Abstract
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.
Highlights
In addition to thymic regulation, peripheral induction of a regulatory phenotype in conventional T (Tconv) cells provides protection from undesirable immune responses to self antigens
This low purity limits the yield of Foxp3+ induced Foxp3+ Treg (iTreg) cells, the contamination with activated Foxp3− T cells that may exert pro-inflammatory effector functions poses a potential risk when used for Treg cell-based immunotherapy
Antigen-specific iTreg cells are commonly generated from murine TCR-transgenic CD4+ T cells through activation with plate-bound anti-CD3 and anti-CD28 antibodies in the presence of TGF-β and IL-2 since this method generates large numbers of Foxp3+ cells at very high purity (Thornton et al, 2010; Verhagen et al, 2013a)
Summary
In addition to thymic regulation, peripheral induction of a regulatory phenotype in conventional T (Tconv) cells provides protection from undesirable immune responses to self antigens. We have demonstrated previously that, in the Tg4 mouse model, iTreg cells can be generated using cognate Myelin Basic Protein (MBP) peptide as a stimulus but the level of conversion lags behind that achieved using antibody stimulation (65–75% vs 90–95%) (Verhagen et al, 2013a). This low purity limits the yield of Foxp3+ iTreg cells, the contamination with activated Foxp3− T cells that may exert pro-inflammatory effector functions poses a potential risk when used for Treg cell-based immunotherapy. We demonstrate that our method induces antigen-specific iTreg cells of high purity that successfully protect against CNS autoimmune disease
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