Tolerance in clinical liver transplantation: The long road ahead
Authors: Jean C. Emond, Adam D. Griesemer
Journal: Hepatology (2016)

Potential conflict of interest: Dr. Griesemer received grants from United Therapeutics. See Article On Page 647 In this issue of Hepatology, Feng et al. report on long‐term immunological, histological, and clinical outcomes of a cohort of 12 pediatric liver transplantation (LT) patients that were weaned off of immunosuppression. This editorial discusses the broader implications of the study and some of the important issues involved in liver tolerance. Despite excellent short‐term results, organ transplants are subject to chronic deterioration over time and the recipients face the consequences of long‐term exposure to immunosuppressive drugs. Nonetheless, it has long been known that some patients may thrive without immunosuppression, a state that is uncovered either because of nonadherence or the removal of immunosuppression to treat infections or lymphoproliferative disorders.1 These observations have led to numerous efforts to withdraw or minimize immunosuppression in clinical LT, with the most optimistic reports suggesting that up to 50% of selected patients might be weaned from immunosuppression.3 The term "operational tolerance" describes a patient with clinically normal graft function without immunosuppression. This practical definition does not specify the complex immunological balance between donor and recipient that characterizes the true tolerant state, and spontaneously tolerant subjects may not share a common mechanism for the maintenance of graft acceptance. The earliest views of tolerance posited ignorance or nonrecognition of alloantigens analogous to clonal deletion occurring during development of the immune system.1 A more‐subtle view suggests that tolerance is a dynamic, self‐replicating state requiring host recognition of the graft antigens with the generation of a stable regulatory environment.4 Events that cause graft inflammation in the early (ischemia‐reperfusion/preservation injury) or late posttransplant period (inflammation attributed to bacterial or viral infections) can alter expression of stimulatory cell‐surface markers,5 and such events have been observed to induce immune responses in previously tolerant subjects, perhaps analogous to development of autoimmunity in otherwise healthy people. On the other hand, the study of immunosuppression withdrawal in hepatitis C virus (HCV)‐positive patients led by Sanchez‐Fueyo suggested that some types of inflammation, in this case HCV‐mediated interferon production, may assist in developing tolerance in certain patients by leading to immune exhaustion.6 In clinical practice, LT patients are followed without liver biopsies as long as they maintain normal liver function, so there is a paucity of information about the histological state of long‐term LTs. Both immunological and nonimmunological damage may occur post‐LT, leading to fibrosis and cholestasis. Chronic rejection is most often associated with cholestatic injury given that the bile duct epithelium richly expresses class II antigens,7 although the bile ducts may be secondarily destroyed by microvascular‐driven humoral mechanisms. LT recipients are also subject to other chronic injury attributed to recurrence of viral or autoimmune diseases that led to their need for transplant in the first place. However, early trials of immunosuppression withdrawal supported by formal histological assessment of protocol biopsies produced the disturbing observation that many apparently well patients have significantly abnormal histology characterized by both inflammation and occult fibrosis. The European trial reported on 8 patients that were operationally tolerant in the presence of HCV in all cases.8 There was a limited immunological analysis of these patients, but a longer‐term follow‐up showed no difference in liver fibrosis in the tolerant patients versus those that failed to be weaned. The Kyoto group found a high incidence of liver fibrosis in pediatric patients in their weaning trial, which regressed if immunosuppression was reinstituted.9 In the current issue of Hepatology, Feng et al. report on a five‐year histological follow‐up of 12 operationally tolerant children who received parental living donor grafts enrolled in the Withdrawal of Immunosuppression in Pediatric Liver Transplant Recipients study.10 In addition to biopsies, humoral markers of immune responses and liver sinusoidal endothelial cell (LSEC) phenotypes were followed prospectively. Overall, none of the tolerant subjects demonstrated progressive increase in inflammation over time, with the exception of a patient with biliary problems. Five of the 12 subjects developed de novo donor‐specific antibodies (DSA) during withdrawal, including both transient and persistent expression of C1q binding that was not consistently associated with class II DSA and was not correlated with histological abnormalities. Interestingly, no patient developed significant levels immunoglobulin G (IgG) 3, the most potent subclass of IgG with regard to ability to fix complement and bind Fc receptors. It is unclear whether this is attributed to a lack of production or more‐efficient clearance by Fc‐expressing cells. Overall, the relationship between the absence of DSA and achievement operational tolerance is inconsistent. The investigators speculate that the highly efficient scavenger system within the liver, with effective clearance of immune complexes, might be one of the features that increase the likelihood of operational tolerance in LT patients. Furthermore, there was decreased fibrosis overall at 5 years, an encouraging finding that suggests reparative mechanisms may reverse fibrosis. Though this study demonstrates the lack of histological progression of graft injury in operationally tolerant children who have received living donor liver transplantation (LDLT), the generalizability of this observation remains in question. These data indicate that operational tolerance can exist in the setting of ongoing B‐cell and plasma cell responses. This, combined with the absence of systemic macrochimerism, makes it likely that so‐called peripheral mechanisms lead to development of operational tolerance post‐LT. The ability of the liver to maintain tolerance to the many foreign food‐derived and bacterial antigens presented in the portal vein has been invoked to explain the "tolerogenicity" of the liver post‐LT. Many cells in the liver have been implicated, including liver dendritic cells, LSECs, and hepatocytes (reviewed in Horst et al.4 and Crispe11). In the liver, CD8 T cells are activated, but unless rescued by interleukin‐2 stimulation,4 this activation leads to exhaustion and cell death.12 Liver antigen‐presenting cells, including LSECs, also promote regulatory T cell (Treg) development and/or recruitment.11 In this report, the investigators examined the phenotype of LSECs in operationally tolerant patients and found no phenotypic changes of activation. Although they did not investigate the expansion of forkhead box P3 (FoxP3)‐positive Tregs in the liver, others have reported an enrichment of FoxP3‐positive cells in biopsies of operationally tolerant livers,9 and Tregs mediate graft survival in mouse LT models.13 The goals in identifying spontaneously tolerant patients are to characterize the regulatory state and eventually identify other patients who might be weaned successfully and to inform interventions to induce tolerance. However, it appears that despite the most optimistic projections, only a small fraction of the overall candidate pool become tolerant. Haploidentical pediatric LDLT recipients are clearly not representative of the overall population attributed to receiving an organ with partial major histocompatibility class matching at a time when the immune system is not fully developed. Characterization of the immune responses in operationally tolerant adults and recipients of fully mismatched livers should place the current findings in perspective. Furthermore, recipients with both primary immunological and viral liver diseases have typically been excluded from withdrawal trials, and the effect of these comorbidities on the potential to develop operational tolerance remains to be determined. It does not necessarily follow that characterizing the operationally tolerant patients will lead to development of effective interventions to induce tolerance. We are hopeful that application of new techniques to define the alloreactive T‐cell repertoire at the genetic level pretransplant and then follow the expansion or contraction of the donor‐reactive T‐cell pool posttransplant will shed light on the active mechanism(s) leading to operational tolerance and potentially help identify patients that are stable on immunosuppression, but who could be withdrawn.14 The chimeric recipients who have tolerance induced by bone marrow grafting have been shown to have regulatory and deletional states that induce graft acceptance.14 Ultimately, both deletional and regulatory mechanisms may be needed to withstand disruption of the homeostasis of tolerance that can be induced by inflammation associated with graft injury or systemic infections. Better mechanistic understanding of how the remaining immune responses are controlled in operationally tolerant patients is required in order to extend the lessons learned from studying these patients to the broader transplant population. Author names in bold designate shared co‐first authorship.