Kidney International, Vol. 65 (2004), pp. 1560–1563

Clinical transplantation tolerance: The promise and challenges

MOHAMMED JAVEED I. ANSARI and MOHAMED H. SAYEGH1

Brigham & Women’s Hospital, Boston, Massachusetts

Clinical transplantation tolerance: The promise and challenges.Organ transplantation is now well established as a preferredoption for the treatment of end-stage organ failure. However,there is a severe shortage of donor organs and continued loss ofa significant number of organ grafts due to chronic allograft dys-function. Induction of tolerance of a transplant recipient towardtheir foreign organ graft, therefore, remains the “Holy Grail”of transplantation immunobiologists. Recently, clinical trials toexplore pilot tolerance protocols in humans have been initi-ated. Defining the ideal strategy(ies) and the role of immuno-suppressive drugs, developing tolerance assay(s), and enhanc-ing cooperation between transplant professionals, industry, andthe government are some of the challenges to achieving clinicaltransplantation tolerance. This article reviews the promise andthe challenges of achieving clinical transplantation tolerance inhuman organ transplant recipients.

Over the last two decades there has been a progressiveimprovement of allograft survival, in particular kidney al-lografts [1]. Intriguingly, this improvement was seen onlyin recipients who never had an acute rejection episode,emphasizing the recipient’s alloimmune response as amajor determinant of overall outcome of the transplant.Furthermore, the increasing demand of organs for trans-plantation [2] creates an urgent need for optimizing theoutcome of transplantation by achieving long-term, drug-free, graft acceptance with normal organ function. Thebaffling array of potential complex treatment combina-tions currently available to the transplant immunobiol-ogists [3] and the vast experimental data, on ways toachieve transplantation tolerance, that has amassed sincethe original description, half a century ago, of the phe-nomenon of tolerance in experimental animals imploresus to evaluate where we stand on the road to achiev-ing clinical transplant tolerance, and underscore the chal-lenges that we face, so that we may choose the best courseof action [4].

T cells are the vital elements of the immune responseand interact with the alloantigen by the direct and indirect

1Participant and speaker.

Key words: transplantation, organ, kidney, islet, tolerance, toleranceassays, tolerance trials, Immune Tolerance Network (ITN).

C© 2004 by the International Society of Nephrology

pathways, recognizing the foreign major histocompati-bility complex (MHC) molecules directly on the donorantigen-presenting cells and processed donor antigenson self antigen-presenting cells, respectively [5]. The Tcells reacting to their specific antigen can undergo a num-ber of different responses, namely activation followed byproliferation and differentiation into effector and mem-ory cells, and termination. Physiologic termination of theT-cell immune response is carried out by a number ofmechanisms, specifically, deletion (central or peripheral);anergy, where T cells are unresponsive to restimulationwith specific antigen; and regulation by regulatory cellsand cytokines [6]. These physiologic mechanisms formthe basis of inducing donor-specific tolerance in clini-cal transplantation [7–9]. Another possible mechanismof immunologic tolerance that is unique to the transplantsetting is microchimerism, the persistence of a small num-ber of donor-derived bone marrow cells in recipients [10–12].

It is imperative to define transplant tolerance at theoutset so that we understand precisely our objective.Transplant tolerance does not mean complete unrespon-siveness of the immune system toward the graft, rather alack of a destructive immune response toward it, in thepresence of generalized immune competence [13]. Anoperational definition of transplant tolerance in the clin-ical setting is the absence of acute and chronic rejectionand indefinite graft survival with normal graft functionin an immunocompetent host. The issue of ongoing im-munosuppression remains to be resolved as to whetherwe should aim for a complete drug-free state or, more re-alistically, accept a minimal amount of ongoing immuno-suppression/immunomodulation [14].

Generally, reports claiming tolerance induction citegraft survival in rodents of over 100 days with donor-specific hyporesponsiveness (indicated by acceptance ofa second graft from the original donor strain and rejec-tion of third-party grafts). It is impractical to confirm tol-erance induction in this way, in humans, leaving a voidin this crucial area. Consequently, devising an assay thatallows us to prospectively follow the status of the im-mune response toward the graft and detect tolerance orearly signs of rejection is an urgent necessity [15–18].Yet, it seems unlikely that a single assay will provide an

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Ansari and Sayegh: Clinical transplantation tolerance 1561

Table 1. Potential tolerance assays [4]

Measuring T-cell alloreactivityProliferation (MLR)Cytotoxicity (CML)Cytokine analyses (ELISA, ELISPOT, flow cytometry)Cell division (CFSE labeling)Delayed-type hypersensitivity (trans-vivo DTH assay)

Humoral immune responseAlloantibody titers

Profiling of immune cellsLymphocyte activation markers by flow cytometry

Blood, urine, graft infiltrating cells“Landscaping” of blood T lymphocytes using transcriptome

technologyPatterns of V-b usage associated with tolerance(“TCR signature” of tolerance)

Profiling circulating DC subsetsPrecursor (p)DC1 and pDC2

Genetic analysesImmune gene polymorphismsDefining “tolerance genes” by GeneChip microarray technology

Other assaysDefining “tolerance proteins” by proteomics technologyGraft morphology and immunohistochemistry

Abbreviations are: MLR, mixed lymphocyte reaction; CML, cell-mediatedlymphocytotoxicity; ELISA, enzyme linked immunosorbent assay; ELISPOT,enzyme-linked immunospot assay; CFSE, carboxy-fluorescein diacetatesuccinimidyl ester; DTH, delayed-type hypersensitivity; DC, dendritic cell; TCR,T cell receptor.

adequate immunologic profile and a panel of assays maybe required. Hitherto, a number of promising assays haveemerged, but their wider clinical validation is still calledfor (Table 1).

Very small minorities of patients, who unwittingly dis-continue their immunosuppression, provide rare exam-ples of clinical transplant tolerance [19]. The basis ofthis immunosuppression-free tolerant state, however, re-mains intriguing and merits further study so that we maylearn how this can be achieved reproducibly (if at all pos-sible). This phenomenon has also been reported in pa-tients receiving total body irradiation as induction ther-apy [20, 21] and in those kidney transplant recipients whohad received a previous bone marrow transplant from thesame donor, first reported by our group [22] and morerecently by others in a patient with multiple myelomacomplicated by end stage renal failure [23].

The utilization of bone marrow transplantation in or-der to induce tolerance through mixed chimerism of theimmune system has been expansively studied in animalmodels and to a lesser extent in humans [24]. A ma-jor challenge that remains, for the induction of lastingchimerism, is the development of clinically applicablenonmyeloablative regimens that can be safely used in hu-man leukocyte antigen (HLA)-mismatched patients [25].

A more novel approach to tolerance induction involvesthe use of in vitro conditioned or immature donor den-dritic cells that have the capacity to induce peripheraland central tolerance [26, 27]. Gene therapy is anothernovel approach [28]. Elucidating the optimal conditions

in non-human primates and clarifying the risks associatedwith such approaches are the first hurdles to be overcomebefore moving on to clinical trials of these strategies.

Other strategies, utilizing T-cell depleting agents orcostimulatory blockade with or without donor-specifictransfusion, appear to achieve tolerance in a variety ofanimal models [7] but not in a true sense of the wordin primate models [29]. In the past several years therehas been great enthusiasm about the potential of trans-lating strategies targeting the CD28/CTLA-4:B7-1/2 andthe CD40:CD154 T-cell costimulatory pathways to theclinic [5, 30]. Our understanding of these importantcostimulatory pathways and their interaction with eachother and other novel pathways such as ICOS:ICOSL,CD134:CD134L, CD27:CD70, and PD-1:PD-L1/2 arestill unfolding. These novel pathways appear to playgreater roles under some circumstances [31–36]. Target-ing of these pathways may however only work when thealloreactive T-cell repertoire is rendered to a manageablesize with adjunctive depleting or deletional therapies [8].The new immunosuppressive drug rapamycin may playsuch a role by inducing T-cell apoptosis [37]. There re-mains a challenge, however, of defining how much dele-tion is enough and how safe it is in humans. Further, theprecise impact of the conventional immunosuppressivedrugs on tolerizing strategies needs to be reevaluated,since the initial suggestion that certain drugs impair thegeneration of tolerance in some models [37, 38] have notproven founded in others [39, 40].

Another major challenge is the resolution of the re-lationship of tolerance with chronic allograft dysfunc-tion. There are conflicting data from experimental modelson the impact of alloantigen-dependent and alloantigen-independent mechanisms on chronic allograft dysfunc-tion [41–43]. However, some data indicate that donor-specific hyporesponsiveness is associated with protectionfrom chronic rejection in humans [44].

The impact of tolerizing regimens on the risk of in-fectious complications and likewise the detrimental ef-fect of previous, ongoing or later infections on the induc-tion or maintenance of tolerance and also on the courseof infection itself is uncertain. Indeed, certain tolerizingstrategies are ineffective if performed during ongoinginfectious episodes [45] and a recent study sheds morelight on the possible mechanism responsible for this phe-nomenon, suggesting that individuals harboring virallyinduced memory T cells that are crossreactive with donoralloantigen (a phenomenon termed heterologous immu-nity) are resistant to tolerance induction [12, 46]. Onthe other hand, attempting to use a tolerizing regimenin the presence of a latent infectious agent may allowtolerance to develop toward it too. Therefore, it seemsprudent to exclude patients with certain chronic or la-tent infections (e.g., hepatitis B or C, cytomegalovirus,Epstein-Barr virus) from initial tolerance trials.

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Table 2. Ongoing clinical trials of protolerogenic therapies under theauspices of Immune Tolerance Network (ITN) [47, 48]

Transplant Therapy

1 Bone marrow andkidney formultiple myelomaand end-stagerenal failure

Non-myeloablative conditioning regimen(cyclophosphamide, anti-thymocyteglobulin, and thymic irradiation) formixed chimerism and induction oftolerance

2 Kidney Campath 3 combined with sirolimus andtapering mycophenolate mofetil

3 Bone marrow andkidney

Cyclosporine, cyclophosphamide,MEDI-507, and thymic irradiation

4 Kidney Campath-1H, sirolimus, and short courseof tacrolimus

5 Islet Edmonton protocol of steroid-freeimmunosuppression: daclizumab,sirolimus, and low-dose tacrolimus

6 Islet Campath-1H and sirolimus maintenancetherapy

7 Islet hOKT3c1(Ala-Ala) and sirolimusmonotherapy

The choice of which patient population will be thefirst to be enrolled into such trials is a very difficultone, especially when the clinicians are faced with theethical issue of risking possible rejection from a failedtolerance protocol in an era when 1-year graft survivalrates exceed 90% and few grafts are lost to rejection.There is added convolution, due to conflict of interest ofpharmaceutical companies manufacturing immunosup-pressive agents currently used, because such tolerizingstrategies may not benefit them. An altruistic coopera-tion between the biotechnology industry and the trans-plant biologist is needed to successfully achieve the nec-essary development of the appropriate tolerizing agents.Finally, the proper conduct and execution of the clini-cal trials cannot be overemphasized and will need to beoverseen by a suitably appointed regulatory (governmen-tal) agency. The Immune Tolerance Network (ITN), NIH(USA) (http://www.immunetolerance.org) was expresslyinstituted for this sole purpose. It provides a platform forsharing of ideas as well as core facilities and provides afocus for the development of the most suitable strate-gies. The ITN is sponsoring several research projects in-volving islet transplantation, solid organ transplantation(Table 2) [47], autoimmune disease, allergy/asthma, toler-ance assay studies and special projects such as “The ITNTolerant Kidney Transplant Patient Registry,” to estab-lish a world-wide registry of kidney transplant recipientswho are off all immunosuppression.

CONCLUSION

We have learned that the goal of clinical transplant tol-erance is achievable especially in animal models but alsoin a few humans. Identifying the most successful of thesestrategies and then translating them to larger animals totest their suitability for the patients is the next step. This

demands persistence and meticulous investigation to con-firm the robustness and longevity as well as safety of thetolerance inducing regimens. If we are successful in do-ing this, then we may still arrive at our chosen destination,although it may seem very distant.

Reprint requests to Mohamed H. Sayegh, Brigham & Women’s Hos-pital, 75 Francis Street, Boston, MA 02115.E-mail: msayegh@rics.bwh.harvard.edu

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