María-Carlota Londoño

Prospective multicenter clinical trial of immunosuppressive drug withdrawal in stable adult liver transplant recipients

Lifelong immunosuppression increases morbidity and mortality in liver transplantation. Discontinuation of immunosuppressive drugs could lessen this burden, but the safety, applicability, and clinical outcomes of this strategy need to be carefully defined. We enrolled 102 stable liver recipients at least 3 years after transplantation in a single‐arm multicenter immunosuppression withdrawal trial. Drugs were gradually discontinued over a 6 to 9‐month period. The primary endpoint was the development of operational tolerance, defined as successful immunosuppressive drug cessation maintained for at least 12 months with stable graft function and no histopathologic evidence of rejection. Out of the 98 recipients evaluated, 57 rejected and 41 successfully discontinued all immunosuppressive drugs. In nontolerant recipients rejection episodes were mild and resolved over 5.6 months (two nontolerant patients still exhibited mild gradually improving cholestasis at the end of follow‐up). In tolerant recipients no progressive clinically significant histological damage was apparent in follow‐up protocol biopsies performed up to 3 years following drug withdrawal. Tolerance was independently associated with time since transplantation (odds ratio [OR] 1.353; P = 0.0001), recipient age (OR 1.073; P = 0.009), and male gender (OR 4.657; P = 0.016). A predictive model incorporating the first two clinical variables identified subgroups of recipients with very high (79%), intermediate (30%‐38%), and very low (0%) likelihood of successful withdrawal. Conclusion: When conducted at late timepoints after transplantation, immunosuppression withdrawal is successful in a high proportion of carefully selected liver recipients. A combination of clinical parameters could be useful to predict the success of this strategy. Additional prospective studies are now needed to confirm these results and to validate clinically applicable diagnostic biomarkers. (Hepatology 2013;58:1824–1835)

Intra-graft expression of genes involved in iron homeostasis predicts the development of operational tolerance in human liver transplantation.

Following organ transplantation, lifelong immunosuppressive therapy is required to prevent the host immune system from destroying the allograft. This can cause severe side effects and increased recipient morbidity and mortality. Complete cessation of immunosuppressive drugs has been successfully accomplished in selected transplant recipients, providing proof of principle that operational allograft tolerance is attainable in clinical transplantation. The intra-graft molecular pathways associated with successful drug withdrawal, however, are not well defined. In this study, we analyzed sequential blood and liver tissue samples collected from liver transplant recipients enrolled in a prospective multicenter immunosuppressive drug withdrawal clinical trial. Before initiation of drug withdrawal, operationally tolerant and non-tolerant recipients differed in the intra-graft expression of genes involved in the regulation of iron homeostasis. Furthermore, as compared with non-tolerant recipients, operationally tolerant patients exhibited higher serum levels of hepcidin and ferritin and increased hepatocyte iron deposition. Finally, liver tissue gene expression measurements accurately predicted the outcome of immunosuppressive withdrawal in an independent set of patients. These results point to a critical role for iron metabolism in the regulation of intra-graft alloimmune responses in humans and provide a set of biomarkers to conduct drug-weaning trials in liver transplantation.

Comparison of Transcriptional and Blood Cell‐Phenotypic Markers Between Operationally Tolerant Liver and Kidney Recipients

A proportion of transplant recipients can spontaneously accept their grafts in the absence of immunosuppression (operational tolerance). Previous studies identified blood transcriptional and cell‐phenotypic markers characteristic of either liver or kidney tolerant recipients. However, the small number of patients analyzed and the use of different transcriptional platforms hampered data interpretation. In this study we directly compared samples from kidney and liver tolerant recipients in order to identify potential similarities in immune‐related parameters. Liver and kidney tolerant recipients differed in blood expression and B‐cell immunophenotypic patterns and no significant overlaps were detectable between them. Whereas some recipients coincided in specific NK‐related transcripts, this observation was not reproducible in all cohorts analyzed. Our results reveal that certain immune features, but not others, are consistently present across all cohorts of operationally tolerant recipients. This provides a set of reproducible biomarkers that should be explored in future large‐scale immunomonitoring trials.

Comparative Transcriptional and Phenotypic Peripheral Blood Analysis of Kidney Recipients Under Cyclosporin A or Sirolimus Monotherapy

Due to its low level of nephrotoxicity and capacity to harness tolerogenic pathways, sirolimus (SRL) has been proposed as an alternative to calcineurin inhibitors in transplantation. The exact mechanisms underlying its unique immunosuppressive profile in humans, however, are still not well understood. In the current study, we aimed to depict the in vivo effects of SRL in comparison with cyclosporin A (CSA) by employing gene expression profiling and multiparameter flow cytometry on blood cells collected from stable kidney recipients under immunosuppressant monotherapy. SRL recipients displayed an increased frequency of CD4 + CD25highFoxp3 + T cells. However, this was accompanied by an increased number of effector memory T cells and by enrichment in NFkB‐related pro‐inflammatory expression pathways and monocyte and NK cell lineage‐specific transcripts. Furthermore, measurement of a transcriptional signature characteristic of operationally tolerant kidney recipients failed to detect differences between SRL and CSA‐treated recipients. In conclusion, we show here that the blood transcriptional profile induced by SRL monotherapy in vivo does not resemble that of operationally tolerant recipients and is dominated by innate immune cells and NFkB‐related pro‐inflammatory events. These data provide novel insights on the complex effects of SLR on the immune system in clinical transplantation.

ATG-Fresenius Treatment and Low-Dose Tacrolimus: Results of a Randomized Controlled Trial in Liver Transplantation

We report the results of a prospective randomized controlled trial in liver transplantation assessing the efficacy and safety of antithymocyte globulin (ATG‐Fresenius) plus tacrolimus monotherapy at gradually decreasing doses. Patients were randomized to either: (a) standard‐dose tacrolimus plus steroids;or (b) peritransplant ATG‐Fresenius plus reduced‐dose tacrolimus monotherapy followed by weaning of tacrolimus starting 3 months after transplantation. The primary end‐point was the achievement of very low‐dose tacrolimus (every‐other‐day or once daily dose with <5 ng/mL trough levels) at 12 months after transplantation. Acute rejection occurring during the first 3 months after transplantation was more frequent in the ATG group (52.4% vs. 25%). Moreover, late acute rejection episodes occurred in all recipients in whom weaning was attempted and no recipients reached the primary end‐point. This motivated the premature termination of the trial. Tacrolimus trough levels were lower in the ATG‐Fresenius group but no benefits in terms of improved renal function, lower metabolic complications or increased prevalence of tolerance‐related biomarkers were observed. In conclusion, the use of ATG‐Fresenius and tacrolimus at gradually decreasing doses was associated with a high rate of rejection, did not allow for the administration of very low doses of tacrolimus and failed to provide detectable clinical benefits. ClinicalTrials.gov identifier:NCT00436722.

HCV-Induced Immune Responses Influence the Development of Operational Tolerance After Liver Transplantation in Humans

Persistent HCV infection does not preclude the establishment of liver allograft tolerance in humans. Turning the Tables on Tolerance When it comes to transplantation, chronic viral infection may not be so bad after all. Bohne et al. demonstrate that chronic infection with hepatitis C virus (HCV) does not prevent the development of tolerance in liver transplant patients. Animal studies have previously indicated that immune responses to infection prevent the establishment of transplantation tolerance, which has resulted in the exclusion of individuals with persistent infection from transplantation tolerance trials. This prospective trial of immunosuppression withdrawal in HCV-infected liver transplant recipients found that the HCV-induced immune response did not influence outcome. Rather, tolerance induction was associated with expression of immunoregulatory genes that were specific for HCV infection. Thus, HCV infection not only does not always prevent the development of transplantation tolerance but also may actually contribute to restraining antitransplant immune responses. Pathogen-induced immune responses prevent the establishment of transplantation tolerance in experimental animal models. Whether this occurs in humans as well remains unclear. The development of operational tolerance in liver transplant recipients with chronic hepatitis C virus (HCV) infection allows us to address this question. We conducted a clinical trial of immunosuppression withdrawal in HCV-infected adult liver recipients to elucidate (i) the mechanisms through which allograft tolerance can be established in the presence of an ongoing inflammatory response and (ii) whether anti-HCV heterologous immune responses influence this phenomenon. Of 34 enrolled liver recipients, drug withdrawal was successful in 17 patients (50%). Tolerance was associated with intrahepatic overexpression of type I interferon and immunoregulatory genes and with an expansion of exhausted PD1/CTLA4/2B4-positive HCV-specific circulating CD8+ T cells. These findings were already present before immunosuppression was discontinued and were specific for HCV infection. In contrast, the magnitude of HCV-induced proinflammatory gene expression and the breadth of anti-HCV effector T cell responses did not influence drug withdrawal outcome. Our data suggest that in humans, persistent viral infections exert immunoregulatory effects that could contribute to the restraining of alloimmune responses, and do not necessarily preclude the development of allograft tolerance.

Immunosuppression minimization vs. complete drug withdrawal in liver transplantation

Despite the increase in long-term survival, liver transplant recipients still exhibit higher morbidity and mortality than the general population. This is in part attributed to the lifelong administration of immunosuppression and its associated side effects. Several studies reported in the last decades have evaluated the impact of immunosuppression minimization in liver transplant recipients, but results have been inconsistent due to the heterogeneity of study designs and insufficient sample sizes. On the other hand, complete immunosuppression withdrawal has proven to be feasible in approximately 20% of carefully selected liver transplant recipients, especially in older patients and those with longer duration after transplantation. The long-term risks and clinical benefits of this strategy, however, also need to be clarified. As a consequence, and despite the general perception that a large proportion of liver recipients are over-immunosuppressed, it is currently not possible to derive evidence-based guidelines on how to manage long-term immunosuppression to improve clinical outcomes. Large clinical trials of drug minimization and/or withdrawal focused on clinically-relevant long-term outcomes are required. Development of personalized medicine tools and a deeper understanding of the pathogenesis of idiopathic inflammatory graft lesions will be pre-requisites to achieve these goals.

Efficacy and tolerability of interferon-free antiviral therapy in kidney transplant recipients with chronic hepatitis C

BACKGROUND & AIMS The development of direct-acting antiviral agents (DAAs) is a major step forward in the treatment of hepatitis C (HCV). The aims of the study were to evaluate the efficacy and tolerability of DAAs in kidney transplant (KT) recipients. METHODS Hepa-C is a Spanish registry of patients treated with DAAs in which clinical, virological and analytical data were prospectively included. We report on the data from 103 KT recipients who received DAAs. RESULTS The most commonly used DAAs combinations were sofosbuvir/ledipasvir (n=59, 57%) and sofosbuvir+daclatasvir (n=18, 17%). Ribavirin was used in 41% of patients. Sustained viral response after 12weeks (SVR12) rate was 98%. Grade 2 or 3 anemia appeared in 14 (33%) of patients receiving ribavirin and in 9 (15%) without (p=0.03). There were three episodes of acute humoral graft rejection. No patient discontinued therapy due to adverse events. Importantly, 57 (55%) patients required immunosuppression dose adjustment. Overall, there were no statistically significant differences in the mean level of serum creatinine, eGFR and proteinuria before and after treatment. Nonetheless, seventeen (16%) patients experienced renal dysfunction (increase in serum creatinine >25%) during antiviral therapy, of whom 65% were cirrhotic in comparison with only 29% cirrhotic patients who did not develop significant renal dysfunction (p=0.004). CONCLUSIONS Antiviral therapy with DAAs was highly efficacious and safe in KT recipients. Nevertheless, a non-negligible number of patients, most of them cirrhotic, developed mild allograft dysfunction and a significant proportion of patients required immunosuppression dose adjustment, warranting a close follow-up during therapy. LAY SUMMARY Infection by hepatitis C virus is often found in kidney transplant patients and its presence increases mortality and graft failure. We investigated the efficacy and safety of the new direct-acting hepatitis C antivirals in this population, in which previous information is scarce. Our data shows that, as occurs in the non-transplant setting, new anti-HCV antivirals are highly efficacious kidney transplant patients. Overall, this therapy is also quite safe, although worsening of renal function is observed in 16% of patients warranting a close follow-up observation of graft function during antiviral therapy.

Liver Stiffness 1 Year After Transplantation Predicts Clinical Outcomes in Patients With Recurrent Hepatitis C

The value of transient elastography (TE) to assess clinical outcomes in hepatitis C recurrence after liver transplantation (LT) has not been explored so far. We studied 144 hepatitis C–infected and 48 non–hepatitis C virus (HCV)‐infected LT recipients and evaluated the prognostic value of TE 1 year after transplantation to predict clinical decompensations and graft and patient survival. In HCV patients, cumulative probabilities of liver decompensation 5 years after LT were 8% for patients with liver stiffness measurement (LSM) <8.7 kilopascals (kPa) versus 47% for patients with LSM ≥ 8.7 kPa (p < 0.001). Five‐year graft and patient cumulative survival were 90% and 92% in patients with LSM < 8.7 kPa (p < 0.001) and 63% and 64% in patients with LSM ≥ 8.7 kPa, respectively (p < 0.001). Patients with low LSM 1 year after LT had excellent outcomes independently from receiving antiviral treatment or achieving sustained virological response (SVR). In contrast, graft survival significantly improved in patients with LSM ≥ 8.7 kPa who achieved SVR. No association between outcomes and LSM at 12 months was observed in non‐HCV patients. In conclusion, LSM 1 year after LT is a valuable tool to predict hepatitis C‐related outcomes in recurrent hepatitis C and can be used in clinical practice to identify the best candidates for antiviral therapy.

Virologic, Clinical, and Immune Response Outcomes of Patients With Hepatitis C Virus–Associated Cryoglobulinemia Treated With Direct-Acting Antivirals

BACKGROUND & AIMS Cryoglobulins (circulating immune complexes of polyclonal IgG, monoclonal IgM, and rheumatoid factor) are detected in the circulation of 40% to 60% of patients with chronic hepatitis C virus infection, and cryoglobulinemic vasculitis (CV) is observed in approximately 10% of patients. We aimed to assess the clinical and immune effects of direct‐acting antiviral treatment. METHODS We performed a prospective study of 64 patients with HCV infection with circulating cryoglobulins receiving direct‐acting antiviral therapy at a single center in Barcelona, Spain, from January 2014 through April 2016. Patients were classified as having CV (n = 35) or asymptomatic circulating cryoglobulins (ACC, n = 29). Clinical response was considered complete if a patient’s Birmingham Vasculitis Activity Score (version 3) was 0, or if all affected organs improved 12 weeks after the end of therapy. A complete immunologic response (CIR) was defined as no detection of circulating cryoglobulins and normalized levels of complement and/or rheumatoid factor. RESULTS Clinical manifestations of CV included purpura (65%), weakness (70%), arthralgia (31%), myalgia (20%), peripheral neuropathy (50%), and renal involvement (20%). At baseline, patients with CV had significantly higher levels of rheumatoid factor and lower levels of C4 complement than patients with ACC, whereas cryocrits were similar between groups (3.2% vs 2.6%). Overall, 60 patients (94%) had a sustained viral response 12 weeks after therapy. Among patients with CV, the median Birmingham Vasculitis Activity Score (version 3) decreased from 9 (range, 2–31) to 3 (range, 0–12) (P < .001). Twenty‐five patients with CV (71%) achieved a complete clinical response. Immune‐suppressive therapy was reduced for 4 of 13 patients and withdrawn for 6 of 13. Overall, 48% of patients achieved a CIR. A low baseline cryocrit level (<2.7%) was the only factor associated with CIR (odds ratio, 9.8; 95% confidence interval, 2.2–44; P = .03). CONCLUSIONS Viral eradication was associated with clinical improvement in most patients with CV. Markers of immune activation, including circulating cryoglobulins, persisted in 52% of patients with CV or ACC, despite a sustained viral response 12 weeks after therapy. A longer follow‐up period after viral eradication might be necessary to ensure a normal immune response.