Kyle Soltys

Barriers to the successful treatment of liver disease by hepatocyte transplantation

Management of patients with hepatic failure and liver-based metabolic disorders is complex and expensive. Hepatic failure results in impaired coagulation, altered consciousness and cerebral function, a heightened risk of multiple organ system failure, and sepsis [1]. Such manifold problems are only treatable today and for the foreseeable future by transplantation. In fact, whole or auxiliary partial liver transplantation is often the only available treatment option for severe, even if transient, hepatic failure. Patients with life-threatening liver-based metabolic disorders similarly require organ transplantation even though their metabolic diseases are typically the result of a single enzyme deficiency, and the liver otherwise functions normally. For all of the benefits it may confer, liver transplantation is not an ideal therapy, even for severe hepatic failure. More than 17,000 patients currently await liver transplantation in the United States, a number that seriously underestimates the number of patients that need treatment [2], as it has been estimated that more than a million patients could benefit from transplantation [3]. Unfortunately, use of whole liver transplantation to treat these disorders is limited by a severe shortage of donors and by the risks to the recipient associated with major surgery [4].

Allospecific CD154+ T cells identify rejection-prone recipients after pediatric small-bowel transplantation.

BACKGROUNDnUp to 70% of children with small bowel transplantation (SBTx) experience acute cellular rejection (ACR). Allospecific CD154+ T cells predict liver ACR in children in a novel, 16-hour mixed leukocyte response (MLR) assay, but remain untested in SBTx.nnnMETHODSnThe expression of CD154 was measured in 4 subsets-naive (N) and memory (M) CD154+ T-helper (Th) and T-cytotoxic (Tc) cells (ie, CD154+ ThN, CD154+ ThM, CD154+ TcN, and CD154+ TcM, respectively)-in the MLR of single blood samples obtained from 32 children with SBTx within 60 days of SBTx biopsy. Children showing ACR in these biopsies were termed Rejectors. The ratio of donor-induced to third-party-induced CD154+ T cells was called the immunoreactivity index (IR). We hypothesized that IR >1 denoted increased donor-specific alloreactivity and increased risk of rejection; in contrast, IR <1 implied decreased risk. CD154 expression was correlated with the expression of CTLA4, a negative T-cell costimulator that antagonizes and is inversely related to CD154 (n = 18).nnnRESULTSnRejectors showed significantly greater numbers of donor-specific CD154+ T-cell subsets. Logistic regression analysis and leave-one-out cross validation followed by receiver operating characteristic analysis showed that, among the 4 subsets, IR > or =1.23 for CD154+ TcM identified Rejectors with a sensitivity and specificity of 93% and 88%. Also, a significant negative correlation was observed between CD154 expression and CTLA4 expression in allospecific Tc (Spearmans rho = -0.616, P = .006) but not in Th.nnnCONCLUSIONnAllospecific CD154+ TcM identify rejection-prone children with SBTx.

New potential cell source for hepatocyte transplantation: discarded livers from metabolic disease liver transplants.

UNLABELLEDnDomino liver transplantation is a method used to increase the number of liver grafts available for orthotopic liver transplantation (OLT). Reports indicate that livers from patients with metabolic liver disease can be safely transplanted into select recipients if the donors defect and the recipients metabolic needs are carefully considered. The liver of patients with many types of metabolic liver disease is morphologically and biochemically normal, except for the mutation that characterizes that disease. Other biochemical functions normally performed by the liver are present and presumably normal in these hepatocytes. Hepatocytes were isolated from the liver of 35 organ donors and 35 liver tissues taken at OLT from patients with liver disease were analyzed for 9 different measures of viability and function. The data indicate that cells isolated from some diseased livers performed as well or better than those isolated from organ donors with respect to viability, cell yield, plating efficiency and in assays of liver function, including drug metabolism, conjugation reactions and ammonia metabolism. Cells from metabolic diseased livers rapidly and efficiently repopulated a mouse liver upon transplantation.nnnCONCLUSIONSnAs with domino liver transplantation, domino cell transplantation deserves consideration as method to extend the pool of available organs and cells for transplantation.

Analysis of surgical interruption of the enterohepatic circulation as a treatment for pediatric cholestasis

To evaluate the efficacy of nontransplant surgery for pediatric cholestasis, 58 clinically diagnosed children, including 20 with Alagille syndrome (ALGS), 16 with familial intrahepatic cholestasis‐1 (FIC1), 18 with bile salt export pump (BSEP) disease, and 4 others with low γ‐glutamyl transpeptidase disease (levels <100 U/L), were identified across 14 Childhood Liver Disease Research Network (ChiLDReN) centers. Data were collected retrospectively from individuals who collectively had 39 partial external biliary diversions (PEBDs), 11 ileal exclusions (IEs), and seven gallbladder‐to‐colon (GBC) diversions. Serum total bilirubin decreased after PEBD in FIC1 (8.1 ± 4.0 vs. 2.9 ± 4.1 mg/dL, preoperatively vs. 12‐24 months postoperatively, respectively; P = 0.02), but not in ALGS or BSEP. Total serum cholesterol decreased after PEBD in ALGS patients (695 ± 465 vs. 457 ± 319 mg/dL, preoperatively vs. 12‐24 months postoperatively, respectively; P = 0.0001). Alanine aminotransferase levels increased in ALGS after PEBD (182 ± 70 vs. 260 ± 73 IU/L, preoperatively vs. 24 months; P = 0.03), but not in FIC1 or BSEP. ALGS, FIC1, and BSEP patients experienced less severely scored pruritus after PEBD (ALGS, 100% vs. 9% severe; FIC1, 64% vs. 10%; BSEP, 50% vs. 20%, preoperatively vs. >24 months postoperatively, respectively; P < 0.001). ALGS patients experienced a trend toward greater freedom from xanthomata after PEBD. There was a trend toward decreased pruritus in FIC1 after IE and GBC. Vitamin K supplementation increased in ALGS after PEBD (33% vs. 77%; P = 0.03). Overall, there were 15 major complications after surgery. Twelve patients (3 ALGS, 3 FIC1, and 6 BSEP) subsequently underwent liver transplantation. Conclusion: This was a multicenter analysis of nontransplant surgical approaches to intrahepatic cholestasis. Approaches vary, are well tolerated, and generally, although not uniformly, result in improvement of pruritus and cholestasis. (Hepatology 2017;65:1645‐1654).

Liver transplantation for treatment of severe S-adenosylhomocysteine hydrolase deficiency.

A child with severe S-adenosylhomocysteine hydrolase (AHCY) deficiency (AHCY c.428A>G, p.Tyr143Cys; c.982T>G, p.Tyr328Asp) presented at 8 months of age with growth failure, microcephaly, global developmental delay, myopathy, hepatopathy, and factor VII deficiency. Plasma methionine, S-adenosylmethionine (AdoMet), and S-adenosylhomocysteine (AdoHcy) were markedly elevated and the molar concentration ratio of AdoMet:AdoHcy, believed to regulate a myriad of methyltransferase reactions, was 15% of the control mean. Dietary therapy failed to normalize biochemical markers or alter the AdoMet to AdoHcy molar concentration ratio. At 40 months of age, the proband received a liver segment from a healthy, unrelated living donor. Mean AdoHcy decreased 96% and the AdoMet:AdoHcy concentration ratio improved from 0.52±0.19 to 1.48±0.79 mol:mol (control 4.10±2.11 mol:mol). Blood methionine and AdoMet were normal and stable during 6 months of follow-up on an unrestricted diet. Average calculated tissue methyltransferase activity increased from 43±26% to 60±22%, accompanied by signs of increased transmethylation in vivo. Factor VII activity increased from 12% to 100%. During 6 postoperative months, head growth accelerated 4-fold and the patient made promising gains in gross motor, language, and social skills.

Intestinal Transplantation in Children

This review summarizes the outcomes and known adverse effects of current immunosuppression strategies in use in pediatric intestinal transplantation. Intestinal transplantation has evolved from an experimental therapy to a highly successful treatment for children with intestinal failure who have complications with total parenteral nutrition. Because of continued success with intestinal transplantation over the past decade, the focus of clinicians and researchers is shifting from short-term patient survival to optimizing long-term outcomes.Current 5-year patient and graft survival rates after intestinal transplantation are 58% and 40%, respectively, in the US; single centers have reported nearly 80% patient and 60% graft survival rates at 5 years. The immunosuppression strategy in intestinal transplantation includes a tacrolimus-based regimen, usually in conjunction with an antibody induction therapy such as rabbit-antithymocyte globulin, interleukin-2 receptor antagonists, or alemtuzumab. The use of these immunosuppressive regimens, along with improved medical and surgical care, has contributed significantly toward improved outcomes. Optimization of post-transplant immunosuppression strategies to reduce adverse effects while minimizing acute and chronic graft rejection is a strong clinical and research focus.

Host conditioning and rejection monitoring in hepatocyte transplantation in humans

BACKGROUND & AIMSnHepatocyte transplantation partially corrects genetic disorders and has been associated anecdotally with reversal of acute liver failure. Monitoring for graft function and rejection has been difficult, and has contributed to limited graft survival. Here we aimed to use preparative liver-directed radiation therapy, and continuous monitoring for possible rejection in an attempt to overcome these limitations.nnnMETHODSnPreparative hepatic irradiation was examined in non-human primates as a strategy to improve engraftment of donor hepatocytes, and was then applied in human subjects. T cell immune monitoring was also examined in human subjects to assess adequacy of immunosuppression.nnnRESULTSnPorcine hepatocyte transplants engrafted and expanded to comprise up to 15% of irradiated segments in immunosuppressed monkeys preconditioned with 10Gy liver-directed irradiation. Two patients with urea cycle deficiencies had early graft loss following hepatocyte transplantation; retrospective immune monitoring suggested the need for additional immunosuppression. Preparative radiation, anti-lymphocyte induction, and frequent immune monitoring were instituted for hepatocyte transplantation in a 27year old female with classical phenylketonuria. Post-transplant liver biopsies demonstrated multiple small clusters of transplanted cells, multiple mitoses, and Ki67+ hepatocytes. Mean peripheral blood phenylalanine (PHE) level fell from pre-transplant levels of 1343±48μM (normal 30-119μM) to 854±25μM (treatment goal ≤360μM) after transplant (36% decrease; p<0.0001), despite transplantation of only half the target number of donor hepatocytes. PHE levels remained below 900μM during supervised follow-up, but graft loss occurred after follow-up became inconsistent.nnnCONCLUSIONSnRadiation preconditioning and serial rejection risk assessment may produce better engraftment and long-term survival of transplanted hepatocytes. Hepatocyte xenografts engraft for a period of months in non-human primates and may provide effective therapy for patients with acute liver failure.nnnLAY SUMMARYnHepatocyte transplantation can potentially be used to treat genetic liver disorders but its application in clinical practice has been impeded by inefficient hepatocyte engraftment and the inability to monitor rejection of transplanted liver cells. In this study, we first show in non-human primates that pretreatment of the host liver with radiation improves the engraftment of transplanted liver cells. We then used this knowledge in a series of clinical hepatocyte transplants in patients with genetic liver disorders to show that radiation pretreatment and rejection risk monitoring are safe and, if optimized, could improve engraftment and long-term survival of transplanted hepatocytes in patients.

Invasive candidiasis in liver transplant patients: Incidence and risk factors in a pediatric cohort

Prolonged OR, re‐transplantation, and high‐volume intraoperative transfusion have been associated with increased risk for IC in adult LT recipients. Antifungal prophylaxis is recommended for adult patients with these risk factors. There are limited data on the incidence of and risk factors for IC in pediatric LT recipients. A retrospective cohort study of all pediatric LT patients at the CHOP between 2000 and 2012 and the CHP between 2004 and 2012 was performed to define the incidence of IC within 30 days of LT. A 3:1 matched case–control study with incidence density sampling was performed. Conditional logistic regression analyses were used to explore risk factors associated with IC. Among 397 recipients, the incidence of IC was 2.5%. Bivariate analyses showed that ICU admission prior to transplant, OR > 10 h, intraoperative volume infusion of >300 mL/kg, and broad‐spectrum antibiotics were significantly associated with IC. In a multivariate model, only ICU admission remained significantly associated with IC. Antifungal prophylaxis was not significantly protective against IC. The low incidence of IC and lack of an identified protective effect from antifungal prophylaxis suggest that prophylaxis in pediatric LT recipients should not be routinely recommended to prevent IC events in the first 30 days post‐transplant.

Living related transplantation for MSUD--caution, or a new path forward?

Since we introduced elective orthotopic liver transplantation to treat severe branched-chain ketoacid dehydrogenase deficiency (BCKDH; “classical” maple syrup urine disease, MSUD) (1, 2), more than 100 patients have undergone transplant for this indication in the United States (UNOS.org). The fundamental goal of transplantation is metabolic cure, which requires provision of sufficient enzyme mass to handle unrestricted dietary branched-chain amino acid (BCAA) loads as well as the dynamic metabolic fluctuations that occur during illnesses, when BCAAs rapidly enter the circulation as a consequence of protein degradation in muscle. Based on the prevailing understanding of BCKDH enzyme distribution in humans, we initially selected whole deceased donor livers for the treatment protocol. The only study to address this quantitatively indicates that in vivo, the liver may account for only 9–13% of “whole-body” BCKDH activity as compared to 54–66% in muscle (3). We hypothesized that 9–13% wholebody BCKDH activity would suffice to respond to dietary and physiological leucine flux, minimizing or eliminating the risk of neurological sequelae. Our results to date bear this out. Since 2004, 60 individuals with classical MSUD have been transplanted under our protocol; graft and recipient survival are 100%, and we have observed no neurological crisis nor progression of neurological illness post-transplant. Interestingly, the broad distribution of BCKDH activity among various tissues (e.g., liver, muscle, kidney, brain) explains why an explanted MSUD liver can be safely transplanted into a non-MSUD recipient (i.e., “domino” transplant) (4, 5), but these same data initially deterred us from using living related donors to treat classical MSUD. Our fear was that transplanted liver tissue heterozygous for one severe MSUD mutation (i.e., expressing only 50% normal BCKDH activity) would replace only 4–7% of BCKDH activity on a whole-body basis, an amount that might not be adequate to ensure good metabolic outcome. Classical MSUD can be treated effectively with stringent monitoring and rigorous adherence to a complex dietary regimen (6). However, it is not possible to fully safeguard the brain using this strategy, and in developing countries, resources that allow for good dietary treatment are scarce. Thus, MSUD remains a volatile and dangerous disease, and in many parts of the world, it is uniformly associated with death or severe neurological disability. These same regions often face a dire shortage of deceased donor livers. Technical variant grafts such as split grafts have been subsequently used in our series, and Feier et al. (7) recently described the successful transplantation of a partial liver allograft from a heterozygous mother to her child with MSUD. In parallel, we have learned that a handful of living related donor transplants (LRDTs) for MSUD from heterozygote donors were performed at other centers, accompanied by anecdotal reports of initial success (G. Mazariegos, personal communication). The evidence provided by Patel et al. in this issue of Pediatric Transplantation (8) further supports the efficacy of LRDT for the treatment of MSUD, but also underscores the kind of additional detail required to bolster confidence in this strategy. First, whereas the quantitative amino acid profiles of the recipient pretransplant are consistent with classic MSUD, additional data on preand post-transplant leucine tolerance in mg/kg/day are necessary to determine disease severity (e.g., classical, intermediate, intermittent, etc.) and quantify the change in leucine tolerance as a result of the graft. In our series of 60 children and adults with MSUD, leucine tolerance improved ≥10-fold post-transplant, from 10–25 mg/kg/day to >150 mg/kg/day. Data by Patel et al. do not allow for such conclusions. Second, BCAA profiles during illness should be monitored to document the graft’s ability to

Right diaphragmatic hernia after liver transplant in pediatrics: A case report and review of the literature

Diaphragmatic hernias (DH) are an unusual complication after pediatric liver transplantation; however, they have been reported with increased frequency in the past few years. DHs are responsible for nearly half of the small bowel obstructions requiring surgical intervention in this patient population. It has been suggested that the use of a left lobe liver graft, surgical trauma, malnourishment, elevated intra‐abdominal pressures, and mTor inhibitors may predispose to development of DH. The use of a segmental graft may increase the recognition of diaphragmatic hernia because the surgically damaged right hemi‐diaphragm often remains exposed to underlying viscera, instead of being covered by the right hepatic lobe. Treatment is surgical reduction, with up to 20% of the patients requiring resection of the herniated intestine. Herein we describe a case of DH after left segmental liver transplant in a two‐ yr‐old boy that presented one month post left lobe split liver transplant with abdominal pain, anorexia, and respiratory distress. Just like in the majority of the reported cases, an urgent laparotomy with primary repair was performed. No resection of the herniated segment of intestine was required. For pediatric patients with otherwise unexplained respiratory or gastrointestinal symptoms after a left lateral segment liver transplant, right‐sided diaphragmatic hernias should always be high in the differential diagnosis.