter Gorer Department of Immunobiology, School of Immunology & Microbial Sciences, King's College London, London, SE1 9RT, UK.

B cells have been implicated in transplant rejection via antibody-mediated mechanisms and more recently by presenting donor antigens to T cells. We have shown in patients with chronic antibody-mediated rejection that B cells control the indirect T cell alloresponses. To understand more about the role of B cells as antigen-presenting cells for CD4 T cell with indirect allospecificity, B cells were depleted in C57BL/6 mice, using an anti-CD20 antibody, prior to receiving MHC class I-mismatched (K ) skin. The absence of B cells at the time of transplantation prolonged skin graft survival. To study the mechanisms behind this observation, T cells with indirect allospecificity were transferred in mice receiving a K skin transplant. T cell proliferation was markedly inhibited in the absence of recipient B cells, suggesting that B cells contribute to indirect pathway sensitization. Furthermore, we have shown that a possible way in which B cells present alloantigens is via acquisition of MHC-peptide complexes. Finally, we demonstrate that the addition of B cell depletion to the transfer of regulatory T cells (Tregs) with indirect alloresponse further prolonged skin graft survival. This study supports an important role for B cells in indirect T cell priming and further emphasizes the advantage of combination therapies in prolonging transplant survival.
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http://dx.doi.org/10.1111/ajt.16088DOI Listing
June 2020

Regulatory cell therapy in kidney transplantation (The ONE Study): a harmonised design and analysis of seven non-randomised, single-arm, phase 1/2A trials.

Lancet 2020 05;395(10237):1627-1639

Department of Surgery, University of Regensburg, University Hospital Regensburg, Regensburg, Germany; Division of Personalized Tumor Therapy, Fraunhofer Institute for Experimental Medicine and Toxicology, Regensburg, Germany; Regensburg Center for Interventional Immunology, University of Regensburg, Regensburg, Germany. Electronic address:

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.
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http://dx.doi.org/10.1016/S0140-6736(20)30167-7DOI Listing
May 2020

Treg sensitivity to FasL and relative IL-2 deprivation drive idiopathic aplastic anemia immune dysfunction.

Blood 2020 Aug;136(7):885-897

Department of Haematology, School of Cancer and Pharmaceutical Sciences, Faculty of Life Sciences and Medicine, King's College London, London, United Kingdom.

Idiopathic aplastic anemia (AA) has 2 key characteristics: an autoimmune response against hematopoietic stem/progenitor cells and regulatory T-cells (Tregs) deficiency. We have previously demonstrated reduction in a specific subpopulation of Treg in AA, which predicts response to immunosuppression. The aims of the present study were to define mechanisms of Treg subpopulation imbalance and identify potential for therapeutic intervention. We have identified 2 mechanisms that lead to skewed Treg composition in AA: first, FasL-mediated apoptosis on ligand interaction; and, second, relative interleukin-2 (IL-2) deprivation. We have shown that IL-2 augmentation can overcome these mechanisms. Interestingly, when high concentrations of IL-2 were used for in vitro Treg expansion cultures, AA Tregs were able to expand. The expanded populations expressed a high level of p-BCL-2, which makes them resistant to apoptosis. Using a xenograft mouse model, the function and stability of expanded AA Tregs were tested. We have shown that these Tregs were able to suppress the macroscopic clinical features and tissue manifestations of T-cell-mediated graft-versus-host disease. These Tregs maintained their suppressive properties as well as their phenotype in a highly inflammatory environment. Our findings provide an insight into the mechanisms of Treg reduction in AA. We have identified novel targets with potential for therapeutic interventions. Supplementation of ex vivo expansion cultures of Tregs with high concentrations of IL-2 or delivery of IL-2 directly to patients could improve clinical outcomes in addition to standard immunosuppressive therapy.
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http://dx.doi.org/10.1182/blood.2019001347DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7532000PMC
August 2020

Beyond bacterial killing: NADPH oxidase 2 is an immunomodulator.

Immunol Lett 2020 05 21;221:39-48. Epub 2020 Feb 21.

King's College London, Division of Transplantation Immunology and Mucosal Biology, London, United Kingdom.

NADPH oxidase 2 is a superoxide-generating enzymatic complex based on the catalytic subunit gp91phox that is also known as Nox2. Initially identified in neutrophils, NADPH oxidase 2 was long considered responsible only for the killing of phagocytized microorganisms. However, advances in knowledge about redox signalling and the discovery of Nox2 expression in different cell types, including macrophages, endothelial cells (ECs), dendritic cells (DCs), B and T lymphocytes, have changed this paradigm. For instance, Nox2 expressed in macrophages and neutrophils limits the transcription of cytokines and toll-like receptors (TLRs) induced by lipopolysaccharide (LPS), whereas DC Nox2 facilitates antigen cross-presentation to T cells. More recently, our group observed that Nox2 inhibits the suppressive ability of regulatory T cells (Tregs) by limiting NF-κB and FoxP3 activation. In this review, we discuss non-canonical microbicidal functions and redox-signalling-associated roles of Nox2 in different cell types, emphasizing its roles in the innate and adaptive immune system.
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http://dx.doi.org/10.1016/j.imlet.2020.02.009DOI Listing
May 2020

Getting to the Heart of the Matter: The Role of Regulatory T-Cells (Tregs) in Cardiovascular Disease (CVD) and Atherosclerosis.

Front Immunol 2019 28;10:2795. Epub 2019 Nov 28.

Peter Gorer Department of Immunobiology, School of Immunology and Microbiological Sciences, King's College London, London, United Kingdom.

Cardiovascular diseases (CVD) are the leading cause of mortality worldwide. Atherosclerosis is directly associated with CVD and is characterized by slow progressing inflammation which results in the deposition and accumulation of lipids beneath the endothelial layer in conductance and resistance arteries. Both chronic inflammation and disease progression have been associated with several risk factors, including but not limited to smoking, obesity, diabetes, genetic predisposition, hyperlipidemia, and hypertension. Currently, despite increasing incidence and significant expense on the healthcare system in both western and developing countries, there is no curative therapy for atherosclerosis. Instead patients rely on surgical intervention to avoid or revert vessel occlusion, and pharmacological management of the aforementioned risk factors. However, neither of these approaches completely resolve the underlying inflammatory environment which perpetuates the disease, nor do they result in plaque regression. As such, immunomodulation could provide a novel therapeutic option for atherosclerosis; shifting the balance from proatherogenic to athero-protective. Indeed, regulatory T-cells (Tregs), which constitute 5-10% of all CD4 T lymphocytes in the peripheral blood, have been shown to be athero-protective and could function as new targets in both CVD and atherosclerosis. This review aims to give a comprehensive overview about the roles of Tregs in CVD, focusing on atherosclerosis.
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http://dx.doi.org/10.3389/fimmu.2019.02795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6894511PMC
November 2020

Applicability, safety, and biological activity of regulatory T cell therapy in liver transplantation.

Am J Transplant 2020 04 3;20(4):1125-1136. Epub 2020 Feb 3.

MRC Centre for Transplantation, Peter Gorer Department of Immunobiology, Faculty of Life Sciences & Medicine, King's College London, London, UK.

Regulatory T cells (Tregs) are a lymphocyte subset with intrinsic immunosuppressive properties that can be expanded in large numbers ex vivo and have been shown to prevent allograft rejection and promote tolerance in animal models. To investigate the safety, applicability, and biological activity of autologous Treg adoptive transfer in humans, we conducted an open-label, dose-escalation, Phase I clinical trial in liver transplantation. Patients were enrolled while awaiting liver transplantation or 6-12 months posttransplant. Circulating Tregs were isolated from blood or leukapheresis, expanded under good manufacturing practices (GMP) conditions, and administered intravenously at either 0.5-1 million Tregs/kg or 3-4.5 million Tregs/kg. The primary endpoint was the rate of dose- limiting toxicities occurring within 4 weeks of infusion. The applicability of the clinical protocol was poor unless patient recruitment was deferred until 6-12 months posttransplant. Thus, only 3 of the 17 patients who consented while awaiting liver transplantation were dosed. In contrast, all six patients who consented 6-12 months posttransplant received the cell infusion. Treg transfer was safe, transiently increased the pool of circulating Tregs and reduced anti-donor T cell responses. Our study opens the door to employing Treg immunotherapy to facilitate the reduction or complete discontinuation of immunosuppression following liver transplantation.
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http://dx.doi.org/10.1111/ajt.15700DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7154724PMC
April 2020

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition).

Authors:
Andrea Cossarizza Hyun-Dong Chang Andreas Radbruch Andreas Acs Dieter Adam Sabine Adam-Klages William W Agace Nima Aghaeepour Mübeccel Akdis Matthieu Allez Larissa Nogueira Almeida Giorgia Alvisi Graham Anderson Immanuel Andrä Francesco Annunziato Achille Anselmo Petra Bacher Cosima T Baldari Sudipto Bari Vincenzo Barnaba Joana Barros-Martins Luca Battistini Wolfgang Bauer Sabine Baumgart Nicole Baumgarth Dirk Baumjohann Bianka Baying Mary Bebawy Burkhard Becher Wolfgang Beisker Vladimir Benes Rudi Beyaert Alfonso Blanco Dominic A Boardman Christian Bogdan Jessica G Borger Giovanna Borsellino Philip E Boulais Jolene A Bradford Dirk Brenner Ryan R Brinkman Anna E S Brooks Dirk H Busch Martin Büscher Timothy P Bushnell Federica Calzetti Garth Cameron Ilenia Cammarata Xuetao Cao Susanna L Cardell Stefano Casola Marco A Cassatella Andrea Cavani Antonio Celada Lucienne Chatenoud Pratip K Chattopadhyay Sue Chow Eleni Christakou Luka Čičin-Šain Mario Clerici Federico S Colombo Laura Cook Anne Cooke Andrea M Cooper Alexandra J Corbett Antonio Cosma Lorenzo Cosmi Pierre G Coulie Ana Cumano Ljiljana Cvetkovic Van Duc Dang Chantip Dang-Heine Martin S Davey Derek Davies Sara De Biasi Genny Del Zotto Gelo Victoriano Dela Cruz Michael Delacher Silvia Della Bella Paolo Dellabona Günnur Deniz Mark Dessing James P Di Santo Andreas Diefenbach Francesco Dieli Andreas Dolf Thomas Dörner Regine J Dress Diana Dudziak Michael Dustin Charles-Antoine Dutertre Friederike Ebner Sidonia B G Eckle Matthias Edinger Pascale Eede Götz R A Ehrhardt Marcus Eich Pablo Engel Britta Engelhardt Anna Erdei Charlotte Esser Bart Everts Maximilien Evrard Christine S Falk Todd A Fehniger Mar Felipo-Benavent Helen Ferry Markus Feuerer Andrew Filby Kata Filkor Simon Fillatreau Marie Follo Irmgard Förster John Foster Gemma A Foulds Britta Frehse Paul S Frenette Stefan Frischbutter Wolfgang Fritzsche David W Galbraith Anastasia Gangaev Natalio Garbi Brice Gaudilliere Ricardo T Gazzinelli Jens Geginat Wilhelm Gerner Nicholas A Gherardin Kamran Ghoreschi Lara Gibellini Florent Ginhoux Keisuke Goda Dale I Godfrey Christoph Goettlinger Jose M González-Navajas Carl S Goodyear Andrea Gori Jane L Grogan Daryl Grummitt Andreas Grützkau Claudia Haftmann Jonas Hahn Hamida Hammad Günter Hämmerling Leo Hansmann Goran Hansson Christopher M Harpur Susanne Hartmann Andrea Hauser Anja E Hauser David L Haviland David Hedley Daniela C Hernández Guadalupe Herrera Martin Herrmann Christoph Hess Thomas Höfer Petra Hoffmann Kristin Hogquist Tristan Holland Thomas Höllt Rikard Holmdahl Pleun Hombrink Jessica P Houston Bimba F Hoyer Bo Huang Fang-Ping Huang Johanna E Huber Jochen Huehn Michael Hundemer Christopher A Hunter William Y K Hwang Anna Iannone Florian Ingelfinger Sabine M Ivison Hans-Martin Jäck Peter K Jani Beatriz Jávega Stipan Jonjic Toralf Kaiser Tomas Kalina Thomas Kamradt Stefan H E Kaufmann Baerbel Keller Steven L C Ketelaars Ahad Khalilnezhad Srijit Khan Jan Kisielow Paul Klenerman Jasmin Knopf Hui-Fern Koay Katja Kobow Jay K Kolls Wan Ting Kong Manfred Kopf Thomas Korn Katharina Kriegsmann Hendy Kristyanto Thomas Kroneis Andreas Krueger Jenny Kühne Christian Kukat Désirée Kunkel Heike Kunze-Schumacher Tomohiro Kurosaki Christian Kurts Pia Kvistborg Immanuel Kwok Jonathan Landry Olivier Lantz Paola Lanuti Francesca LaRosa Agnès Lehuen Salomé LeibundGut-Landmann Michael D Leipold Leslie Y T Leung Megan K Levings Andreia C Lino Francesco Liotta Virginia Litwin Yanling Liu Hans-Gustaf Ljunggren Michael Lohoff Giovanna Lombardi Lilly Lopez Miguel López-Botet Amy E Lovett-Racke Erik Lubberts Herve Luche Burkhard Ludewig Enrico Lugli Sebastian Lunemann Holden T Maecker Laura Maggi Orla Maguire Florian Mair Kerstin H Mair Alberto Mantovani Rudolf A Manz Aaron J Marshall Alicia Martínez-Romero Glòria Martrus Ivana Marventano Wlodzimierz Maslinski Giuseppe Matarese Anna Vittoria Mattioli Christian Maueröder Alessio Mazzoni James McCluskey Mairi McGrath Helen M McGuire Iain B McInnes Henrik E Mei Fritz Melchers Susanne Melzer Dirk Mielenz Stephen D Miller Kingston H G Mills Hans Minderman Jenny Mjösberg Jonni Moore Barry Moran Lorenzo Moretta Tim R Mosmann Susann Müller Gabriele Multhoff Luis Enrique Muñoz Christian Münz Toshinori Nakayama Milena Nasi Katrin Neumann Lai Guan Ng Antonia Niedobitek Sussan Nourshargh Gabriel Núñez José-Enrique O'Connor Aaron Ochel Anna Oja Diana Ordonez Alberto Orfao Eva Orlowski-Oliver Wenjun Ouyang Annette Oxenius Raghavendra Palankar Isabel Panse Kovit Pattanapanyasat Malte Paulsen Dinko Pavlinic Livius Penter Pärt Peterson Christian Peth Jordi Petriz Federica Piancone Winfried F Pickl Silvia Piconese Marcello Pinti A Graham Pockley Malgorzata Justyna Podolska Zhiyong Poon Katharina Pracht Immo Prinz Carlo E M Pucillo Sally A Quataert Linda Quatrini Kylie M Quinn Helena Radbruch Tim R D J Radstake Susann Rahmig Hans-Peter Rahn Bartek Rajwa Gevitha Ravichandran Yotam Raz Jonathan A Rebhahn Diether Recktenwald Dorothea Reimer Caetano Reis e Sousa Ester B M Remmerswaal Lisa Richter Laura G Rico Andy Riddell Aja M Rieger J Paul Robinson Chiara Romagnani Anna Rubartelli Jürgen Ruland Armin Saalmüller Yvan Saeys Takashi Saito Shimon Sakaguchi Francisco Sala-de-Oyanguren Yvonne Samstag Sharon Sanderson Inga Sandrock Angela Santoni Ramon Bellmàs Sanz Marina Saresella Catherine Sautes-Fridman Birgit Sawitzki Linda Schadt Alexander Scheffold Hans U Scherer Matthias Schiemann Frank A Schildberg Esther Schimisky Andreas Schlitzer Josephine Schlosser Stephan Schmid Steffen Schmitt Kilian Schober Daniel Schraivogel Wolfgang Schuh Thomas Schüler Reiner Schulte Axel Ronald Schulz Sebastian R Schulz Cristiano Scottá Daniel Scott-Algara David P Sester T Vincent Shankey Bruno Silva-Santos Anna Katharina Simon Katarzyna M Sitnik Silvano Sozzani Daniel E Speiser Josef Spidlen Anders Stahlberg Alan M Stall Natalie Stanley Regina Stark Christina Stehle Tobit Steinmetz Hannes Stockinger Yousuke Takahama Kiyoshi Takeda Leonard Tan Attila Tárnok Gisa Tiegs Gergely Toldi Julia Tornack Elisabetta Traggiai Mohamed Trebak Timothy I M Tree Joe Trotter John Trowsdale Maria Tsoumakidou Henning Ulrich Sophia Urbanczyk Willem van de Veen Maries van den Broek Edwin van der Pol Sofie Van Gassen Gert Van Isterdael René A W van Lier Marc Veldhoen Salvador Vento-Asturias Paulo Vieira David Voehringer Hans-Dieter Volk Anouk von Borstel Konrad von Volkmann Ari Waisman Rachael V Walker Paul K Wallace Sa A Wang Xin M Wang Michael D Ward Kirsten A Ward-Hartstonge Klaus Warnatz Gary Warnes Sarah Warth Claudia Waskow James V Watson Carsten Watzl Leonie Wegener Thomas Weisenburger Annika Wiedemann Jürgen Wienands Anneke Wilharm Robert John Wilkinson Gerald Willimsky James B Wing Rieke Winkelmann Thomas H Winkler Oliver F Wirz Alicia Wong Peter Wurst Jennie H M Yang Juhao Yang Maria Yazdanbakhsh Liping Yu Alice Yue Hanlin Zhang Yi Zhao Susanne Maria Ziegler Christina Zielinski Jakob Zimmermann Arturo Zychlinsky

Eur J Immunol 2019 Oct;49(10):1457-1973

Max Planck Institute for Infection Biology, Berlin, Germany.

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.
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http://dx.doi.org/10.1002/eji.201970107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350392PMC
October 2019

Regulatory B cells: Development, phenotypes, functions, and role in transplantation.

Immunol Rev 2019 11 27;292(1):164-179. Epub 2019 Sep 27.

Peter Gorer Department of Immunobiology, MRC Centre for Transplantation, School of Immunology & Mucosal Biology, King's College London, King's Health Partners, Guy's Hospital, London, UK.

The interest in regulatory B cells (Bregs) began in the 1970s with the evidence that B cells could downregulate the immune system by the production of "inhibitory" antibodies. Subsequently, a series of results from different studies have emphasized that B cells have antibody-independent immunoregulatory functions. Since then, different subsets of B cells with regulatory functions and their development and mechanisms of action have been identified both in human and in animal models of inflammation, transplantation, and autoimmunity. The present review outlines the suggested pathways by which Bregs develop, describes the different subsets of Bregs with their phenotypes and function as well as their role in transplantation, highlighting the differences between human and animal studies throughout.
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http://dx.doi.org/10.1111/imr.12800DOI Listing
November 2019

IL-36 Promotes Systemic IFN-I Responses in Severe Forms of Psoriasis.

J Invest Dermatol 2020 04 17;140(4):816-826.e3. Epub 2019 Sep 17.

Department of Medical and Molecular Genetics, School of Basic & Medical Biosciences, King's College London, London, United Kingdom. Electronic address:

Psoriasis is an immune-mediated skin disorder associated with severe systemic comorbidities. Whereas IL-36 is a key disease driver, the pathogenic role of this cytokine has mainly been investigated in skin. Thus, its effects on systemic immunity and extracutaneous disease manifestations remain poorly understood. To address this issue, we investigated the consequences of excessive IL-36 activity in circulating immune cells. We initially focused our attention on generalized pustular psoriasis (GPP), a clinical variant associated with pervasive upregulation of IL-36 signaling. By undertaking blood and neutrophil RNA sequencing, we demonstrated that affected individuals display a prominent IFN-I signature, which correlates with abnormal IL-36 activity. We then validated the association between IL-36 deregulation and IFN-I over-expression in patients with severe psoriasis vulgaris (PV). We also found that the activation of IFN-I genes was associated with extracutaneous morbidity, in both GPP and PV. Finally, we undertook mechanistic experiments, demonstrating that IL-36 acts directly on plasmacytoid dendritic cells, where it potentiates toll-like receptor (TLR)-9 activation and IFN-α production. This effect was mediated by the upregulation of PLSCR1, a phospholipid scramblase mediating endosomal TLR-9 translocation. These findings identify an IL-36/ IFN-I axis contributing to extracutaneous inflammation in psoriasis.
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http://dx.doi.org/10.1016/j.jid.2019.08.444DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7097848PMC
April 2020

Protease Activated Receptor 4 as a Novel Modulator of Regulatory T Cell Function.

Front Immunol 2019 18;10:1311. Epub 2019 Jun 18.

MRC Centre for Transplantation, Guy's Hospital, King's College London, London, United Kingdom.

Regulatory T cells (Tregs) are a subpopulation of T cells that maintain immunological tolerance. In inflammatory responses the function of Tregs is tightly controlled by several factors including signaling through innate receptors such as Toll like receptors and anaphylatoxin receptors allowing an effective immune response to be generated. Protease-activated receptors (PARs) are another family of innate receptors expressed on multiple cell types and involved in the pathogenesis of autoimmune disorders. Whether proteases are able to directly modulate Treg function is unknown. Here, we show using two complimentary approaches that signaling through PAR-4 influences the expression of CD25, CD62L, and CD73, the suppressive capacity, and the stability of Tregs phosphorylation of FoxO1 and negative regulation of PTEN and FoxP3. Taken together, our results demonstrate an important role of PAR4 in tuning the function of Tregs and open the possibility of targeting PAR4 to modulate immune responses.
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http://dx.doi.org/10.3389/fimmu.2019.01311DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6591367PMC
October 2020

Mesenchymal stem cells inhibit T-cell function through conserved induction of cellular stress.

PLoS One 2019 14;14(3):e0213170. Epub 2019 Mar 14.

MRC Centre for Transplantation, School of Immunology and Microbial Sciences, King's College London, United Kingdom.

The physiological role of mesenchymal stem cells (MSCs) is to provide a source of cells to replace mesenchymal-derivatives in stromal tissues with high cell turnover or following stromal tissue damage to elicit repair. Human MSCs have been shown to suppress in vitro T-cell responses via a number of mechanisms including indoleamine 2,3-dioxygenase (IDO). This immunomodulatory capacity is likely to be related to their in vivo function in tissue repair where local, transient suppression of immune responses would benefit differentiation. Further understanding of the impact of locally modulated immune responses by MSCs is hampered by evidence that IDO is not produced or utilized by mouse MSCs. In this study, we demonstrate that IDO-mediated tryptophan starvation triggered by human MSCs inhibits T-cell activation and proliferation through induction of cellular stress. Significantly, we show that despite utilizing different means, immunomodulation of murine T-cells also involves cellular stress and thus is a common strategy of immunoregulation conserved between mouse and humans.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0213170PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6417714PMC
December 2019

Ways Forward for Tolerance-Inducing Cellular Therapies- an AFACTT Perspective.

Front Immunol 2019 22;10:181. Epub 2019 Feb 22.

Division of Immunology, Germans Trias i Pujol University Hospital, LCMN, IGTP, Badalona, Spain.

Clinical studies with cellular therapies using tolerance-inducing cells, such as tolerogenic antigen-presenting cells (tolAPC) and regulatory T cells (Treg) for the prevention of transplant rejection and the treatment of autoimmune diseases have been expanding the last decade. In this perspective, we will summarize the current perspectives of the clinical application of both tolAPC and Treg, and will address future directions and the importance of immunomonitoring in clinical studies that will result in progress in the field.
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http://dx.doi.org/10.3389/fimmu.2019.00181DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6395407PMC
January 2020

Past, Present, and Future of Regulatory T Cell Therapy in Transplantation and Autoimmunity.

Front Immunol 2019 31;10:43. Epub 2019 Jan 31.

Immunoregulation Laboratory, MRC Centre for Transplantation, School of Immunology & Microbial Sciences, King's College London, London, United Kingdom.

Regulatory T cells (Tregs) are important for the induction and maintenance of peripheral tolerance therefore, they are key in preventing excessive immune responses and autoimmunity. In the last decades, several reports have been focussed on understanding the biology of Tregs and their mechanisms of action. Preclinical studies have demonstrated the ability of Tregs to delay/prevent graft rejection and to control autoimmune responses following adoptive transfer . Due to these promising results, Tregs have been extensively studied as a potential new tool for the prevention of graft rejection and/or the treatment of autoimmune diseases. Currently, solid organ transplantation remains the treatment of choice for end-stage organ failure. However, chronic rejection and the ensuing side effects of immunosuppressants represent the main limiting factors for organ acceptance and patient survival. Autoimmune disorders are chronic diseases caused by the breakdown of tolerance against self-antigens. This is triggered either by a numerical or functional Treg defect, or by the resistance of effector T cells to suppression. In this scenario, patients receiving high doses of immunosuppressant are left susceptible to life-threatening opportunistic infections and have increased risk of malignancies. In the last 10 years, a few phase I clinical trials aiming to investigate safety and feasibility of Treg-based therapy have been completed and published, whilst an increasing numbers of trials are still ongoing. The first results showed safety and feasibility of Treg therapy and phase II clinical trials are already enrolling. In this review, we describe our understanding of Tregs focussing primarily on their ontogenesis, mechanisms of action and methods used in the clinic for isolation and expansion. Furthermore, we will describe the ongoing studies and the results from the first clinical trials with Tregs in the setting of solid organ transplantation and autoimmune disorders. Finally, we will discuss strategies to further improve the success of Treg therapy.
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http://dx.doi.org/10.3389/fimmu.2019.00043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6371029PMC
December 2019

IL-2 therapy preferentially expands adoptively transferred donor-specific Tregs improving skin allograft survival.

Am J Transplant 2019 07 15;19(7):2092-2100. Epub 2019 Mar 15.

MRC Centre for Transplantation, Peter Gorer Department of Immunobiology, Faculty of Life Sciences & Medicine, King's College London, London, UK.

Regulatory T cells (Tregs) have unique immunosuppressive properties and are essential to ensure effective immunoregulation. In animal models, Tregs have been shown to prevent autoimmune disorders and establish transplantation tolerance. Therefore, the prospect of harnessing Tregs, either by increasing their frequency or by conferring allospecificity, has prompted a growing interest in the development of immunotherapies. Here, employing a well-established skin transplant model with a single major histocompatibility complex mismatch, we compared the therapeutic efficacy of adoptively transfer Treg with or without donor specificity and the administration of IL-2 to promote in vivo expansion of Treg. We showed that IL-2 treatment preferentially enhances the proliferation of the allospecific Tregs adoptively transferred in an antigen-dependent manner. In addition, donor-specific Tregs significantly increased the expression of regulatory-related marker, such as CTLA4 and inducible costimulator (ICOS), in the skin allograft and draining lymph nodes compared to endogenous and polyclonal transferred Tregs. Importantly, by combining IL-2 with donor-specific Tregs, but not with polyclonal Tregs, a synergistic effect in prolonging skin allograft survival was observed. Altogether, our data suggest that this combination therapy could provide the appropriate conditions to enhance the immunoregulation of alloimmune responses in clinical transplantation.
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http://dx.doi.org/10.1111/ajt.15306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6618286PMC
July 2019

Correction of Defective T-Regulatory Cells From Patients With Crohn's Disease by Ex Vivo Ligation of Retinoic Acid Receptor-α.

Gastroenterology 2019 05 30;156(6):1775-1787. Epub 2019 Jan 30.

School of Immunology and Microbial Sciences, King's College London, London, UK; National Institute for Health Research Biomedical Research Centre, Guy's and St Thomas' NHS Trust and King's College London, London, UK. Electronic address:

Background & Aims: Crohn's disease (CD) is characterized by an imbalance of effector and regulatory T cells in the intestinal mucosa. The efficacy of anti-adhesion therapies led us to investigate whether impaired trafficking of T-regulatory (Treg) cells contributes to the pathogenesis of CD. We also investigated whether proper function could be restored to Treg cells by ex vivo expansion in the presence of factors that activate their regulatory activities.

Methods: We measured levels of the integrin α4β7 on Treg cells isolated from peripheral blood or lamina propria of patients with CD and healthy individuals (controls). Treg cells were expanded ex vivo and incubated with rapamycin with or without agonists of the retinoic acid receptor-α (RARA), and their gene expression profiles were analyzed. We also studied the cells in cytokine challenge, suppression, and flow chamber assays and in SCID mice with human intestinal xenografts.

Results: We found that Treg cells from patients with CD express lower levels of the integrin α4β7 than Treg cells from control patients. The pathway that regulates the expression of integrin subunit α is induced by retinoic acid (RA). Treg cells from patients with CD incubated with rapamycin and an agonist of RARA (RAR568) expressed high levels of integrin α4β7, as well as CD62L and FOXP3, compared with cells incubated with rapamycin or rapamycin and all-trans retinoic acid. These Treg cells had increased suppressive activities in assays and migrated under conditions of shear flow; they did not produce inflammatory cytokines, and RAR568 had no effect on cell stability or lineage commitment. Fluorescently labeled Treg cells incubated with RAR568 were significantly more likely to traffic to intestinal xenografts than Treg cells expanded in control medium.

Conclusions: Treg cells from patients with CD express lower levels of the integrin α4β7 than Treg cells from control patients. Incubation of patients' ex vivo expanded Treg cells with rapamycin and an RARA agonist induced expression of α4β7 and had suppressive and migratory activities in culture and in intestinal xenografts in mice. These cells might be developed for treatment of CD. ClinicalTrials.gov, Number: NCT03185000.
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http://dx.doi.org/10.1053/j.gastro.2019.01.025DOI Listing
May 2019

Application of carbon nanotubes in cancer vaccines: Achievements, challenges and chances.

J Control Release 2019 03 16;297:79-90. Epub 2019 Jan 16.

Institute of Pharmaceutical Science, School of Cancer and Pharmaceutical Science, Faculty of Life Sciences & Medicine, King's College London, Franklin-Wilkins Building, London SE1 9NH, United Kingdom. Electronic address:

Tumour-specific, immuno-based therapeutic interventions can be considered as safe and effective approaches for cancer therapy. Exploitation of nano-vaccinology to intensify the cancer vaccine potency may overcome the need for administration of high vaccine doses or additional adjuvants and therefore could be a more efficient approach. Carbon nanotube (CNT) can be described as carbon sheet(s) rolled up into a cylinder that is nanometers wide and nanometers to micrometers long. Stemming from the observed capacities of CNTs to enter various types of cells via diversified mechanisms utilising energy-dependent and/or passive routes of cell uptake, the use of CNTs for the delivery of therapeutic agents has drawn increasing interests over the last decade. Here we review the previous studies that demonstrated the possible benefits of these cylindrical nano-vectors as cancer vaccine delivery systems as well as the obstacles their clinical application is facing.
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http://dx.doi.org/10.1016/j.jconrel.2019.01.017DOI Listing
March 2019

Human retinoic acid-regulated CD161 regulatory T cells support wound repair in intestinal mucosa.

Nat Immunol 2018 12 5;19(12):1403-1414. Epub 2018 Nov 5.

MRC, Centre for Transplantation, King's College London, London, UK.

Repair of tissue damaged during inflammatory processes is key to the return of local homeostasis and restoration of epithelial integrity. Here we describe CD161 regulatory T (T) cells as a distinct, highly suppressive population of T cells that mediate wound healing. These T cells were enriched in intestinal lamina propria, particularly in Crohn's disease. CD161 T cells had an all-trans retinoic acid (ATRA)-regulated gene signature, and CD161 expression on T cells was induced by ATRA, which directly regulated the CD161 gene. CD161 was co-stimulatory, and ligation with the T cell antigen receptor induced cytokines that accelerated the wound healing of intestinal epithelial cells. We identified a transcription-factor network, including BACH2, RORγt, FOSL2, AP-1 and RUNX1, that controlled expression of the wound-healing program, and found a CD161 T cell signature in Crohn's disease mucosa associated with reduced inflammation. These findings identify CD161 T cells as a population involved in controlling the balance between inflammation and epithelial barrier healing in the gut.
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http://dx.doi.org/10.1038/s41590-018-0230-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6474659PMC
December 2018

Immunomodulatory role of Keratin 76 in oral and gastric cancer.

Nat Commun 2018 08 24;9(1):3437. Epub 2018 Aug 24.

Centre for Stem Cells & Regenerative Medicine, King's College London, Guy's Hospital, Great Maze Pond, London, SE1 9RT, UK.

Keratin 76 (Krt76) is expressed in the differentiated epithelial layers of skin, oral cavity and squamous stomach. Krt76 downregulation in human oral squamous cell carcinomas (OSCC) correlates with poor prognosis. We show that genetic ablation of Krt76 in mice leads to spleen and lymph node enlargement, an increase in regulatory T cells (Tregs) and high levels of pro-inflammatory cytokines. Krt76 Tregs have increased suppressive ability correlated with increased CD39 and CD73 expression, while their effector T cells are less proliferative than controls. Loss of Krt76 increases carcinogen-induced tumours in tongue and squamous stomach. Carcinogenesis is further increased when Treg levels are elevated experimentally. The carcinogenesis response includes upregulation of pro-inflammatory cytokines and enhanced accumulation of Tregs in the tumour microenvironment. Tregs also accumulate in human OSCC exhibiting Krt76 loss. Our study highlights the role of epithelial cells in modulating carcinogenesis via communication with cells of the immune system.
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http://dx.doi.org/10.1038/s41467-018-05872-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6109110PMC
August 2018

Expanded Regulatory T Cells Induce Alternatively Activated Monocytes With a Reduced Capacity to Expand T Helper-17 Cells.

Front Immunol 2018 20;9:1625. Epub 2018 Jul 20.

Immunoregulation Laboratory, MRC Centre for Transplantation, School of Immunology & Microbial Sciences, King's College London, London, United Kingdom.

Regulatory T cells (Tregs) are essential in maintaining peripheral immunological tolerance by modulating several subsets of the immune system including monocytes. Under inflammatory conditions, monocytes migrate into the tissues, where they differentiate into dendritic cells or tissue-resident macrophages. As a result of their context-dependent plasticity, monocytes have been implicated in the development/progression of graft-vs-host disease (GvHD), autoimmune diseases and allograft rejection. In the last decade, Tregs have been exploited for their use in cell therapy with the aim to induce tolerance after solid organ transplantation and for the treatment of autoimmune diseases and GvHD. To date, safety and feasibility of Treg infusion has been demonstrated; however, many questions of how these cells induce tolerance have been raised and need to be answered. As monocytes constitute the major cellular component in inflamed tissues, we have developed an model to test how Tregs modulate their phenotype and function. We demonstrated that expanded Tregs can drive monocytes toward an alternatively activated state more efficiently than freshly isolated Tregs. The effect of expanded Tregs on monocytes led to a reduced production of pro-inflammatory cytokines (IL-6 and tumor necrosis factor-α) and NF-κB activation. Furthermore, monocytes co-cultured with expanded Tregs downregulated the expression of co-stimulatory and MHC-class II molecules with a concomitant upregulation of M2 macrophage specific markers, CD206, heme oxygenase-1, and increased interleukin-10 production. Importantly, monocytes co-cultured with expanded Tregs showed a reduced capacity to expand IL-17-producing T cells compared with monocyte cultured with freshly isolated Tregs and conventional T cells. The capacity to decrease the expansion of pro-inflammatory Th-17 was not cytokine mediated but the consequence of their lower expression of the co-stimulatory molecule CD86. Our data suggest that expanded Tregs have the capacity to induce phenotypical and functional changes in monocytes that might be crucial for tolerance induction in transplantation and the prevention/treatment of GvHD and autoimmune diseases.
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http://dx.doi.org/10.3389/fimmu.2018.01625DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6062605PMC
July 2018

Optimizing regulatory T cells for therapeutic application in human organ transplantation.

Curr Opin Organ Transplant 2018 10;23(5):516-523

Department of Immunoregulation and Immune Intervention, Faculty of Life Sciences & Medicine, King's College London, Guy's Hospital, London, United Kingdom.

Purpose Of Review: Initial clinical trials of adoptive regulatory T-cell (Treg) therapy in solid organ transplantation have proven to be both feasible and well tolerated. With Phase 2 trials underway, efforts have been focused on the optimization of the Treg product.

Recent Findings: With science and our knowledge on the biology of these cells constantly advancing, we have been able to refine our search for a Treg population that would be ideally suited for therapeutic application. This idealized population must be readily isolated, allow for in-vitro expansion, demonstrate potent and specific suppressor function, maintain lineage stability and demonstrate a relevant homing profile. With the advent of innovative cell analysis/isolation techniques and genetic modifications, we are able to choose and design Tregs to fulfil these criteria.

Summary: By utilizing advances in science and technology, we can optimize Treg therapy in human organ transplantation maximizing their prospects in the arena of transplantation tolerance.
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http://dx.doi.org/10.1097/MOT.0000000000000561DOI Listing
October 2018

Nox2 in regulatory T cells promotes angiotensin II-induced cardiovascular remodeling.

J Clin Invest 2018 07 11;128(7):3088-3101. Epub 2018 Jun 11.

King's College London British Heart Foundation Centre, School of Cardiovascular Medicine and Sciences, London, United Kingdom.

The superoxide-generating enzyme Nox2 contributes to hypertension and cardiovascular remodeling triggered by activation of the renin-angiotensin system. Multiple Nox2-expressing cells are implicated in angiotensin II-induced (Ang II-induced) pathophysiology, but the importance of Nox2 in leukocyte subsets is poorly understood. Here, we investigated the role of Nox2 in T cells, particularly Tregs. Mice globally deficient in Nox2 displayed increased numbers of Tregs in the heart at baseline, whereas Ang II-induced effector T cell (Teff) infiltration was inhibited. To investigate the role of Treg Nox2, we generated a mouse line with CD4-targeted Nox2 deficiency (Nox2fl/flCD4Cre+). These animals showed inhibition of Ang II-induced hypertension and cardiac remodeling related to increased tissue-resident Tregs and reduction in infiltrating Teffs, including Th17 cells. The protection in Nox2fl/flCD4Cre+ mice was reversed by anti-CD25 antibody depletion of Tregs. Mechanistically, Nox2-/y Tregs showed higher in vitro suppression of Teff proliferation than WT Tregs, increased nuclear levels of FoxP3 and NF-κB, and enhanced transcription of CD25, CD39, and CD73. Adoptive transfer of Tregs confirmed that Nox2-deficient cells had greater inhibitory effects on Ang II-induced heart remodeling than WT cells. These results identify a previously unrecognized role of Nox2 in modulating suppression of Tregs, which acts to enhance hypertension and cardiac remodeling.
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http://dx.doi.org/10.1172/JCI97490DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025997PMC
July 2018