IL-2 as a therapeutic target for the restoration of Foxp3+ regulatory T cell function in organ-specific autoimmunity: implications in pathophysiology and translation to human disease

Peripheral immune tolerance requires a finely controlled balance between tolerance to self-antigens and protective immunity against enteric and invading pathogens. Self-reactive T cells sometimes escape thymic clonal deletion, and can subsequently provoke autoimmune diseases such as type 1 diabetes (T1D) unless they are controlled by a network of tolerance mechanisms in the periphery, including CD4+ regulatory T cells (Treg) cells. CD4+ Treg cells are characterized by the constitutive expression of the IL-2Rα chain (CD25) and preferentially express the forkhead winged helix transcriptional regulator Foxp3. These cells have been shown to possess immunosuppressive properties towards various immune cell subsets and their defects are thought to contribute to many autoimmune disorders. Strong evidence shows that IL-2 is one of the important stimulatory signals for the development, function and fitness of Treg cells. The non-obese diabetic (NOD) mouse model, a prototypic model of spontaneous autoimmunity, mimics many features of human T1 D. Using this model, the contribution of the IL-2-IL-2R pathway to the development of T1 D and other autoimmune disorders has been extensively studied. In the past years, strong genetic and molecular evidence has indicated an essential role for the IL-2/IL-2R pathway in autoimmune disorders. Thus, the major role of IL-2 is to maintain immune tolerance by promoting Treg cell development, functional fitness and stability. Here we first summarize the genetic and experimental evidence demonstrating a role for IL-2 in autoimmunity, mainly through the study of the NOD mouse model, and analyze the cellular and molecular mechanisms of its action on Treg cells. We then move on to describe how this data can be translated to applications for human autoimmune diseases by using IL-2 as a therapeutic agent to restore Treg cell fitness, numbers and functions

The role of FOXP3 in human CD4+CD25+ regulatory T cell phenotype and function

Naturally occurring CD4+CD25High regulatory T cells (nTreg) have emerged as an essential component of self-tolerance. These cells arise in the thymus and represent 1-10% of CD4+ T cells in humans and mice. They constitutively express the FOXP3 transcription factor. The importance of FOXP3 for Treg cell differentiation and function is illustrated by the Immune-dysregulation, Polyendocrinopathy, Enteropathy, X-linked (IPEX) syndrome; a rare and severe systemic autoimmune disorder caused by mutations in the FOXP3 gene. Treg cell defects are suspected to underlie most other human autoimmune diseases. However the characterization of human nTreg cells has been hindered by the failure to discriminate them from other CD4+ T cells expressing similar markers in states of inflammation, as FOXP3 is expressed by non-suppressive T effector (Teff) cells upon activation. Thus the functional correlates of FOXP3 expression in human Treg cells are ill-defined. We hypothesize that human CD4+CD25HighFOXP3+ T cells represent a heterogeneous population comprised of several Treg and T cell subsets in which the expression of FOXP3 has different functional and phenotypic outcomes. Using a novel single-cell cloning approach, we show that indeed about 30% of memory FOXP3+ T cells are not suppressive, thus revealing a functional heterogeneity amongst CD4+CD25High T cells. These non-suppressive cells are phenotypically identical to their suppressive counterparts, but are found to down-regulate FOXP3 upon activation. By examining the transcriptome of these unstable FOXP3+ T cells, we show that they constitute a genuine, Treg-related, T cell subset, endowed with a specific gene expression signature. Moreover, we find evidence that the stability of the Treg lineage and FOXP3 expression are orchestrated by a higher-level of T-helper cell lineage regulation. Finally, we show that the IPEX-derived A384T mutation in FOXP3 abrogates the capacity of FOXP3 to elicit the suppressive function of Treg cells, while preserving its ability to control their phenotype. We thus provide the first demonstration of a functional dichotomy in the role of FOXP3 in human Treg cells. In conclusion, we demonstrate that FOXP3 is not an absolute marker of suppressive function in human T cells, precluding its sole use as a clinical marker for the monitoring of immunoregulation. Furthermore, we delineate subsets enclosed within FOXP3+ T cells which vary in function and stability, and characterize the hallmarks of these unstable FOXP3+ T cells. These findings will allow for the exploration of their contribution to immune homeostasis, and of their defects in autoimmune disorders.Les cellules T CD4+CD25+++ régulatrices dites « naturelles » (nTreg) constituent une composante essentielle de la tolérance immune au soi. Ces cellules se développent dans le thymus, et représentent 1-10% des cellules T CD4+ périphériques chez l'humain et la souris. Elles expriment constitutivement le facteur de transcription FOXP3, dont l'importance dans la différenciation et la fonction des cellules Treg est illustrée par le syndrome d'IPEX (Immuno-dérégulation, Poly-endocrinopathie, Entéropathie, liée a l'X), une condition rare et sévère d'auto-immunité systémique, causée par une mutation du gène foxp3. Des défaillances des cellules Treg sont aussi soupçonnées d'être à l' origine de la plupart des autres maladies auto-immunes chez l'humain. Cependant, la caractérisation de ces cellules chez l'humain a été entravée par l'incapacité de les distinguer d'autres cellules T CD4+ exprimant des marqueurs similaires en état d'inflammation. En effet, FOXP3 est exprimé par les cellules T effectrices non-suppressives après activation. Ainsi, les corrélats fonctionnels de l'expression de FOXP3 dans les cellules Treg humaines sont mal définis. Nous émettons l'hypothèse que les cellules T CD4+CD25+++FOXP3+ humaines constituent une population hétérogène, composée de plusieurs sous-populations de cellules T et Treg, dans lesquels l'expression de FOXP3 a différentes conséquences fonctionnelles et phénotypiques.Grâce à une nouvelle approche de clonage cellulaire, nous démontrons que chez l'humain environ 30% des cellules T FOXP3+ ne sont pas suppressives, révélant ainsi une hétérogénéité fonctionnelle parmi les lymphocytes T CD4+CD25+++ humains. Ces cellules non-suppressives sont phénotypiquement identiques à leurs homologues suppressives, mais perdent l'expression de FOXP3 à l'activation. En examinant le transcriptome de ces cellules T FOXP3+ instables, nous montrons qu'elles constituent une population de lymphocytes T à part entière, apparentée aux cellules Treg, et dotée d'une signature d'expression génique spécifique. En outre, nous observons que la stabilité de la lignée Treg et de l'expression de FOXP3 sont orchestrés à un niveau supérieur de la régulation du lignage cellulaire. Enfin, nous montrons que la mutation A384T dans FOXP3, dérivée de l'IPEX, abroge la capacité de FOXP3 à générer la fonction suppressive des cellules Treg, tout en préservant sa capacité à contrôler leur phénotype. Nous offrons ainsi la première démonstration d'une dichotomie fonctionnelle dans le rôle de FOXP3 dans les cellules Treg humains. En conclusion, nous démontrons que FOXP3 n'est pas un marqueur absolu de la fonction suppressive dans les cellules T CD4+ chez l'homme, excluant la possibilité d'utiliser FOXP3 comme seul marqueur clinique pour le suivi de l'immuno-régulation. En outre, nous définissons des sous-populations parmi les cellules T FOXP3+ qui diffèrent en fonction et en stabilité, et déterminons les caractéristiques de ces cellules instables. Ces résultats permettront l'exploration de leur contribution à l'homéostasie immunitaire et de leurs défauts dans des maladies auto-immunes