Uniquely, using a short-term minimal immunosuppressive (IS) drug regimen comprising costimulation blockade (CoSB; CTLA4Ig/belatacept) + tapered rapamycin, we have shown that maturation-resistant, donor-derived DCreg, infused 1w before transplant, can safely prolong renal allograft survival in rhesus macaques, accompanied by selective attenuation of donor-reactive memory T cell (Tmem) responses, a mechanism that may help overcome a critical barrier to transplant tolerance induction in NHP and humans. We will now ascertain whether a novel, modified, CNI-free IS regimen that is (i) more permissive to extended graft survival and (ii) that we hypothesize will enhance the immunomodulatory function of DCreg, can achieve donor-specific tolerance. There is recent evidence that lymphocyte depletion followed by CoSB (belatacept) and rapamycin maintenance (the 2-drug regimen we used to demonstrate DCreg efficacy in monkeys) can control CoSB-resistant rejection in transplant patients, with reduction in CoSB (belatacept)-resistant Tmem. However, operational tolerance was not achieved. Notably, combination of donor DCreg infusion (a week before transplant) with perioperative lymphodepletion, promotes permanent, donor-specific allograft survival in rodents. This indicates that lymphodepletion after DCreg infusion does not interfere with their therapeutic effect. Moreover, DCreg infusion post-transplant following lymphodepletion, can also promote indefinite graft survival. We therefore hypothesize that maturation-resistant rhesus DCreg, administered to ATG- lymphodepleted, CoSB and rapamycin-treated renal graft recipients, will induce immunological changes and selective attenuation of donor-reactive Tmem conducive to donor-specific tolerance. We further hypothesize that novel biomarker analyses of host alloreactive Tmem responses (in particular, their expression of Eomes) will correlate with and be predictive of safe withdrawal of IS. Our Specific Aims are: Aim 1: To determine the influence of donor-derived DCreg infusion before transplant on renal allograft survival in NHP given combined lymphodepletion (ATG), belatacept and rapamycin (ABR). Aim 2: To determine the influence of donor-derived DCreg infusion before transplant on renal allograft survival in NHP given combined ATG, non-depleting αCD40 mAb and rapamycin (AAR). Aim 3: To compare the influence of donor-versus recipient-derived DCreg infusion post-transplant on renal allograft survival in NHP given combined ATG, CoSB and rapamycin (ABR or AAR). Each Aim will be accompanied by comprehensive, rationally-designed mechanistic studies that will elucidate the trafficking, fate and immune regulatory function of the adoptively-transferred DCreg and underlying mechanisms.