Since the concept of immunologic tolerance was discovered in the 1940s, the pursuit of tolerance induction in human transplantation has led to a rapid development of pharmacologic and biologic agents. Short-term graft survival remains an all-time high, but successful withdrawal of immunosuppression to achieve operational tolerance rarely occurs outside of liver transplantation. Collaborative efforts through the NIH sponsored Immune Tolerance Network and the European Commission sponsored Reprogramming the Immune System for Establishment of Tolerance consortia have afforded researchers opportunity to evaluate the safety and efficacy of tolerogenic strategies, investigate mechanisms of tolerance, and identify molecular and genetic markers that distinguish the tolerance phenotype. In this article, we review traditional and novel approaches to inducing tolerance for organ transplantation, with an emphasis on their translation into clinical trials.

Chronic allograft rejection is a major impediment to long-term transplant success. Humoral immune responses to alloantigens are a growing clinical problem in transplantation, with mounting evidence associating alloantibodies with the development of chronic rejection. Nearly a third of transplant recipients develop de novo antibodies, for which no established therapies are effective at preventing or eliminating, highlighting the need for a nonhuman primate model of antibody-mediated rejection. In this report, we demonstrate that depletion using anti-CD3 immunotoxin (IT) combined with maintenance immunosuppression that included tacrolimus with or without alefacept reliably prolonged renal allograft survival in rhesus monkeys. In these animals, a preferential skewing towards CD4 repopulation and proliferation was observed, particularly with the addition of alefacept. Furthermore, alefacept-treated animals demonstrated increased alloantibody production (100%) and morphologic features of antibody-mediated injury. In vitro, alefacept was found to enhance CD4 effector memory T cell proliferation. In conclusion, alefacept administration after depletion and with tacrolimus promotes a CD4+memory T cell and alloantibody response, with morphologic changes reflecting antibody-mediated allograft injury. Early and consistent de novo alloantibody production with associated histological changes makes this nonhuman primate model an attractive candidate for evaluating targeted therapeutics.

Depletional strategies directed toward achieving tolerance induction in organ transplantation have been associated with an increased incidence and risk of antibody-mediated rejection (AMR) and graft injury. Our clinical data suggest correlation of increased serum B cell activating factor/survival factor (BAFF) with increased risk of antibody-mediated rejection in alemtuzumab treated patients. In the present study, we tested the ability of BAFF blockade (TACI-Ig) in a nonhuman primate AMR model to prevent alloantibody production and prolong allograft survival. Three animals received the AMR inducing regimen (CD3-IT/alefacept/tacrolimus) with TACI-Ig (atacicept), compared to five control animals treated with the AMR inducing regimen only. TACI-Ig treatment lead to decreased levels of DSA in treated animals at 2 and 4 weeks posttransplantation (p < 0.05). In addition, peripheral B cell numbers were significantly lower at 6 weeks posttransplantation. However, it provided only a marginal increase in graft survival (59 ± 22 vs. 102 ± 47 days; p = 0.11). Histological analysis revealed a substantial reduction in findings typically associated with humoral rejection with atacicept treatment. More T cell rejection findings were observed with increased graft T cell infiltration in atacicept treatment, likely secondary to the graft prolongation. We show that BAFF/APRIL blockade using concomitant TACI-Ig treatment reduced the humoral portion of rejection in our depletion-induced preclinical AMR model.

Cross-talk between B and T cells in transplantation is increasingly recognized as being important in the alloimmune response. T cell activation of B cells occurs by a 3-stage pathway, culminating with costimulation signals. We review the distinct T cell subtypes required for B cell activation, and discuss the formation of the Germinal Center (GC) after transplantation, with particular reference to the repopulation of the GC following depletional induction, and the subsequent effect of immunosuppressive manipulation of T-B cell interactions. Additionally, ectopic GCs are seen in transplantation, but their role is not fully understood. Therapeutic options to target T-B cell interactions are of considerable interest, both as immunosuppressive tools, and to aid further understanding of these important alloimmune mechanisms.

Even though the etiology of chronic rejection (CR) is multifactorial, donor specific antibody (DSA) is considered to have a causal effect on CR development. Currently the antibody-mediated mechanisms during CR are poorly understood due to lack of proper animal models and tools. In a clinical setting, we previously demonstrated that induction therapy by lymphocyte depletion, using alemtuzumab (anti-human CD52), is associated with an increased incidence of serum alloantibody, C4d deposition and antibody-mediated rejection in human patients. In this study, the effects of T cell depletion in the development of antibody-mediated rejection were examined using human CD52 transgenic (CD52Tg) mice treated with alemtuzumab. Fully mismatched cardiac allografts were transplanted into alemtuzumab treated CD52Tg mice and showed no acute rejection while untreated recipients acutely rejected their grafts. However, approximately half of long-term recipients showed increased degree of vasculopathy, fibrosis and perivascular C3d depositions at posttransplant day 100. The development of CR correlated with DSA and C3d deposition in the graft. Using novel tracking tools to monitor donor-specific B cells, alloreactive B cells were shown to increase in accordance with DSA detection. The current animal model could provide a means of testing strategies to understand mechanisms and developing therapeutic approaches to prevent chronic rejection.

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Because lymphocytes were shown to mediate transplant rejection, their depletion has been studied as a mechanism of preventing rejection and perhaps inducing immunologic tolerance. Agents that profoundly deplete lymphocytes have included monoclonal antibodies, cytotoxic drugs, and radiation. We have studied several such agents but focused on antibodies that deplete not only peripheral blood lymphocytes, but also lymph node lymphocytes. Depletion of lymph node T lymphocytes appears to permit peripheral tolerance at least for T cells in animal models. Nevertheless, B-cell responses may be resistant to such approaches, and T memory cells are likewise relatively resistant to depleting antibodies. We review the experimental and clinical approaches to depletion strategies and outline some of the pitfalls of depletion, such as limitations of currently available agents, duration of tolerance, infection, and malignancy. It is notable that most tolerogenic strategies that have been attempted experimentally and clinically include depleting agents even when they are not named as the underlying strategy. Thus, there is an implicitly acknowledged role for reducing the precursor frequency of donor antigen-specific lymphocytes when approaching the daunting goal of transplant tolerance.

Summary The last two decades have witnessed a pandemic in antibody development, with over 600 entering clinical studies and a total of 28 approved by the FDA and European Union. The incorporation of biologics in transplantation has made a significant impact on allograft survival. Herein, we review the armamentarium of clinical and preclinical biologics used for organ transplantation – with the exception of belatacept – from depleting and IL‐2R targeting induction agents to costimulation blockade, B‐cell therapeutics, BAFF and complement inhibition, anti‐adhesion, and anti‐cytokine approaches. While individual agents may be insufficient for tolerance induction, they provide possibilities for reduction of steroid or calcineurin inhibitor use, alternatives to rejection episodes refractory to conventional therapies, and specialized immunosuppression for highly sensitized patients.