Combination of IL-2, rapamycin, DNA methyltransferase and histone deacetylase inhibitors for the expansion of human regulatory T cells

Abstract

FOXP3⁺ regulatory T cell (Treg) based cellular therapies represent promising therapeutic options in autoimmunity, allergy, transplantation and prevention of Graft Versus Host (GVH) Disease. Among human FOXP3-expressing CD4⁺T cells, only the CD45RA⁺ naïve Treg (nTreg) subset is suitable for <i>in vitro</i> expansion. However, FoxP3 expression decays in cells using currently described culture protocols. Rapamycin alone was not able to prevent FOXP3 loss in nTregs cells, as only a half of them maintained FOXP3 expression after 14 days of culture. In contrast we report a novel combined drug regimen that can drastically stabilize FOXP3 expression in cultured Tregs. IL-2, rapamycin, histone deacetylase and DNA methyltransferase inhibitors act in synergy to allow expansion of human regulatory T cells with sustained high expression of FOXP3 and CD15s with potent suppressive capacities <i>in vitro</i> and control of murine xeno-GVH reactions. Of note, an additional subsequent infusion of expanded nTreg cells did not improve survival of mice. Combination of IL-2, rapamycin, histone deacetylase and DNA methyltransferase inhibitors is optimal for the expansion <i>in vitro</i> of pure effective nTreg maintaining high levels of FOXP3 for therapeutic purposes.