2007 JASN IMPACT FACTOR 7.111	HOME   AUTHOR INFO   EDITORIAL BOARD   SUBSCRIBE   FEEDBACK   ALERTS   HELP
advanced	CURRENT ISSUE	ARCHIVES	JASN Express	ONLINE SUBMISSION

This Article Abstract Full Text (PDF) All Versions of this Article: ASN.2007020180v1 18/8/2242    most recent Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Girlanda, R. Articles by Kirk, A. D. Search for Related Content PubMed PubMed Citation Articles by Girlanda, R. Articles by Kirk, A. D.

Frontiers in Nephrology: Immune Tolerance to Allografts in Humans

Transplantation Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Department of Health and Human Services, Bethesda, Maryland

Correspondence: Dr. Allan D. Kirk, Room 5-5752, Building 10CRC, Center Drive, Bethesda, MD 20892. Phone: 301-496-3047; Fax: 301-451-6989; E-mail: allank{at}intra.niddk.nih.gov

	Abstract

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
Vascularized allografts are rejected unless some indefinite modification to the recipient's immune system is made. This modification is typically achieved through the long-term administration of immunosuppressive drugs. Patients thus trade their end-stage organ failure for dependence on daily drug therapy and the accompanying chronic condition of immunodeficiency. However, it is clear from studies in experimental animals that rejection can be prevented through the use of several therapeutic approaches, including donor hematopoietic cell infusion, chimerism, T cell depletion, and/or co-stimulation blockade. Successfully treated animals avoid rejection beyond the period of therapy without a phenotype of chronic immunosuppression and are thus considered to be tolerant of their grafts. Although intriguing, this success in animals has yet to be reproducibly translated to the clinic, and human transplant recipients remain tethered to immunosuppressive drugs with rare exceptions. This article provides an overview of the existing, largely anecdotal, clinical experience with organ allograft tolerance. It reviews the various approaches that are being applied in pilot human trials and suggests avenues for future clinical investigation.

	Introduction

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
It has been commonly recognized since the earliest days of clinical skin grafting^1,2 that transplanted nonautologous tissues fail unless the recipient's immune system is modified in some sustained manner.^3,4 In the past century, the underlying immune mechanisms of allograft rejection have become increasingly defined and exploited through immunosuppressive drugs, permitting organ transplantation to become a successful therapy for most end-stage organ diseases. Experimentally, certain modifications of the recipient have also been shown to permit "actively acquired tolerance"⁴ of allografted organs—that is, graft acceptance without an ongoing requirement for therapy or apparent immune compromise. These methods have been studied with the intent of facilitating clinical transplantation without immunosuppression-associated adverse effects.

Experimental allograft tolerance is now commonly achieved in rodents and has been demonstrated in higher animals, including pigs,^5,6 dogs^7–9 and nonhuman primates,^10,11 with increasing regularity. Thus, rejection is not a biologic inevitability. Indeed, in humans, tolerance to transplanted allografts has been achieved, although its occurrence remains uncommon and unpredictable.

In this article, we review the clinical allograft tolerance literature and survey the general mechanisms that are applicable to kidney transplantation. We discuss the anecdotal reports of human tolerance and recent attempts to induce prospectively tolerance clinically and outline efforts to make tolerance a more common and predictable clinical outcome.

	DEFINING TOLERANCE

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
Although there are many definitions of tolerance, clinically it is the maintenance of stable allograft function without clinically evident immunosuppression. This can include true tolerance, the absence of any detectable detrimental immune response and no immunocompromise, and operational tolerance, the gross phenotype of tolerance with an immune response or deficit that has no significant clinical impact. These scenarios manifest through a variety of immunologic processes that are simultaneously at play, including modulations in effector cell precursor frequency, antigen presentation efficiency, effector cell activation threshold, regulation, and altered cell trafficking. Thus, tolerance should be considered a mosaic phenotype—the result of multiple competing variables with the end outcome achieved via many potential combinations.

Tolerance is also a product of one's environment. Most immunocompetent individuals are capable of mounting an alloimmune response under facilitating conditions, just as apparently healthy people can develop autoimmunity independent of their genotype.^12,13 Similarly, given the cross-reactive nature of alloimmunity, responses to environmental pathogens can evoke an alloimmune response,¹⁴ and an inability to foster alloimmunity suggests some element of immunocompromise, although perhaps inconsequential. Thus, tolerance strategies and assessments require knowledge not only of an individual's immune competency but also of the environmental threats to homeostasis.

Tolerance also varies as a matter of time and susceptibility. Time may reveal that a patient with operational tolerance, established by an inefficient immune response, is actually rejecting slowly. An organ's functional resilience may also alter a clinician's view such that an organ with regenerative capacity, such as the liver, may silently endure an immune attack that would lead to loss of a more sensitive allograft, such as a coronary artery. In all, the ultimate definition of tolerance rests with the sensitivity and accuracy of monitoring strategies used and the investigator's persistence.

	TOLERANCE IS, IN GENERAL, POSSIBLE

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
During pregnancy, a mother's immune system tolerates an allogeneic fetus. Indeed, the propagation of mammalian species is to some extent predicated on an inability to muster alloimmunity under certain conditions. Consistent with this, clinical cases have been reported whereby graft function has been maintained indefinitely after the cessation of immunosuppression, demonstrating that operational tolerance can be achieved (Table 1). However, these spontaneous phenomena have typically occurred by way of drug nonadherence or withdrawal mandated by complications, and these conditions more commonly lead to rejection. Thus, tolerance is a stochastic event under current treatment regimens.^15,16

	SPONTANEOUS TOLERANCE AFTER KIDNEY TRANSPLANTATION

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
The number of kidney transplant recipients reported as tolerant is small (<1%) compared with the total number of kidney transplants performed (Table 1). Studies of these rare individuals allow few generalizations. Tolerance typically appears years after transplantation, suggesting that it was achieved via a process rather than through a sudden induction, with some exceptional cases after posttransplantation lymphoproliferative disease. There is no relationship to the recipient's underlying renal disease. Donor age tends to be lower in tolerant patients compared with the general population, suggesting that organ resilience, among other things, may be supplementing any immune properties of the recipient. Donor mismatch also tends to be lower, suggesting the relevance of a lower donor-specific precursor frequency. Most cases of tolerance are the result of noncompliance, although the typical outcome of drug discontinuation is graft loss within 8 mo.²² Similarly, modern surveys suggest a graft loss rate of 20 to 40% in patients who are identified as operationally tolerant with relatively short follow-up.^15,16 Nonetheless, these reports have shown that some patients do not experience rejection within the follow-up period after drug cessation. The rate of success simply fails to meet the success that is seen with continued immunosuppression, but it does demonstrate that some circumstances lead to tolerance.

The small number of tolerant patients has precluded systematic mechanistic analysis in humans. However, a recently established registry formed by the Immune Tolerance Network has embarked on exploratory mechanistic study.¹⁶ Early reports suggest significant differential expression of some candidate genes, particularly those associated with regulatory and effector T cell function, such as IL-10, FoxP3, and CXCR3 in the urinary sediment of tolerant patients. These preliminary findings fit into a general paradigm of regulatory control as one element influencing ultimate graft outcome. Other recent analyses of operationally tolerant liver transplant recipients revealed distinct mononuclear cell profiles, including higher frequency of CD4⁺CD25⁺ T regulatory cells^23,24 and plasmacytoid dendritic cells²⁵ in the peripheral blood compared with patients on maintenance immunosuppression and healthy control subjects. However, in-depth characterization of tolerant patients remains in its infancy, and there is currently no way to predict tolerance in an individual patient. Also, even fully characterized, detailed descriptions of complex circumstances do not equate with control over the outcome.²⁶

Nevertheless, from the clinical experience accumulated so far, it is reasonable to hypothesize that some patients, perhaps most, do not need to take daily immunosuppressive medications at the same dosage for life. In light of the growing recognition that alloimmunity fluctuates with environmental immunity,^14,27 it is likely that the risk for rejection undulates and may be able to be controlled with variable, risk-related dosing regimens. For example, because the pace of a de novo alloimmune response has some time requirement and rejection is typically slow to develop when noncompliance is late,²² intermittent immunosuppression may be completely sufficient in many cases. This is supported by recent results using spaced immunosuppressive therapy after depletional induction.²⁸ However, if the need for nonspecific immunosuppression is transient, then so perhaps is tolerance.

It is interesting to note that the transience of tolerance was established with its first description by Billingham et al.⁴ Although many have cited the classic experiments of Medawar's group as a description of robust and enduring tolerance, their original experiment reported that only three of five mice became tolerant of skin grafts after perinatal injection of donor tissue, and one of these three animals lost the skin graft between days 75 and 91 after transplantation. This is paralleled by reports of late allograft rejection^29,30 and graft function deterioration¹⁵ in long-term tolerant patients.

The reasons for tolerance's instability are likely related to the intrinsic nature of the adaptive immune system; namely, it is adaptive. In keeping with the exceptional insight of Medawar, this, too, was anticipated in the original defining studies.⁴ The phenomenon of alloimmunity derived from immunity against cross-reactive environmental pathogens is referred to as heterologous immunity and has been clearly shown to deter or break experimental tolerance¹⁴ (reviewed by Selin et al.²⁷). It is likely that this process is increasingly at play in the clinic, where transplantation is performed in recipients with mature and varied environmental exposure histories.

	STRATEGIES TO ACHIEVE TOLERANCE IN THE CLINIC

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
An understanding of spontaneous tolerance is useful to reaffirm its credibility as a clinical goal. However, observations must transition to interventional studies if tolerance is to have clinical impact. The transplant literature is replete with successful preclinical strategies from which to draw for clinical investigation. Indeed, tolerance has been achieved in many preclinical large animal models, including dog,^7–9 pig,⁵ and nonhuman primates^31,32 (reviewed by Kean et al.¹⁰ and Kirk¹¹). The rodent literature on tolerance is exhaustive and also well reviewed.^14,33–35 This experience generally shows tolerance to be more readily achieved in small, inbred animals compared with large, outbred animals, and even robust animal models seem less rigorous than adult humans. This likely reflects species-specific differences and differential environmental antigen exposure, T cell repertoire diversity, primed T cell pool size, and rigor in long-term follow-up. Thus, all attempts to achieve tolerance in humans remain decidedly experimental, subject to appropriate oversight, and require informed patient consent.³⁶

Modern clinical tolerance strategies have focused on specific mechanistic principles that primarily target single immune organs or sites of antigen exposure (Figure 1). Each approach has sought control over initial antigen exposure and subsequent immune repertoire development through alterations of the antigen source and route and attenuation of the resultant response. They differ in, among other things, the sustainability of the effect with some approaches, such as depletional induction, dependent on physiologic peripheral maintenance (reviewed by Kirk³⁷) and others, such as chimerism, actively shaping the effector repertoire over time.³⁸ This highlights the role of antigen exposure as a necessary step to activate the immune system for both tolerance and rejection and has been called a "window of opportunity for immune engagement" (WOFIE³⁹). Clinical results are discussed as they relate to the predominant intervention used, with the recognition that there is overlap in the methods and effects.

View larger version (61K): [in this window] [in a new window] [as a PowerPoint slide]   Figure 1. Sites for intervention in immune tolerance, a systemic view. Shown are multiple organs involved in the integrated immune response to the allograft. Because the alloimmune response is systemic, leading to changes in central effector cell output, peripheral effector cell frequency, and alterations in systemic chemotaxis culminating with the destruction of the graft, effective tolerance strategies are likely to influence simultaneously primary lymphoid organs (bone marrow, thymus), secondary lymphoid organs (lymph nodes, spleen), and the graft itself.

This concept of a balance between rejection and tolerance remains theoretical but is conceptually attractive. There is ample experimental evidence that graft-derived antigens foster tolerance (reviewed by Karim et al.⁴²). However, the use of the graft inherently risks organ injury, and the stochastic nature of success has prevented this approach from being ethically testable on any large scale. Whereas true tolerance avoids both acute and chronic allograft injury, there is justifiable concern that operational tolerance will eventually give way to chronic allograft nephropathy if there is even modest intragraft inflammation. Thus, most tolerance strategies have looked to other, more expendable sources of antigen, such as hematopoietic cells, to influence the immune response. Alternatively, tolerance has been encouraged through treatments that facilitate clonal deletion or anergy in a more efficient manner, such as co-stimulation blockade.

Hematopoietic Cells
Several approaches have used hematopoietic cells as a tolerance-facilitating antigen. The methods differ in the intended role of antigen, either to drive the apoptosis of activated effector T cells (activation-induced cell death [AICD]) or to influence thymic and central lymphoid repertoire development (chimerism). These methods are frequently confused because they both involve hematopoietic cell administration but differ dramatically in the role of adjuvant therapies and conceptual implementation.

In general, T cell activation that occurs in the absence of sufficient supporting adhesion, cytokine stimulation, or co-stimulatory signals leads to T cell death predicated on the antigen–T cell receptor interaction (reviewed by Green et al.⁴³). This hypothetically has the effect of selectively eliminating alloreactive T cells without eliminating environmental specificities and is conceptually relevant to the arguments favoring the graft as the tolerizing antigen. It is a peripheral (thymus-independent) mechanism. In this case, expendable hematopoietic cells function by being eliminated and stimulating an effector response, sparing the graft the brunt of the immune attack.

The use of hematopoietic cells as a surrogate antigen is to be distinguished from bone marrow engraftment. Engraftment seeks not to have the cells rejected but rather to have them permanently incorporated into the central lymphoid organs of the host (see the Chimerism section). Given the different mechanisms involved, differing immune therapies can function as either antagonists or protagonists of success. For example, a calcineurin inhibitor prevents marrow rejection, allowing eventual thymic presentation, but inhibits T cell receptor function and AICD.^44–47

The most direct use of hematopoietic elements to prevent rejection involves the use of random or donor-specific blood transfusions. It has been well established that recipients of blood transfusions have somewhat improved allograft survivals.^48–50 Similarly, mixed lymphocyte unresponsiveness and improved allograft survivals have been shown using donor bone marrow infusion, albeit with ongoing maintenance immunosuppression.^50–52 Although these approaches have not produced clinical tolerance, their salutary effects are critically established in the experimental literature (reviewed by Wood et al.⁵³) and clinically well documented.^48–50

It is interesting that the clinical effects of donor blood transfusion became less reproducible in the cyclosporine era. This is perhaps due to the efficacy of calcineurin inhibition's overshadowing minor transfusion-mediated effects but may also reflect inhibitory effects on AICD.^44,45 Interest in the use of nonengrafting donor antigen administration is now resurfacing with calcineurin inhibitor–free protocols.^54,55

Donor antigen infusion and antigen from the organ itself have been associated with microchimerism—trace numbers of donor cells (<1% of circulating cells) residing outside replicating hematopoietic niches. Although, strictly speaking, every transplant recipient is a chimera, microchimerism has been viewed as a special circumstance in many experimental studies.^56–58 Clinically, only one study has suggested an association between microchimerism and tolerance.⁵⁸ In general, microchimerism seems to be a property of transplant recipients and not a mechanistic harbinger of immune tolerance.^59,60

Chimerism
In complete distinction from hematopoietic cell infusion and microchimerism, hematopoietic cell engraftment and establishment of macrochimerism is a durable mechanism for central tolerance. Ideally, full chimerism provides the best condition for allograft tolerance, with successful marrow replacement from a kidney donor ensuring tolerance.^61–66 However, the significant morbidity of marrow transplantation makes it realistically achievable in only a small number of cases, with morbidity clearly exceeding that of standard immunosuppressive regimens.

To achieve the same outcome as marrow transplantation with less morbidity, investigators have developed mixed chimerism^67,68 (reviewed by Sykes³⁸ and Wekerle and Sykes⁵⁹). In mixed chimerism, the recipient marrow is largely preserved but modified such that both donor and recipient hemopoietic components coexist. By avoiding donor marrow rejection, transferred hematopoietic cells populate the recipient thymus and marrow and facilitate central deletion of donor alloreactive T and B cells.^67,69 This nonmyeloablative approach has the advantages of being less toxic, preserving immunocompetence, and lessening the risk for graft-versus-host disease. It does, however, depend on a rigorous early regimen that variably includes T cell depletion, transient maintenance immunosuppression, and thymic or total lymphoid irradiation to prevent marrow rejection.^70,71 Once established, mixed chimerism leads to a continuous influence on the developing T cell repertoire and thus seems more durable than the comparatively acute effects of peripheral AICD. Persistent cells may also promote the peripheral effects described with marrow infusion.

Mixed chimerism has been richly investigated and proved to be a means of achieving durable tolerance in animals (reviewed by Sykes,³⁸ and Wekerle and Sykes,⁵⁹ and Sykes et al.⁷²). Initial clinical success was reported by Strober using a conditioning regimen based on total lymphoid irradiation (TLI; see the TLI section) and has more recently been achieved in patients who require marrow replacement for multiple myeloma.^70,71,73,74 In the latter cases, the marrow donor was HLA identical with the recipient, and the conditioning regimen and marrow transplant were medically indicated regardless of the renal transplant. Pilot trials using haplodisparate donor–recipient pairs without underlying malignancy are now ongoing with cautiously optimistic preliminary results.⁷⁵ Mixed chimerism seems to be a promising but practically complex approach to tolerance. Unlike in small animal models, in which tolerance depends on the persistence of macrochimerism, in HLA-disparate primates (human and nonhuman), transient chimerism has been sufficient to induce tolerance in some patients.^32,70 Thus, the mechanisms involved in the clinical experience may be somewhat different from those in the experimental setting and take on some of the peripheral properties of AICD already discussed and vigorous depletion discussed next.

TLI
Lymph nodes and the spleen are critical to the normal development of an alloimmune response.⁷⁶ As such, targeted irradiation to lymphoid tissue has been used to control the immune response after transplantation in animals^9,31,77 and in patients.^29,78,79 When used alone, this approach has had the effect of vigorous T cell depletion (see next section). However, more recently, TLI has been used as a means of immunosuppression to facilitate marrow engraftment and subsequent mixed chimerism.⁷⁹ As a pretransplantation conditioning regimen, TLI has induced tolerance in chimeric and nonchimeric humans, whereas posttransplantation irradiation has failed to produce stable chimeras or tolerance.⁸⁰

Lymphocyte Depletion
Tolerance strategies all have a common aim of controlling allospecific T cell precursor frequency. Between 1 and 10% of peripheral T cells are able to recognize alloantigen,^81,82 which far exceeds the frequency of T cells for any given nominal antigen. Lymphocyte depletion with polyclonal or monoclonal antibodies has been envisioned as a strategy to reduce nonspecifically the precursor frequency, allowing peripheral mechanisms to proceed with less risk for allograft injury. These agents have been used both as induction and as rescue therapy of acute rejection (reviewed by Kirk³⁷). Although nonspecific, polyclonal depletion is not homogeneous. For example, effector memory T cells seem to be relatively resistant to depletion.⁸³ In addition, remaining T cells after depletion undergo homeostatic repopulation, and this has been recognized as a barrier to the development of tolerance.^84,85 Thus, even vigorous depletion alone is not a reliable means toward tolerance.⁸⁶ Depletion is best viewed as a component of other approaches. However, depletion does facilitate reduced maintenance immunosuppressive requirements in many patients, a condition now described as "prope" or "almost" tolerance.⁸⁷

Co-Stimulation Blockade
There is considerable experimental evidence that costimulation blockade facilitates tolerance induction (reviewed by Gudmundsdottir and Turka,³³ Larsen et al.,⁸⁸ Alegre and Najafian,⁸⁹ Yamada et al.,⁹⁰ and Harlan and Kirk⁹¹). Co-stimulation blockade is based on the paradigm that specific immune responses require two signals for optimal activation.⁹² In the absence of a facilitating co-stimulatory signal, antigen stimulation induces anergy or apoptosis. Thus, antigen exposure, combined with co-stimulation molecule inhibition, has the effect of eliminating cells in an antigen-specific manner. Co-stimulation blockade has been used in robust preclinical models to facilitate the protolerant mechanisms at play during peripheral antigen encounter, either with the graft itself^93–96 or with infused hematopoietic cells.⁹⁷ It has also been used as a means to facilitate chimerism.^98,99

Although many co-stimulatory molecules have been identified and tested experimentally, only one, CD28, is targeted by a clinically available agent. CD28 is the most studied T cell co-stimulatory receptor, and its inhibition has been shown to mediate the classic effects of co-stimulation blockade (reviewed by Salomon and Bluestone¹⁰⁰). Two fusion proteins, abatacept and belatacept, have been developed to bind the ligands for CD28, the B7 molecules CD80 and CD86.¹⁰¹ Neither agent has been tested in clinical tolerance trials, although their promise for this eventual application is immense. A recent randomized clinical trial on the use of belatacept in renal transplantation was published with promising results for its use as an immunosuppressive agent.¹⁰² Although designed to study its efficacy in preventing rejection compared with cyclosporine and not to address tolerance, this early study suggests that co-stimulation blockade will have a major role in future tolerance strategies.

Thymic Manipulation
The thymus has a central role in shaping the T cell repertoire via positive and negative selection. Thymic transplantation has been performed clinically in children with thymic agenesia (DiGeorge syndrome). In this application, it successfully restores immunocompetence¹⁰³ and shapes the T cell repertoire. The maturation of T cells in a thymic microenvironment harboring alloantigen (donor murine islet cells¹⁰⁴ or bone marrow cells¹⁰⁵) induces selective unresponsiveness in rats. Other studies have demonstrated this phenomenon after intrathymic injection of donor antigen.^106–110

In animals, the thymus itself has been transplanted in different ways to induce tolerance: As nonvascularized allogeneic thymic tissue, in composite organs ("thymokidney"¹¹¹), and as vascularized thymic lobe transplants.¹¹² Even adult thymic grafts, after the age of thymic involution, can induce transplantation tolerance in animals.¹¹³ These thymus-dependent strategies of tolerance induction are still in the preclinical phase, but the experience in DiGeorge syndrome has provided some clinical proof of concept.

	MONITORING FOR TOLERANCE

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
A major barrier to clinical tolerance is the absence of a method to detect it prospectively. In animal models, donor and third-party skin grafting has been used as a robust test, but this is not a practical clinical approach for many reasons. In humans, many immunologic assays have been used as surrogate tests (Table 2) to monitor the immune response after transplantation (reviewed by Newel and Larson¹¹⁴ and Najafian et al.¹¹⁵). Tests of antigen-specific T cell responses (mixed lymphocyte reaction, limiting dilution) have not been shown to predict the development of tolerance or been helpful guides for immunosuppression withdrawal. More recent tests of precursor frequency (enzyme-linked immunosorbent spot, tetramer analysis, and others^116,117), although promising during immunosuppression, have not yet been applied in tolerant patients. Very sensitive molecular techniques have become available to quantify relevant gene expression patterns^118–121 and protein signatures^122,123 in biologic samples. Using these tests, the search for a signature of tolerance is very active but remains elusive. It remains to be seen whether definition of tolerance at a given point in time will predict one's future risk of alloimmune activation.

	FUTURE DIRECTIONS

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

Regimens that will likely continue to be investigated include those that are based on mechanisms of chimerism, depletion, and co-stimulation blockade. In addition, continuous monitoring of patients who achieved spontaneous tolerance and of those who did not will improve our understanding of tolerance. Finally, given the almost universal elimination of acute rejection as a cause of graft loss with current immunosuppression regimens, more attention will need to be devoted to the development of tolerance strategies that are aimed at the prevention of chronic allograft damage. Indeed, the relationship between existing immunosuppression strategies and the mechanisms of tolerance cited herein will likely have relevance in developing enduring means of avoiding chronic graft fibrosis and tubular atrophy. Funding mechanisms such as the Immune Tolerance Network¹²⁵ and other concerted efforts to facilitate the attack on this worthy but challenging goal in transplantation will be required well into the future for success to be realized and validated.

	DISCLOSURES

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 
None.

	Acknowledgments

 
This work was supported by the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases of the National Institutes of Health.

	Footnotes

 
Published online ahead of print. Publication date available at www.jasn.org.

	REFERENCES

Top Abstract Introduction DEFINING TOLERANCE TOLERANCE IS, IN GENERAL,... SPONTANEOUS TOLERANCE AFTER... STRATEGIES TO ACHIEVE TOLERANCE... MONITORING FOR TOLERANCE FUTURE DIRECTIONS DISCLOSURES REFERENCES

 

1. Gaspare Tagliacozzi (1545–1599): Surgical Implants for Mutilations, edited by Gaspace Bindoni the Younger, Venice, 1597
2. Padgett ECM: Is iso-skin grafting practicable? South Med J 25 : 895 –900, 1932
3. Brown JB: Homografting of skin: With report of success in identical twins. Surgery 1 : 558 –563, 1937
4. Billingham RE, Brent L, Medawar PB: "Actively acquired" tolerance of foreign cells. Nature 172 : 603 –606, 1953[CrossRef][Medline]
5. Calne RY, Sells RA, Pena JR, Davis DR, Millard PR, Herbertson BM, Binns RM, Davies DA: Induction of immunological tolerance by porcine liver allografts. Lancet 223 : 472 –476, 1969
6. Horner BM, Cina RA, Wikiel KJ, Lima B, Ghazi A, Lo DP, Yamada K, Sachs DH, Huang CA: Predictors of organ allograft tolerance following hematopoietic cell transplantation. Am J Transplant 6 : 2894 –2902, 2006[CrossRef][Medline]
7. Kuhr CS, Allen MD, Junghanss C, Zaucha JM, Marsh CL, Yunusov M, Zellme E, Little MT, Torok-Storb B, Storb R: Tolerance to vascularized kidney grafts in canine mixed hematopoietic chimeras. Transplantation 73 : 1487 –1493, 2002[CrossRef][Medline]
8. Storb R, Yu C, Wagner JL, Deeg HJ, Nash RA, Kiem HP, Leisenring W, Shulman H: Stable mixed hematopoietic chimerism in DLA-identical littermate dogs given sublethal total body irradiation before and pharmacological immunosuppression after marrow transplantation. Blood 89 : 3048 –3054, 1997[Abstract/Free Full Text]
9. Strober S, Modry DL, Hoppe RT, Pennock JL, Bieber CP, Holm BI, Jamieson SW, Stinson EB, Schroder J, Suomalainen H: Induction of specific unresponsiveness to heart allografts in mongrel dogs treated with total lymphoid irradiation and antithymocyte globulin. J Immunol 132 : 1013 –1018, 1984[Abstract]
10. Kean LS, Gangappa S, Pearson TC, Larsen CP: Transplant tolerance in non-human primates: Progress, current challenges and unmet needs. Am J Transplant 6 : 884 –893, 2006[CrossRef][Medline]
11. Kirk AD: Crossing the bridge: Large animal models in translational transplantation research. Immunol Rev 196 : 176 –196, 2003[CrossRef][Medline]
12. Williams A, Eldridge R, McFarland H, Houff S, Krebs H, McFarlin D: Multiple sclerosis in twins. Neurology 30 : 1139 –1147, 1980[Abstract/Free Full Text]
13. Brandrup F, Holm N, Grunnet N, Henningsen K, Hansen HE: Psoriasis in monozygotic twins: Variations in expression in individuals with identical genetic constitution. Acta Derm Venereol 62 : 229 –236, 1982[Medline]
14. Adams AB, Williams MA, Jones TR, Shirasugi N, Durham MM, Kaech SM, Wherry EJ, Onami T, Lanier JG, Kokko KE, Pearson TC, Ahmed R, Larsen CP: Heterologous immunity provides a potent barrier to transplantation tolerance. J Clin Invest 111 : 1887 –1895, 2003[CrossRef][Medline]
15. Roussey-Kesler G, Giral M, Moreau A, Subra JF, Legendre C, Noel C, Pillebout E, Brouard S, Soulillou JP: Clinical operational tolerance after kidney transplantation. Am J Transplant 6 : 736 –746, 2006[CrossRef][Medline]
16. Newell KA, Burlingham WJ, Marks VH, Gisler TD, Seyfert VL, Ding R, Turka LA, Suthanthiran M, Kirk AD: Preliminary analysis of the ITN registry of tolerant kidney transplant recipients. Transplantation 82 : 1055 , 2006
17. United Network for Organ Sharing: Data, Richmond, VA, United Network for Organ Sharing. Available at: www.unos.org/data. Accessed December 14, 2006
18. Lerut J, Sanchez-Fueyo A: An appraisal of tolerance in liver transplantation. Am J Transplant 6 : 1774 –1780, 2006[CrossRef][Medline]
19. Donohoe JA, Andrus L, Bowen KM, Simeonovic C, Prowse SJ, Lafferty KJ: Cultured thyroid allografts induce a state of partial tolerance in adult recipient mice. Transplantation 35 : 62 –67, 1983[Medline]
20. Crispe IN, Giannandrea M, Klein I, John B, Sampson B, Wuensch S: Cellular and molecular mechanisms of liver tolerance. Immunol Rev 213 : 101 –118, 2006[CrossRef][Medline]
21. Asolati M, Testa G, Gangemi A, Sankary H, Oberholzer J, Benedetti E: ‘Prope’ tolerance in a noncompliant living related small bowel transplant recipient after severe rejection. Transplantation 83 : 77 –79, 2007[CrossRef][Medline]
22. Zoller KM: Cessation of immunosuppressive therapy after successful transplantation: A national survey. Kidney Int 18 : 110 –114, 1980[Medline]
23. Li Y, Koshiba T, Yoshizawa A, Yonekawa Y, Masuda K, Ito A, Ueda M, Mori T, Kawamoto H, Tanaka Y, Sakaguchi S, Minato N, Wood KJ, Tanaka K: Analyses of peripheral blood mononuclear cells in operational tolerance after pediatric living donor liver transplantation. Am J Transplant 4 : 2118 –2125, 2004[CrossRef][Medline]
24. Martinez-Llordella M, Puig-Pey I, Orlando G, Ramoni M, Tisone G, Rimola A, Lerut J, Latinne D, Margarit C, Bilbao I, Brouard S, Hernandez-Fuentes M, Soulillou J-P, Sanchez-Fueyo A: Multiparameter immune profiling of operational tolerance in liver transplantation. Am J Transplant 7 : 309 –319, 2007[CrossRef][Medline]
25. Mazariegos GV, Zahorchak AF, Reyes J, Ostrowski L, Flynn B, Zeevi A, Thomson AW: Dendritic cell subset ratio in peripheral blood correlates with successful withdrawal of immunosuppression in liver transplant patients. Am J Transplant 3 : 689 –696, 2003[CrossRef][Medline]
26. Kirk AD: Meteorology and tolerance. Am J Transplant 6 : 645 –646, 2006[CrossRef][Medline]
27. Selin LK, Brehm MA, Naumov YN, Cornberg M, Kim SK, Clute SC, Welsh RM: Memory of mice and men: CD8+ T-cell cross-reactivity and heterologous immunity. Immunol Rev 211 : 164 –181, 2006[CrossRef][Medline]
28. Tan HP, Kaczorowski DJ, Basu A, Unruh M, McCauley J, Wu C, Donaldson J, Dvorchik I, Kayler L, Marcos A, Randhawa P, Smetanka C, Starzl TE, Shapiro R: Living donor renal transplantation using alemtuzumab induction and tacrolimus monotherapy. Am J Transplant 6 : 2409 –2417, 2006[CrossRef][Medline]
29. Strober S: Acquired immune tolerance to cadaveric renal allografts. A study of three patients treated with total lymphoid irradiation. N Engl J Med 321 : 28 –33, 1989[Medline]
30. Girlanda R, Rela M, Williams R, O'Grady JG, Heaton ND: Long-term outcome of immunosuppression withdrawal after liver transplantation. Transplant Proc 37 : 1708 –1709, 2005[CrossRef][Medline]
31. Myburgh JA, Smit JA, Hill RR, Browde S: Transplantation tolerance in primates following total lymphoid irradiation and allogeneic bone marrow injection. II. Renal allografts. Transplantation 29 : 405 –408, 1980[Medline]
32. Kawai T, Cosimi AB, Colvin RB, Powelson J, Eason J, Kozlowski T, Sykes M, Monroy R, Tanaka M, Sachs DH: Mixed allogeneic chimerism and renal allograft tolerance in cynomolgus monkeys. Transplantation 59 : 256 –262, 1995[Medline]
33. Gudmundsdottir H, Turka LA: T cell costimulatory blockade: New therapies for transplant rejection. J Am Soc Nephrol 10 : 1356 –1365, 1999[Abstract/Free Full Text]
34. Dong VM, Womer KL, Sayegh MH: Transplantation tolerance: The concept and its applicability. Pediatr Transplant 3 : 181 –192, 1999[CrossRef][Medline]
35. Thomas W, Megan S: Induction of tolerance. Surgery 135 : 359 –364, 2004[CrossRef][Medline]
36. Kirk AD: Ethics in the quest for transplant tolerance. Transplantation 77 : 947 –951, 2004[CrossRef][Medline]
37. Kirk AD: Induction immunosuppression. Transplantation 82 : 593 –602, 2006[Medline]
38. Sykes M: Mixed chimerism and transplant tolerance. Immunity 14 : 417 –424, 2001[CrossRef][Medline]
39. Roy Calne S: Progress toward tolerance and xenografting. Transplant Proc 29 : 16 –18, 1997[Medline]
40. Starzl TE, Marchioro TL, Wadell WR: The reversal of rejection in human renal homografts with subsequent development of homograft tolerance. Surg Gynecol Obstet 117 : 385 –395, 1963[Medline]
41. Starzl TE, Murase N, Demetris AJ, Trucco M, Abu-Elmagd K, Gray EA, Eghtesad B, Shapiro R, Marcos A, Fung JJ: Lessons of organ-induced tolerance learned from historical clinical experience. Transplantation 77 : 926 –929, 2004[CrossRef][Medline]
42. Karim M, Steger U, Bushell AR, Wood KJ: The role of the graft in establishing tolerance. Front Biosci 7 : e129 –e154, 2002[Medline]
43. Green DR, Droin N, Pinkoski M: Activation-induced cell death in T cells. Immunol Rev 193 : 70 –81, 2003[CrossRef][Medline]
44. Li Y, Li XC, Zheng XX, Wells AD, Turka LA, Strom TB: Blocking both signal 1 and signal 2 of T-cell activation prevents apoptosis of alloreactive T cells and induction of peripheral allograft tolerance. Nat Med 5 : 1298 –1302, 1999[CrossRef][Medline]
45. Wells AD, Li XC, Li Y, Walsh MC, Zheng XX, Wu Z, Nunez G, Tang A, Sayegh M, Hancock WW, Strom TB, Turka LA: Requirement for T-cell apoptosis in the induction of peripheral transplantation tolerance. Nat Med 5 : 1303 –1307, 1999[CrossRef][Medline]
46. Adams AB, Shirasugi N, Jones TR, Williams MA, Durham MM, Ha J, Dong Y, Guo Z, Newell KA, Pearson TC, Larsen CP: Conventional immunosuppression is compatible with costimulation blockade-based, mixed chimerism tolerance induction. Am J Transplant 3 : 895 –901, 2003[CrossRef][Medline]
47. Blaha P, Bigenzahn S, Koporc Z, Schmid M, Langer F, Selzer E, Bergmeister H, Wrba F, Kurtz J, Kiss C, Roth E, Muehlbacher F, Sykes M, Wekerle T: The influence of immunosuppressive drugs on tolerance induction through bone marrow transplantation with costimulation blockade. Blood 101 : 2886 –2893, 2003[Abstract/Free Full Text]
48. Opelz G: Effect of blood transfusions on subsequent kidney transplants. Transplant Proc 5 : 253 –259, 1973[Medline]
49. Flye MW: Donor-specific transfusions have long-term beneficial effects for human renal allografts. Transplantation 60 : 1395 –1401, 1995[Medline]
50. Monaco AP: Possible active enhancement of a human cadaver renal allograft with antilymphocyte serum (ALS) and donor bone marrow: Case report of an initial attempt. Surgery 79 : 384 –392, 1976[Medline]
51. Barber WH: Long-term results of a controlled prospective study with transfusion of donor-specific bone marrow in 57 cadaveric renal allograft recipients. Transplantation 51 : 70 –75, 1991[Medline]
52. Ciancio G, Burke GW, Moon J, Garcia-Morales R, Rosen A, Esquenazi V, Mathew J, Jin Y, Miller J: Donor bone marrow infusion in deceased and living donor renal transplantation. Yonsei Med J 45 : 998 –1003, 2004[Medline]
53. Wood KJ, Jones ND, Bushell AR, Morris PJ: Alloantigen-induced specific immunological unresponsiveness. Phil Trans R Soc B 356 : 665 –680, 2001[CrossRef][Medline]
54. Swanson SJ, Hale DA, Mannon RB, Kleiner DE, Cendales LC, Chamberlain CE, Polly SM, Harlan DM, Kirk AD: Kidney transplantation with rabbit antithymocyte globulin induction and sirolimus monotherapy. Lancet 360 : 1662 –1664, 2002[CrossRef][Medline]
55. Starzl TE, Murase N, Abu-Elmagd K, Gray EA, Shapiro R, Eghtesad B, Corry RJ, Jordan ML, Fontes P, Gayowski T: Tolerogenic immunosuppression for organ transplantation. Lancet 361 : 1502 –1510, 2003[CrossRef][Medline]
56. Starzl TE: Chimerism and tolerance in transplantation. Proc Natl Acad Sci U S A 101 : 14607 –14614, 2004[Abstract/Free Full Text]
57. Starzl TE, Murase N, Ildstad S, Ricordi C, Demetris AJ, Trucco M: Cell migration, chimerism, and graft acceptance. Lancet 339 : 1579 –1582, 1992[CrossRef][Medline]
58. Ciancio G, Miller J, Garcia-Morales RO, Carreno M, Burke GWI, Roth D, Kupin W, Tzakis AG, Ricordi C, Rosen A, Fuller L, Esquenazi V: Six-year clinical effect of donor bone marrow infusions in renal transplant patients. Transplantation 71 : 827 –835, 2001[CrossRef][Medline]
59. Wekerle T, Sykes M: Mixed chimerism and transplantation tolerance. Annu Rev Med 52 : 353 –370, 2001[CrossRef][Medline]
60. Monaco AP, Medawar P: Chimerism in organ transplantation: Conflicting experiments and clinical observations. Transplantation 75 : 13S –16S, 2003[CrossRef][Medline]
61. Sayegh MH: Immunologic tolerance to renal allografts after bone marrow transplants from the same donors. Ann Intern Med 114 : 954 –955, 1991[Medline]
62. Helg C: Renal transplantation without immunosuppression in a host with tolerance induced by allogeneic bone marrow transplantation. Transplantation 58 : 1420 –1422, 1994[Medline]
63. Sorof JM: Renal transplantation without chronic immunosuppression after T cell-depleted, HLA-mismatched bone marrow transplantation. Transplantation 59 : 1633 –1635, 1995[Medline]
64. Butcher JA, Hariharan S, Adams MB, Johnson CP, Roza AM, Cohen EP: Renal transplantation for end-stage renal disease following bone marrow transplantation: A report of six cases, with and without immunosuppression. Clin Transplant 13 : 330 –335, 1999[CrossRef][Medline]
65. Dey BM, Sykes MM, Spitzer TRM: Outcomes of recipients of both bone marrow and solid organ transplants: A review. Medicine 77 : 355 –369, 1998[CrossRef][Medline]
66. Jacobsen N, Taaning E, Ladefoged J, Kvist Kristensen J, Pedersen F: Tolerance to an HLA-B, DR disparate kidney allograft after bone-marrow transplantation from same donor. Lancet 343 : 800 , 1994[CrossRef][Medline]
67. Ildstad ST, Wren SM, Bluestone JA, Barbieri SA, Sachs DH: Characterization of mixed allogeneic chimeras. Immunocompetence, in vitro reactivity, and genetic specificity of tolerance. J Exp Med 162 : 231 –244, 1985[Abstract/Free Full Text]
68. Sykes M, Szot GL, Swenson KA, Pearson DA: Induction of high levels of allogeneic hematopoietic reconstitution and donor-specific tolerance without myelosuppressive conditioning. Nat Med 3 : 783 –787, 1997[CrossRef][Medline]
69. Domenig C, Sanchez-Fueyo A, Kurtz J, Alexopoulos SP, Mariat C, Sykes M, Strom TB, Zheng XX: Roles of deletion and regulation in creating mixed chimerism and allograft tolerance using a nonlymphoablative irradiation-free protocol. J Immunol 175 : 51 –60, 2005[Abstract/Free Full Text]
70. Spitzer TR, Delmonico F, Tolkoff-Rubin N, McAfee S, Sackstein R, Saidman S, Colby C, Sykes M, Sachs DH, Cosimi AB: Combined histocompatibility leukocyte antigen matched donor bone marrow and renal transplantation for multiple myeloma with end stage renal disease: The induction of allograft tolerance through mixed lymphohematopoietic chimerism. Transplantation 68 : 480 –484, 1999[CrossRef][Medline]
71. Millan MT, Shizuru JA, Hoffmann P, Dejbakhsh-Jones S, Scandling JD, Carl Grumet F, Tan JC, Salvatierra O, Hoppe RT, Strober S: Mixed chimerism and immunosuppressive drug withdrawal after HLA-mismatched kidney and hematopoietic progenitor transplantation. Transplantation 73 : 1386 –1391, 2002[CrossRef][Medline]
72. Sykes M, Shimizu I, Kawahara T: Mixed hematopoietic chimerism for the simultaneous induction of T and B cell tolerance. Transplantation 79[Suppl] : S28 –S29, 2005[CrossRef][Medline]
73. Buhler LH, Spitzer TR, Sykes M, Sachs DH, Delmonico FL, Tolkoff-Rubin N, Saidman SL, Sackstein R, McAfee S, Dey B, Colby C, Cosimi AB: Induction of kidney allograft tolerance after transient lymphohematopoietic chimerism in patients with multiple myeloma and end-stage renal disease. Transplantation 74 : 1405 –1409, 2002[CrossRef][Medline]
74. De Pauw L, Toungouz M, Goldman M: Infusion of donor-derived hematopoietic stem cells in organ transplantation: Clinical data. Transplantation 75[Suppl] : 46S –49S, 2003[CrossRef][Medline]
75. Kawai T, Sachs DH, Spitzer T, Tolkoff-Rubin N, Shaffer J, Saidman S, Williams W, Dey B, Goes N, Ko D, Hertl M, Cotter P, Wong W, Colvin RB, Sykes M, Cosimi AB: Successful induction of renal allograft tolerance in HLA-mismatched kidney transplant recipients. Transplantation 82[Suppl 2] : 169 , 2006
76. Lakkis FG, Arakelov A, Konieczny BT, Inoue Y: Immunologic ‘ignorance’ of vascularized organ transplants in the absence of secondary lymphoid tissue. Nat Med 6 : 686 –688, 2000[CrossRef][Medline]
77. Raaf J, Bryan C, Monden M, Bray A, Kim JH, Chu F, Chaganti RS, Shank B, Cahan A, Fortner JG: Bone marrow and renal transplantation in canine recipients prepared by total lymphoid irradiation. Transplant Proc 13 : 429 –433, 1981[Medline]
78. Levin B, Collins G, Waer M, Girinsky T, Hoppe R, Miller E, Bieber C, Strober S: Treatment of cadaveric renal transplant recipients with total lymphoid irradiation, antithymocyte globulin, and low-dose prednisone. Lancet 326 : 1321 –1325, 1985[CrossRef]
79. Strober S, Benike C, Krishnaswamy S, Engleman EG, Grumet FC: Clinical transplantation tolerance twelve years after prospective withdrawal of immunosuppressive drugs: Studies of chimerism and anti-donor reactivity. Transplantation 69 : 1549 –1554, 2000[Medline]
80. Strober S, Lowsky RJ, Shizuru JA, Scandling JD, Millan MT: Approaches to transplantation tolerance in humans. Transplantation 77 : 932 –936, 2004[CrossRef][Medline]
81. Ford WL, Atkins RC: The proportion of lymphocytes capable of recognizing strong transplantation antigens in vivo. Adv Exp Med Biol 29 : 255 –262, 1973[Medline]
82. Suchin EJ, Langmuir PB, Palmer E, Sayegh MH, Wells AD, Turka LA: Quantifying the frequency of alloreactive T cells in vivo: New answers to an old question. J Immunol 166 : 973 –981, 2001[Abstract/Free Full Text]
83. Pearl JP, Parris J, Hale DA, Hoffmann SC, Bernstein WB, McCoy KL, Swanson SJ, Mannon RB, Roederer M, Kirk AD: Immunocompetent T-cells with a memory-like phenotype are the dominant cell type following antibody-mediated T-cell depletion. Am J Transplant 5 : 465 –474, 2005[Medline]
84. Wu Z, Bensinger SJ, Zhang J, Chen C, Yuan X, Huang X, Markmann JF, Kassaee A, Rosengard BR, Hancock WW, Sayegh MH, Turka LA: Homeostatic proliferation is a barrier to transplantation tolerance. Nat Med 10 : 87 –92, 2004[CrossRef][Medline]
85. Neujahr DC, Chen C, Huang X, Markmann JF, Cobbold S, Waldmann H, Sayegh MH, Hancock WW, Turka LA: Accelerated memory cell homeostasis during T cell depletion and approaches to overcome it. J Immunol 176 : 4632 –4639, 2006[Abstract/Free Full Text]
86. Kirk AD, Hale DA, Mannon RB, Kleiner DE, Hoffmann SC, Kampen RL, Cendales LK, Tadaki DK, Harlan DM, Swanson SJ: Results from a human renal allograft tolerance trial evaluating the humanized CD52-specific monoclonal antibody alemtuzumab (CAMPATH-1H). Transplantation 76 : 120 –129, 2003[CrossRef][Medline]
87. Calne RY: Prope tolerance: The future of organ transplantation from the laboratory to the clinic. Transpl Immunol 13 : 83 –86, 2004[CrossRef][Medline]
88. Larsen CP, Knechtle SJ, Adams A, Pearson T, Kirk AD: A new look at blockade of T-cell costimulation: A therapeutic strategy for long-term maintenance immunosuppression. Am J Transplant 6 : 876 –883, 2006[CrossRef][Medline]
89. Alegre ML, Najafian N: Costimulatory molecules as targets for the induction of transplantation tolerance. Curr Mol Med 8 : 843 –857, 2006
90. Yamada A, Sayegh MH: The CD154-CD40 costimulatory pathway in transplantation. Transplantation 73[Suppl] : S36 –S39, 2002[CrossRef][Medline]
91. Harlan DM, Kirk AD: The future of organ and tissue transplantation: Can T-cell costimulatory pathway modifiers revolutionize the prevention of graft rejection? JAMA 282 : 1076 –1082, 1999[Abstract/Free Full Text]
92. Cunningham AJ, Lafferty KJ: A simple conservative explanation of the H-2 restriction of interactions between lymphocytes. Scand J Immunol 6 : 1 –6, 1977[CrossRef][Medline]
93. Kirk AD, Burkly LC, Batty DS, Baumgartner RE, Berning JD, Buchanan K, Fechner JH, Germond RL, Kampen RL, Patterson NB, Swanson SJ, Tadaki DK, TenHoor CN, White L, Knechtle SJ, Harlan DM: Treatment with humanized monoclonal antibody against CD154 prevents acute renal allograft rejection in nonhuman primates. Nat Med 5 : 686 –693, 1999[CrossRef][Medline]
94. Kirk AD, Harlan DM, Armstrong NN, Davis TA, Dong Y, Gray GS, Hong X, Thomas D, Fechner JH Jr, Knechtle SJ: CTLA4-Ig and anti-CD40 ligand prevent renal allograft rejection in primates. Proc Natl Acad Sci U S A 94 : 8789 –8794, 1997[Abstract/Free Full Text]
95. Larsen CP, Elwood ET, Alexander DZ, Ritchie SC, Hendrix R, Tucker-Burden C, Cho HR, Aruffo A, Hollenbaugh D, Linsley PS, Winn KJ, Pearson TC: Long-term acceptance of skin and cardiac allografts after blocking CD40 and CD28 pathways. Nature 381 : 434 –438, 1996[CrossRef][Medline]
96. Kirk AD, Tadaki DK, Celniker A, Scott Batty D, Berning JD, Colonna JO, Cruzata F, Elster EA, Gray GS, Kampen RL, Patterson NB, Szklut P, Swanson J, Xu H, Harlan DM: Induction therapy with monoclonal antibodies specific for CD80 and CD86 delays the onset of acute renal allograft rejection in non-human primates. Transplantation 72 : 377 –384, 2001[CrossRef][Medline]
97. Preston EH, Xu H, Dhanireddy KK, Pearl JP, Leopardi FV, Starost MF, Hale DA, Kirk AD: IDEC-131 (Anti-CD154), Sirolimus and donor-specific transfusion facilitate operational tolerance in non-human primates. Am J Transplant 5 : 1032 –1041, 2005[CrossRef][Medline]
98. Kawai T, Sogawa H, Boskovic S, Abrahamian G, Smith RN, Wee SL, Andrews D, Nadazdin O, Koyama I, Sykes M, Winn HJ, Colvin RB, Sachs DH, Cosimi AB: CD154 Blockade for induction of mixed chimerism and prolonged renal allograft survival in nonhuman primates. Am J Transplant 4 : 1391 –1398, 2004[CrossRef][Medline]
99. Kean LS, Adams AB, Strobert E, Hendrix R, Gangappa S, Jones TR, Shirasugi N, Rigby MR, Hamby K, Jiang J, Bello H, Anderson D, Cardona K, Durham MM, Pearson TC, Larsen CP: Induction of chimerism in rhesus macaques through stem cell transplant and costimulation blockade-based immunosuppression. Am J Transplant 7 : 320 –335, 2007[CrossRef][Medline]
100. Salomon B, Bluestone JA: Complexities of CD28/B7: CTLA-4 costimulatory pathways in autoimmunity and transplantation. Annu Rev Immunol 19 : 225 –252, 2001[CrossRef][Medline]
101. Larsen CP, Pearson TC, Adams AB, Tso P, Shirasugi N, Strobert ME, Anderson D, Cowan S, Price K, Naemura J, Emswiler J, Greene J, Turk LA, Bajorath J, Townsend R, Hagerty D, Linsley PS, Peach RJ: Rational development of LEA29Y (belatacept), a high-affinity variant of CTLA4-Ig with potent immunosuppressive properties. Am J Transplant 5 : 443 –453, 2005[CrossRef][Medline]
102. Vincenti F, Larsen C, Durrbach A, Wekerle T, Nashan B, Blancho G, Lang P, Grinyo J, Halloran PF, Solez K, Hagerty D, Levy E, Zhou W, Natarajan K, Charpentier B; the Belatacept Study Group: Costimulation blockade with belatacept in renal transplantation. N Engl J Med 353 : 770 –781, 2005[Abstract/Free Full Text]
103. Rice HE, Skinner MA, Mahaffey SM, Oldham KT, Ing RJ, Hale LP, Markert ML: Thymic transplantation for complete DiGeorge syndrome: Medical and surgical considerations. J Pediatr Surg 39 : 1607 –1615, 2004[CrossRef][Medline]
104. Posselt AM, Barker CF, Tomaszewski JE, Markmann JF, Choti MA, Naji A: Induction of donor-specific unresponsiveness by intrathymic islet transplantation. Science 249 : 1293 –1295, 1990[Abstract/Free Full Text]
105. Odorico JS: Induction of donor-specific tolerance to rat cardiac allografts by intrathymic inoculation of bone marrow. Surgery 112 : 370 –376, 1992[Medline]
106. Knechtle SJ, Wang J, Jiao S, Geissler EK, Sumimoto R, Wolff J: Induction of specific tolerance by intrathymic injection of recipient muscle cells transfected with donor class I major histocompatibility complex. Transplantation 57 : 990 –996, 1994[Medline]
107. Ohzato H, Monaco AP: Induction of specific unresponsiveness (tolerance) to skin allografts by intrathymic donor-specific splenocyte injection in antilymphocyte serum-treated mice. Transplantation 54 : 1090 –1095, 1992[Medline]
108. Knechtle SJ, Wang J, Graeb C, Zhai Y, Hong X, Fechner JH Jr, Geissler EK: Direct MHC class I complementary DNA transfer to thymus induces donor-specific unresponsiveness, which involves multiple immunologic mechanisms. J Immunol 159 : 152 –158, 1997[Abstract]
109. Goss JA, Nakafusa Y Flye MW: Intrathymic injection of donor alloantigens induces donor-specific vascularized allograft tolerance without immunosuppression. Ann Surg 216 : 414 –416, 1992
110. Remuzzi G, Rossini M, Imberti O, Perico N: Kidney graft survival in rats without immunosuppressants after intrathymic glomerular transplantation. Lancet 337 : 750 –752, 1991[CrossRef][Medline]
111. Yamada K, Shimizu A, Ierino FL, Utsugi R, Barth RN, Esnaola N, Colvin RB, Sachs DH: Thymic transplantation in miniature swine: I. Development and function of the "Thymokidney." Transplantation 68 : 1684 –1692, 1999[CrossRef]