The infusion of ex vivo activated and expanded CD4⁺CD25⁺ immune regulatory cells inhibits graft-versus-host disease lethality

Immune regulatory CD4⁺CD25⁺ cells play a vital role in the induction and maintenance of self-tolerance and the prevention of autoimmunity. Recently, CD4⁺CD25⁺ cells have been shown to be required for the ex vivo induction of tolerance to alloantigen via co-stimulatory blockade and to inhibit allogeneic skin graft rejection. Data presented here demonstrate that CD4⁺CD25⁺ cells play an important role in graft-versus-host disease (GVHD) generation. Depletion of CD4⁺CD25⁺ cells from the donor T-cell inoculum or in vivo CD25-depletion of the recipient before transplantation resulted in increased GVHD mediated by CD4⁺ or whole T cells in several strain combinations irrespective of the total body irradiation conditioning regime. The infusion of freshly purified donor CD4⁺CD25⁺ cells modestly inhibited GVHD when administered in equal numbers with whole CD4⁺ cells. Because CD4⁺CD25⁺ cells only account for 5% to 10% of the total CD4⁺ population, the administration of high numbers of fresh donor CD4⁺CD25⁺ cells may not be clinically practical. However, we found that large numbers of CD4⁺CD25⁺ cells can be obtained by ex vivo activation and expansion. Cultured CD4⁺CD25⁺ cells, administered in equal numbers with CD4⁺ T cells or CD25-depleted whole T cells, resulted in significant inhibition of rapidly lethal GVHD. To our knowledge, this study is the first to demonstrate that activated, cultured CD4⁺CD25⁺ cells can offer substantial protection in a relevant in vivo animal model of disease. These data have important ramifications for clinical bone marrow and solid organ transplantation. CD4⁺CD25⁺ cells warrant consideration as an exciting new modality of cellular therapy for the inhibition of undesirable autologous and allogeneic responses.