Antigen-specific Immune Suppression Research Articles

Organ- and Cell-Selective Delivery of mRNA In Vivo Using Guanidinylated Serinol Charge-Altering Releasable Transporters.

Selective RNA delivery is required for the broad implementation of RNA clinical applications, including prophylactic and therapeutic vaccinations, immunotherapies for cancer, and genome editing. Current polyanion delivery relies heavily on cationic amines, while cationic guanidinium systems have received limited attention due in part to their strong polyanion association, which impedes intracellular polyanion release. Here, we disclose a general solution to this problem in which cationic guanidinium groups are used to form stable RNA complexes upon formulation but at physiological pH undergo a novel charge-neutralization process, resulting in RNA release. This new delivery system consists of guanidinylated serinol moieties incorporated into a charge-altering releasable transporter (GSer-CARTs). Significantly, systematic variations in structure and formulation resulted in GSer-CARTs that exhibit highly selective mRNA delivery to the lung (∼97%) and spleen (∼98%) without targeting ligands. Illustrative of their breadth and translational potential, GSer-CARTs deliver circRNA, providing the basis for a cancer vaccination strategy, which in a murine model resulted in antigen-specific immune responses and effective suppression of established tumors.

Regulatory T Cells for Control of Autoimmunity.

Regulatory T (Treg) cells, which specifically express the master transcription factor FoxP3, are indispensable for the maintenance of immunological self-tolerance and homeostasis. Their functional or numerical anomalies can be causative of autoimmune and other inflammatory diseases. Recent advances in the research of the cellular and molecular basis of how Treg cells develop, exert suppression, and maintain their function have enabled devising various ways for controlling physiological and pathological immune responses by targeting Treg cells. It is now envisaged that Treg cells as a "living drug" are able to achieve antigen-specific immune suppression of various immune responses and reestablish immunological self-tolerance in the clinic.

Regulatory T cells in autoimmune kidney diseases and transplantation.

Regulatory T (Treg) cells that express the transcription factor forkhead box protein P3 (FOXP3) are naturally present in the immune system and have roles in the maintenance of immunological self-tolerance and immune system and tissue homeostasis. Treg cells suppress T cell activation, expansion and effector functions by various mechanisms, particularly by controlling the functions of antigen-presenting cells. They can also contribute to tissue repair by suppressing inflammation and facilitating tissue regeneration, for example, via the production of growth factors and the promotion of stem cell differentiation and proliferation. Monogenic anomalies of Treg cells and genetic variations of Treg cell functional molecules can cause or predispose patients to the development of autoimmune diseases and other inflammatory disorders, including kidney diseases. Treg cells can potentially be utilized or targeted to treat immunological diseases and establish transplantation tolerance, for example, by expanding natural Treg cells in vivo using IL-2 or small molecules or by expanding them in vitro for adoptive Treg cell therapy. Efforts are also being made to convert antigen-specific conventional T cells into Treg cells and to generate chimeric antigen receptorTreg cells from natural Treg cells for adoptive Treg cell therapies with the aim of achieving antigen-specific immune suppression and tolerance in the clinic.

Opportunities for Treg cell therapy for the treatment of human disease.

Regulatory T (Treg) cells are essential for maintaining peripheral tolerance, preventing autoimmunity, and limiting chronic inflammatory diseases. This small CD4+ T cell population can develop in the thymus and in the peripheral tissues of the immune system through the expression of an epigenetically stabilized transcription factor, FOXP3. Treg cells mediate their tolerogenic effects using multiple modes of action, including the production of inhibitory cytokines, cytokine starvation of T effector (e.g., IL-2), Teff suppression by metabolic disruption, and modulation of antigen-presenting cell maturation or function. These activities together result in the broad control of various immune cell subsets, leading to the suppression of cell activation/expansion and effector functions. Moreover, these cells can facilitate tissue repair to complement their suppressive effects. In recent years, there has been an effort to harness Treg cells as a new therapeutic approach to treat autoimmune and other immunological diseases and, importantly, to re-establish tolerance. Recent synthetic biological advances have enabled the cells to be genetically engineered to achieve tolerance and antigen-specific immune suppression by increasing their specific activity, stability, and efficacy. These cells are now being tested in clinical trials. In this review, we highlight both the advances and the challenges in this arena, focusing on the efforts to develop this new pillar of medicine to treat and cure a variety of diseases.

125 Mechanism of treg cells exerting antigen-specific immune suppression under UVB treatment: From patients to translational study

125 Mechanism of treg cells exerting antigen-specific immune suppression under UVB treatment: From patients to translational study

Treg-based immunotherapy for antigen-specific immune suppression and stable tolerance induction: a perspective.

FoxP3-expressing regulatory T cells (Tregs), whether naturally generated in the immune system or unnaturally induced from conventional T cells (Tconvs) in the laboratory, have much therapeutic value in treating immunological diseases and establishing transplantation tolerance. Natural Tregs (nTregs) can be selectively expanded in vivo by administration of low-dose IL-2 or IL-2 muteins for immune suppression. For adoptive Treg cell therapy, nTregs can be expanded in vitro by strong antigenic stimulation in the presence of IL-2. Synthetic receptors such as CAR can be expressed in nTregs to equip them with a particular target specificity for suppression. In addition, antigen-specific Tconvs can be converted in vitro to functionally stable Treg-like cells by a combination of antigenic stimulation, FoxP3 induction, and establishment of the Treg-type epigenome. This review discusses current and prospective strategies for Treg-based immune suppression and the issues to be resolved for achieving stable antigen-specific immune suppression and tolerance induction in the clinic by targeting Tregs.

Research Highlights.

Research Highlights.

OR22-4 Homing of Antigen-specific Engineered Regulatory T Cells To Human Pancreatic Islets

Abstract The therapeutic application of regulatory T cells (Tregs) for the treatment of autoimmune disorders, although efficacious, has been limited by the scarcity of antigen-specific Tregs. If antigen-specific Tregs, capable of reaching the desired autoimmune target, could be produced on demand, antigen-specific immune suppression of autoimmune diseases would be achievable. One approach to endow T cells with a desired antigen-specificity uses chimeric T cell antigen-receptors (CAR) with antibody-type specificity. Adoptive cell transfer therapies with CAR-re-directed cytotoxic T cells, have shown impressive efficacy in the treatment of hematologic malignancies. Accordingly, employing such technology to re-direct Tregs to sites of autoimmune attack may be a useful therapeutic approach to alleviate a broad spectrum of diseases in which uncontrolled auto and alloimmune responses play a major role. We recently developed pancreatic beta-cell, antigen-specific, CAR Tregs (1) and explored their therapeutic potential against T1D in our humanized mouse model (2). Ours is the first successful, antigen-specific CAR-Treg treatment of T1D in a humanized mouse model that closely resembles the human disease. Based on our mice data, we believe treatment with pancreatic beta-cell, antigen-specific CAR-Tregs will allow for recovery and reconstitution of beta cells in human T1D patients as well. The purpose of this study was to determine if antigen-specific human CAR Tregs could also identify target and home to human pancreatic islets in culture as they do in mice. The study involved drawing 10 cc of blood 1-2 weeks prior to pancreas surgery; followed by collection of a small piece of pancreas (5 cc wedge) once the pancreas was removed for a clinical indication (cancer, pancreatitis). We first isolated Tregs from peripheral blood of these human donors and expanded them in vitro. Treg cells were genetically modified to express either a beta-cell antigen-specific (GAD65) CAR or an irrelevant (EPCAM) CAR construct together with GFP marker. CAR Tregs were selectively expanded in the presence of rhGAD65 antigen. Once pancreas tissue became available, it was processed for islet separation using the collagenase method. Pancreatic islets were then co-cultured with syngeneic CAR Tregs for 7 days. IncuCyte S3 Live-cell System (Sartorius) is a real-time system that automatically acquires and analyzes HD, phase and fluorescent images of cell cultures, around the clock, for 7 days, while cells remain undisturbed. Live immunofluorescence microscopy demonstrated the distinct homing of GAD65 CAR Tregs to islets as compared to control EPCAM CAR Tregs as early as 24 hours of co-culture. Importantly, proliferation of GAD65 CAR Tregs was clearly demonstrated within 72 hours. GAD65 CAR Treg cytokine profile from co-culture supernatant was characteristic of activated Tregs. 1 PATENT #WO2020097546 2 PATENT #US20200037586 Presentation: Monday, June 13, 2022 11:45 a.m. - 12:00 p.m.

Pancreatic islet-specific engineered Tregs exhibit robust antigen-specific and bystander immune suppression in type 1 diabetes models.

Adoptive transfer of regulatory T cells (Tregs) is therapeutic in type 1 diabetes (T1D) mouse models. Tregs that are specific for pancreatic islets are more potent than polyclonal Tregs in preventing disease. However, the frequency of antigen-specific natural Tregs is extremely low, and ex vivo expansion may destabilize Tregs, leading to an effector phenotype. Here, we generated durable, antigen-specific engineered Tregs (EngTregs) from primary human CD4+ T cells by combining FOXP3 homology-directed repair editing and lentiviral T cell receptor (TCR) delivery. Using TCRs derived from clonally expanded CD4+ T cells isolated from patients with T1D, we generated islet-specific EngTregs that suppressed effector T cell (Teff) proliferation and cytokine production. EngTregs suppressed Teffs recognizing the same islet antigen in addition to bystander Teffs recognizing other islet antigens through production of soluble mediators and both direct and indirect mechanisms. Adoptively transferred murine islet-specific EngTregs homed to the pancreas and blocked diabetes triggered by islet-specific Teffs or diabetogenic polyclonal Teffs in recipient mice. These data demonstrate the potential of antigen-specific EngTregs as a targeted therapy for preventing T1D.

Nanoparticle-Based Drug Delivery Systems for Induction of Tolerance and Treatment of Autoimmune Diseases.

Autoimmune disease is a chronic inflammatory disease caused by disorders of immune regulation. Antigen-specific immunotherapy has the potential to inhibit the autoreactivity of inflammatory T cells and induce antigen-specific immune suppression without impairing normal immune function, offering an ideal strategy for autoimmune disease treatment. Tolerogenic dendritic cells (Tol DCs) with immunoregulatory functions play important roles in inducing immune tolerance. However, the effective generation of tolerogenic DCs in vivo remains a great challenge. The application of nanoparticle-based drug delivery systems in autoimmune disease treatment can increase the efficiency of inducing antigen-specific tolerance in vivo. In this review, we discuss multiple nanoparticles, with a focus on their potential in treatment of autoimmune diseases. We also discuss how the physical properties of nanoparticles influence their therapeutic efficacy.

New Treg cell-based therapies of autoimmune diseases: towards antigen-specific immune suppression.

Naturally occurring FoxP3+CD4+ regulatory T (Treg) cells indispensable for the maintenance of immunological self-tolerance and homeostasis are instrumental in treating autoimmune and other immunological disorders. Stable function of natural Treg cells requires not only the expression of Foxp3 and other Treg signature genes such as CD25 and CTLA-4 but also the generation of Treg-specific epigenetic changes, especially Treg-specific DNA hypomethylation, at these gene loci. Recent studies have shown that the Treg-specific transcriptional and epigenetic changes can be induced in antigen-specific conventional T cells in vivo and in vitro, converting them to functionally stable Treg cells. Such natural or induced Treg cells bear the potential to achieve stable antigen-specific immune suppression and reestablish immunological self-tolerance in treating and preventing autoimmune diseases.

Polymeric Nanoparticles Containing Both Antigen and Vitamin D3 Induce Antigen-Specific Immune Suppression.

The active form of vitamin D3, 1,25-dihydroxyvitamin D3 (aVD3), is known to exert beneficial effects in the treatment of autoimmune diseases because of its immunosuppressive effects. However, clinical application of aVD3 remains limited because of the potential side effects, particularly hypercalcemia. Encapsulation of aVD3 within biodegradable nanoparticles (NPs) would enhance the delivery of aVD3 to antigen presenting cells, while preventing the potential systemic side effects of aVD3. In the present study, polymeric NPs containing ovalbumin (OVA) and aVD3 (NP[OVA+aVD3]) were prepared via the water-in-oil-in-water double emulsion solvent evaporation method, after which their immunomodulatory effects were examined. Bone marrow-derived immature dendritic cells (DCs) treated with NP(OVA+aVD3) did not mature into immunogenic DCs but were converted into tolerogenic DCs, which express low levels of co-stimulatory molecules and MHC class II molecules, produce lower levels of pro-inflammatory cytokines while increasing the production of IL-10 and TGF-β, and induce the generation of Tregs. Intravenous injection with NP(OVA+aVD3) markedly suppressed the generation of OVA-specific CTLs in mice. Furthermore, OVA-specific immune tolerance was induced in mice orally administered with NP(OVA+aVD3). These results show that biodegradable NPs encapsulating both antigen and aVD3 can effectively induce antigen-specific immune suppression.

Pemphigus.

Pemphigus is a group of IgG-mediated autoimmune diseases of stratified squamous epithelia, such as the skin and oral mucosa, in which acantholysis (the loss of cell adhesion) causes blisters and erosions. Pemphigus has three major subtypes: pemphigus vulgaris, pemphigus foliaceus and paraneoplastic pemphigus. IgG autoantibodies are characteristically raised against desmoglein 1 and desmoglein 3, which are cell-cell adhesion molecules found in desmosomes. The sites of blister formation can be physiologically explained by the anti-desmoglein autoantibody profile and tissue-specific expression pattern of desmoglein isoforms. The pathophysiological roles of T cells and B cells have been characterized in mouse models of pemphigus and patients, revealing insights into the mechanisms of autoimmunity. Diagnosis is based on clinical manifestations and confirmed with histological and immunochemical testing. The current first-line treatment is systemic corticosteroids and adjuvant therapies, including immunosuppressive agents, intravenous immunoglobulin and plasmapheresis. Rituximab, a monoclonal antibody against CD20+ B cells, is a promising therapeutic option that may soon become first-line therapy. Pemphigus is one of the best-characterized human autoimmune diseases and provides an ideal paradigm for both basic and clinical research, especially towards the development of antigen-specific immune suppression treatments for autoimmune diseases.

Relevance of Interferon Regulatory Factor-8 Expression in Myeloid-Tumor Interactions.

Perturbations in myelopoiesis are a common feature in solid tumor biology, reflecting the central premise that cancer is not only a localized affliction but also a systemic disease. Because the myeloid compartment is essential for the induction of adaptive immunity, these alterations in myeloid development contribute to the failure of the host to effectively manage tumor progression. These "dysfunctional" myeloid cells have been coined myeloid-derived suppressor cells (MDSCs). Interestingly, such cells not only arise in neoplasia but also are associated with many other inflammatory or pathologic conditions. MDSCs affect disease outcome through multiple mechanisms, including their ability to mediate generalized or antigen-specific immune suppression. Consequently, MDSCs pose a significant barrier to effective immunotherapy in multiple disease settings. Although much interest has been devoted to unraveling mechanisms by which MDSCs mediate immune suppression, a large gap has remained in our understanding of the mechanisms that drive their development in the first place. Investigations into this question have identified an unrecognized role of interferon regulatory factor-8 (IRF-8), a member of the IRF family of transcription factors, in tumor-induced myeloid dysfunction. Ordinarily, IRF-8 is involved in diverse stages of myelopoiesis, namely differentiation and lineage commitment toward monocytes, dendritic cells, and granulocytes. Several recent studies now support the hypothesis that IRF-8 functions as a "master" negative regulator of MDSC formation in vivo. This review focuses on IRF-8 as a potential target suppressed by tumors to cripple normal myelopoiesis, redirecting myeloid differentiation toward the emergence of MDSCs. Understanding the bases by which neoplasia drives MDSC accumulation has the potential to improve the efficacy of therapies that require a competent myeloid compartment.

Macrophages play an essential role in antigen-specific immune suppression mediated by T CD8⁺ cell-derived exosomes.

Murine contact sensitivity (CS) reaction could be antigen-specifically regulated by T CD8(+) suppressor (Ts) lymphocytes releasing microRNA-150 in antibody light-chain-coated exosomes that were formerly suggested to suppress CS through action on macrophages (Mφ). The present studies investigated the role of Mφ in Ts cell-exosome-mediated antigen-specific suppression as well as modulation of Mφ antigen-presenting function in humoral and cellular immunity by suppressive exosomes. Mice depleted of Mφ by clodronate liposomes could not be tolerized and did not produce suppressive exosomes. Moreover, isolated T effector lymphocytes transferring CS were suppressed by exosomes only in the presence of Mφ, demonstrating the substantial role of Mφ in the generation and action of Ts cell regulatory exosomes. Further, significant decrease of number of splenic B cells producing trinitrophenyl (TNP) -specific antibodies with the alteration of the ratio of serum titres of IgM to IgG was observed in recipients of exosome-treated, antigen-pulsed Mφ and the significant suppression of CS was demonstrated in recipients of exosome-treated, TNP-conjugated Mφ. Additionally, exosome-pulsed, TNP-conjugated Mφ mediated suppression of CS in mice pre-treated with a low-dose of cyclophosphamide, suggesting de novo induction of T regulatory (Treg) lymphocytes. Treg cell involvement in the effector phase of the studied suppression mechanism was proved by unsuccessful tolerization of DEREG mice depleted of Treg lymphocytes. Furthermore, the inhibition of proliferation of CS effector cells cultured with exosome-treated Mφ in a transmembrane manner was observed. Our results demonstrated the essential role of Mφ in antigen-specific immune suppression mediated by Ts cell-derived exosomes and realized by induction of Treg lymphocytes and inhibition of T effector cell proliferation.

Plant-based oral tolerance to hemophilia therapy employs a complex immune regulatory response including LAP+CD4+ T cells

AbstractCoagulation factor replacement therapy for the X-linked bleeding disorder hemophilia is severely complicated by antibody (“inhibitor”) formation. We previously found that oral delivery to hemophilic mice of cholera toxin B subunit-coagulation factor fusion proteins expressed in chloroplasts of transgenic plants suppressed inhibitor formation directed against factors VIII and IX and anaphylaxis against factor IX (FIX). This observation and the relatively high concentration of antigen in the chloroplasts prompted us to evaluate the underlying tolerance mechanisms. The combination of oral delivery of bioencapsulated FIX and intravenous replacement therapy induced a complex, interleukin-10 (IL-10)–dependent, antigen-specific systemic immune suppression of pathogenic antibody formation (immunoglobulin [Ig] 1/inhibitors, IgE) in hemophilia B mice. Tolerance induction was also successful in preimmune mice but required prolonged oral delivery once replacement therapy was resumed. Orally delivered antigen, initially targeted to epithelial cells, was taken up by dendritic cells throughout the small intestine and additionally by F4/80+ cells in the duodenum. Consistent with the immunomodulatory responses, frequencies of tolerogenic CD103+ and plasmacytoid dendritic cells were increased. Ultimately, latency-associated peptide expressing CD4+ regulatory T cells (CD4+CD25−LAP+ cells with upregulated IL-10 and transforming growth factor-β (TGF-β) expression) as well as conventional CD4+CD25+ regulatory T cells systemically suppressed anti-FIX responses.

Suppression of antigen-specific adaptive immunity by IL-37 via induction of tolerogenic dendritic cells.

IL-1 family member IL-37 limits innate inflammation in models of colitis and LPS-induced shock, but a role in adaptive immunity remains unknown. Here, we studied mice expressing human IL-37b isoform (IL-37tg) subjected to skin contact hypersensitivity (CHS) to dinitrofluorobenzene. CHS challenge to the hapten was significantly decreased in IL-37tg mice compared with wild-type (WT) mice (-61%; P < 0.001 at 48 h). Skin dendritic cells (DCs) were present and migrated to lymph nodes after antigen uptake in IL-37tg mice. When hapten-sensitized DCs were adoptively transferred to WT mice, antigen challenge was greatly impaired in mice receiving DCs from IL-37tg mice compared with those receiving DCs from WT mice (-60%; P < 0.01 at 48 h). In DCs isolated from IL-37tg mice, LPS-induced increase of MHC II and costimulatory molecule CD40 was reduced by 51 and 31%, respectively. In these DCs, release of IL-1β, IL-6, and IL-12 was reduced whereas IL-10 secretion increased (37%). Consistent with these findings, DCs from IL-37tg mice exhibited a lower ability to stimulate syngeneic and allogeneic naive T cells as well as antigen-specific T cells and displayed enhanced induction of T regulatory (Treg) cells (86%; P < 0.001) in vitro. Histological analysis of CHS skin in mice receiving hapten-sensitized DCs from IL-37tg mice revealed a marked reduction in CD8(+) T cells (-74%) but an increase in Treg cells (2.6-fold). Together, these findings reveal that DCs expressing IL-37 are tolerogenic, thereby impairing activation of effector T-cell responses and inducing Treg cells. IL-37 thus emerges as an inhibitor of adaptive immunity.

CD28 signals the differential control of regulatory T cells and effector T cells

One possible means of driving antigen-specific immune suppression is to expand or induce antigen-specific FoxP3-expressing Treg cells. One way of activating and expanding these specialized cells, both in vitro and in vivo, is by strong costimulation via CD28 with an agonistic anti-CD28 monoclonal antibody, called anti-CD28 superagonist (CD28SA). However, CD28SA also strongly activates conventional T (Tconv) cells to secrete proinflammatory cytokines and, under certain conditions, causes serious cytokine release syndrome. In this issue of European Journal of Immunology, Tabares etal. [Eur. J. Immunol. 2014. 44: 1225-1236] address how CD28SA can be used for the differential control of human Treg and Tconv cells to suppress immune responses without serious adverse effects. They show that, depending on the dose of the antibody or by comedication of cortico-steroid, the selective expansion of Treg cells can be achieved without significantly activating Tconv cells to produce inflammatory cytokines. This difference in CD28 signal sensitivity between the two populations can be exploited for better control of immune responses.

Regulatory T cells get nervous in the melanoma microenvironment

Regulatory T cells get nervous in the melanoma microenvironment

Neonatal lymph node stromal cells drive myelodendritic lineage cells into a distinct population of CX3CR1+CD11b+F4/80+ regulatory macrophages in mice

AbstractBeyond providing a scaffold for immune cells, recent studies indicate that lymph node stromal cells provide potent regulatory capacities that affect the quality of adaptive immune responses. In this study, we provide evidence that neonatal lymph node stromal cells (nnLNSCs) consistently promote the differentiation of macrophage dendritic cell progenitors as well as mature and immature dendritic cells into a distinct population of CX3CR1+ CD11b+F4/80+ regulatory macrophages (regMΦ). These cells possess remarkably low levels of T cell costimulatory molecules as well as MHC class II molecules. regMΦ do not interfere with early T-cell activation but, via nitric oxide secretion, efficiently suppress T-cell proliferation. Furthermore, CD4+ T cells proliferating in the presence of regMΦ gain immunosuppressive capacity and MΦ isolated from day 3 nnLNs are T-cell immunosuppressive. Adoptive transfer of antigen-loaded regMΦ induce a profound antigen-specific immune suppression in vivo. Together our data show that nnLNSCs skew the differentiation of dendritic cells and their progenitors toward regMΦ, thus revealing a novel mechanism for local immune regulation.