Current approaches to prevent or treat transplant rejection, graft-versus-host disease and severe autoimmunity rely on non-specific immunosuppressive drugs. Ongoing efforts aimed at harnessing regulatory T (Treg) cells hold promise for revolutionizing the current therapeutic options, reducing if not abandoning immune suppression in favor of immune tolerance. This paradigm shift carries the potential of dramatically enhancing efficacy, persistency, and specificity while reducing side effects. Here, we review the various strategies devised to manipulate Treg cells in vitro and in vivo, the clinical progress achieved to date, and critical issues that still need to be addressed.