Antigen-specific immunotherapy is considered the holy-grail for treatment of autoimmune diseases. However, unlike the unattainable myth of Arthurian legend, effective antigen-specific immunotherapy is now being realised through clinical trials in patients. This review describes the various approaches being taken, how antigens are being designed for therapy and carriers created for their delivery. A critical assessment is made concerning the need for such carrier systems.

In multiple sclerosis (MS) T cells aberrantly recognize self-peptides of the myelin sheath and attack the central nervous system (CNS). Antigen-specific peptide immunotherapy, which aims to restore tolerance while avoiding the use of non-specific immunosuppressive drugs, is a promising approach to combat autoimmune disease, but the cellular mechanisms behind successful therapy remain poorly understood. Myeloid-derived suppressor cells (MDSCs) have been studied intensively in the field of cancer and to a lesser extent in autoimmunity. Because of their suppressive effect on the immune system in cancer, we hypothesized that the development of MDSCs and their interaction with CD4(+) T cells could be beneficial for antigen-specific immunotherapy. Hence, changes in the quantity, phenotype and function of MDSCs during tolerance induction in our model of MS were evaluated. We reveal, for the first time, an involvement of a subset of MDSCs, known as polymorphonuclear (PMN)-MDSCs, in the process of tolerance induction. PMN-MDSCs were shown to adopt a more suppressive phenotype during peptide immunotherapy and inhibit CD4(+) T-cell proliferation in a cell-contact-dependent manner, mediated by arginase-1. Moreover, increased numbers of tolerogenic PMN-MDSCs, such as observed over the course of peptide immunotherapy, were demonstrated to provide protection from disease in a model of experimental autoimmune encephalomyelitis.

A key concern for the RTS,S/AS01 malaria vaccine is the higher meningitis incidence in children aged 5–17 months who were given the intervention than in age-matched controls,1RTS,S Clinical Trials PartnershipEfficacy and safety of RTS,S/AS01 malaria vaccine with or without a booster dose in infants and children in Africa: final results of a phase 3, individually randomised, controlled trial.Lancet. 2015; 386: 31-45Summary Full Text Full Text PDF PubMed Scopus (808) Google Scholar with 11 cases in the R3R group (who were given three doses of RTS,S/AS01 plus a booster dose), ten cases in the R3C group (who were given three doses of RTS,S/AS01 plus a comparator vaccine), and one case in the control group (who were given four doses of a comparator vaccine). Characteristics of the meningitis safety signal include disease development up to roughly 1100 days after doses 1, 2, and 3, and 500 days after dose 4; signal absence in infants aged 6–12 weeks; occurrence of cases in eight of 11 trial sites;1RTS,S Clinical Trials PartnershipEfficacy and safety of RTS,S/AS01 malaria vaccine with or without a booster dose in infants and children in Africa: final results of a phase 3, individually randomised, controlled trial.Lancet. 2015; 386: 31-45Summary Full Text Full Text PDF PubMed Scopus (808) Google Scholar, 2Smith P on behalf of the Joint Technical Expert Group (JTEG)JTEG's summary of RTS,S/AS01 clinical trial data.http://www.who.int/immunization/sage/meetings/2015/october/Smith_Malaria_Review_RTS_S_AS01_print.pdfGoogle Scholar and low incidence (relative to baseline) in the control group, rather than high incidence (relative to baseline) in the intervention group.3WHOEstimating meningitis hospitalization rates for sentinel hospitals conducting surveillance of invasive bacterial vaccine-preventable diseases.Wkly Epidemiol Rep. 2013; 88: 471PubMed Google Scholar Additionally, more cerebral malaria cases were reported in children aged 5–17 months in the R3R and R3C groups (43 cases) than in the control group (10 cases). Calculations again suggest that, compared with baseline, cerebral malaria incidence was lower than expected in the control group, rather than higher than expected in the intervention group.4Roca-Feltrer A Carneiro I Armstrong Schellenberg JRM Estimates of the burden of malaria morbidity in Africa in children under the age of 5 years.Trop Med Int Health. 2008; 13: 771-783Crossref PubMed Scopus (69) Google Scholar Interpretation of these findings has so far focused on three possibilities: first, a chance finding, since meningitis was one of dozens of adverse events monitored; second, systematic bias despite the randomised, blinded trial design; and third, an adverse effect from the RTS,S/AS01 vaccine—eg, through effects on the blood–brain barrier or choroid plexus. The third hypothesis would unify the meningitis and cerebral malaria signals into an overall CNS infection signal and explain some epidemiological findings. However, it would not explain the signal absence in infants aged 6–12 weeks or the relatively low incidence in children aged 5–17 months in the control group. A fourth possibility exists. Unlike young infants, who received the meningococcal conjugate vaccine, older controls received rabies vaccine. The hypothesis that rabies vaccine provided non-specific protection against CNS disease, rather than that RTS,S/AS01 increased risk, is consistent with the main epidemiological features of the safety signal. Indirect vaccine effects have been reported previously—eg, reduced childhood mortality after immunisation with live measles virus.5Aaby P Jensen H Samb B et al.Differences in female-male mortality after high-titre measles vaccine and association with subsequent vaccination with diphtheria-tetanus-pertussis and inactivated poliovirus: reanalysis of West African studies.Lancet. 2003; 361: 2183-2188Summary Full Text Full Text PDF PubMed Scopus (175) Google Scholar Although the absence of a biological mechanism for rabies vaccine to reduce CNS infection risk undermines such a hypothesis, if proven the implications would be profound for trial design, understanding of non-specific vaccine effects, and rabies vaccine use. Therefore, we recommend accelerated research, including studies of animal and human responses to rabies vaccine and, if results are consistent with our hypothesis, randomised trials of rabies vaccine use. BDG works for Agence de Médecine Preventive, which receives grants from GlaxoSmithKline for work on malaria and malaria vaccines. AF reports grants and personal fees from GlaxoSmithKline, Sanofi-Pasteur MSD, Novartis, Pfizer, and Alios; and personal fees from Takeda, all of which are outside the submitted work and paid to his employers. DCW declares no competing interests.

Background & Aims: It is well-known that the liver can induce immune tolerance, yet this knowledge could, thus far, not be translated into effective treatments for autoimmune diseases. We have previously shown that liver sinusoidal endothelial cells (LSECs) could substantially contribute to hepatic tolerance through their ability to induce CD4+ Foxp3+ regulatory T cells (Tregs). Here, we explored whether the Treg-inducing potential of LSECs could be harnessed for the treatment of autoimmune disease.Methods: We engineered a polymeric nanoparticle (NP) carrier for the selective delivery of autoantigen peptides to LSECs in vivo. In the well-characterized autoimmune disease model of experimental autoimmune encephalomyelitis (EAE), we investigated whether administration of LSEC-targeting autoantigen peptide- loaded NPs could protect mice from autoimmune disease.Results: We demonstrate that NP-based autoantigen delivery to LSECs could completely and permanently prevent the onset of clinical EAE. More importantly, in a therapeutic approach, mice with already established EAE improved rapidly and substantially following administration of a single dose of autoantigen peptideloaded NPs, whereas the control group deteriorated. Treatment efficacy seemed to depend on Tregs. The Treg frequencies in the spleens of mice treated with autoantigen peptide-loaded NPs were significantly higher than those in vehicle-treated mice. Moreover, NP-mediated disease control was abrogated after Treg depletion by repeated administration of Treg-depleting antibody.Conclusion: Our findings provide proof of principle that the selective delivery of autoantigen peptides to LSECs by NPs can induce antigen-specific Tregs and enable effective treatment of autoimmune disease. These findings highlight the importance of Treg induction by LSECs for immune tolerance. (C) 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Adoptive transfer of antigen-specific, in vitro-induced Foxp3+ Treg (iTreg) cells protects against autoimmune disease. To generate antigen-specific iTreg cells at high purity, however, remains a challenge. Whereas polyclonal T cell stimulation with anti-CD3 and anti-CD28 antibody yields Foxp3+ iTreg cells at a purity of 90–95%, antigen-induced iTreg cells typically do not exceed a purity of 65–75%, even in a TCR-transgenic model. In a similar vein to thymic Treg cell selection, iTreg cell differentiation is influenced not only by antigen recognition and the availability of TGF-β but also by co-factors including costimulation and adhesion molecules. In this study, we demonstrate that blockade of the T cell integrin Leukocyte Function-associated Antigen-1 (LFA-1) during antigen-mediated iTreg cell differentiation augments Foxp3 induction, leading to approximately 90% purity of Foxp3+ iTreg cells. This increased efficacy not only boosts the yield of Foxp3+ iTreg cells, it also reduces contamination with activated effector T cells, thus improving the safety of adoptive transfer immunotherapy.

Clinically effective antigen-based immunotherapy must silence antigen-experienced effector T cells (Teff) driving ongoing immune pathology. Using CD4+ autoimmune Teff cells, we demonstrate that peptide immunotherapy (PIT) is strictly dependent upon sustained T cell expression of the co-inhibitory molecule PD-1. We found high levels of 5-hydroxymethylcytosine (5hmC) at the PD-1 (Pdcd1) promoter of non-tolerant T cells. 5hmC was lost in response to PIT, with DNA hypomethylation of the promoter. We identified dynamic changes in expression of the genes encoding the Ten-Eleven-Translocation (TET) proteins that are associated with the oxidative conversion 5-methylcytosine and 5hmC, during cytosine demethylation. We describe a model whereby promoter demethylation requires the co-incident expression of permissive histone modifications at the Pdcd1 promoter together with TET availability. This combination was only seen in tolerant Teff cells following PIT, but not in Teff that transiently express PD-1. Epigenetic changes at the Pdcd1 locus therefore determine the tolerizing potential of TCR-ligation.

In vitro induced Foxp3+ T regulatory (iTreg) cells form a novel and promising target for therapeutic tolerance induction. However, the potential of these cells as a target for the treatment of various immune diseases, as well as the factors involved in their development and function, remain debated. Here, we demonstrate in a myelin basic protein (MBP)-specific murine model of CNS autoimmune disease that adoptive transfer of antigen-specific iTreg cells ameliorates disease progression. Moreover, we show that the co-stimulatory molecule CTLA-4 mediates in vitro differentiation of iTreg cells. Finally, we demonstrate that the secreted, immunosuppressive cytokine IL-10 controls the ability of antigen-specific iTreg cells to suppress autoimmune disease. Overall, we conclude that antigen-specific iTreg cells, which depend on various immune regulatory molecules for their differentiation and function, represent a major target for effective immunotherapy of autoimmune disease.

Tissues of the CNS, such as the brain, optic nerves, and spinal cord, may be affected by a range of insults including genetic, autoimmune, infectious, or neurodegenerative diseases and cancer. The immune system is involved in the pathogenesis of many of these, either by causing tissue damage or alternatively by responding to disease and contributing to repair. It is clearly vital that cells of the immune system patrol the CNS and protect against infection. However, in contrast to other tissues, damage caused by immune pathology in the CNS can be irreparable. The nervous and immune systems have, therefore, coevolved to permit effective immune surveillance while limiting immune pathology. Here we will consider aspects of adaptive immunity in the CNS and the retina, both in the context of protection from infection as well as cancer and autoimmunity, while focusing on immune responses that compromise health and lead to significant morbidity.

Recent studies have expanded our understanding of the role of the anti-inflammatory cytokine interleukin (IL)-10, produced by multiple lineages of both human and murine T cells, in regulating the immune response. Here, we demonstrate that the small percentage of circulating CD4(+) T cells that secrete IL-10 can be isolated from human peripheral blood and, importantly, we have optimized a protocol to expand these cells in both antigen-specific and polyclonal manners. Expanded CD4(+)IL-10(+) T cells abrogate proliferation and T helper (Th) 1-like cytokine production in an antigen-specific manner, and to a lesser extent exhibit bystander suppressive capacity. CD4(+)IL-10(+) T cells are suppressive in a cell contact-dependent way, though they do not require secretion of IL-10 for their suppressive role in vitro. CD4(+)IL-10(+) T cells have an activated phenotype, with high expression of CD25, CD69, and cytotoxic T-lymphocyte antigen-4, and are largely FoxP3 negative. This novel method for the isolation and expansion of suppressive IL-10-secreting T cells has important implications both for further research and clinical therapeutic development.

Mesenchymal stem cells (MSCs) can abrogate the animal model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE), but whether this therapeutic effect occurs entirely through systemic immune modulation and whether CNS infiltration occurs after peripheral delivery are uncertain. We studied the clinical and neuropathologic effects of intravenously administered human MSCs (hMSCs) in C57BL/6 mice with EAE. Human MSCs significantly reduced the clinical disease severity, particularly in later disease. Large numbers of hMSCs migrated into gray and white matter at all levels of the spinal cord in both naive mice and mice with EAE. In the latter, hMSCs accumulated over time in demyelinated areas. There were 2 distinct morphological appearances of the hMSCs in the tissue, that is, rounded and less numerous process-bearing forms; very few expressed neural markers. The number of spinal cord white matter lesions and areas of white matter demyelination were reduced after hMSC treatment compared with control treatment. These findings show that central nervous system infiltration occurs after peripheral delivery of hMSCs, that they accumulate where there is myelin damage, and that they are associated with a reduced extent of demyelination. These data support a potential role for hMSCs in autologous cell therapy in multiple sclerosis.

It is generally acknowledged that cytotoxic T-lymphocyte–associated antigen-4 (CTLA-4/CD152) plays a pivotal role in the regulation of T-cell activation and the establishment of self-tolerance in the periphery. CTLA-4–deficient (CTLA-4KO) mice develop a lymphoproliferative disorder and die within 4 weeks of birth, suggesting a role for CTLA-4 in T-cell homeostasis or the development and activity of T-regulatory (Treg) cells. To study the role of CTLA-4 in the control of experimental autoimmune encephalomyelitis (EAE), we have generated a CTLA-4KO mouse in which >90% of all CD4 + T cells bear a Vβ8.2 transgenic T-cell receptor that is specific for myelin basic protein peptide Ac1–9 (ASQKRPSQR). These mice do not develop spontaneous lymphoproliferative disease or EAE and are resistant to disease induction. This correlates with a higher frequency of functional FoxP3 + Treg cells in the spleen and thymus of CTLA-4KO mice. The absence of CTLA-4–mediated suppression of CD28 signaling resulted in the early expression of FoxP3 on double-positive cells in the thymic cortex. We conclude that CTLA-4 is not essential for the peripheral function of FoxP3 + Treg cells but plays a pivotal role in their thymic selection.

Recent advances in immunology have led to exciting new possibilities in medicine. We now understand how to design vaccine capable of resetting the balance between immune effector and regulatory cells in people suffering from autoimmune and allergic diseases. These approaches are effective while avoiding the side effects associated with non-specific immune suppression.

The neuropeptide galanin is widely expressed by many differing subsets of neurons in the nervous system. There is a marked upregulation in the levels of the peptide in a variety of nerve injury models and in the basal forebrain of humans with Alzheimer's disease. Here we demonstrate that galanin expression is specifically and markedly upregulated in microglia both in multiple sclerosis (MS) lesions and shadow plaques. Galanin expression is also upregulated in the experimental autoimmune encephalomyelitis (EAE) model of MS, although solely in oligodendrocytes. To study whether the observed increase in expression of galanin in inflammatory demyelination might modulate disease activity, we applied the EAE model to a panel of galanin transgenic lines. Over-expression of galanin in transgenic mice (Gal-OE) abolishes disease in the EAE model, whilst loss-of-function mutations in galanin or galanin receptor-2 (GalR2) increase disease severity. The pronounced effects of altered endogenous galanin or GalR2 expression on EAE disease activity may reflect a direct neuroprotective effect of the neuropeptide via activation of GalR2, similar to that previously described in a number of neuronal injury paradigms. Irrespective of the mechanism(s) by which galanin alters EAE disease activity, our findings imply that galanin/GalR2 agonists may have future therapeutic implications for MS.

Maintaining tolerance of T cells to self-antigens is essential to avoid autoimmune disease. How self-reactive T cells are kept functionally inactive is, however, unknown. In this study, we show that early growth response gene 2 (Egr-2), a zinc-finger transcription factor, is expressed in CD44(high) T cells and controls their proliferation and activation. In the absence of Egr-2, CD44(high), but not CD44(low) T cells, are hyperreactive and hyperproliferative in vivo. The accumulation of activated CD4(+)CD44(high) T cells leads to the development of a late onset lupuslike autoimmune disease characterized by the accumulation of interferon (IFN)-gamma and interleukin (IL)-17-producing CD4(+) T cells, loss of tolerance to nuclear antigens, massive infiltration of T cells into multiple organs and glomerulonephritis. We found that the expression of cyclin-dependent kinase inhibitor p21cip1 was impaired in Egr-2-deficient T cells, whereas the expression of IFN-gamma and IL-17 in response to T cell receptor ligation was significantly increased, suggesting that Egr-2 activates the expression of genes involved in the negative regulation of T cell proliferation and inflammation. These results demonstrate that Egr-2 is an intrinsic regulator of effector T cells and controls the expansion of self-reactive T cells and development of autoimmune disease.

Tregs are important mediators of immune tolerance to self antigens, and it has been suggested that Treg inactivation may cause autoimmune disease. Therefore, immunotherapy approaches that aim to restore or expand autoantigen-specific Treg activity might be beneficial for the treatment of autoimmune disease. Here we report that Treg-mediated suppression of autoimmune disease can be achieved in vivo by taking advantage of the ability of the liver to promote immune tolerance. Expression of the neural autoantigen myelin basic protein (MBP) in the liver was accomplished stably in liver-specific MBP transgenic mice and transiently using gene transfer to liver cells in vivo. Such ectopic MBP expression induced protection from autoimmune neuroinflammation in a mouse model of multiple sclerosis. Protection from autoimmunity was mediated by MBP-specific CD4+CD25+Foxp3+ Tregs, as demonstrated by the ability of these cells to prevent disease when adoptively transferred into nontransgenic mice and to suppress conventional CD4+CD25- T cell proliferation after antigen-specific stimulation with MBP in vitro. The generation of MBP-specific CD4+CD25+Foxp3+ Tregs in vivo depended on expression of MBP in the liver, but not in skin, and occurred by TGF-beta-dependent peripheral conversion from conventional non-Tregs. Our findings indicate that autoantigen expression in the liver may generate autoantigen-specific Tregs. Thus, targeting of autoantigens to hepatocytes may be a novel approach to prevention or treatment of autoimmune diseases.

Multiple sclerosis is a currently incurable inflammatory demyelinating syndrome. Recent reports suggest that bone marrow derived mesenchymal stem cells may have therapeutic potential in experimental models of demyelinating disease, but various alternative mechanisms, ranging from systemic immune effects to local cell replacement, have been proposed. Here we used intraperitoneal delivery of human mesenchymal stem cells to help test (a) whether human cells can indeed suppress disease, and (b) whether CNS infiltration is required for any beneficial effect. We found pronounced amelioration of clinical disease but profoundly little CNS infiltration. Our findings therefore help confirm the therapeutic potential of mesenchymal stem cells, show that this does indeed extend to human cells, and are consistent with a peripheral or systemic immune effect of human MSCs in this model.

CD4+ CD25+ Foxp3+ regulatorische T Zellen (Treg) sind wichtige Mediatoren der Immuntoleranz gegen Selbst-Antigene, die im Thymus, aber auch durch noch nicht vollständig geklärte Mechanismen in der Peripherie gebildet werden. Eine Inaktivierung von Treg kann Autoimmunerkrankungen auslösen. Eine mögliche Immuntherapie von Autoimmunerkrankungen bestünde deshalb darin, Autoantigen-spezifische Treg zu aktivieren bzw. zu expandieren. Hier zeigen wir, dass die ektope Expression eines Autoantigens Myelin Basisches Protein (MBP) in der Leber zur Generierung von Autoantigen-spezifischen Treg führt, die in der Lage sind, eine Autoimmunerkrankung zu unterdrücken. Die ektope Expression des neuralen Autoantigens Myelin Basisches Protein (MBP) erfolgte dabei entweder konstitutiv in MBP-transgenen Mäusen oder transient durch hydrodynamischen Gentransfer eines MBP-Expressionsplasmids in Leberzellen. Beide Arten der MBP-Expression in der Leber induzierte die selektive Expansion MBP-spezifischer Treg, die einen vollständigen Schutz vor der Auslösung einer autoimmunen Encephalomyelitis, dem Tiermodell der humanen Multiplen Sklerose, vermittelten. Der Schutz konnte durch Transfer von Treg aus der Milz der Mäuse nach hepatischer MBP-Expression in Wildtyp-Mäuse übertragen werden. In vitro supprimierten die durch MBP-Expression in der Leber generierten Treg erheblich effektiver die Stimulation MBP-spezifischer CD4 Effektorzellen als Treg aus Wildtyp-Mäusen. Diese Befunde zeigen, dass die Leber eine physiologische Rolle bei der peripheren Generierung von Tregs zu spielen scheint. Die so induzierten Treg sind in der Lage, Autoimmunerkrankungen zu unterdrücken. Die Expression von Autoantigenen in Hepatozyten könnte ein neuartiger therapeutischer Ansatz zur Behandlung oder Prävention von Autoimmunerkrankungen sein.

It has recently been proposed that experimental autoimmune encephalomyelitis, once considered the classical Th1 disease, is predominantly Th17 driven. In this study we show that myelin-reactive Th1 preparations devoid of contaminating IL-17(+) cells are highly pathogenic. In contrast, Th17 preparations lacking IFN-gamma(+) cells do not cause disease. Our key observation is that only Th1 cells can access the noninflamed CNS. Once Th1 cells establish the experimental autoimmune encephalomyelitis lesion, Th17 cells appear in the CNS. These data shed important new light on the ability of Th1 vs Th17 cells to access inflamed vs normal tissue. Because the IL-17-triggered release of chemokines by stromal cells could attract many other immune cells, allowing Th17 cells to access the tissues only under conditions of inflammation may be a key process limiting (auto)immune pathology. This has major implications for the design of therapeutic interventions, many of which are now aiming at Th17 rather than Th1 cells.