Experience With Molecular Adsorbent Recirculating System Treatment in 20 Children Listed for High-Urgency Liver Transplantation

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1 LIVER TRANSPLANTATION 21: , 2015 ORIGINAL ARTICLE Experience With Molecular Adsorbent Recirculating System Treatment in 20 Children Listed for High-Urgency Liver Transplantation Willem S. Lexmond, 1 Carin M. L. Van Dael, 1 Rene Scheenstra, 2 Joanne F. Goorhuis, 3 Egbert Sieders, 4 Henkjan J. Verkade, 2 Patrick F. Van Rheenen, 2 and Martin K omhoff 1 1 Division of Pediatric Nephrology, Beatrix Children s Hospital, 2 Division of Pediatric Gastroenterology and Hepatology, Beatrix Children s Hospital, 3 Division of Pediatric Intensive Care, Beatrix Children s Hospital, and 4 Division of Hepatobiliary Surgery and Liver Transplantation, Department of Surgery, University Medical Center Groningen, Groningen, the Netherlands For more than 10 years, children at our national center for pediatric liver transplantation (LT) have been treated with Molecular Adsorbent Recirculating System (MARS) liver dialysis as a bridging therapy to high-urgency LT. Treatment was reserved for 20 patients with the highest degrees of hepatic encephalopathy (HE; median grade 5 3.5). Death from neurological sequelae was considered imminent for these patients, and this was further reflected in significantly higher international normalized ratios and ammonia levels and worse prognostic liver indices (Model for End-Stage Liver Disease/Pediatric End-Stage Liver Disease scores and liver injury units) in comparison with 32 wait-listed patients who did not receive MARS dialysis. MARS therapy was generally well tolerated, with a reduction in thrombocytes and hemorrhaging as the most common side effects. HE improvement was documented in 30% of the treated patients, but progression to grade IV encephalopathy occurred in 45% of the patients despite the treatment. Serum ammonia, bilirubin, bile acid, and creatinine levels significantly decreased during treatment. Eighty percent of MARS-treated patients survived to undergo LT, and their survival was equivalent to that of non MARStreated patients with severe liver failure (69%, P ). The heterogeneity between MARS-treated patients and non MARStreated patients in our cohort precluded a statistical evaluation of a benefit from MARS for patient survival. Our data demonstrate the safety of MARS even in the most severely ill patients awaiting LT, but strategies that promote the more rapid and widespread availability of high-quality donor organs remain of critical importance for improving patient survival in cases of severe acute liver failure. Liver Transpl 21: , VC 2015 AASLD. Received July 28, 2014; accepted October 13, Abbreviations: AI, acetaminophen intoxication; ACLF, acute-on-chronic liver failure; ALF, acute liver failure; DBLT, death before liver transplantation; ELS, extracorporeal liver support; GD, graft dysfunction; HE, hepatic encephalopathy; HepB, hepatitis B infection; HULT, high-urgency liver transplantation; IE, indeterminate etiology; INR, international normalized ratio; LRLT, living related liver transplantation; LT, liver transplantation; MARS, Molecular Adsorbent Recirculating System; MELD, Model for End- Stage Liver Disease; NS, not significant; PALF, pediatric acute liver failure; PELD, Pediatric End-Stage Liver Disease; PICU, pediatric intensive care unit; SD, standard deviation; SR, spontaneous recovery. Potential conflict of interest: Carin M. L. van Dael has received speaking fees from Gambro-Hospal, the manufacturer of the Molecular Adsorbent Recirculating System and filters. Gambro-Hospal was in no way involved in the treatment of patients or in the preparation of this report, nor did it provide Molecular Adsorbent Recirculating System equipment at a discounted rate. Data from this study were presented in September 2012 at the 7th International Conference on Pediatric Continuous Renal Replacement Therapy in Cincinnati, OH and in September 2013 at the 16th Congress of the International Pediatric Nephrology Association in Shanghai, China. Carin M. L. Van Dael is currently affiliated with the Department of Pediatrics, VieCuri Medical Center, Venlo, the Netherlands. Address reprint requests to Martin K omhoff, M.D., Ph.D., Division of Pediatric Nephrology, Beatrix Children s Hospital, University Medical Center Groningen, Hanzeplein 1, Post Box 30001, 9700 RB Groningen, the Netherlands. m.komhoff@bkk.umcg.nl DOI /lt View this article online at wileyonlinelibrary.com. LIVER TRANSPLANTATION.DOI /lt. Published on behalf of the American Association for the Study of Liver Diseases VC 2015 American Association for the Study of Liver Diseases.

2 370 LEXMOND ET AL. LIVER TRANSPLANTATION, March 2015 See Editorial on Page 277 Progressive acute liver failure (ALF) will lead to death within days without liver transplantation (LT). 1-3 Together with severe graft dysfunction (GD) after transplantation, 4 this condition qualifies for highpriority donor liver allocation. Despite the existence of emergency channels for receiving an organ, data from the United States, England, and the Eurotransplant region indicate that 15% to 22% of patients die while awaiting high-urgency liver transplantation (HULT). 5-7 Extracorporeal liver support (ELS) is designed to improve the survival of patients with severe liver failure by replacing the detoxification function of the liver and thereby creating a bridge to transplantation. 8 ELS modalities include the fractionated plasma separation, adsorption, and dialysis system (Prometheus), 9,10 single-pass albumin dialysis, 11 and Molecular Adsorbent Recirculating System (MARS) albumin dialysis, which was introduced by Stange and Mitzner in the 1990s In 1999, MARS was shown to be effective in the clearance of protein-bound and water-soluble toxins and to have a positive effect on hepatic encephalopathy (HE) in 13 adult patients with acute-on-chronic liver failure (ACLF). 15 Since then, MARS has been introduced and accepted as ELS in Europe and Asia. 16,17 MARS dialysis was shown to improve HE significantly in a randomized controlled study of adults with advanced liver cirrhosis. 18 A recent meta-analysis confirmed a beneficial effect of MARS therapy on the clearance of protein-bound toxins and HE. 19 This analysis, however, did not reveal a significant effect of MARS treatment on patient survival. Furthermore, a beneficial effect on survival could not be demonstrated in 2 recent randomized controlled trials in adult patients with ALF 20 and ACLF, 21 with preliminary data from the latter already being included in the meta-analysis of Vaid et al. 19 In contrast to the adult population, scientific data on the use of ELS in children are scarce. 22,23 Although MARS dialysis is considered the most extensively studied ELS modality in pediatric liver failure, 2 the current body of literature remains limited to case reports and small case series. Controlled pediatric trials that compare MARS dialysis with standard medical care or other ELS modalities have not been published. Because both the etiology of severe liver failure and the clinical sequelae that result from failing hepatic detoxification differ between children and adults, 24,25 conclusions from adult trials cannot be extrapolated to the pediatric population. It, therefore, remains unknown whether MARS therapy is associated with a beneficial effect on the survival of children listed for HULT. Since 2002, our Dutch national center for pediatric LT has applied MARS dialysis to the treatment of a subset of children with severe ALF as a bridging strategy to transplantation. The decision to initiate treatment is made on a case-to-case basis, with MARS dialysis generally being reserved for patients with the most severe clinical conditions. Here we report on our 10-year experience in treating patients awaiting HULT, with a particular focus on the subset of patients who underwent MARS dialysis during the course of their treatment. PATIENTS AND METHODS Transplantation Center and Treatment Setting Beatrix Children s Hospital is the quaternary care academic hospital for pediatric LT in the Netherlands. Patients who meet the criteria to be listed for HULT are treated by a multidisciplinary team of pediatric intensivists, hepatologists, nephrologists, neurologists, and transplantation surgeons. The Netherlands is a member of the Eurotransplant International Foundation for donor organ allocation. MARS Unit and Indication MARS treatment in children has been performed at our center since 2001 with an AK-200 dialysis machine (Gambro Lundia, Lund, Sweden). According to their body weight, the children were treated with either the conventional adult filter size of 2 m 2 (>20 kg) or the MiniMARS system with a filter size of 0.6 m 2 (<20 kg). Heparin was used only when the activated clotting time was shorter than 140 seconds. ALF in children can be defined according to Squires et al. 24 as (1) evidence of acute hepatic cell necrosis and (2) coagulopathy [international normalized ratio (INR) 2.0], regardless of the degree of HE. However, to qualify for HULT in the Eurotransplant region, patients require a diagnosis of ALF according to the King s College or Clichy criteria or must suffer from acute graft failure less than 15 days after transplantation. Therefore, only patients who met these conditions were included in this study. Patients This study was performed according to the guidelines of the medical ethics committee of University Medical Center Groningen. MARS treatment was not performed as part of a consented randomized research protocol, although consent was obtained from parents in individual cases before the initiation of dialysis. The costs of treatment were covered under Dutch medical insurance regulations as part of the pediatric intensive care costs associated with ALF. Charts of all patients who were listed for HULT for ALF or GD between 2002 and 2011 were reviewed for the following: date and time of the intensive care unit admission, hepatic diagnosis, laboratory tests, written consultations performed by the pediatric neurologist (including the degree of HE scored according to the West Haven criteria or its adapted classification for infants/children as described elsewhere 2,24,26 ), daily clinical assessments by the pediatric intensivists, individual dialysis reports, cerebral imaging reports, transfusion of blood products, and outcomes and complications of therapy. The time and duration of each treatment cycle were registered. Biochemical

3 LIVER TRANSPLANTATION, Vol. 21, No. 3, 2015 LEXMOND ET AL. 371 TABLE 1. Clinical and Biochemical Characteristics of Patients Listed for HULT MARS Non-MARS P Value Patients, n Age in years, mean 6 SD (range) (0-15) (0-15) 0.18 Sex: male, n (%) 14 (70) 17 (53) 0.26 Patients < 90 days old, n (%) 2 (10) 3 (9) >0.99 Etiology, n (%) ALF 17 (85) 18 (56) IE 9 12 Viral 1 2 Metabolic disease 4 2 Intoxication 2 2 Autoimmune 1 0 GD 3 (15) 14 (44) Primary nonfunction 1 6 Thrombotic complication 2 8 Patient characteristics at time of HULT listing, mean 6 SD (range) HE grade (0-3.5) (0-3) 0.14 NH 3 (lmol/l) (40-220) (3-215) 0.19 INR (1.5-10) (1.2-10) 0.55 PELD/MELD score (5-54) (0-56) 0.40 Liver injury unit score ( ) (73-672) 0.59 Peak values, mean 6 SD (range) HE grade (2-4) (0-4) <0.001 NH 3 (lmol/l) (84-330) (20-236) <0.001 INR (3.1-10) (1.3-10) 0.02 Total bilirubin (lmol/l) (63-851) (2-1300) 0.20 Creatinine (lmol/l) (14-391) (13-263) 0.54 PELD/MELD score (22-63) (7-59) 0.04 Liver injury unit score ( ) (87-771) parameters obtained 6 hours before 1 hour after the start of treatment were considered pretreatment values and those collected 2 hours before the end of treatment until 6 hours afterward were accepted as posttreatment values. Prognostic Indices Several predictive models have been proposed to estimate the risk of death in children with ALF from biochemical parameters, including the liver injury unit score 27 and the Model for End-Stage Liver Disease (MELD; 12 years old)/pediatric End-Stage Liver Disease (PELD; <12 years old) score. 28,29 For statistical purposes, MELD/PELD scores were allowed to exceed 40, even though this is the upper limit for use in clinical practice. Review of Literature A PubMed search was conducted to identify all publications reporting on MARS treatment in children. In addition, a manual search of cited publications was performed. Reports that described the outcomes of children together with adults were included in our analysis only when information specific to the outcomes of pediatric patients could be retrieved. Results of single case reports were excluded from the analysis to limit the risk of publication bias. Statistical Analysis A comparison of MARS-treated and non MARStreated cohorts was performed with Fisher s exact test for categorical data and with either the Student t test or the Mann-Whitney U test for continuous variables (depending on the distribution). Differences in biochemical parameters before and after treatment were assessed with the paired Student t test for normally distributed values and with the Wilcoxon signed rank test for nonnormally distributed values. The normality of distribution was assessed with the Shapiro-Wilk test. Statistical tests were performed with Prism 5.0 (GraphPad Software, La Jolla, CA) or Stata 12.1 (StataCorp, College Station, TX). Results were considered significant with a 2-sided P value < RESULTS Between 2002 and 2011, 52 children with ALF (n 5 35 or 67%) or GD (n 5 17 or 33%) were listed for HULT. The median age of the patients was 4.5 years, and their ages ranged from 1 week to 15.5 years. The median time between listing and HULT was 2 days, and all surviving patients had undergone

4 372 LEXMOND ET AL. LIVER TRANSPLANTATION, March 2015 TABLE 2. Clinical Characteristics of Patients Undergoing MARS Dialysis and Technical Treatment Details Pediatric Filter (0.6 m 2 ) Adult Filter (2 m 2 ) Total Clinical parameters Patients, n MARS runs, n Age in years, median (range) 2 (0-14) 12 (3-15) 5.8 (0-15) Weight in kg, median (range) 12.5 (6-40) 44 (23-70) 19 (6-70) Continuous venovenous hemofiltration/impaired 3/11 (27) 3/9 (33) 6/20 (30) renal function, n/n (%) Sedation, n/n (%) 10/16 (63) 3/17 (18) 13/33 (39) Assisted ventilation, n/n (%) 16/16 (100) 17/17 (100) 33/33 (100) Inotropy, n/n (%) 16/16 (100) 11/17 (65) 27/33 (82) Technical parameters, median (range) Runs per patient 1 (1-2) 2 (1-4) 1.5 (1-4) Treatment time (hours) 8 (4-26) 8 (4-8) 8 (4-26) Average blood flow (ml/minute) 78 (28-138) 150 (71-200) 84 (28-200) Heparin maintenance, n/n (%) 7/16 (44) 2/17 (12) 9/33 (27) transplantation within 8 days after registration except for 1 infant with gestational alloimmune liver disease, who ultimately underwent transplantation 22 days after HULT listing. Clinical and Biochemical Characteristics of Patients Undergoing MARS Dialysis Twenty patients (38%) were treated with MARS dialysis: 17 were suffering from ALF, and 3 were suffering from GD (Table 1). In every case, the decision to initiate MARS dialysis was made by a multidisciplinary team that included pediatric intensivists, hepatologists, nephrologists, neurologists, and transplant surgeons. Advanced HE (grade III-IV) with the inherent risk of cerebellar herniation formed the primary indication for starting MARS treatment. This was reflected in the significant difference between the peak HE grades of MARS-treated patients and untreated patients (mean grade: 3.4 versus 2.0, P < 0.001; Table 1). Peak biochemical parameters further confirmed more severe liver failure in patients who underwent MARS dialysis: MARS patients developed significantly higher peak levels of serum ammonia and higher INRs (Table 1). Importantly, at the time of HULT listing, the severity of liver failure as assessed by the HE grade and biochemical parameters did not yet predict which patients would ultimately develop high-grade HE that qualified for MARS therapy; this implied that the heterogeneity between MARS-treated and non MARStreated patients arose during the most advanced stage of the disease when the children had already been listed for HULT (Table 1). Peak levels of serum bilirubin and creatinine were also higher in MARS-treated patients, but these differences did not reach statistical significance. Because patients with GD were relatively underrepresented in the group of MARS-treated patients, we repeated the comparison in the subset of ALF patients. Both the degree of HE and the serum ammonia levels were significantly higher (P < and P , respectively) in MARS-treated patients versus non MARS-treated ALF patients, and this suggested that the greater disease severity in patients who underwent dialysis did not result from etiological differences between cohorts. We observed significantly worse prognostic index scores for the MARS-treated patients: the mean peak liver injury unit scores were 568 for the MARS-treated group and 420 for the non MARS-treated group (P ). Similarly, the mean peak MELD/PELD scores were higher for patients who underwent MARS therapy (47 versus 38, P ). Both of these prognostic scoring systems thus predicted a higher risk of mortality and need for LT in the patients undergoing MARS dialysis versus those receiving standard medical treatment only. In this respect, it is important to note that a subset of patients did not receive MARS treatment despite advancement to HE grade III-IV, either because of the rapid progression of disease or the timely availability of a donor organ. Regardless, these results show that MARS dialysis was applied to a distinct subset of pediatric acute liver failure (PALF) and GD patients characterized by clinical and biochemical signs of more advanced hepatic failure. Efficacy and Safety of MARS Treatment We performed a combined total of 33 MARS runs in 20 patients who were awaiting HULT (Table 2). Most patients received 1 or 2 cycles of MARS dialysis before liver (re)transplantation, although 1 patient underwent 4 runs. In the great majority of cases, MARS dialysis was applied for 8 consecutive hours and was repeated daily until transplantation, although 3 runs were successfully extended beyond 8 hours, and 4 runs were terminated before 6 hours had passed because of either clotting of the system or emergency cerebral imaging. All patients undergoing MARS

5 LIVER TRANSPLANTATION, Vol. 21, No. 3, 2015 LEXMOND ET AL. 373 therapy were mechanically ventilated, and 19 of 20 required vasopressors. Effect on HE A standardized neurological assessment by a pediatric neurologist right before and directly after MARS dialysis was not performed in the majority of children, and this hampered quantification of the effect of MARS on HE. Concurrent sedation (in 39% of runs) during mechanical ventilation often precluded formal scoring of the degree of HE or EMV scores. Regardless, improvement of encephalopathy (increase in the Glasgow Coma Scale, responsiveness, or pupillary reflexes) was documented in the charts of a total of 6 patients (30%) and occurred with the use of both adult (n 5 3) and pediatric filters (n 5 3) and in children suffering from ALF (n 5 5) or GD (n 5 1). In 5 other patients (25%), no change in the neurological condition could be objectified after treatment. In all, 9 children (45%) had progression of HE to grade IV despite MARS, and this was accompanied by clinical signs of cerebellar herniation. Ten children underwent emergency cerebral imaging during or after MARS treatment in the 24 hours before LT to rule out cerebellar herniation. Effect on Biochemical Parameters MARS dialysis with both adult and pediatric filters led to the removal of albumin-bound and water-soluble toxins (Fig. 1A). Absolute ammonia levels decreased significantly, with a median of 52 lmol/l (range lmol/l) for pediatric-sized filters and a median of 12 lmol/l (range 5 35 to 206 lmol/l) for adult-sized filters. Similarly, other levels decreased, with medians of 33 lmol/l (28 to 331 lmol/l) and 81 lmol/l (210 to 369 lmol/l) for total bilirubin, with medians of 19 lmol/l (213 to 106 lmol/l) and 30 lmol/l (8-211 lmol/l) for bile acids, and with medians of 9 lmol/l (23 to 120 lmol/l) and 14 lmol/l (22 to 220 lmol/ L) for creatinine with pediatric and adult filters, respectively. The median changes in lactate levels before and after treatment were 0.15 and 0.2 mmol/l, but they ranged from a decrease of 2.9 mmol/l to an increase of 2.7 mmol/l. Combined, these results show that both MARS and MiniMARS were capable of clearing albumin-bound and water-soluble toxins in patients suffering from liver failure. A subanalysis furthermore revealed that MARS dialysis decreased ammonia and bilirubin regardless of whether patients suffered from ALF of an indeterminate etiology (IE); from GD; or from ALF resulting from intoxication or metabolic or viral causes. To demonstrate the effect of MARS on the development of serum levels of bilirubin and ammonia, Fig. 1D depicts the complete set of values of these parameters as obtained in the course of treatment at the pediatric intensive care unit (PICU) with respect to the intervals of MARS dialysis. These graphs clearly show that MARS therapy affected the serum concentrations of bilirubin and ammonia in these 2 patients who underwent >2 MARS cycles while awaiting HULT. Side Effects of MARS Treatment MARS treatment led to a significant reduction in thrombocyte counts with a median decrease of /L (range 5 16 to /L; Fig. 1B,D). Overall, no significant differences were seen in INR before and after treatment, although both large increases and decreases were seen in a number of patients. Serious bleeding that required multiple transfusions of erythrocytes, thrombocytes, or freshfrozen plasma occurred in 5 patients treated with MARS (25%) and contributed directly to pretransplant mortality in 1 patient. However, coagulopathy and hemorrhaging are inherent to severe ALF, and 5 non MARS-treated patients awaiting HULT also suffered from significant bleeding, including 2 fatal events (P > 0.99 versus MARS-treated patients). Nevertheless, before undergoing HULT, MARS-treated patients had received significantly more thrombocyte transfusions ( versus , P ). In addition, there was a trend toward increased erythrocyte ( versus , P ) and fresh-frozen plasma transfusions ( versus , P ) in MARS-treated patients. On average, there was no change in the mean arterial blood pressure with the use of either adult or pediatric filters (Fig. 1B), although variations in respiratory and heart rates were typically observed during MARS therapy (Fig. 1C). These results demonstrate a safety profile of MARS treatment in children that is comparable to that in the adult population. 18,20,21 Outcomes of Patients Listed for HULT Spontaneous recovery (SR) was rare in both MARStreated (1 of 20) and untreated patients (5 of 32, P ). Overall, 38 HULT procedures were performed (73% of registrations). In the remaining 8 cases, death occurred before a donor liver became available (Table 3). Similar percentages of MARStreated and untreated children survived until HULT could be performed (80% versus 69%, P ). In 2 patients in both cohorts, death resulted from cerebellar herniation from HE. Another 4 children died from progressive cardiorespiratory failure, which was preceded by severe hemorrhaging in 3 patients. The 3- month survival rates were 65% for MARS-treated patients and 62% for non MARS-treated patients, and the timing of the availability of donor organs was not different between patients who underwent MARS and those who did not (Fig. 2). Almost all cases of death after transplantation were due to circulatory and multiorgan failure, with 1 additional death from hemorrhaging in a MARS-treated patient. Comparison With Published Case Series Our PubMed search strategy identified 9 MEDLINEindexed reports on the outcomes of MARS therapy in a series of pediatric patients (Table 4). Combined, these reports describe the outcomes of MARS therapy in a total of 44 children. 7,30-37 Almost all of these

6 374 LEXMOND ET AL. LIVER TRANSPLANTATION, March 2015 Figure 1. (A) Effects of MARS dialysis with either adult (2 m 2 ) or pediatric dialysis filters (0.6 m 2 ) on serum levels of water-soluble and albumin-bound toxins. Values obtained before and after dialysis are plotted and compared with the paired t test or Wilcoxon signed rank test. (B) Dialysis significantly reduced platelet counts but did not affect INR or mean arterial blood pressure. (C) Changes in hemodynamic parameters (systolic and diastolic blood pressures and heart rates) over the course of an 8-hour MARS cycle in 2 representative patients treated with either the pediatric or adult filter. (D) Complete set of laboratory data from the time of PICU admission to the time of HULT for serum bilirubin, ammonia, and thrombocytes in 2 patients who underwent multiple MARS cycles (denoted by gray areas). *P < 0.05; **P < 0.01; ***P < patients (92%) suffered from ALF or ACLF, and only 4 patients (8%) underwent MARS treatment for GD. Recovery of the native liver was reported for 10 children with ALF (21%), including 4 patients with mushroom intoxication and 3 with an acetaminophen overdose. LT was the most common outcome (45%).

7 LIVER TRANSPLANTATION, Vol. 21, No. 3, 2015 LEXMOND ET AL. 375 TABLE 3. Outcomes of Patients Listed to Undergo HULT MARS Non-MARS P Value Patients, n SR, n (%) 1 (5) 5 (16) 0.39 Death before HULT, n (%) 3 (15) 5 (16) >0.99 Sepsis 0 0 >0.99 Circulatory/multiorgan failure 0 1 >0.99 Hemorrhage 1 2 >0.99 Neurological/cerebral herniation Transplantation, n (%) 16 (80) 22 (69) 0.52 Time between listing and HULT (days) 2.5 (1-22) 2 (1-8) 0.73 Death within 3 months of HU listing, n (%) 7 (35) 12 (38) 0.77 Sepsis Circulatory/multiorgan failure Hemorrhage Neurological/cerebral herniation Figure 2. (A) Kaplan-Meier curves depicting 3-month survival from the time of wait listing for patients awaiting HULT who were either MARS-treated (solid curve) or not MARS-treated (dashed curve). (B) Time from listing to HULT for MARS-treated patients (solid curve) and non MARS-treated patients (dashed curve). Subjects were censored at the time of death before HULT or at the time of removal from the wait list in the case of SR. In all, 16 children (34%) died before transplantation could be performed. HE was the most common indication for MARS treatment, although a disparity existed with respect to the degree of severity, with some centers starting treatment at an HE grade of I. Four studies showed a significant decrease in serum bilirubin and ammonia, 30,31,34,37 but in the largest series of 10 children, this effect was not seen. 36 There were no reported major adverse effects of treatment in any of the studies, except for 1 case of hemorrhaging after vascular injury upon catheter insertion. Four reports described reductions in thrombocytes upon treatment. Prognostic liver indices (MELD/PELD) were reported for only 1 study. 32 DISCUSSION In the early 2000s, articles on the successful application of MARS dialysis in children started to accumulate in the medical literature These early reports focused on practicality and efficacy in single patients with different underlying morbidities and illustrated that MARS dialysis in children was technically feasible. Limited numbers of small case series have since become available on the efficacy of MARS treatment as a bridge to transplantation in children with ALF, and they have illustrated the potential of MARS dialysis in removing protein-bound toxins and improving neurological conditions in at least a subset of these patients. With our current study, we report the largest series thus far of pediatric patients undergoing MARS therapy while awaiting HULT. The decision to initiate MARS therapy was made on a case-to-case basis, and we started treatment only when clinical judgment indicated a profound risk of death from the sequelae of hyperammonemia despite maximally supportive medical treatment. As a result, dialysis was reserved for those patients who were in the worst clinical condition, which was reflected in higher degrees of HE,

8 376 LEXMOND ET AL. LIVER TRANSPLANTATION, March 2015 TABLE 4. Overview of Published Case Series on the Use of MARS in the Pediatric Population Sex/Age Reference Year Patients (Range) Disease Etiology MARS Indication Number of MARS Treatments Biochemical and Clinical Effect Reported Outcome Covic et al males and 4 females/7-16 years Da Costa males and 1 et al. 31 female/5-15 years Auth et al females/6-14 years PALF (mushroom poisoning) PALF (2 Wilson s disease, 1 HepB, 1 IE) PALF (Wilson s disease) Nadalin et al Not specified PALF (not specified) Wai et al (analyzed with 46 adults) Not specified PALF (1 IE, 1 Wilson s disease, 2 not specified) HE I (4), HE II (1), HE IV (1) HE III-IV (4) hours (4, average) HE III-IV, PELD scores of 23 and 24 INR > 3.0 and HE 2 or renal insufficiency or hepatorenal syndrome hours (6) Decrease in NH 3, bilirubin, and Child-Turcotte- Pugh score, neurological improvement (4) hours for MiniMARS (1), 6 hours for Mini- MARS 1 6 hours for MARS (1) Not specified for duration of 3 days (range days) (5) Decrease in bilirubin, no effect on INR MELD score of 23! 10 (1), MELD score of 24! 19 (1); improved Fisher ratio and lactate level; constantly high bilirubin (1); increasing NH 3 and copper levels (1) SR (4) or DBLT (2) LRLT (2) and DBLT (2 multiorgan failures) LT (2) with neurologic recovery Not specified Survival after LT (5) Not specified Not specified Not specified LRLT (2) and DBLT (2) Novelli et al females and 2 males/3-15 years PALF (3 IE, 2 AI, 1 HepB) HE > 2, INR > 2.5, bilirubin > 15 m- g/dl, Continuous, filter change every 8 hours (6) Significant decrease in bilirubin, NH 3, creatinine, and lactate; beneficial LT (3) with 2 survivals, SR (2), DBLT (1) Adverse Effects None; no hypotension or drop in thrombocytes None reported Slight drop in thrombocytes not requiring substitution None reported Not specified; decrease in hemoglobin and platelet count in total study population None reported

9 LIVER TRANSPLANTATION, Vol. 21, No. 3, 2015 LEXMOND ET AL. 377 TABLE 4. Continued Sex/Age Reference Year Patients (Range) Disease Etiology MARS Indication creatinine > 2 m- g/dl Rusu et al (analyzed with 20 adults) Not specified 4 GD, 3 ALF (1 AI, 1 HepB, 1 not specified) Bilirubin > 5 mg/ dl, HE 1 Schaefer (8 also et al. 36 received plasma exchange and hemodialysis) Not specified/ years (9.7 years) ALF 1 ACLF (not specified) Hyperammonemia (7) and hyperbilirubinemia (6); prolonged INR (6); HE II (1), HE III (1), HE IV (4) Rustom females and 1 et al. 37 male ALF (4 Wilson s disease) Wilson index- > 11 (range ) Total ALF, 4 GD, 10 ACLF Number of MARS Treatments Mean time of hours, though not specified for children MARS (7) 1 MiniMARS (3), 1-4 sessions per patient hours (1), hours (3) Biochemical and Clinical Effect effect on hemodynamic indices (mean arterial pressure, heart rate, systemic vascular resistance index), Glasgow coma scale, and intracranial pressure Not specified for children No significant decrease in bilirubin or NH3, inferior efficacy of MARS versus plasma exchange and hemodialysis HE 2! 1 (1) or remained stable (3); improved Fisher index; average decreases of 26% in bilirubin and 19% in NH3 and copper levels Reported Outcome GD: re-lt (1), DBLT (3) ALF: SR (2, AI 1 HepB), DBLT (1) SR (2), LT (3), DBLT (5) (multiorgan failure and sepsis, including all 3 MiniMARStreated children) Survival with LT (4) SR (10 or 21%), DBLT (16 or 33%), LT (22 or 46%) Adverse Effects Not specified for children Fresh frozen plasma (3) and thrombocyte transfusion (3); no hemorrhage Hemorrhage from catheter (1), average 42% drop in thrombocytes requiring transfusion

10 378 LEXMOND ET AL. LIVER TRANSPLANTATION, March 2015 higher levels of serum ammonia, higher INRs, higher MELD/PELD scores, and a liver injury unit score derived >35% higher risk of death without transplantation in comparison with non MARS-treated patients. Neurological improvement was documented in 30% of the patients, and statistically significant beneficial effects of therapy were observed for serum ammonia and bilirubin. Eighty percent of MARStreated patients were successfully bridged to HULT after a median of 2.5 days, with a 3-month patient survival rate of 65%. We have presented these data in conjunction with the clinical characteristics and outcomes of patients at our center who were awaiting HULT but did not undergo MARS therapy. This allowed a number of important observations. First, MARS therapy did not prevent transplantation in the patients who received it. SR of the native liver was exceedingly rare in our entire patient cohort, and only in a single case did a MARS-treated child survive without undergoing HULT. Because we reserved dialysis for only those patients for whom death from neurological sequelae was considered imminent, we cannot rule out that its wider application to patients with less severe degrees of liver failure could have resulted in increased rates of SR. Similarly, we cannot say whether prolongation of MARS dialysis would perhaps have prevented HULT in a subset of patients, although we consider this unlikely because all patients still met Eurotransplant s criteria for HULT at the time of transplantation despite MARS therapy. Second, MARS was generally well tolerated, and patients undergoing dialysis were not more likely to die from acute complications (most notably hemorrhaging) than their nontreated counterparts, although according to our results, it is likely that MARS contributed partially to increased transfusion requirements. Third, the survival of MARStreated patients was not worse than that of nontreated patients. Death before transplantation could be performed occurred in 15% of MARS-treated patients versus 16% of non MARS-treated patients. Likewise, 3-month survival rates were equivalent between treated and nontreated patient groups. Combined, our data therefore show that MARS dialysis as a bridging strategy to HULT was generally well tolerated in even the sickest ALF patients, and we have no indication that treatment has resulted in additional mortality in our patient cohort. The question of whether MARS therapy confers a survival benefit remains, however, unanswered. We emphasize that the differences between our MARStreated and non MARS-treated patients with respect to disease etiology, age, and clinical and biochemical parameters of disease severity preclude the inference of an effect of MARS on patient survival. Although it is tempting to assume that even modest improvements in the clinical and biochemical conditions of these critically ill patients awaiting transplantation are beneficial, a positive effect of MARS dialysis on the survival of ALF patients remains to be demonstrated for either pediatric or adult populations. Two large multicenter trials in adults that compared the survival of ALF patients and ACLF patients treated with either MARS or standard medical care did not detect a statistically significant survival benefit of MARS therapy. 20,21 In the study by Saliba et al., patients with ALF were randomized to receive standard medical therapy alone or in association with MARS. There was no statistically significant effect of MARS on 6- month survival, but the short period between randomization and transplantation led the authors to report the study as inconclusive. In the RELIEF trial, HE 2, the baseline MELD score, and an increase in serum bilirubin on day 4 were the only significant predictors of mortality in a multivariate logistic regression, and after adjustments for these confounders, no significant effect of MARS therapy on 28-day survival in ACLF could be observed (odds ratio , 95% confidence interval ). 21 In the absence of randomized pediatric trials, the results from these 2 studies in the adult population must be carefully weighed against all factors that distinguish children from adult patients awaiting HULT, such as differences in the etiology of liver disease, the pathophysiological responses to hyperammonemia, the comorbidity profiles, and the dialysis-filter-tobody-size ratio. 22,24,25,42 The exceedingly rare nature of ALF and GD in children renders the feasibility of prospective trials low. Our results indicate that on average only 5 or 6 children per year would be eligible in the Netherlands (with a population of 16.5 million); this means that a trial should ideally be internationally coordinated. Besides the challenge associated with low disease incidence, inclusion in such a study will likely be further hampered by the narrow time window in which informed consent has to be obtained after listing for HULT. 20 As an alternative, the collection of standardized data and the international registration of pediatric patients awaiting HULT could further help define the effects of MARS dialysis and other modalities of ELS 22,36 in as many patients as possible. On the basis of the data presented here, including our summary of previously published data on the use of MARS in pediatric patients, it is clear that such initiatives are critically important in answering remaining questions about which patients will benefit most from MARS dialysis; what the optimal timing, duration, and frequency of treatment should be; and how MARS therapy compares with alternative modalities such as plasmapheresis with hemodialysis. 36 Although our data suggest that MARS therapy is not harmful to pediatric patients who are awaiting HULT, the currently available clinical data are insufficient to assess whether its potential benefits justify the high costs involved. It is important to recognize that ALF survival is dependent not only on the pretransplant clinical condition of the recipient but also in large part on the timeliness of donor organ availability and the quality and age of the donor organ as well as intra- and postoperative factors. 6 Therefore, strategies that promote the more rapid and widespread

11 LIVER TRANSPLANTATION, Vol. 21, No. 3, 2015 LEXMOND ET AL. 379 availability of high-quality donor organs, including public awareness campaigns to raise active donor registration, promotion of living related donor programs, 43 international organ sharing networks, 44 and technical advances in artificial organ generation, could potentially have a greater impact on patient survival than optimizing ELS bridging techniques. Until the implementation of such strategies has led to a reduction in HULT waiting times, we will continue to administer MARS dialysis in the care of these critically ill pediatric patients. In conclusion, MARS albumin dialysis has been successfully applied to children listed for HULT at our national LT center for more than a decade and has resulted in significant reductions of albumin-bound toxins. MARS therapy was reserved for patients with the most severe clinical condition, who nevertheless had survival rates similar to those of non MARStreated children. 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