Killer Artificial Antigen Presenting Cells (KaAPC) for Efficient In Vitro Depletion of Human Antigen-specific T Cells

Current treatment of T cell mediated autoimmune diseases relies mostly on strategies of global immunosuppression, which, in the long term, is accompanied by adverse side effects such as a reduced ability to control infections or malignancies. Therefore, new approaches need to be developed that target only the disease mediating cells and leave the remaining immune system intact. Over the past decade a variety of cell based immunotherapy strategies to modulate T cell mediated immune responses have been developed. Most of these approaches rely on tolerance-inducing antigen presenting cells (APC). However, in addition to being technically difficult and cumbersome, such cell-based approaches are highly sensitive to cytotoxic T cell responses, which limits their therapeutic capacity. Here we present a protocol for the generation of non-cellular killer artificial antigen presenting cells (KaAPC), which allows for the depletion of pathologic T cells while leaving the remaining immune system untouched and functional. KaAPC is an alternative solution to cellular immunotherapy which has potential for treating autoimmune diseases and allograft rejections by regulating undesirable T cell responses in an antigen specific fashion.     

Autoimmunity: basic mechanisms and implications in endocrine diseases. Part II

Regulation of the immune response to self-antigens is a complex process that involves maintaining self-tolerance while preserving the capacity to exert an effective immune response. The primary mechanism that leads to self-tolerance is central tolerance. However, potential pathogenic autoreactive lymphocytes are normally present in the periphery of all individuals. This suggests the existence of mechanisms of peripheral tolerance that prevent the initiation of autoimmune diseases by limiting the activation of autoreactive lymphocytes. If these mechanisms of peripheral tolerance are impaired, the autoreactive lymphocytes may be activated and autoimmune diseases can develop. Several processes are involved in the maintenance of peripheral tolerance: the active suppression mediated by regulatory T cell populations, the different maturation state of antigen-presenting cells presenting the autoantigen to autoreactive lymphocytes, inducing tolerance instead of cell activation, the characteristics of B cell populations. A deeper comprehension of these mechanisms may lead to important therapeutic applications, such as the development of cellular vaccines for organ-specific autoimmune diseases. In addition, autoimmunity does not always have pathological consequences, but may exert a protective function, as suggested by several observations on the beneficial role of autoreactive T cells in central nervous system injury.     

Dendritic cells,chemokine receptors and autoimmune inflammatory diseases

Dendritic cells (DC) have been implicated in the induction of autoimmune diseases and have been identified in lesions associated with several autoimmune inflammatory diseases. Since DC are regarded as the professional antigen-presenting cell (APC) of the immune system and the only APC capable of activating na?ve T cells, they are likely to play a significant role in breaking tolerance of self-reactive lymphocytes and in supporting autoimmune responses in these diseases. A number of studies have revealed that small molecular weight chemotactic proteins known as chemokines are present within the autoimmune lesions and may contribute to the recruitment not only of DC populations, but also of immune cells such as T cells, B cells, neutrophils and monocytes into the site, and to the formation of organized lymphoid tissue structures within the target organ. The focus of this review will be a discussion of the role of chemokines in the recruitment of DC in human autoimmune inflammatory disorders, specifically the trafficking of DC into the inflammatory sites and the subsequent migration of differentiated DC from the inflammatory sites into the draining lymph nodes. Once DC are properly positioned within the lymph nodes, circulating antigen specific na?ve T cells can interact with DC and become activated, clonally expanded and stimulated to undergo differentiation into antigen-experienced memory T cells. Subsequent reactivation of memory T cells that enter the autoimmune lesions by DC present in the inflammatory lesion is thought to play a central role in tissue inflammation.     

Emerging role of immune cell network in autoimmune skin disorders: An update on pemphigus,vitiligo and psoriasis

Autoimmune skin diseases are a group of disorders that arise due to a deregulated immune system resulting in skin tissue destruction. In the majority of these conditions, either autoreactive immune cells or the autoantibodies are generated against self-antigens of the skin. Although the etiology of these diseases remains elusive, biochemical, genetic, and environmental factors such as infectious agents, toxins damage the skin tissue leading to self-antigen generation, autoantibody attack and finally results in autoimmunity of skin. Immune dysregulation, which involves predominantly T helper 1/17 (Th1/Th17) polarization and the inability of regulatory T cells to regress immune response, is implicated in autoimmune skin diseases.The emerging roles of immune cells, cytokines, and chemokines in the pathogenesis of common autoimmune skin diseases like pemphigus, vitiligo, and psoriasis are discussed in this review. The main focus is on the interplay between immune cell network including the innate and adaptive immune system, regulatory cells, immune checkpoints and recently identified tissue-resident memory cells (TRMs) in disease pathogenesis and relapse. We also attempt to highlight on the immune mechanisms common to these diseases which can be targeted for designing novel therapeutics.     

Restoring the balance: immunotherapeutic combinations for autoimmune disease

Autoimmunity occurs when T cells, B cells or both are inappropriately activated, resulting in damage to one or more organ systems. Normally, high-affinity self-reactive T and B cells are eliminated in the thymus and bone marrow through a process known as central immune tolerance. However, low-affinity self-reactive T and B cells escape central tolerance and enter the blood and tissues, where they are kept in check by complex and non-redundant peripheral tolerance mechanisms. Dysfunction or imbalance of the immune system can lead to autoimmunity, and thus elucidation of normal tolerance mechanisms has led to identification of therapeutic targets for treating autoimmune disease. In the past 15 years, a number of disease-modifying monoclonal antibodies and genetically engineered biologic agents targeting the immune system have been approved, notably for the treatment of rheumatoid arthritis, inflammatory bowel disease and psoriasis. Although these agents represent a major advance, effective therapy for other autoimmune conditions, such as type 1 diabetes, remain elusive and will likely require intervention aimed at multiple components of the immune system. To this end, approaches that manipulate cells ex vivo and harness their complex behaviors are being tested in preclinical and clinical settings. In addition, approved biologic agents are being examined in combination with one another and with cell-based therapies. Substantial development and regulatory hurdles must be overcome in order to successfully combine immunotherapeutic biologic agents. Nevertheless, such combinations might ultimately be necessary to control autoimmune disease manifestations and restore the tolerant state.KEY WORDS: Tolerance, Autoimmune, Biologic     

Immunoregulation by Vbeta specific antibodies in myasthenia gravis: mining physiological T cell homeostasis for TCR specific therapy.

Besides regulatory T cells, also comprising T cell receptor (CR)-specific T cells, it is increasingly evident that natural autoantibodies, among which anti-TCR antibodies represent additional immunomodulators in the immune system. We took advantage of myasthenia gravis (MG), a well-characterized antibody-mediated autoimmune disease, to demonstrate that without prior vaccination against TCR determinants, patients with MG present increased circulating anti-TCR antibodies directed to the dominant TCR used by pathogenic T cells. These findings, pointing to a regulatory protective role of anti-TCR antibodies, are discussed in the context of the mechanisms of action and the physiological role of anti-TCR antibodies in T cell homeostasis, and of the puzzling world of regulatory T cells. Natural anti-TCR antibodies are found in the serum of all individuals, with prevalence in physiological and pathological situations such as ageing, pregnancy, allograft transplantation, retroviral infection, and autoimmune diseases, including MG. The common link is the mounting of immune responses against alloantigens, pathogens or autoantigens, conferring on anti-TCR antibodies a broader role in controlling responses to any antigen (self or non-self) and more generally in T cell homeostasis. This homeostasis mechanism may well be exploited in therapeutic strategies based on TCR peptide vaccination in autoimmune diseases.     

Indexation as a novel mechanism of lymphocyte homeostasis: the number of CD4+CD25+ regulatory T cells is indexed to the number of IL-2-producing cells

To fulfill its mission, the immune system must maintain a complete set of different cellular subpopulations that play specific roles in immune responses. We have investigated the mechanisms regulating CD4+CD25+ regulatory T (Treg) cell homeostasis. We show that the expression of the high-affinity IL-2Ralpha endows these cells with the capacity to explore the IL-2 resource, ensuring their presence while keeping their number tied to the number of CD4+ T cells that produce IL-2. We show that such a homeostatic mechanism allows the increased expansion of T cells without causing disease. The indexing of Treg cells to the number of activated IL-2-producing cells may constitute a feedback mechanism that controls T cell expansion during immune responses, thus preventing autoimmune or lymphoproliferative diseases. The present study highlights that maintenance of proportions between different lymphocyte subsets may also be critical for the immune system and are under strict homeostatic control.     

Current understanding of the role of tyrosine kinase 2 signaling in immune responses

Immune system is a complex network that clears pathogens,toxic substrates,and cancer cells.Distinguishing self-antigens from non-self-antigens is critical for the immune cell-mediated response against foreign antigens.The innate immune system elicits an early-phase response to various stimuli,whereas the adaptive immune response is tailored to previously encountered antigens.During immune responses,B cells differentiate into antibody-secreting cells,while na?ve T cells differentiate into functionally specific effector cells[T helper 1(Th1),Th2,Th17,and regulatory T cells].However,enhanced or prolonged immune responses can result in autoimmune disorders,which are characterized by lymphocytemediated immune responses against self-antigens.Signal transduction of cytokines,which regulate the inflammatory cascades,is dependent on the members of the Janus family of protein kinases.Tyrosine kinase 2(Tyk2)is associated with receptor subunits of immune-related cytokines,such as type I interferon,interleukin(IL)-6,IL-10,IL-12,and IL-23.Clinical studies on the therapeutic effects and the underlying mechanisms of Tyk2 inhibitors in autoimmune or chronic inflammatory diseases are currently ongoing.This review summarizes the findings of studies examining the role of Tyk2 in immune and/or inflammatory responses using Tyk2-deficient cells and mice.     

Nucleic Acid Scavenging Polymers Inhibit Extracellular DNA-Mediated Innate Immune Activation without Inhibiting Anti-Viral Responses

Toll-like receptor (TLR) family members, 3, 7 and 9 are key components in initiation and progression of autoimmune disorders such as systemic lupus erythematosus (SLE). These TLRs are often referred to as nucleic acid-sensing TLRs based on their ability to recognize DNAs or RNAs produced by pathogens or damaged cells. During autoimmune disease progression these receptors recognize self nucleic acids as well as self nucleic acid-containing complexes and contribute to inflammatory cytokine production and subsequent enhancement of serum autoantibody levels. We have recently discovered that nucleic-acid scavenging polymers (NASPs) can neutralize the proinflammatory effects of nucleic acids. Here, we begin to explore what effects such NASPs have on normal immune function. We show that such NASPs can inhibit TLR activation without affecting nucleic acid-independent T cell activation. Moreover, we observe that stimulation of immune cells by encapsulated nucleic acids, such as those found in viral particles, is unaffected by NASPs. Thus NASPs only limit the activation of the immune system by accessible extra-cellular nucleic acid and do not engender non-specific immune suppression. These important findings suggest that NASPs represent a new approach toward anti-inflammatory drug development as these agents can potentially be utilized to block overt autoimmune disorders and inflammation while allowing normal immune responses to occur.     

"2-6-11" motif in heat shock protein 60 and central nervous system antigens: a preliminary study in multiple sclerosis patients

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS) with unknown etiology and pathogenesis. A local autoimmune process involving activation of autoreactive T cells against CNS protein components is likely crucial in the development of MS lesions. Myelin-reactive T cells are believed to be primed in the periphery during infections by antigens of bacterial or viral origin via molecular mimicry, a postulated mechanism that might account for the trigger of an autoimmune response on the basis of sequence homology between foreign and self determinants. Immune responses to heat shock proteins (hsp) have been implicated in the initiation or progression of a number of autoimmune diseases. Hsp may function as immunodominant targets during the immune response evoked by pathogens, and theoretically a cross-reactive response to sequences shared by these immunogens and autoantigens in the CNS may contribute to the pathogenesis of MS. We examined the immune response of peripheral blood mononuclear cells (PBMNc) from MS patients and healthy subjects elicited by peptides derived from hsp60 containing a common structural motif ("2-6-11" motif) already described, which is also present in CNS putative antigens. This structural pattern consists of an apolar residue or Lys at position 2, Pro always at position 6, and Glu, Asp or Lys at residue 11. Results reported here are indicative of maturation of peripheral blood monocytes towards a differentiated CD14(+)CD16(+)DR(+) cell and release of pro-inflammatory cytokines consistent with a Th1-like pattern. These are typical features exhibited by immune cells implicated in autoimmune responses.     

T cell hyporesponsiveness induced by oral administration of ovalbumin is associated with impaired NFAT nuclear translocation and p27kip1 degradation

Oral tolerance is an important physiological component of the immune system whereby the organism avoids dangerous reactions such as hypersensitivity to ingested food proteins and other luminal Ags which may cause tissue damage and inflammation. In addition, it has been shown in animal models and in humans that oral tolerance can be applied to controlling undesired immune responses, including autoimmune diseases, allergies, and organ transplant rejections. However, the molecular mechanisms of oral tolerance have been poorly defined. In this study, we investigated the molecular basis underlying the hyporesponsiveness of orally tolerant CD4 T cells using a TCR transgenic mouse system in which oral tolerance was induced by long-term feeding with high dose Ag. We demonstrate that the hyporesponsive state of the CD4 T cells was maintained by a selective impairment in the TCR-induced calcium/NFAT signaling pathway and in the IL-2R-induced degradation of p27(kip1) and cell cycle progression. Thus, physiological mucosal tolerance is revealed to be associated with a unique type of T cell hyporesponsiveness which differs from previously described anergic T cells.     

The enigma of arrival: [corrected] the entrance of auto-immune T lymphocytes in central nervous tissues and of their attack against the myelin structures

This paper describes the conditions under which autoimmune T cells enter the central nervous system to attack the local tissue. During a prodromal period preceding the onset of neurological disease, the CNS tissue is converted from its physiological immune-hostile state to an immune-friendly state. At the same time, in the peripheral immune system, activated autoimmune T cells undergo profound changes that allow them to enter through the blood-brain barrier and to finally interact with local autoantigen-presenting cells. Antigen recognition leads to T cell activation and the secretion of inflammatory mediators that are responsible for the development of the inflammatory lesions and the neurological deficits.     

From T to B and back again: positive feedback in systemic autoimmune disease

Systemic lupus erythematosus, a prototypical systemic autoimmune disease, is the result of a series of interactions within the immune system that ultimately lead to the loss of self-tolerance to nuclear autoantigens. Here, we present an integrated model that explains how self-tolerance is initially lost and how the loss of tolerance is then amplified and maintained as a chronic autoimmune state. Key to this model are the self-reinforcing interactions of T and B cells, which we suggest lead to perpetuation of autoimmunity as well as its spread to multiple autoantigen targets.     

Regulatory roles for NKT cell ligands in environmentally induced autoimmunity

The development of autoimmune diseases is frequently linked to exposure to environmental factors such as chemicals, drugs, or infections. In the experimental model of metal-induced autoimmunity, administration of subtoxic doses of mercury (a common environmental pollutant) to genetically susceptible mice induces an autoimmune syndrome with rapid anti-nucleolar Ab production and immune system activation. Regulatory components of the innate immune system such as NKT cells and TLRs can also modulate the autoimmune process. We examined the interplay among environmental chemicals and NKT cells in the regulation of autoimmunity. Additionally, we studied NKT and TLR ligands in a tolerance model in which preadministration of a low dose of mercury in the steady state renders animals tolerant to metal-induced autoimmunity. We also studied the effect of Sphingomonas capsulata, a bacterial strain that carries both NKT cell and TLR ligands, on metal-induced autoimmunity. Overall, NKT cell activation by synthetic ligands enhanced the manifestations of metal-induced autoimmunity. Exposure to S. capsulata exacerbated autoimmunity elicited by mercury. Although the synthetic NKT cell ligands that we used are reportedly similar in their ability to activate NKT cells, they displayed pronounced differences when coinjected with environmental agents or TLR ligands. Individual NKT ligands differed in their ability to prevent or break tolerance induced by low-dose mercury treatment. Likewise, different NKT ligands either dramatically potentiated or inhibited the ability of TLR9 agonistic oligonucleotides to disrupt tolerance to mercury. Our data suggest that these differences could be mediated by the modification of cytokine profiles and regulatory T cell numbers.     

Noble metals strip peptides from class II MHC proteins

Class II major histocompatibility complex (MHC) proteins are essential for normal immune system function but also drive many autoimmune responses. They bind peptide antigens in endosomes and present them on the cell surface for recognition by CD4(+) T cells. A small molecule could potentially block an autoimmune response by disrupting MHC-peptide interactions, but this has proven difficult because peptides bind tightly and dissociate slowly from MHC proteins. Using a high-throughput screening assay we discovered a class of noble metal complexes that strip peptides from human class II MHC proteins by an allosteric mechanism. Biochemical experiments indicate the metal-bound MHC protein adopts a 'peptide-empty' conformation that resembles the transition state of peptide loading. Furthermore, these metal inhibitors block the ability of antigen-presenting cells to activate T cells. This previously unknown allosteric mechanism may help resolve how gold(I) drugs affect the progress of rheumatoid arthritis and may provide a basis for developing a new class of anti-autoimmune drugs.     

Insights into the role of Toll-like receptors in modulation of T cell responses

The innate immune receptors, such as Toll-like receptors (TLRs), are intimately involved in the early sensing of invading microorganisms or their structural components. Engagement of TLRs with their ligands results in activation of several downstream intracellular pathways leading to activation of innate and adaptive immune system cells. It was initially thought that TLRs are primarily expressed by antigen-presenting cells (APCs), such as macrophages and dendritic cells, and that interactions between microbial ligands and TLRs in these cells will indirectly result in activation of cells of the adaptive immune system, especially T cells. However, it has now become evident that TLRs are also expressed by various T cell subsets, such as conventional αβT cells, regulatory T cells, and γδT cells as well as natural killer T cells. Importantly, it appears that at least in some of these T cell subsets, TLRs are functionally active, because stimulation of these cells with TLR agonists in the absence of APCs results in exertion of effector or regulatory functions of T cells. The present review attempts to summarize the recent findings related to TLR expression in different T cell subsets and the direct role of TLRs in the induction and regulation of T cell responses, including those responses that occur at mucosal surfaces. In addition, the potential use of TLR agonists for steering T cell responses as a prophylactic or therapeutic strategy in the context of infectious, allergic or autoimmune diseases is explored.     

Dendritic cells and interferon-mediated autoimmunity

Dendritic cells (DCs) are central cells of the immune responses. They can be considered as the most influential antigen-presenting cells in the body because of their unique role in initiating immunity against most types of antigens. Recent studies have clearly established that the state of maturation of DC can be crucial for the ability of these antigen-presenting cells to inhibit or induce T-cell-mediated autoimmune diseases. Type I interferon has been shown to be produced at very high amounts by a specific type of DC (pDC). In recent years, the study of multiple autoimmune diseases has pointed to a central role for type I interferon (IFN-I) in disease pathogenesis, in particular through the IFN-molecular signature deciphered in some of these diseases. One hypothesis would be that IFN directly affects multiple actors of the immune reaction such as T cells and B cells and that it can induce the unabated activation of peripheral dendritic cells. On the other hand, type II IFN has been considered as pathogenic in multiple autoimmune diseases leading to the paradigm of TH-1 type autoimmune diseases. The discovery of the TH-17 type of cells and the protective role IFN-gamma can exert on particular phases of these diseases urge one to re-evaluate this assumption.     

Immunological tolerance using synthetic peptides--basic mechanisms and clinical application

Dysregulation of T lymphocyte function underpins the development of autoimmune and allergic diseases. These autoantigen-, or allergen-reactive pathogenic T cells are rare within the entire immune repertoire and it is therefore desirable to develop more specific therapies than are currently in use to directly target these cells and avoid adverse side effects. The obvious approach is to use the antigens that are recognized to impose a state of T cell tolerance. T cells recognize antigens as peptide fragments and we can therefore produce the relevant antigens as synthetic peptides. It has been known for many years that the decision of the T cell to mount a productive response (immunity) or to remain silent (tolerance) is controlled by the form in which the antigen is administered. Antigen with adjuvant leads to immunity, whereas soluble antigen without adjuvant leads to tolerance. This paradigm has been used successfully to induce tolerance with soluble peptides, preventing several animal models of autoimmune and allergic disease. These findings obviously have exciting potential for translation to human diseases. However, the basic immune mechanisms that lead to tolerance versus immunity are only beginning to be unravelled. The "effector" phase of tolerance also remains controversial with evidence for T cell death, anergy and the development of immunoregulatory function. This latter possibility of specifically generating autoantigen- or allergen-reactive regulatory T cells is particularly attractive. Here we review recent advances in our understanding of the requirements for tolerance induction and the potential for establishing dominant immune-regulation with peptide therapy.     

Immune ablation and stem-cell therapy in autoimmune disease: Immunological reconstitution after high-dose immunosuppression and haematopoietic stem-cell transplantation

Studies on immunological reconstitution after immune ablation and stem-cell therapy may yield important clues to our understanding of the pathogenesis of human autoimmune disease, due to the profound effects of function and organization of the immune system. Such studies are also indispensable when linking clinical sequelae such as opportunistic infections to the state of immune deficiency that ensues after the treatment. Much has been learnt on these issues from comparable studies in haemato-oncological diseases, although it remains to be proven that the data obtained from these studies can be extrapolated to rheumatological autoimmune diseases. Preliminary results from pilot studies in various rheumatological conditions not only pointed to clinical efficacy of the new treatment modality but also unveiled marked effects on T-cell receptor repertoires of circulating T lymphocytes, on titres of autoantibodies and T- and B-cell subsets.     

Immune ablation and stem-cell therapy in autoimmune disease. Immunological reconstitution after high-dose immunosuppression and haematopoietic stem-cell transplantation

Studies on immunological reconstitution after immune ablation and stem-cell therapy may yield important clues to our understanding of the pathogenesis of human autoimmune disease, due to the profound effects of function and organization of the immune system. Such studies are also indispensable when linking clinical sequelae such as opportunistic infections to the state of immune deficiency that ensues after the treatment. Much has been learnt on these issues from comparable studies in haemato-oncological diseases, although it remains to be proven that the data obtained from these studies can be extrapolated to rheumatological autoimmune diseases. Preliminary results from pilot studies in various rheumatological conditions not only pointed to clinical efficacy of the new treatment modality but also unveiled marked effects on T-cell receptor repertoires of circulating T lymphocytes, on titres of autoantibodies and T- and B-cell subsets.