Evaluation of Graft Fibrosis, Inflammation, and Donor-specific Antibodies at Protocol Liver Biopsies in Pediatric Liver Transplant Patients: A Single-center Experience.

BACKGROUND: The impact of graft fibrosis and inflammation on the natural history of pediatric liver transplants is still debated. Our objectives were to evaluate the evolution of posttransplant fibrosis and inflammation over time at protocol liver biopsies (PLBs), risk factors for fibrosis, presence of donor-specific antibodies (DSAs), and/or their correlation with graft and recipient factors. METHODS: A single-center, retrospective (2000-2019) cross-sectional study on pediatric liver transplant recipients who had at least 1 PLB, followed by a longitudinal evaluation in those who had at least 2 PLBs, was conducted. Fibrosis was assessed by the Liver Allograft Fibrosis Semiquantitative score, inflammation by the rejection activity index, DSAs by Luminex. RESULTS: A total of 134 PLBs from 94 patients were included. Fibrosis was detected in 87% (30% mild, 45% moderate, and 12% severe), 80% in the portal tracts. There was an increase in fibrosis between the 1-3 and the 4-6 y group (P = 0.01), then it was stable. Inflammation was observed in 44% (30% mild, 13% moderate, and 1% severe), 90% in the portal tracts. Anti-HLA II (IgG) DSAs were detected in 14 of 40 (35%). Portal fibrosis was associated with portal inflammation in the 1-3 y group (P = 0.04). Low immunosuppression levels were correlated with sinusoidal fibrosis (P = 0.04) and DSA positivity (P = 0.006). There was no statistically significant correlation between DSA positivity and the presence of graft fibrosis or inflammation. CONCLUSIONS: This study corroborates the concept of an early evolution of silent graft fibrosis. Suboptimal immunosuppression may play a role in the development of fibrosis and DSAs.

Combined liver kidney transplantation for primary hyperoxaluria type 1: Will there still be a future? Current transplantation strategies and monocentric experience.

Combined liver-kidney transplantation is a therapeutic option for children affected by type 1 primary hyperoxaluria. Persistently high plasma oxalate levels may lead to kidney graft failure. It is debated whether pre-emptive liver transplantation, followed by kidney transplantation, might be a better strategy to reduce kidney graft loss. Our experience of 6 pediatric combined liver-kidney transplants for primary hyperoxaluria type 1 in pediatric recipients was retrospectively analyzed. Plasma oxalate levels were monitored before and after transplantation. All the recipients were on hemodialysis at transplantation. Median [IQR] recipient's age at transplantation was 11 [1-14] years; in all cases, a compatible graft from a pediatric brain-dead donor aged 8 [2-16] years was used. In a median follow-up of 7 [2-19] years after combined liver-kidney transplantation, no child died and no liver graft failure was observed; three kidney grafts were lost, due to chronic rejection, primary non-function, and early renal oxalate accumulation. Liver and kidney graft survival remained stable at 1, 3, and 5 years, at 100% and 85%, respectively. Kidney graft loss was the major complication in our series. Risk is higher with very young, low-weight donors. The impact of treatment with glyoxalate pathway enzyme inhibitors treatment in children with advanced disease as well as of donor kidney preservation by ex vivo machine perfusion needs to be evaluated. At present, a case-by-case discussion is needed to establish an optimal treatment strategy.

Early impact of donor CYP3A5 genotype and Graft-to-Recipient Weight Ratio on tacrolimus pharmacokinetics in pediatric liver transplant patients.

Tacrolimus (TAC) pharmacokinetics is influenced by the donor CYP3A5 genotype and the age of pediatric liver recipients. However, an optimization of a genotype-based algorithm for determining TAC starting is needed to earlier achieve stable target levels. As the graft itself is responsible for its metabolism, the Graft-to-Recipient Weight Ratio (GRWR) might play a role in TAC dose requirements. A single-center study was carried out in a cohort of 49 pediatric recipients to analyse the impact of patient and graft characteristics on TAC pharmacokinetics during the first 15 post-transplant days. Children < 2 years received grafts with a significantly higher GRWR (4.2%) than children between 2-8 (2.6%) and over 8 (2.7%). TAC concentration/weight-adjusted dose ratio was significantly lower in recipients from CYP3A5*1/*3 donors or with extra-large (GRWR > 5%) or large (GRWR 3-5%) grafts. The donor CYP3A5 genotype and GRWR were the only significant predictors of the TAC weight adjusted doses. Patients with a GRWR > 4% had a higher risk of acute rejection, observed in 20/49 (41%) patients. In conclusion, TAC starting dose could be guided according to the donor CYP3A5 genotype and GRWR, allowing for a quicker achievement of target concentrations and eventually reducing the risk of rejection.

Management of Hepatitis-B Virus Infection in Immunocompromised Children: A Single Center Experience.

OBJECTIVES: The aims of the study was to expand the pediatric experience on hepatitis-B virus (HBV) reactivation, a known complication in patients with hematologic malignancies or on immunosuppression. METHODS: Retrospective appraisal of HBV therapy/prophylaxis in immunocompromised children, studied from April 2006 to March 2020. RESULTS: Eighteen HBV-positive patients, 5 girls, median age 11.1 (4.1--17.9) years were included. Seventeen of 18 were immunosuppressed at HBV-infection diagnosis. Seventeen were at high risk of reactivation, 1 at moderate risk. Five of 18 had acute hepatitis B as first infection or reactivation, 6 had HBeAg-positive infection, 1 an HBeAg-negative infection and 6 HBsAg-negative infection. Median follow-up was 2.7 (0.7--12.5) years. No HBV-related mortality was observed. Prophylaxis had to be repeated in 1. Lamivudine was used in 6/12 viremic patients and HBV-DNA negativization obtained in 2/6 (33%). Tenofovir-DF was used in 2/12 and entecavir in 4/12: 100% attained HBV-DNA negativization. Therapy had to be switched from tenofovir-DF to entecavir in 1 patient because of renal impairment. Virological breakthroughs were observed in 1 lamivudine-treated patient, leading to a hepatitis flare; 1 patient on entecavir had a hepatitis flare at immunoreconstitution. Mortality was 33% in the HBsAg-positive group. Seven prophylactic treatments were administered to 6 patients with HBsAg-negative infection: tenofovir-DF in 2 HBV-DNA-positive, lamivudine in 5 HBV-DNA-negative, without reverse HBsAg seroconversion, morbidity or mortality. CONCLUSIONS: There is a residual risk of acute hepatitis B in immunocompromised children, mortality rate was substantial, potentially related to the delays in commencing chemotherapy caused by liver dysfunction. Tenofovir-DF or entecavir are the drugs of choice for HBV treatment in immunocompromised children.

Carrying on with liver transplantation during the COVID-19 emergency: Report from piedmont region.

BACKGROUND: The COVID-19 pandemic is an emergency worldwide. In Italy, liver transplant activity was carried on, but despite all efforts, a 25% reduction of procured organs has already been observed during the first 4 weeks of the outbreak. AIMS: To analyze if our strategy and organization of LT pathway during the first two months of the COVID-19 emergency succeeded in keeping a high level of LT activity, comparing the number of LT in the first two months with the same period of time in 2019. METHODS: We compared the liver transplants performed in our Center between February 24th and April 17th, 2020 with liver transplants performed in the same period in 2019. RESULTS: In 2020, 21 patients underwent liver transplantation from deceased donors, exactly as the year before, without statistically significant difference. All patients survived in both groups, and the rate of early graft dysfunction was 24% in 2020 and 33% in 2019. In 2020 Median MELD was higher (17 vs 13). We were able to perform 3 multiorgan transplants and one acute liver failure. Nobody died on waiting list. The performance of our Center, despite the maxi-emergency situation, was steady and this was the result of a tremendous team working within the hospital and in our region. CONCLUSIONS: Team working allowed our Center to maintain its activity level, taking care of patients before and after liver transplantation. Sharing our experience, we hope to be helpful to other Centers that are facing the pandemic and, if another pandemic comes, to be more prepared to deal with it.

Transient elastography for non-invasive evaluation of post-transplant liver graft fibrosis in children.

As graft survival in pediatric LT is often affected by progressive fibrosis, numerous centers carry out protocol liver biopsies. Follow-up biopsy protocols differ from center to center, but all biopsies are progressively spaced out, as time from transplant increases. Therefore, there is a need for non-invasive techniques to evaluate graft fibrosis progression in those children who have no clinical or serological signs of liver damage. Indirect markers, such as the APRI, should be relied on with caution because their sensitivity in predicting fibrosis can be strongly influenced by the etiology of liver disease, severity of fibrosis, and patient age. A valid alternative could be TE, a non-invasive technique already validated in adults, which estimates the stiffness of the cylindrical volume of liver tissue, 100-fold the size of a standard needle biopsy sample. The aims of this study were to evaluate the reliability of TE in children after LT and to compare both the TE and the APRI index results with the histological scores of fibrosis on liver biopsies. A total of 36 pediatric LT recipients were studied. All patients underwent both TE and biopsy within a year (median interval -0.012 months) at an interval from LT of 0.36 to 19.47 years (median 3.02 years). Fibrosis was assessed on the biopsy specimens at histology and staged according to METAVIR. There was a statistically significant correlation between TE stiffness values and METAVIR scores (P = .005). The diagnostic accuracy of TE for the diagnosis of significant fibrosis (F ≥ 2) was measured as the area under the curve (AUROC = 0.865), and it demonstrated that the method had a good diagnostic performance. APRI was not so accurate in assessing graft fibrosis when compared to METAVIR (AUROC = 0.592). A liver stiffness cutoff value of 5.6 kPa at TE was identified as the best predictor for a significant graft fibrosis (METAVIR F ≥ 2) on liver biopsy, with a 75% sensitivity, a 95.8% specificity, a 90% positive predictive value, and an 88.5% negative predictive value. These data suggest that TE may represent a non-invasive, reliable tool for the assessment of graft fibrosis in the follow-up of LT children, alerting the clinicians to the indication for a liver biopsy, with the aim of reducing the number of protocol liver biopsies.

Successful Sequential Liver and Hematopoietic Stem Cell Transplantation in a Child With CD40 Ligand Deficiency and Cryptosporidium-Induced Liver Cirrhosis.

BACKGROUND: Hematopoietic stem cell transplantation (HSCT) is curative in patients with primary immunodeficiencies. However, pre-HSCT conditioning entails unacceptably high risks if the liver is compromised. The presence of a recurrent opportunistic infection affecting the biliary tree and determining liver cirrhosis with portal hypertension posed particular decisional difficulties in a 7-year-old child with X-linked CD40-ligand deficiency. We aim at adding to the scanty experience available on such rare cases, as successful management with sequential liver transplantation (LT) and HSCT has been reported in detail only in 1 young adult to date. METHODS: A closely sequential strategy, with a surgical complication-free LT, followed by reduced-intensity conditioning, allowed HSCT to be performed only one month after LT, preventing Cryptosporidium parvum recolonization of the liver graft. RESULTS: Combined sequential LT and HSCT resolved the cirrhotic evolution and corrected the immunodeficiency so that the infection responsible for the progressive sclerosing cholangitis did not recur. CONCLUSIONS: Hopefully, this report of the successful resolution of a potentially fatal combination of immunodeficiency and chronic opportunistic infection with end-stage organ damage in a child will encourage others to adapt a sequential transplant approach to this highly complex pathology. However, caution is to be exercised to carefully balance the risks intrinsic to transplant surgery and immunosuppression in primary immunodeficiencies.

Negativization of viremia prior to liver transplant reduces early allograft dysfunction in hepatitis C-positive recipients.

Although early allograft dysfunction (EAD) negatively impacts survival from the first months following liver transplantation (LT), direct-acting antiviral agents (DAAs) have revolutionized hepatitis C virus (HCV) therapy. We investigated the EAD definition best predicting 90-day graft loss and identified EAD risk factors in HCV-positive recipients. From November 2002 to June 2016, 603 HCV-positive patients (hepatocellular carcinoma, 53.4%) underwent a first LT with HCV-negative donors. The median recipient Model for End-Stage Liver Disease (MELD) score was 15, and the median donor age was 63 years. At LT, 77 (12.8%) patients were HCV RNA negative; negativization was achieved and maintained by pre-LT antiviral therapy (61 patients) or pre-LT plus a pre-emptive post-LT course (16 patients); 60 (77.9%) patients received DAAs and 17 (22.1%) interferon. We compared 3 different EAD definitions: (1) bilirubin ≥ 10 mg/dL or international normalized ratio ≥ 1.6 on day 7 after LT or aspartate aminotransferase or alanine aminotransferase > 2000 IU/L within 7 days of LT; (2) bilirubin > 10 mg/dL on days 2-7 after LT; and (3) MELD ≥ 19 on day 5 after LT. EAD defined by MELD ≥ 19 on day 5 after LT had the lowest negative (0.1) and the highest positive (1.9) likelihood ratio to predict 90-day graft loss. At 90 days after LT, 9.2% of recipients with EAD lost their graft as opposed to 0.7% of those without EAD (P < 0.001). At multivariate analysis, considering variables available at LT, MELD at LT of >25 (OR = 7.4) or 15-25 (OR = 3.2), graft macrovesicular steatosis ≥ 30% (OR = 6.7), HCV RNA positive at LT (OR = 2.7), donor age > 70 years (OR = 2.0), earlier LT era (OR = 1.8), and cold ischemia time ≥ 8 hours (OR = 1.8) were significant risk factors for EAD. In conclusion, in HCV-positive patients, MELD ≥ 19 on day 5 after LT best predicts 90-day graft loss. Preventing graft infection by pre-/peri-LT antiviral therapy reduces EAD incidence and could be most beneficial in high-MELD patients and recipients of suboptimal grafts. Liver Transplantation 23 915-924 2017 AASLD.

Impact of viral eradication with sofosbuvir-based therapy on the outcome of post-transplant hepatitis C with severe fibrosis.

BACKGROUND & AIMS: Several studies have shown that new direct-acting antivirals maintain their efficacy in liver transplant (LT) recipients with severe hepatitis C virus (HCV) recurrence. We determined the clinical impact of sofosbuvir/ribavirin in LT through the changes in liver function and fibrosis state at 24 and 48 weeks after treatment. METHODS: Between June 2014 and July 2015, 126 patients (30 F3, 96 F4 Metavir stage) were enrolled to receive sofosbuvir + ribavirin (24 weeks, 118 patients) or sofosbuvir + simeprevir + ribavirin (12 weeks, 8 patients); treatment was initiated at a median time of 4.3 years from LT. Median follow-up after therapy completion was 461 days. RESULTS: All 30 F3 patients achieved a sustained virological response at week 24 after treatment (SVR24) and showed a distinct amelioration of the AST-to-platelet ratio index (APRI), FIB-4 and liver stiffness at elastography by week 24 post-therapy, which were maintained at week 48. Of the 96 F4 cirrhotic patients, 72 (75%) achieved SVR24 accompanied by significant improvement of liver function, which was maintained at week 48 (Child B-C 22% baseline, 11% week 24, 7% week 48); APRI, FIB-4 and liver stiffness further improved significantly between weeks 24 and 48 of follow-up. Among the 77 responders (27 F3, 50 F4) who underwent elastography at baseline and at the end of follow-up, 39 (50.6%; 18 F3, 21 F4) exhibited a regression in fibrosis stage. CONCLUSION: At about 1 year from the completion of successful sofosbuvir-based therapy, patients with post-LT HCV and severe fibrosis experienced a long-term liver function improvement accompanied by a regression of fibrosis stage in half of them.

Early reduced liver graft survival in hepatitis C recipients identified by two combined genetic markers.

HLA and IL-28B genes were independently associated with severity of HCV-related liver disease. We investigated the effects of these combined genetic factors on post-transplant survival in HCV-infected recipients, aiming to provide new data to define the optimal timing of novel antiviral therapies in the transplant setting. HLA-A/B/DRB1 alleles and IL-28B rs12979860 (C > T) polymorphism frequencies were determined in 449 HCV viremic recipients and in their donors. Median follow-up was 10 years; study outcome was graft survival. HLA-DRB1*11 phenotype and IL-28B C/C genotype were significantly less frequent in recipients than donors (27.8% vs. 45.9% and 27.4% vs. 44.9%, respectively, P < 0.00001). Ten-year graft survival was better in patients with HLA-DRB1*11 (P = 0.0183) or IL-28B C/C (P = 0.0436). Conversely, concomitant absence of HLA-DRB1*11 and IL-28B C/C in 228 (50.8%) predicted worse survival (P = 0.0006), which was already evident at the first post-transplant year (P = 0.0370). In multivariable Cox analysis, absence of both markers ranked second as risk factor for survival (HR = 1.74), following donor age ≥ 70 years (HR = 1.77). In the current era of direct-acting antiviral agents, the negative effects of this common immunogenetic profile in HCV-infected recipients could be most effectively neutralized by peri-transplant treatment. This should be particularly relevant in countries where elderly donors represent an unavoidable resource.

Early Liver Transplantation for Neonatal-Onset Methylmalonic Acidemia.

With conventional dietary treatment, the clinical course of methylmalonic acidemia due to cobalamin-unresponsive methylmalonyl-CoA mutase (MCM) deficiency is characterized by the persistent risk of recurrent life-threatening decompensation episodes with metabolic acidosis, hyperammonemia, and coma. Liver transplant has been proposed as an alternative treatment and anecdotally attempted in the last 2 decades with inconsistent results. Most criticisms of this approach have been directed at the continuing risk of neurologic and renal damage after transplant. Here, we report the perioperative and postoperative clinical and biochemical outcomes of 2 patients with severe MCM deficiency who underwent early liver transplant. In both cases, liver transplant allowed prevention of decompensation episodes, normalization of dietary protein intake, and a marked improvement of quality of life. No serious complications have been observed at 12 years' and 2 years' follow-up, respectively, except for mild kidney function impairment in the older patient. On the basis of our experience, we strongly suggest that liver transplant should be offered as a therapeutic option for children with cobalamin-unresponsive MCM deficiency at an early stage of the disease.

Analysis of viral testing in nonacetaminophen pediatric acute liver failure.

OBJECTIVE: Viral infections are often suspected to cause pediatric acute liver failure (PALF), but large-scale studies have not been performed. We analyzed the results of viral testing among nonacetaminophen PALF study participants. METHODS: Participants were enrolled in the PALF registry. Diagnostic evaluation and final diagnosis were determined by the site investigator and methods for viral testing by local standard of care. Viruses were classified as either causative viruses (CVs) or associated viruses (AVs). Supplemental testing for CV was performed if not done clinically and serum was available. Final diagnoses included "viral," "indeterminate," and "other." RESULTS: Of 860 participants, 820 had at least 1 test result for a CV or AV. A positive viral test was found in 166/820 (20.2%) participants and distributed among "viral" (66/80 [82.5%]), "indeterminate" (52/420 [12.4%]), and "other" (48/320 [15.0%]) diagnoses. CVs accounted for 81/166 (48.8%) positive tests. Herpes simplex virus (HSV) was positive in 39/335 (11.6%) who were tested 26/103 (25.2%) and 13/232 (5.6%) among infants 0 to 6 and >6 months, respectively. HSV was not tested in 61.0% and 53% of the overall cohort and those 0 to 6 months, respectively. Supplemental testing yielded 17 positive, including 5 HSV. CONCLUSIONS: Viral testing in PALF occurs frequently but is often incomplete. The evidence for acute viral infection was found in 20.2% of those tested for viruses. HSV is an important viral cause for PALF in all age groups. The etiopathogenic role of CV and AV in PALF requires further investigation.

Immunosuppressants: what's new?

PURPOSE OF REVIEW: Advances in surgical techniques and combinations of conventional immunosuppressants have made paediatric liver transplantation the success story it is today. However, the increasing numbers of survivors reaching adulthood highlight important issues of long-term quality of life and drug induced complications. The aim of this review is to describe the trends and advances in immunosuppression for paediatric liver transplantation over the last 12 months. RECENT DEVELOPMENTS: As our knowledge of the immune cell populations and intracellular mechanisms involved in alloreactivity improves, induction immunosuppression has emerged as a powerful therapeutic manoeuvre to counter the initial alloimmune response. Many centres have adopted a more focused use of biological agents at induction to improve immunosuppression in the critical peritransplant period and to reduce the level of subsequent maintenance requirements. Corticosteroid avoidance and calcineurin inhibitors minimization trials have obtained encouraging results. New immunosuppressive strategies have evolved towards the goal of inducing operational tolerance, and paediatric liver transplant recipients seem to be a particularly promising target. New strategies are being developed also to improve quality of life and reduce nonadherence in adolescents and young adults who underwent liver transplantation. New drugs target B-cell and complement driven rejection and new monoclonal antibodies and small molecules are under trial to inhibit specific signals in the immune response. SUMMARY: We review current trends of immunosuppressive protocols in paediatric liver transplantation, focusing on induction agents, corticosteroid avoidance and calcineurin inhibitor sparing protocols, protocols for adult transition and new drugs currently under development.

Pattern of diagnostic evaluation for the causes of pediatric acute liver failure: an opportunity for quality improvement.

OBJECTIVE: To describe the frequency of diagnostic testing for the 4 most common causes of pediatric acute liver failure (PALF) (drugs, metabolic disease, autoimmune process, and infections) in indeterminate PALF within the PALF Study Group Database. STUDY DESIGN: PALF was defined by severe hepatic dysfunction within 8 weeks of onset of illness, with no known underlying chronic liver disease in patients from birth through 17 years of age. RESULTS: Of the 703 patients in the database, 329 (47%) had indeterminate PALF. In this group, a drug history was obtained in 325 (99%) urine toxicology screenings performed in 118 (36%) and acetaminophen level measured in 124 (38%) patients. No testing for common metabolic diseases was done in 179 (54%) patients. Anti-nuclear antibody, anti-smooth muscle antibody, and anti-liver kidney microsomal autoantibodies associated with autoimmunity were determined in 239 (73%), 233 (71%), and 208 (63%) patients, and no tests were obtained in 70 (21%). Testing was performed for hepatitis A virus, hepatitis B virus, and Epstein Barr virus in 80%, 86%, and 68%, respectively. CONCLUSIONS: Current practice indicates that investigation for metabolic and autoimmune causes of PALF are infrequent in patients ultimately given a diagnosis of indeterminate acute liver failure. This offers an opportunity to improve diagnosis and potential treatment options in children with acute liver failure.

Calcineurin inhibitor minimization in pediatric liver allograft recipients.

The use of CNI in pediatric LTx has dramatically improved the outcome for children with end-stage liver disease by significantly reducing the rate of acute and chronic rejection. Long-term concerns about CNI-induced nephrotoxicity and other adverse effects remain an issue, particularly as the emphasis moves from short-term survival to long-term quality of life. This review summarizes lessons learnt from pediatric and adult solid organ transplantation in minimizing CNI use in immunosuppression protocols in children following LTx. There are three models for CNI minimization: dose reduction, withdrawal or avoidance, supplemented by the use of IL-2 receptor blocking antibodies in the peri-transplant period, and early transition to alternate drugs such as MMF or SRL. Prospective studies evaluating reduction or withdrawal protocols in adult and pediatric LTx indicate that rejection rates are comparable with traditional CNI-based immunosuppression and that two and five yr patient and graft survival are similar, with recovery in renal function. There are few studies evaluating complete avoidance of CNI, apart from that in renal transplantation, although the benefits of long-term reduction in cardiovascular, metabolic, and possibly neoplastic side effects may justify this approach. It is not clear yet how CNI minimization will affect the development of tolerance but experimental and preliminary clinical studies indicate that CNI and steroid avoidance or minimization in the peri-operative period may favor the development of long-term graft tolerance. In summary, CNI minimization may be safe and effective in the short term but large-scale pediatric randomized studies are required to evaluate the long-term efficacy of these regimes in the development of chronic rejection, PTLD, and graft tolerance.