Acute rheumatic fever (ARF) is an autoimmune disease triggered by antibodies and T cells targeting the group A Streptococcus (GAS, Strep A) bacterium, often leading to rheumatic heart disease (RHD) and Sydenham's chorea. Long-term monthly penicillin injections are recognized as a cornerstone of public health programs to prevent Strep A reinfection and progression of ARF. However, compliance is poor, and better tools are required to slow disease progression. Preclinical evidence suggests that this can be achieved. Using a rat model that replicates post-streptococcal autoimmune complications, we explored the potential of low-dose interleukin-2 (LD-IL-2) as an immunotherapeutic intervention for ARF/RHD. In this model, injections of recombinant M protein from Strep A type 5 (rM5) to Lewis rats induce cardiac tissue inflammation, conduction abnormalities, and cross-reactive antibodies against cardiac and brain proteins central to disease pathogenesis. In animals injected with rM5 and treated with LD-IL-2, no cardiac functional or histological changes was observed. LD-IL-2 therapy effectively reduced the production of cross-reactive antibodies raised against host proteins and significantly increased regulatory T cells in the mediastinal lymph nodes. These novel findings suggest that LD-IL-2 will be an effective immunotherapeutic agent for treating ARF and has the potential to replace the standard monthly penicillin injections.

Post-streptococcal autoimmune syndromes, including acute rheumatic fever, rheumatic heart disease, and Sydenham's chorea, represent a significant yet often under-recognized health and economic burden. This is especially true in low-income countries and among Indigenous populations in high-income nations, where the disease burden is most severe. These conditions arise from an autoimmune response to group A Streptococcus infections, leading to long-term health complications, disability, and premature death. Despite their widespread impact, no vaccine is currently available to prevent reinfections, and no specific therapy exists to treat the resulting autoimmune process. This study uses a rat model of rheumatic heart disease to evaluate the potential of low-dose interleukin 2 therapy in improving clinical outcomes and reducing the incidence of autoimmune diseases triggered by streptococcal infections.

The discovery of interleukin-2 (IL-2) and its receptor (IL-2R) almost 50 years ago revolutionized immunology, marking a pivotal moment in understanding T cell biology and immune regulation. Initially identified as a T cell growth factor, IL-2 unveiled critical insights into cytokine-mediated immune cell proliferation and differentiation.

This review highlighted the characterization of IL-2R as a multi-chain receptor complex set a precedent for decoding cytokine receptor signaling. The unique interplay between IL-2 and its high-affinity receptor component, IL-2Rα, epitomizes the principle of specificity and efficiency in cytokine signaling, enabling precise immune modulation. Regulatory T cells (Tregs) exploit IL-2Rα high affinity to outcompete effector T cells for IL-2, ensuring immune tolerance and preventing autoimmunity.

Despite its foundational role in immune homeostasis, leveraging IL-2 for therapeutic purposes has proven challenging.

IL-2-based therapies hold transformative potential in autoimmunity, cancer immunology, and transplantation, yet they remain elusive due to the complex balance between immunostimulatory and immunosuppressive effects. This review explores the milestones in IL-2 biology, its dualistic functions, and the ongoing quest to harness its therapeutic promise.

Antigen-specific therapies have a long history in the treatment of allergy but have not been successful in autoimmunity. However, in the past 20 years, advances in the definition of the self-antigens that promote autoimmunity and the growing understanding of the mechanisms that maintain tolerance in health but fail in autoimmunity have led to antigen-specific approaches being considered for the treatment of autoimmune diseases. The core goal of each antigen-specific treatment approach is to remove the immune response that promotes autoimmunity whilst sparing protective responses. Approaches to antigen-specific therapy range from targeted deletion of autoreactive lymphocytes to tolerization of autoreactive T cells and active inhibition of autoimmune responses. Technologies such as vaccines, nanoparticles, cell-based therapies and gene editing are being harnessed to achieve these goals. Remaining challenges include the selection of the best antigen to target, modality and timing of administration of these therapies and the disease in which the therapies are used; overcoming these challenges will be vital to move antigen-specific therapies forward. Once established, antigen-specific therapy has the potential to be applied broadly in the area of autoimmunity.

Regulatory T (Treg) cells are a suppressive subset of CD4+ T cells that maintain immune homeostasis and restrain inflammation. Three decades after their discovery, the promise of strategies to harness Treg cells for therapy has never been stronger. Multiple clinical trials seeking to enhance endogenous Treg cells or deliver them as a cell-based therapy have been performed and hint at signs of success, as well as to important limitations and unanswered questions. Strategies to deplete Treg cells in cancer are also in active clinical testing. Furthermore, multi-dimensional methods to interrogate the biology of Treg cells are leading to a refined understanding of Treg cell biology and new approaches to harness tissue-specific functions for therapy. A new generation of Treg cell clinical trials is now being fuelled by advances in nanomedicine and synthetic biology, seeking more precise ways to tailor Treg cell function. This Review will discuss recent advances in our understanding of human Treg cell biology, with a focus on mechanisms of action and strategies to assess outcomes of Treg cell-targeted therapies. It highlights results from recent clinical trials aiming to enhance or inhibit Treg cell activity in a variety of diseases, including allergy, transplantation, autoimmunity and cancer, and discusses ongoing strategies to refine these approaches.

Regulatory T (Treg) cells are pivotal in immune tolerance to allergens. Low-dose IL-2 (IL-2LD) activates Treg cells.

Our aim was to assess IL-2LD efficacy for controlling clinical responses to allergen exposures.

RHINIL-2 was a phase-2a, randomized, double-blind, placebo-controlled trial. Patients with allergic rhinitis to birch pollen (BP) were included; 66% of them had concomitant asthma. All had a total nasal symptom score (TNSS) of 5 or more following nasal exposure to BP in an environmental exposure chamber. Patients received 1 MUI per day of IL-2 (n = 12) or placebo (n = 12) for 5 days, followed by weekly injections for 4 weeks. Clinical responses to subsequent BP exposures in the environmental exposure chamber were evaluated by using TNSS, the rhinitis visual analog scale (VAS), and spirometry. The primary efficacy end point was the difference in TNSS area under the curve (AUC) between inclusion and day 40.

IL-2LD treatment induced a significant expansion of Treg cells. The difference in TNSS AUC between inclusion and day 40 AUC in the IL-2 and placebo groups was not significant. TNSS and visual analog scale AUCs were significantly reduced from baseline to day 40 in the IL-2LD group only (P = .04 and P = .01, respectively). The ratio of FEV1 to forced vital capacity (FEV1P) and the forced midexpiratory flow (FEF25%-75%) showed improvement in the IL-2LD-treated versus in the groups given placebo at day 40 (P = .04 and P = .04, respectively). However, the short treatment duration used in this study could not have effects on specific IgE or IgG4 levels given their half-life. There were no severe treatment-related adverse events.

IL-2LD is well tolerated in patients with allergy, even in those with asthma, thus clearing the path for further therapeutic development. Our work suggests that Treg cells can safely attenuate an ongoing allergic response. It paves the way for larger studies with longer treatment periods, which are needed to properly evaluate the therapeutic potential of IL-2 in allergy.

Immune abnormalities including an insufficiency of regulatory T cells (Treg) and increased blood-based inflammatory markers have been observed in bipolar disorders (BD), particularly during depression. As Tregs are pivotal to control inflammation, Treg stimulation by low-dose IL-2 (IL-2LD) could have a therapeutic impact on bipolar depression. We performed a randomized, double-blind, placebo-controlled (2 active: 1 placebo) proof-of-concept trial of add-on IL-2LD in patients with bipolar depression. Patients received a placebo or IL-2LD (1MIU) once a day for 5 days, and then once a week for 4 weeks starting on week 2. The primary objective was to demonstrate a biological Treg response to IL-2LD assessed by fold increase in Treg percentage of CD4 + cells from baseline to day 5. Secondary objectives included safety assessment and mood improvement throughout the study period. This trial is registered with ClinicalTrials.gov, number NCT04133233. Fourteen patients with bipolar depression were included, with 4 receiving placebo and 10 IL-2LD. Baseline clinical and biological characteristics were balanced between groups. The primary evaluation criterion was met, with IL-2LD expanding 1.17 [95 % CI 1.01-1.34] vs 1.01 [95 % CI 0.90-1.12] (p = 0.0421) and activating Tregs. Secondary evaluation criteria were also met with significant improvements of depressive symptoms and global functioning from day-15 onwards in the IL-2LD treated patients. The treatment was well-tolerated, with no serious adverse events related to treatment. This proof-of-concept trial shows that stimulating Tregs in patients with bipolar depression is safe and associated with clinical improvements. This supports a pathophysiological role of inflammation in BD and warrants pursuing the evaluation of IL-2LD as an adjunct treatment of major mood disorders.

Wiskott-Aldrich syndrome (WAS) is an X-linked immunodeficiency condition caused by ablation of functional WAS protein (WASP) expression, and associated with susceptibility to infections, eczema, and autoimmunity. Regulatory T cell (Treg) defects are an important cause of autoimmunity in WAS. Currently, the mechanisms underlying cytoskeleton involvement in Treg-regulated autoimmunity remain unclear, and WAS is an excellent model for investigation of this question. Here, we examined patients with WAS and WASP knockout (WASp-/-) mice to uncover a new mechanism involving the actin nucleation promoting factor, WASP, in regulating Treg tolerance by modulating their surface IL-2 receptor (IL-2R) levels. Surface expression levels of IL-2R and its downstream signaling molecules, phosphoinositide 3-kinase/pSTAT5, are decreased in WASp-/- Tregs. Low dosage IL-2 combined with anti-IL-2 monoclonal antibody (IL2 complex) treatment can compensate for Treg deficiency in WAS in vitro and in vivo. Moreover, IL2 complex treatment relieved autoimmune colitis in WASp-/- mice. Reduced surface IL-2R is primarily caused by elevated IL-2R internalization and degradation, and lysosomal and endosomal genes associated with these processes are upregulated in WASp-/- Tregs. Finally, spatiotemporal analysis of dynamin and Neural Wiskott Aldrich Syndrome Protein (N-WASP) recruitment, by generating lipid bilayers and total internal reflection fluorescence microscopy, showed that WASP deficiency promoted IL-2R internalization and degradation by enhancing N-WASP activation. Consistently, N-WASP inhibition in Tregs using wiskostatin reduced IL-2R internalization. Together, our results reveal a novel intrinsic role of WASP in regulation of surface IL-2R dynamics in Tregs, highlighting a potential new therapeutic approach for autoimmune diseases.

Regulatory T cells (Tregs) are reduced in the peripheral blood and skin lesions of patients with bullous pemphigoid (BP). Low-dose interleukin 2 (IL-2) therapy can stimulate Tregs specifically, suggesting potential for the treatment of BP.

To evaluate the response to low-dose IL-2 therapy in the treatment of moderate-to-severe BP.

Forty-three patients with moderate-to-severe BP were included. The therapy included systemic corticosteroids with an initial dose of 0.5 mg/kg/d for moderate and 1.0 mg/kg/d for severe disease, respectively, combined with allowed immunosuppressants for the control group, whereas in addition to the same corticosteroid therapy, IL-2 (half million IU) was administered subcutaneously every other day for the treatment group for 8 weeks. The primary outcome was the number of days required to achieve disease control. Secondary outcomes included other clinical responses.

The number of days required to achieve disease control with the treatment group was (7.60 ± 3.00), which was shorter than in the control group (10.43 ± 3.06) (P = .008). The total amount of systemic corticosteroids was less, and no serious infections were detected in the treatment group.

Single center, open-label study with short duration and small size.

Our trial supports the potential of low-dose IL-2 therapy for patients with moderate-to-severe BP, which showed earlier treatment responses.

Atherosclerosis is the primary underlying cause of myocardial infarction and stroke. Atherosclerotic cardiovascular disease is characterized by a chronic inflammatory reaction in medium-to-large-sized arteries, with its onset and perpetuation driven by leukocytes infiltrating the subendothelial space. Activation of endothelial cells triggered by hyperlipidaemia and lipoprotein retention in the arterial intima initiates the accumulation of pro-inflammatory leukocytes in the arterial wall, fostering the progression of atherosclerosis. This inflammatory response is coordinated by an array of soluble mediators, namely cytokines and chemokines, that amplify inflammation both locally and systemically and are complemented by tissue-specific molecules that regulate the homing, adhesion and transmigration of leukocytes. Despite abundant evidence from mouse models, only a few therapies targeting leukocytes in atherosclerosis have been assessed in humans. The major challenges for the clinical translation of these therapies include the lack of tissue specificity and insufficient selectivity of inhibition strategies. In this Review, we discuss the latest research on receptor-ligand pairs and interactors that regulate leukocyte influx into the inflamed artery wall, primarily focusing on studies that used pharmacological interventions. We also discuss mechanisms that promote the resolution of inflammation and highlight how major findings from these research areas hold promise as potential therapeutic strategies for atherosclerotic cardiovascular disease.

Lactic acid bacteria (LAB) probiotics are health-promoting but their characteristics, safety profile and functional mechanisms are not fully understood. Hence, this study aimed to characterize some hypothetical proteins of the chicken-isolated Limosilactobacillus reuteri genome and unravel their IL-2 and IL-10-inducing potential to understand mechanisms of their immunological functionality for sustainable applications. The selected proteins were subjected to in silico analyses for transmembrane topology, sub-cellular localization, IL-2 and IL-10-inducing ability and IL-2 and IL-10 gene expression across various conditions. IL-2 and IL-10-inducing mutants were statistically analyzed using a one-way analysis of variance of a general linear model of SAS and statistical significance was set at p < 0.05. The analyzed proteins are stable under a wide temperature range. All the hypothetical proteins are IL-2 and IL-10-inducing but QHPv.2.12, QHPv.2.13 and QHPv.2.15 are non-immunogenic. The evaluated mutants are IL-2 and IL-10-inducers but QHPv.2.13 and QHPv.2.15 are not immunogenic. This study sheds light on understanding the functional mechanisms of chicken-isolated L. reuteri and suggests it or its proteins as potent candidates for feed additive and therapeutic purposes.

Interleukin-2 (IL-2) was one of the first cytokines discovered and its central role in T cell function soon led to the notion that the cytokine could specifically activate immune cells to combat cancer cells. Recombinant human IL-2 (recIL-2) belonged to the first anti-cancer immunotherapeutics that received marketing authorization and while it mediated anti-tumor effects in some cancer entities, treatment was associated with severe and systemic side effects. RecIL-2 holds an exceptional therapeutic potential, which can either lead to stimulation of the immune system - favorable during cancer treatment - or immunosuppression - used for treatment of inflammatory diseases such as autoimmunity. Due to these pleiotropic immune effects, recIL-2 therapy is still a hot topic in research and modified recIL-2 drug candidates show ameliorated efficacy and safety in pre-clinical and clinical studies. The Immune Safety Avatar (imSAVAR) consortium aims to systemically assess mechanisms leading to adverse events provoked by recIL-2 immunotherapy as a use case in order to aid safety evaluation of future recIL-2-based therapies. Here, we summarize the historical use of recIL-2 therapy, associated side effects, and describe the molecular basis of the dual role of IL-2. Finally, an overview of new recIL-2 compounds and delivery systems, which are currently being developed, will be given, highlighting a possible comeback of recIL-2 therapy.

Due to its stimulatory potential for immunomodulatory CD4+ regulatory T (Treg) cells, low-dose interleukin-2 (IL-2) immunotherapy has gained considerable attention for the treatment of autoimmune diseases. In this investigator-initiated single-arm non-placebo-controlled phase-2 clinical trial of low-dose IL-2 immunotherapy in systemic lupus erythematosus (SLE) patients, we generated a comprehensive atlas of in vivo human immune responses to low-dose IL-2. We performed an in-depth study of circulating and cutaneous immune cells by imaging mass cytometry, high-parameter flow cytometry, transcriptomics, and targeted serum proteomics. Low-dose IL-2 stimulated various circulating immune cells, including Treg cells with a skin-homing phenotype that appeared in the skin of SLE patients in close interaction with endothelial cells. Analysis of surface proteins and transcriptomes revealed different IL-2-driven Treg cell activation programs, including gut-homing CD38+, skin-homing HLA-DR+, and highly proliferative inflammation-homing CD38+ HLA-DR+ Treg cells. Collectively, these data define the distinct human Treg cell subsets that are responsive to IL-2 immunotherapy.

In the last decade, cell therapies have revolutionized the treatment of some diseases, earning the definition of being the "third pillar" of therapeutics. In particular, the infusion of regulatory T cells (Tregs) is explored for the prevention and control of autoimmune reactions and acute/chronic allograft rejection. Such an approach represents a promising new treatment for autoimmune diseases to recover an immunotolerance against autoantigens, and to prevent an immune response to alloantigens. The efficacy of the in vitro expanded polyclonal and antigen-specific Treg infusion in the treatment of a large number of autoimmune diseases has been extensively demonstrated in mouse models. Similarly, experimental work documented the efficacy of Treg infusions to prevent acute and chronic allograft rejections. The Treg therapy has shown encouraging results in the control of type 1 diabetes (T1D) as well as Crohn's disease, systemic lupus erythematosus, autoimmune hepatitis and delaying graft rejection in clinical trials. However, the best method for Treg expansion and the advantages and pitfalls with the different types of Tregs are not fully understood in terms of how these therapeutic treatments can be applied in the clinical setting. This review provides an up-to-date overview of Treg infusion-based treatments in autoimmune diseases and allograft transplantation, the current technical challenges, and the highlights and disadvantages of this therapeutic approaches."

Regulatory T cells (Treg), a subset of CD4 + T cells, are potent regulators of immune reactions, which have been shown to be a promising therapeutic alternative to toxic immunosuppressive drugs. Data support the utility of Treg in managing immunopathologies, including solid organ transplant rejection, graft-versus-host disease, and autoimmune disorders. Notably, reports suggest that interleukin-2 (IL-2) is critical to survival of Treg, which constitutively express high levels of CD25, that is, the IL-2 receptor α-chain, and are exquisitely sensitive to IL-2, even at very low concentrations in contrast to effector T cells, which only upregulate IL-2 receptor α-chain on activation. This has led to the notion of using low doses of exogenous IL-2 therapeutically to modulate the immune system, specifically Treg numbers and function. Here, we summarize developments of clinical experience with low-dose IL-2 (LD-IL-2) as a therapeutic agent. So far, no clinical data are available to support the therapeutic use of LD-IL-2 therapy in the solid organ transplant setting. For the latter, fine-tuning by biotechnological approaches may be needed because of the narrow therapeutic window and off-target effects of LD-IL-2 therapy and so to realize the therapeutic potential of this molecule.

Regulatory T cells (Tregs) are essential for maintaining immune homeostasis by serving as negative regulators of adaptive immune system effector cell responses. Reduced production or function of Tregs has been implicated in several human autoimmune diseases. The cytokine interleukin 2 plays a central role in promoting Treg differentiation, survival, and function in vivo and may therefore have therapeutic benefits for autoimmune diseases. mRNA-6231 is an investigational, lipid nanoparticle-encapsulated, mRNA-based therapy that encodes a modified human interleukin 2 mutein fused to human serum albumin (HSA-IL2m). Herein, we report the development of a semi-mechanistic kinetic-pharmacodynamic model to quantify the relationship between subcutaneous dose(s) of mRNA-6231, HSA-IL2m protein expression, and Treg expansion in nonhuman primates. The nonclinical kinetic-pharmacodynamic model was extrapolated to humans using allometric scaling principles and the physiological basis of pharmacological mechanisms to predict the clinical response to therapy a priori. Model-based simulations were used to inform the dose selection and design of the first-in-human clinical study (NCT04916431). The modeling approach used to predict human responses was validated when data became available from the phase I clinical study. This validation indicates that the approach is valuable in informing clinical decision-making.

CD4+CD25+Foxp3+ regulatory T cells (Tregs), a vital component of the immune system, are responsible for maintaining immune homeostasis and preventing excessive immune responses. This review explores the signaling pathways of the cytokines that regulate Treg cells, including transforming growth factor beta (TGF-β), interleukin (IL)-2, IL-10, and IL-35, which foster the differentiation and enhance the immunosuppressive capabilities of Tregs. It also examines how, conversely, signals mediated by IL-6 and tumor necrosis factor -alpha (TNF-α) can undermine Treg suppressive functions or even drive their reprogramming into effector T cells. The B7 family comprises indispensable co-stimulators for T cell activation. Among its members, this review focuses on the capacity of CTLA-4 and PD-1 to regulate the differentiation, function, and survival of Tregs. As Tregs play an essential role in maintaining immune homeostasis, their dysfunction contributes to the pathogenesis of autoimmune diseases. This review delves into the potential of employing Treg-based immunotherapy for the treatment of autoimmune diseases, transplant rejection, and cancer. By shedding light on these topics, this article aims to enhance our understanding of the regulation of Tregs by cytokines and their therapeutic potential for various pathological conditions.

Therapeutic strategies for autoimmune diseases have been based on the use of glucocorticoids and immunosuppressive agents that broadly suppress immune responses. Therefore, organ damage from long-term use and infections due to immunocompromised status have been significant issues. Safer immunosuppressants and biological agents are now available, but there is still an urgent need to develop specific drugs to replace glucocorticoids. T-lymphocytes, central players in immune responses, could be crucial targets in the treatment of autoimmune diseases. Extensive research has been conducted on the phenotypic changes of T-cells in systemic lupus erythematosus, which has led to the discovery of various therapeutic strategies. In this comprehensive review, we discuss novel treatment approaches and target molecules with expected effectiveness in humans and mice, based on research for lymphocytes involved in autoimmune diseases, especially T-cells in SLE.

Regulatory T (Treg) cells are essential for maintaining self-immune tolerance. Reduced numbers or functions of Treg cells have been involved in the pathogenesis of various autoimmune diseases and allograft rejection. Therefore, the approaches that increase the pool or suppressive function of Treg cells in vivo could be a general strategy to treat different autoimmune diseases and allograft rejection. Interleukin-2 (IL-2) is essential for the development, survival, maintenance, and function of Treg cells, constitutively expressing the high-affinity receptor of IL-2 and sensitive response to IL-2 in vivo. And low-dose IL-2 therapy in vivo could restore the imbalance between autoimmune response and self-tolerance toward self-tolerance via promoting Treg cell expansion and inhibiting follicular helper T (Tfh) and IL-17-producing helper T (Th17) cell differentiation. Currently, low-dose IL-2 treatment is receiving extensive attention in autoimmune disease and transplantation treatment. In this review, we summarize the biology of IL-2/IL-2 receptor, the mechanisms of low-dose IL-2 therapy in autoimmune diseases, the application in the progress of different autoimmune diseases, including Systemic Lupus Erythematosus (SLE), Type 1 Diabetes (T1D), Rheumatoid Arthritis (RA), Autoimmune Hepatitis (AIH), Alopecia Areata (AA), Immune Thrombocytopenia (ITP) and Chronic graft-versus-host-disease (GVHD). We also discuss the future directions to optimize low-dose IL-2 treatments.

Immune-inflammatory mechanisms are promising targets for antidepressant pharmacology. Immune cell abnormalities have been reported in mood disorders showing a partial T cell defect. Following this line of reasoning we defined an antidepressant potentiation treatment with add-on low-dose interleukin 2 (IL-2). IL-2 is a T-cell growth factor which has proven anti-inflammatory efficacy in autoimmune conditions, increasing thymic production of naïve CD4 + T cells, and possibly correcting the partial T cell defect observed in mood disorders. We performed a single-center, randomised, double-blind, placebo-controlled phase II trial evaluating the safety, clinical efficacy and biological responses of low-dose IL-2 in depressed patients with major depressive (MDD) or bipolar disorder (BD). 36 consecutively recruited inpatients at the Mood Disorder Unit were randomised in a 2:1 ratio to receive either aldesleukin (12 MDD and 12 BD) or placebo (6 MDD and 6 BD). Active treatment significantly potentiated antidepressant response to ongoing SSRI/SNRI treatment in both diagnostic groups, and expanded the population of T regulatory, T helper 2, and percentage of Naive CD4+/CD8 + immune cells. Changes in cell frequences were rapidly induced in the first five days of treatment, and predicted the later improvement of depression severity. No serious adverse effect was observed. This is the first randomised control trial (RCT) evidence supporting the hypothesis that treatment to strengthen the T cell system could be a successful way to correct the immuno-inflammatory abnormalities associated with mood disorders, and potentiate antidepressant response.

Systemic sclerosis (SSc) is a chronic autoimmune disease, with impaired immune response, increased fibrosis and endothelial dysfunction. Regulatory T cells (Tregs), which are essential to control inflammation, tissue repair and autoimmunity, have a decreased frequency and impaired function in SSc patients. Low-dose interleukin-2 (IL-2LD) can expand and activate Tregs and has, therefore, a therapeutic potential in SSc.

We aimed to assess the safety and biological efficacy of IL-2LD in patients with SSc.

As part of the TRANSREG open-label phase IIa basket trial in multiple autoimmune diseases, we studied nine patients with SSc without severe organ involvement. Patients received 1 million international units (MIU)/day of IL-2 for 5 days, followed by fortnightly injections for 6 months. Laboratory and clinical evaluations were performed between baseline and month 6.

At day 8, the primary endpoint (Treg frequency) was reached with a 1.8±0.5-fold increase of Treg levels among CD4+ T lymphocytes (p=0.0015). There were no significant changes in effector T cells nor in B cells. IL-2LD was well tolerated, and no serious adverse events related to treatment occurred. There was a globally stable measurement in the modified Rodnan skin score and Valentini score at month 6. Disease activity and severity measures, the quality of life evaluated by EuroQL-5D-5L and pulmonary function test parameters remained stable during the study period.

IL-2LD at a dosage of 1 MIU/day safely and selectively activates and expands Tregs. Clinical signs remain stable during the study period. This opens the door to properly powered phase II efficacy trials investigating IL-2LD therapeutic efficacy in SSc.

A Tregs insufficiency is central to autoimmune and inflammatory diseases pathophysiology and low dose interleukin-2 (IL-2LD) can specifically activate Tregs.

To assess IL-2LD therapeutic potential and select diseases for further clinical development, we performed an open-label, phase 2a, disease-finding, "basket trial" involving patients with one of 13 different autoimmune diseases.

81 patients treated with IL-2LD (1 million IU/day) for 5 days, followed by fortnightly injections. The first 48 patients received diluted Proleukin®, while the subsequent 33 received ready-to-use ILT-101®. The primary endpoint was the change in Tregs at day-8 compared to baseline. Key secondary endpoints included clinical efficacy assessments using the Clinical Global Impression (CGI) scale, disease-specific scores, and EuroQL-5D-5L.

Our study unveiled a universal and significant expansion and activation of Tregs, without concomitant Teffs activation, across all 13 autoimmune diseases. Both Proleukin® and ready-to-use ILT-101® demonstrated identical effects on Tregs. CGI scores reflecting activity, severity, and efficacy were significantly reduced in the overall patient population. Disease-specific clinical scores improved in five of the six disease cohorts with at least six patients, namely ankylosing spondylitis, systemic lupus erythematosus, Behçet's disease, Sjögren's syndrome, and systemic sclerosis. Urticaria was the only severe adverse event related to treatment.

IL-2LD was well-tolerated, exhibiting specific Treg activation and clinical improvements across the 13 autoimmune diseases.

Tregs stimulation by IL-2LD is a promising therapeutic strategy and IL-2LD holds considerable promise for integration into combinatorial therapeutic approaches.

Recently, increased interest in the use of Tregs as adoptive cell therapy for the treatment of autoimmune diseases and transplant rejection had led to several advances in the field. However, Treg cell therapies, while constantly advancing, indiscriminately suppress the immune system without the permanent stabilization of certain diseases. Genetically modified Tregs hold great promise towards solving these problems, but, challenges in identifying the most potent Treg subtype, accompanied by the ambiguity involved in identifying the optimal Treg source, along with its expansion and engineering in a clinical-grade setting remain paramount. This review highlights the recent advances in methodologies for the development of genetically engineered Treg cell-based treatments for autoimmune, inflammatory diseases, and organ rejection. Additionally, it provides a systematized guide to all the recent progress in the field and informs the readers of the feasibility and safety of engineered adoptive Treg cell therapy, with the aim to provide a framework for researchers involved in the development of engineered Tregs.

Natural cytokines are poorly suited as therapeutics for systemic administration due to suboptimal pharmacological and pharmacokinetic (PK) properties. Recombinant human interleukin-2 (rhIL-2) has shown promise for treatment of autoimmune (AI) disorders yet exhibits short systemic half-life and opposing immune responses that negate an appropriate therapeutic index.

A semi-synthetic microbial technology platform was used to engineer a site-specifically pegylated form of rhIL-2 with enhanced PK, specificity for induction of immune-suppressive regulatory CD4 + T cells (Tregs), and reduced stimulation of off-target effector T and NK cells. A library of rhIL-2 molecules was constructed with single site-specific, biorthogonal chemistry-compatible non-canonical amino acids installed near the interface where IL-2 engages its cognate receptor βγ (IL-2Rβγ) signaling complex. Biorthogonal site-specific pegylation and functional screening identified variants that retained engagement of the IL-2Rα chain with attenuated potency at the IL-2Rβγ complex.

Phenotypic screening in mouse identifies SAR444336 (SAR'336; formerly known as THOR-809), rhIL-2 pegylated at H16, as a potential development candidate that specifically expands peripheral CD4+ Tregs with upregulation of markers that correlate with their suppressive function including FoxP3, ICOS and Helios, yet minimally expands CD8 + T or NK cells. In non-human primate, administration of SAR'336 also induces dose-dependent expansion of Tregs and upregulated suppressive markers without significant expansion of CD8 + T or NK cells. SAR'336 administration reduces inflammation in a delayed-type hypersensitivity mouse model, potently suppressing CD4+ and CD8 + T cell proliferation.

SAR'336 is a specific Treg activator, supporting its further development for the treatment of AI diseases.

Regulatory T cells (Tregs) are a specialized subgroup of T-cell lymphocytes that is crucial for maintaining immune homeostasis and preventing excessive immune responses. Depending on their differentiation route, Tregs can be subdivided into thymically derived Tregs (tTregs) and peripherally induced Tregs (pTregs), which originate from conventional T cells after extrathymic differentiation at peripheral sites. Although the regulatory attributes of tTregs and pTregs partially overlap, their modes of action, protein expression profiles, and functional stability exhibit specific characteristics unique to each subset. Over the last few years, our knowledge of Treg differentiation, maturation, plasticity, and correlations between their phenotypes and functions has increased. Genetic and functional studies in patients with numeric and functional Treg deficiencies have contributed to our mechanistic understanding of immune dysregulation and autoimmune pathologies. This review provides an overview of our current knowledge of Treg biology, discusses monogenetic Treg pathologies and explores the role of Tregs in various other autoimmune disorders. Additionally, we discuss novel approaches that explore Tregs as targets or agents of innovative treatment options.

Engineered regulatory T (Treg) cells have emerged as precision therapeutics aimed at inducing immune tolerance while reducing the risks associated with generalized immunosuppression. This Viewpoint highlights the opportunities and challenges for engineered Treg cell therapies in treating autoimmune and other inflammatory diseases.

Interleukin-2 (IL-2) is a critical cytokine for T cell peripheral tolerance and immunity. Here, we review how IL-2 interaction with the high-affinity IL-2 receptor (IL-2R) supports the development and homeostasis of regulatory T cells and contributes to the differentiation of helper, cytotoxic, and memory T cells. A critical element for each T cell population is the expression of CD25 (Il2rα), which heightens the receptor affinity for IL-2. Signaling through the high-affinity IL-2R also reinvigorates CD8+ exhausted T (Tex) cells in response to checkpoint blockade. We consider the molecular underpinnings reflecting how IL-2R signaling impacts these various T cell subsets and the implications for enhancing IL-2-dependent immunotherapy of autoimmunity, other inflammatory disorders, and cancer.

The occurrence of many autoimmune diseases takes root on the disrupted balance among Treg cells, Teff cells, etc. Low-dose interleukin-2 (IL-2) cytokine demonstrates promising clinical efficacy in the expansion of Treg cells and the treatment of autoimmune diseases. However, its clinical application is hindered by the small therapeutic index and short half-life. Previous studies have shown that non-covalent complex of human IL-2 and anti-IL-2 antibody biases cytokine activity towards Treg cells and extends IL-2's half-life. The clinical translation of such complex is non-trivial. In this study, we discover an anti-human IL-2 antibody and engineer a covalently-linked single-agent fusion of human IL-2 and its antibody that selectively expands Treg cells and exhibits superior disease control activity in animal models of ulcerative colitis and systemic lupus erythematosus, with proper safety profile and good developability. These studies pave the road for its clinical development in diverse autoimmune diseases.

The interleukin-2 (IL-2) cytokine plays a crucial role in regulating immune responses and maintaining immune homeostasis. Its immunosuppressive effects have been harnessed therapeutically via administration of low cytokine doses. Low-dose IL-2 has shown promise in the treatment of various autoimmune and inflammatory diseases; however, the clinical use of IL-2 is complicated by its toxicity, its pleiotropic effects on both immunostimulatory and immunosuppressive cell subsets, and its short serum half-life, which collectively limit the therapeutic window. As a result, there remains a considerable need for IL-2-based autoimmune disease therapies that can selectively target regulatory T cells with minimal off-target binding to immune effector cells in order to prevent cytokine-mediated toxicities and optimize therapeutic efficacy. In this review, we discuss exciting advances in IL-2 engineering that are empowering the development of novel therapies to treat autoimmune conditions. We describe the structural mechanisms of IL-2 signaling, explore current applications of IL-2-based compounds as immunoregulatory interventions, and detail the progress and challenges associated with clinical adoption of IL-2 therapies. In particular, we focus on protein engineering approaches that have been employed to optimize the regulatory T-cell bias of IL-2, including structure-guided or computational design of cytokine mutants, conjugation to polyethylene glycol, and the development of IL-2 fusion proteins. We also consider future research directions for enhancing the translational potential of engineered IL-2-based therapies. Overall, this review highlights the immense potential to leverage the immunoregulatory properties of IL-2 for targeted treatment of autoimmune and inflammatory diseases.

Viral infection poses a significant threat to human health. In addition to the damage caused by viral replication, the immune response it triggers often leads to more serious adverse consequences. After the occurrence of viral infection, in addition to the adverse consequences of infection, chronic infections can also lead to virus-related autoimmune diseases and tumours. At the same time, the immune response triggered by viral infection is complex, and dysregulated immune response may lead to the occurrence of immune pathology and macrophage activation syndrome. In addition, it may cause secondary immune suppression, especially in patients with compromised immune system, which could lead to the occurrence of secondary infections by other pathogens. This can often result in more severe clinical outcomes. Therefore, regarding the treatment of viral infections, restoring the balance of the immune system is crucial in addition to specific antiviral medications. In recent years, scientists have made an interesting finding that low dose IL-2 (ld-IL-2) could potentially have a crucial function in regulating the immune system and reducing the chances of infection, especially viral infection. Ld-IL-2 exerts immune regulatory effects in different types of viral infections by modulating CD4+ T subsets, CD8+ T cells, natural killer cells, and so on. Our review summarised the role of IL-2 or IL-2 complexes in viral infections. Ld-IL-2 may be an effective strategy for enhancing host antiviral immunity and preventing infection from becoming chronic; additionally, the appropriate use of it can help prevent excessive inflammatory response after infection. In the long term, it may reduce the occurrence of infection-related autoimmune diseases and tumours by promoting the restoration of early immune homeostasis. Furthermore, we have also summarised the application of ld-IL-2 in the context of autoimmune diseases combined with viral infections; it may be a safe and effective strategy for restoring immune homeostasis without compromising the antiviral immune response. In conclusion, focusing on the role of ld-IL-2 in viral infections may provide a new perspective for regulating immune responses following viral infections and improving prognosis.

Long-term organ transplant survival remains suboptimal, and life-long immunosuppression predisposes transplant recipients to an increased risk of infection, malignancy, and kidney toxicity. Promoting the regulatory arm of the immune system by expanding Tregs may allow immunosuppression minimization and improve long-term graft outcomes. While low-dose IL-2 treatment can expand Tregs, it has a short half-life and off-target expansion of NK and effector T cells, limiting its clinical applicability. Here, we designed a humanized mutein IL-2 with high Treg selectivity and a prolonged half-life due to the fusion of an Fc domain, which we termed mIL-2. We showed selective and sustainable Treg expansion by mIL-2 in 2 murine models of skin transplantation. This expansion led to donor-specific tolerance through robust increases in polyclonal and antigen-specific Tregs, along with enhanced Treg-suppressive function. We also showed that Treg expansion by mIL-2 could overcome the failure of calcineurin inhibitors or costimulation blockade to prolong the survival of major-mismatched skin grafts. Validating its translational potential, mIL-2 induced a selective and sustainable in vivo Treg expansion in cynomolgus monkeys and showed selectivity for human Tregs in vitro and in a humanized mouse model. This work demonstrated that mIL-2 can enhance immune regulation and promote long-term allograft survival, potentially minimizing immunosuppression.

Although FOXP3+ regulatory T cells (Treg) depend on IL-2 produced by other cells for their survival and function, the levels of IL-2 in inflamed tissue are low, making it unclear how Treg access this critical resource. Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2. HPSE-/- Treg have impaired stability and function in vivo, including in the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis. Conversely, endowing monoclonal antibody-directed chimeric antigen receptor (mAbCAR) Treg with HPSE enhances their ability to access HS-sequestered IL-2 and their ability to suppress neuroinflammation in vivo. Together, these data identify a role for HPSE and the ECM in immune tolerance, providing new avenues for improving Treg-based therapy of autoimmunity.

Primary biliary cholangitis (PBC) is a chronic autoimmune disease characterized by immune-mediated destruction of intrahepatic small bile ducts. CD8 T cells play a critical role in biliary destruction. However, regulatory T cells (Tregs) have also been identified in the portal tracts of PBC patients. This study tested the hypothesis that hepatic Tregs in PBC were dysfunctional in suppressing immune responses in disease by using our human PBC-like autoimmune cholangitis (AIC) mouse model induced by 2-octynoic acid-conjugated ovalbumin (2-OA-OVA). Our results showed that female and male mice immunized with 2-OA-OVA developed AIC; however, female AIC mice had more severe liver inflammation and fibrosis than male AIC mice. Levels of functional effector CD8 T cells and their chemoattractants, CXCL9 and CXCL10, in the liver were markedly elevated in female AIC mice than in male AIC mice. These results reinforce that CD8 T cells are the primary effector cells in PBC. The number of hepatic Tregs in AIC mice was also higher than in saline-treated mice, but there was no difference between male and female AIC mice. The suppressive function of AIC Tregs was evident despite a discrepancy in the changes in their co-inhibitory receptors and inhibitory cytokines. However, the expansion of hepatic Tregs by low-dose IL-2 treatment did not reduce immune responses to AIC, which may be due to the dysfunction of Tregs in inhibiting T cells. In conclusion, the function of Tregs in the inflamed liver of PBC was insufficient, and low-dose IL-2 treatment could not restore their function to suppress pathological immune responses. Transferring normal Tregs or directly targeting effector CD8 T cells may be beneficial for treating PBC.

Patients with chronic graft versus host disease (cGVHD) have low circulating regulatory T cells (Tregs). Interleukin-2(IL-2) is a growth factor for Tregs, and clinical trials have explored its use in cGVHD patients.

Here we will discuss the biology of IL-2, its rationale for use and results of clinical trials in cGVHD. We also describe its mechanisms of action and alteration in gene expression in T-cell subsets after treatment with low dose IL-2 and photopheresis.

Clinical trials using Low dose IL-2 have been done at single centers in small patient series. The majority of the clinical responses seen with IL-2 in cGVHD are classified as partial responses and efficacy as a single agent is limited. Compared to currently approved oral therapies, it has to be administered subcutaneously and requires specialized processing for compounding and storage limiting its widespread use. Its use is associated with constitutional symptoms and local injection site reactions. Local reactions can be easily managed by supportive care practices like rotation of injection sites and premeditations, constitutional symptoms resolve with, dose reduction (25-50%) allowing for continued therapy. Additional studies are needed to define optimal combination strategies with approved agents. Longer acting formulations of IL-2 that require less frequent dosing may also improve patient compliance.

FLT3-L-dependent classical dendritic cells (cDCs) recruit anti-tumor and tumor-protecting lymphocytes. We evaluate cancer growth in mice with low, normal, or high levels of cDCs. Paradoxically, both low or high numbers of cDCs improve survival in mice with melanoma. In low cDC context, tumors are restrained by the adaptive immune system through influx of effector T cells and depletion of Tregs and NK cells. High cDC numbers favor the innate anti-tumor response, with massive recruitment of activated NK cells, despite high Treg infiltration. Anti CTLA-4 but not anti PD-1 therapy synergizes with FLT3-L therapy in the cDCHi but not in the cDCLo context. A combination of cDC boost and Treg depletion dramatically improves survival of tumor-bearing mice. Transcriptomic data confirm the paradoxical effect of cDC levels on survival in several human tumor types. cDCHi-TregLo state in such patients predicts best survival. Modulating cDC numbers via FLT3 signaling may have therapeutic potential in human cancer.

Regulatory T cells (Tregs) are fundamental to maintaining immune homeostasis by inhibiting immune responses to self-antigens and preventing the excessive activation of the immune system. Their functions extend beyond immune surveillance and subpopulations of tissue-resident Treg cells can also facilitate tissue repair and homeostasis. The unique ability to regulate aberrant immune responses has generated the concept of harnessing Tregs as a new cellular immunotherapy approach for reshaping undesired immune reactions in autoimmune diseases and allo-responses in transplantation to ultimately re-establish tolerance. However, a number of issues limit the broad clinical applicability of Treg adoptive immunotherapy, including the lack of antigen specificity, heterogeneity within the Treg population, poor persistence, functional Treg impairment in disease states, and in vivo plasticity that results in the loss of suppressive function. Although the early-phase clinical trials of Treg cell therapy have shown the feasibility and tolerability of the approach in several conditions, its efficacy has remained questionable. Leveraging the smart tools and platforms that have been successfully developed for primary T cell engineering in cancer, the field has now shifted towards "next-generation" adoptive Treg immunotherapy, where genetically modified Treg products with improved characteristics are being generated, as regards antigen specificity, function, persistence, and immunogenicity. Here, we review the state of the art on Treg adoptive immunotherapy and progress beyond it, while critically evaluating the hurdles and opportunities towards the materialization of Tregs as a living drug therapy for various inflammation states and the broad clinical translation of Treg therapeutics.

Sjögren's Disease, SjD, is a systemic autoimmune disorder characterized by reduced function of the salivary and lacrimal glands. Patients suffer from dryness, fatigue, and pain and may present with or without extra-glandular organ involvement. Symptoms limit SjD patients' quality of life and are the most difficult to improve with therapy. SjD patients are heterogeneous and clustering them into biologically similar subgroups might improve the efficacy of therapies. The need for therapies that address both the symptoms and extra glandular organ involvement of SjD presents an unmet opportunity that has recently attracted a growing interest in the pharmaceutical industry.

The goal of this report is to review recent phase II/III studies in SjD. To accomplish our goal, we performed a literature search for phase II/III studies and abstracts recently presented at conferences.

This review allows updates the reader on the multitude of recent phase II/III clinical trials. We speculate on how subtypes of SjD will drive future therapeutic targeting and inform pathogenesis.

The cytokine interleukin-2 (IL-2) has the potential to treat autoimmune disease but is limited by its modest specificity toward immunosuppressive regulatory T (Treg) cells. IL-2 receptors consist of combinations of α, β, and γ chains of variable affinity and cell specificity. Engineering IL-2 to treat autoimmunity has primarily focused on retaining binding to the relatively Treg-selective, high-affinity receptor while reducing binding to the less selective, low-affinity receptor. However, we found that refining the designs to focus on targeting the high-affinity receptor through avidity effects is key to optimizing Treg selectivity. We profiled the dynamics and dose dependency of signaling responses in primary human immune cells induced by engineered fusions composed of either wild-type IL-2 or mutant forms with altered affinity, valency, and fusion to the antibody Fc region for stability. Treg selectivity and signaling response variations were explained by a model of multivalent binding and dimer-enhanced avidity-a combined measure of the strength, number, and conformation of interaction sites-from which we designed tetravalent IL-2-Fc fusions that had greater Treg selectivity in culture than do current designs. Biasing avidity toward IL2Rα with an asymmetrical multivalent design consisting of one α/β chain-binding and one α chain-binding mutant further enhanced Treg selectivity. Comparative analysis revealed that IL2Rα was the optimal cell surface target for Treg selectivity, indicating that avidity for IL2Rα may be the optimal route to producing IL-2 variants that selectively target Tregs.

Engineered cytokine-based approaches for immunotherapy of cancer are poised to enter the clinic, with IL-12 being at the forefront. However, little is known about potential mechanisms of resistance to cytokine therapies. We found that orthotopic murine lung tumors were resistant to systemically delivered IL-12 fused to murine serum albumin (MSA, IL12-MSA) because of low IL-12 receptor (IL-12R) expression on tumor-reactive CD8+ T cells. IL2-MSA increased binding of IL12-MSA by tumor-reactive CD8+ T cells, and combined administration of IL12-MSA and IL2-MSA led to enhanced tumor-reactive CD8+ T cell effector differentiation, decreased numbers of tumor-infiltrating CD4+ regulatory T cells, and increased survival of lung tumor-bearing mice. Predictably, the combination of IL-2 and IL-12 at therapeutic doses led to significant dose-limiting toxicity. Administering IL-12 and IL-2 analogs with preferential binding to cells expressing Il12rb1 and CD25, respectively, led to a significant extension of survival in mice with lung tumors while abrogating dose-limiting toxicity. These findings suggest that IL-12 and IL-2 represent a rational approach to combination cytokine therapy whose dose-limiting toxicity can be overcome with engineered cytokine variants.