The immunosuppressive drugs rapamycin and cyclosporin A (CsA) are widely used to prevent allograft rejection. Moreover, they were shown to be instrumental in experimental models of tolerance induction. However, it remains to be elucidated whether these drugs have an effect on the CD4 ؉ CD25 ؉ regulatory T-cell (T REG ) population, which plays an important role in allograft tolerance. Recently, we reported that alloantigen-driven expansion of human CD4 ؉ CD25 ؉ T REG s gives rise to a distinct highly suppressive CD27 ؉ T REG subset next to a moderately suppressive CD27 ؊ T REG subset. In the current study we found that rapamycin and CsA do not interfere with the suppressive activity of human naturally occurring CD4 ؉ CD25 ؉ T cells. However, in contrast to CsA, rapamycin preserved the dominance of the potent CD27 ؉ T REG subset over the CD27 ؊ T REG subset after alloantigen-driven expansion of CD4 ؉ CD25 ؉ T REG s in vitro. Accordingly, CD4 ؉ CD25 ؉ T REG s cultured in the presence of rapamycin displayed much stronger suppressive capacity than

Graft-versus-host-disease (GVHD) is the most common cause of poor outcome after allogeneic stem cell transplantation (SCT). Of late, exploitation of FOXP3 þ regulatory T-cell (T REG ) function is emerging as a promising strategy in suppression of GVHD, while preserving graft-versus-leukemia (GVL). Cyclosporine and rapamycin reduce the expansion of effector T cells by blocking interleukin (IL)-2, but signaling by IL-2 is pivotal for T REG homeostasis. The resolution of GVHD is critically dependent on thymus-dependent reconstitution of the immunoregulatory system. Thus, there has been concern about the impact of blocking IL-2 signaling by immunosuppressive agents on T REG homeostasis. Here we demonstrate in a mouse model that in contrast to rapamycin, cyclosporine compromises not only the thymic generation of CD4 þ CD25 þ FoxP3 þ T cells but also their homeostatic behavior in peripheral immune compartments. Treatment with cyclosporine resulted in a sharp reduction of peripheral CD25 þ FoxP3 þ T cells in all immune compartments studied. Prolonged rapamycin treatment allowed for thymic generation of CD4 þ FoxP3 þ T cells, whereas treatment with cyclosporine led to a reduced generation of these cells. In conclusion, cyclosporine and rapamycin differentially affect homeostasis of CD4 þ FoxP3 þ T REG in vivo. As peripheral tolerance induction is a prerequisite for successful treatment outcome after allogeneic SCT, these findings are of potential clinical relevance.

The authors therefore conclude that co-stimulation blockade contributes to functional dominance of regulatory T cells by preventing expansion of alloreactive effector T cells. Tolerance-inducing protocols should ideally facilitate this phenomenon.

Blockade of costimulatory signals is a promising therapeutic target to prevent allograft rejection. In this study, we sought to characterize to what extent CTLA-4 engagement contributes to the development of transplantation tolerance under the cover of CD40/CD40L and CD28/CD86 blockade. In vitro, we found that inhibition of the primary alloresponse and induction of alloantigen hyporesponsiveness by costimulation blockade was abrogated by anti-CTLA-4 mAb. In addition, regulatory CD4+CD25+ T cells (TREG) were confirmed to play a critical role in the induction of hyporesponsiveness by anti-CD40L and anti-CD86 mAb. Our data indicated that CTLA-4 engagement is not required for activation or suppressor function of TREG. Instead, in the absence of either CTLA-4 signaling or TREG, CD8+ T cell division was enhanced, whereas the inhibition of CD4+ T cell division by costimulation blockade remained largely unaffected. In vivo, the administration of additional anti-CTLA-4 mAb abrogated anti-CD40L- and anti-CD86 mAb-induced cardiac allograft survival. Correspondingly, rejection was accompanied by enhanced allograft infiltration of CD8+ cells. We conclude that CTLA-4 signaling and TREG independently cooperate in the inhibition of CD8+ T cell expansion under costimulation blockade.

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