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Background and aims: Alagille syndrome (ALGS) is a multisystem disorder, characterized by cholestasis. Existing outcome data are largely derived from tertiary centers, and real-world data are lacking. This study aimed to elucidate the natural history of liver disease in a contemporary, international cohort of children with ALGS. Approach and results: This was a multicenter retrospective study of children with a clinically and/or genetically confirmed ALGS diagnosis, born between January 1997 and August 2019. Native liver survival (NLS) and event-free survival rates were assessed. Cox models were constructed to identify early biochemical predictors of clinically evident portal hypertension (CEPH) and NLS. In total, 1433 children (57% male) from 67 centers in 29 countries were included. The 10 and 18-year NLS rates were 54.4% and 40.3%. By 10 and 18 years, 51.5% and 66.0% of children with ALGS experienced >= 1 adverse liver-related event (CEPH, transplant, or death). Children (>6 and = 5.0 and = 10.0 mg/dl had an 8.0-fold (95% CI, 3.4-18.4) increased risk of developing CEPH compared with those 10.0 mg/dl were associated with a 4.8 (95% CI, 2.4-9.7) and 15.6 (95% CI, 8.7-28.2) increased risk of transplantation relative to = 5.0 mg/dl, with 79% reaching adulthood with native liver (p < 0.001). Conclusions: in this large international cohort of ALGS, only 40.3% of children reach adulthood with their native liver. A TB, This study received funding support from the following agencies: The Alagille Syndrome Alliance, Mirum Pharmaceuticals Inc. and Albireo Pharma, Inc. who provided unrestricted educational grants to the Hospital for Sick Children (SickKids Foundation). The study sponsors were not involved in the conduct of the research study or preparation of the manuscript.

With the application of modern investigative technologies, cholestatic liver diseases of genetic etiology are increasingly identified as the root cause of previously designated "idiopathic" adult and pediatric liver diseases. Here, we review advances in the field enhanced by a deeper understanding of the phenotypes associated with specific gene defects that lead to cholestatic liver diseases. There are evolving areas for clinicians in the current era specifically regarding the role for biopsy and opportunities for a "sequencing first" approach. Risk stratification based on the severity of the genetic defect holds promise to guide the decision to pursue primary liver transplantation versus medical therapy or nontransplant surgery, as well as early screening for HCC. In the present era, the expanding toolbox of recently approved therapies for hepatologists has real potential to help many of our patients with genetic causes of cholestasis. In addition, there are promising agents under study in the pipeline. Relevant to the current era, there are still gaps in knowledge of causation and pathogenesis and lack of fully accepted biomarkers of disease progression and pruritus. We discuss strategies to overcome the challenges of genotype-phenotype correlation and draw attention to the extrahepatic manifestations of these diseases. Finally, with attention to identifying causes and treatments of genetic cholestatic disorders, we anticipate a vibrant future of this dynamic field which builds upon current and future therapies, real-world evaluations of individual and combined therapeutics, and the potential incorporation of effective gene editing and gene additive technologies.

Objective: To evaluate neurodevelopmental status among children with inherited cholestatic liver diseases with native liver and variables predictive of impairment. Methods: Participants with Alagille syndrome (ALGS), progressive familial intrahepatic cholestasis (PFIC), and alpha 1 antitrypsin deficiency (A1AT) enrolled in a longitudinal, multicenter study and completed the Wechsler Preschool and Primary Scale of Intelligence-III or Intelligence Scale for Children-IV. Full Scale Intelligence Quotient (FSIQ) was analyzed continuously and categorically (>100, 85–99, 70–84, 1 standard deviation [SD] below average) was highest in ALGS (29%) versus 18.6% in PFIC and 12.8% in A1AT, and was greater than expected in ALGS based on normal distribution (29% vs 15.9%, P = 0.003). ALGS scored significantly lower than test norms in almost all Wechsler composites; A1AT scored lower on Working Memory and Processing Speed; PFIC was not different from test norms. Total bilirubin, alkaline phosphatase, albumin, hemoglobin, and parental education were significantly associated with FSIQ. Conclusions: Patients with ALGS are at increased risk of lower FSIQ, whereas our data suggest A1AT and PFIC are not. A1AT and ALGS appear vulnerable to working memory and processing speed deficits suggestive of attention/executive function impairment. Malnutrition, liver disease severity, and sociodemographic factors appear related to FSIQ deficits, potentially identifying targets for early interventions.

X-linked myotubular myopathy (XLMTM) is a rare congenital myopathy characterized by profound hypotonia and poor respiratory effort at birth. The condition is associated with multiple morbidities including chronic respiratory insufficiency, feeding tube dependence, and rarely, vitamin K deficiency leading to bleeding and coagulopathy. We report a case of a 6-month-old boy with X-linked myotubular myopathy who experienced a fatal intracranial hemorrhage due to vitamin K deficiency without prior clinical evidence of cholestasis or micronutrient deficiency. We propose clinically non-apparent cholestasis in combination with acute illness and poor weight gain led to his vitamin K deficiency and intracranial hemorrhage. However, the etiology and mechanism of his cholestasis remains unclear. We conclude that children with X-linked myotubular myopathy, especially with gene therapy on the horizon, may benefit from routine hepatic, coagulation, and nutritional screening to prevent potentially catastrophic bleeding.

BACKGROUND & AIMS: Detailed investigation of the biological pathways leading to hepatic fibrosis and identification of liver fibrosis biomarkers may facilitate early interventions for pediatric cholestasis. APPROACH & RESULTS: A targeted ELISA-based panel of 9 biomarkers (lysyl oxidase (LOX), tissue inhibitor matrix metalloproteinase 1 (TIMP1), connective tissue growth factor (CTGF), interleukin-8 (IL-8), endoglin, periostin, mac-2-binding protein (mac2-BP), matrix metalloproteinase-3 and −7 (MMP-3, MMP-7) was examined in children with biliary atresia (BA, n=187), alpha-1 antitrypsin deficiency (A1AT, n=78) and Alagille syndrome (ALGS, n=65) and correlated with liver stiffness (LSM) and biochemical measures of liver disease. Median age and LSM were 9 years and 9.5 kPa. After adjusting for covariates, there were positive correlations between LSM and endoglin (p=0.04), IL-8 (p

Ring‐finger protein 213 (RNF213) encodes a protein of unknown function believed to play a role in cellular metabolism and angiogenesis. Gene variants are associated with susceptibility to moyamoya disease. Here, we describe two children with moyamoya disease who also demonstrated kidney disease, elevated aminotransferases, and recurrent skin lesions found by exome sequencing to have de novo missense variants in RNF213. These cases highlight the ability of RNF213 to cause Mendelian moyamoya disease in addition to acting as a genetic susceptibility locus. The cases also suggest a new, multi‐organ RNF213‐spectrum disease characterized by liver, skin, and kidney pathology in addition to severe moyamoya disease caused by heterozygous, de novo C‐terminal RNF213 missense variants.

Purpose Detection of all major classes of genomic variants in a single test would decrease cost and increase the efficiency of genomic diagnostics. Genome sequencing (GS) has the potential to provide this level of comprehensive detection. We sought to demonstrate the utility of GS in the molecular diagnosis of 18 patients with clinically defined Alagille syndrome (ALGS), who had a negative or inconclusive result by standard-of-care testing. Methods We performed GS on 16 pathogenic variant-negative probands and two probands with inconclusive results (of 406 ALGS probands) and analyzed the data for sequence, copy-number, and structural variants in JAG1 and NOTCH2. Results GS identified four novel pathogenic alterations including a copy-neutral inversion, a partial deletion, and a promoter variant in JAG1, and a partial NOTCH2 deletion, for an additional diagnostic yield of 0.9%. Furthermore, GS resolved two complex rearrangements, resulting in identification of a pathogenic variant in 97.5% (n = 396/406) of patients after GS. Conclusion GS provided an increased diagnostic yield for individuals with clinically defined ALGS who had prior negative or incomplete genetic testing by other methods. Our results show that GS can detect all major classes of variants and has potential to become a single first-tier diagnostic test for Mendelian disorders.

We describe 10 females with ornithine transcarbamylase (OTC) deficiency and liver dysfunction, revealing a unique pattern of hepatocyte injury in which initial hyperammonemia and coagulopathy is followed by a delayed peak in aminotransferase levels. None of the patients required urgent liver transplantation, though five eventually underwent transplant for recurrent metabolic crises. We intend that this novel observation will initiate further investigations into the pathophysiology of liver dysfunction in OTC-deficient patients, and ultimately lead to the development of therapies and prevent the need for liver transplant.

Approximately 50% of infants with biliary atresia (BA) undergoing Kasai portoenterostomy show survival with native liver (SNL) at age 2 years. Predictors of disease progression after age 2 years are unknown, despite estimates of 20%‐30% undergoing liver transplant (LT) between age 2 and 18 years. We sought to address this knowledge gap by developing prognostic models in participants of the multicenter prospective National Institutes of Health‐supported Childhood Liver Disease Research Network. We extracted 14 clinical and biochemical variables at age 2 years to develop two models for future outcomes: 1) LT or death (LTD) and 2) first sentinel event (SE), either new onset ascites, hepatopulmonary syndrome (HPS), or gastrointestinal (GI) bleed. A total of 240 participants, enrolled between 2004 and 2017, were followed until a median age of 5.1 years (range, 2.0‐13.3 years). Of these participants, 38 underwent LT (n = 37) or death (n = 1); cumulative incidence, 23.7% (95% confidence interval [CI], 16.2%‐32.0%). Twenty‐seven experienced either new‐onset ascites (n = 13), HPS (n = 1), or GI bleed (n = 14). One participant had ascites and GI bleed concurrently; cumulative incidence, 21.5% (95% CI, 14.2%‐29.8%) by age 10 years. The Cox proportional hazard model predicted risk of LTD, using total bilirubin, albumin, platelet count, and history of either ascites or cholangitis (BA LTD model), with a C‐index of 0.88 (range, 0.86‐0.89). A cause‐specific hazard competing risk model predicted SE using platelet count and gamma glutamyltransferase levels (BA SE model) with a C‐index of 0.81 (range, 0.80‐0.84). Internal model validity was assessed using Harrell’s C‐index with cross‐validation. Conclusion: Stratification using these models identified risk of poor outcomes in patients with BA SNL after age 2 years. The models may identify those who would benefit from enhanced clinical surveillance and prioritization in clinical trials.

Objectives In many pediatric acute liver failure (PALF) cases, a diagnosis is not identified, and the etiology is indeterminate (IND-PALF). Our pilot study found dense CD8 T-cell infiltrates and increased T-cell clonality in liver specimens from IND-PALF patients. We aimed to validate these findings in a multicenter cohort with investigators blinded to diagnosis. Methods PALF Study Group registry subjects with IND-PALF (n = 37) and known diagnoses (DX-PALF) (n = 18), ages 1 to 17 years, with archived liver tissue were included. Liver tissue slides were stained for T cells (CD8 and CD4), B cells (CD20), macrophages (CD163), perforin, and tissue resident-memory T cells (Trm, CD103), and scored as minimal, moderate, or dense. Lymphocytes were isolated from frozen liver tissue for T-cell receptor beta (TCRβ) sequencing. Results Dense hepatic CD8 staining was found in significantly more IND-PALF (n = 29, 78%) compared with DX-PALF subjects (n = 5, 28%) (P = 0.001). IND-PALF subjects were more likely to have dense or moderate perforin (88% vs 50%, P = 0.03) and CD103 (82% vs 40%, P = 0.02) staining compared with DX-PALF subjects. TCRβ sequencing of 15 IND-PALF cases demonstrated increased clonal overlap compared with 6 DX-PALF cases (P = 0.002). Conclusions Dense infiltration of effector Trm CD8 T cells characterizes liver tissue from IND-PALF subjects. Increased clonality suggests the T-cell expansion is antigen(s)-driven as opposed to a nonspecific inflammatory response. These findings support CD8 staining as a new biomarker of the activated CD8 T-cell PALF phenotype. Future studies are needed to characterize potential antigens, host risk factors, and inflammatory pathways with the goal of developing targeted therapies.

Alagille syndrome (ALGS) and progressive familial intrahepatic cholestasis (PFIC) are inherited cholestatic disorders with risk of developing end‐stage liver disease requiring liver transplantation (LT). We investigated aspartate aminotransferase‐to‐platelet ratio index (APRI), Fibrosis‐4 score (FIB‐4), and conjugated bilirubin as biomarkers to assess fibrosis severity and risk for LT among children with ALGS and PFIC. This multicenter, cross‐sectional study included 64 children with ALGS or PFIC (per genetics or strict clinical criteria) with APRI, FIB‐4, and conjugated bilirubin levels collected within ±90 days of their most recent liver biopsy. A single, blinded pathologist staged all biopsies (metavir; F0‐F2: nonsevere, F3‐F4: severe). Logistic regression and area under the receiver operating characteristic curve analysis (AUC) were used to assess biomarker associations with fibrosis severity and risk for LT. In ALGS, only APRI distinguished F3‐F4 (AUC 0.72, P = 0.012), with a cutoff greater than 2.97 demonstrating a sensitivity of 61.5% (95% confidence interval 0.32, 0.86) and specificity of 81.5% (0.62, 0.94). In ALGS, a 50% increase of APRI increased the odds of F3‐F4 by 1.31‐fold (1.04, 1.65; P = 0.023). In ALGS, APRI (AUC 0.87; P

Background Entrustable professional activities (EPAs) are critical activities performed by medical professionals, which can be observed and assessed. Adding on to common EPAs for all pediatric subspecialty trainees, specialty-specific EPAs for pediatric gastroenterology, hepatology, and nutritional fellowship were developed by the North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition (NASPGHAN) EPA Task Force. Methods Having developed specialty-specific EPAs, building EPA assessments is the next logical step, as EPAs are included under a larger umbrella of competency-based assessment. Thus, the NASPGHAN EPA Task Force and Training Committee collaborated on an assessment tool and associated curricular resources to aid in tracking trainees' progression to entrustment within individual EPAs and readiness for independent practice. Results This manuscript reports the development of an EPA assessment tool, including guiding principles and the theory behind the assessment tool, with a focus on simple, meaningful assessments that can provide crucial performance feedback to trainees. In addition, curricular resources were developed, based on the assessment tool, to support training. Ultimately, it is the hope of the NASPGHAN EPA Task Force and Training Committee that this tool can aid training programs in providing formative feedback for trainees, and can be used by training programs and clinical competency committees for summative evaluation.

Quality training in pediatric gastroenterology, hepatology, and nutrition is essential for the future of our specialty from advancing the science through research to providing clinical care for children with gastrointestinal, hepatic and nutritional disorders. As educational theory has developed, both the American Board of Pediatrics (ABP) and the Accreditation Council for Graduate Medical Education (ACGME) have commissioned projects to better define training including core competencies, and milestones with the goal of competency-based assessment. Seeking to provide a clinical context for these competencies and milestones, the ABP commissioned a project for each pediatric subspecialty to develop entrustable professional activities (EPA) while at the same time developing EPAs that are common to all pediatric subspecialties. North American Society for Pediatric Gastroenterology, Hepatology, Nutrition (NASPGHAN) commissioned an EPA Task Force to develop the pediatric gastroenterology, hepatology and nutrition EPAs. This document serves as an introduction to EPAs, including their historical background, underlying educational theory, and the process used to develop the pediatric gastroenterology, hepatology and nutrition EPAs in the United States of America.

Biliary atresia (BA) is a severe pediatric liver disease resulting in necroinflammatory obliteration of the extrahepatic biliary tree. BA presents within the first few months of life as either an isolated finding or with additional syndromic features. The etiology of isolated BA is unknown, with evidence for infectious, environmental, and genetic risk factors described. However, to date, there are no definitive causal genes identified for isolated BA in humans, and the question of whether single gene defects play a major role remains open. We performed exome-sequencing in 101 North American patients of European descent with isolated BA (including 30 parent-child trios) and considered several experimental designs to identify potentially deleterious protein-altering variants that may be involved in the disease. In a case-only analysis, we did not identify genes with variants shared among more than two probands, and burden tests of rare variants using a case-case control design did not yield significant results. In the trio analysis of 30 simplex families (patient and parent trios), we identified 66 de novo variants in 66 genes including potentially deleterious variants in STIP1 and REV1. STIP1 is a co-chaperone for the heat-shock protein, HSP90, and has been shown to have diverse functions in yeast, flies and mammals, including stress-responses. REV1 is known to be a key player in DNA repair pathway and to interact with HSP90. In conclusion, our results do not support the hypothesis that a simple genetic model is responsible for the majority of cases of isolated BA. Our finding of de novo variants in genes linked to evolutionarily conserved stress responses (STIP1 and REV1) suggests that exploration of how genetic susceptibility and environmental exposure may interact to cause BA is warranted.

The natural history of gastroesophageal variceal hemorrhage (VH) in biliary atresia (BA) is not well characterized. We analyzed risk factors, incidence, and outcomes of VH in a longitudinal multicenter study.Participants enrolled in either an incident (Prospective Database of Infants with Cholestasis [PROBE]) or prevalent (Biliary Atresia Study of Infants and Children [BASIC]) cohort of BA were included. Variceal hemorrhage (VH) was defined based on gastrointestinal bleeding in the presence of varices accompanied by endoscopic or nontransplant surgical intervention. Cumulative incidence of VH and transplant-free survival was compared based on features of portal hypertension (e.g., splenomegaly, thrombocytopenia) and clinical parameters at baseline in each cohort (PROBE: 1.5 to 4.5 months after hepatoportoenterostomy [HPE]; BASIC: at enrollment3 years of age). Analyses were conducted on 869 children with BA enrolled between June 2004 and December 2020 (521 in PROBE [262 (51%) with a functioning HPE] and 348 in BASIC). The overall incidence of first observed VH at 5 years was 9.4% (95% CI: 7.0-12.4) in PROBE and 8.0% (5.2-11.5) in BASIC. Features of portal hypertension, platelet count, total bilirubin, aspartate aminotransferase (AST), albumin, and AST-to-platelet ratio index at baseline were associated with an increased risk of subsequent VH in both cohorts. Transplant-free survival at 5 years was 45.1% (40.5-49.6) in PROBE and 79.2% (74.1-83.4) in BASIC. Two (2.5%) of 80 participants who had VH died, whereas 10 (12.5%) underwent transplant within 6 weeks of VH.The low risk of VH and associated mortality in children with BA needs to be considered in decisions related to screening for varices and primary prophylaxis of VH.

Poster-Abstract

Background and Aims Recurrent primary sclerosing cholangitis (rPSC) following liver transplant (LT) has a negative impact on graft and patient survival; little is known about risk factors for rPSC or disease course in children. Approach and Results We retrospectively evaluated risk factors for rPSC in 140 children from the Pediatric PSC Consortium, a multicenter international registry. Recipients underwent LT for PSC and had >90 days of follow-up. The primary outcome, rPSC, was defined using Graziadei criteria. Median follow-up after LT was 3 years (interquartile range 1.1-6.1). rPSC occurred in 36 children, representing 10% and 27% of the subjects at 2 years and 5 years following LT, respectively. Subjects with rPSC were younger at LT (12.9 vs. 16.2 years), had faster progression from PSC diagnosis to LT (2.5 vs. 4.1 years), and had higher alanine aminotransferase (112 vs. 66 IU/L) at LT (all P < 0.01). Inflammatory bowel disease was more prevalent in the rPSC group (86% vs. 66%; P = 0.025). After LT, rPSC subjects had more episodes of biopsy-proved acute rejection (mean 3 vs. 1; P < 0.001), and higher prevalence of steroid-refractory rejection (41% vs. 20%; P = 0.04). In those with rPSC, 43% developed complications of portal hypertension, were relisted for LT, or died within 2 years of the diagnosis. Mortality was higher in the rPSC group (11.1% vs. 2.9%; P = 0.05). Conclusions The incidence of rPSC in this cohort was higher than previously reported, and was associated with increased morbidity and mortality. Patients with rPSC appeared to have a more aggressive, immune-reactive phenotype. These findings underscore the need to understand the immune mechanisms of rPSC, to lay the foundation for developing new therapies and improve outcomes in this challenging population.

Mutations in ATP8B1 can lead to familial intrahepatic cholestasis type 1 (FIC1) deficiency, or progressive familial intrahepatic cholestasis type 1 (PFIC1). The rarity of FIC1 deficiency has largely prevented a detailed analysis of its natural history, effects of predicted protein truncating mutations (PPTMs), and possible associations of serum bile acid (sBA) concentrations and surgical biliary diversion (SBD) with long-term outcome. We aimed to provide novel insights by using the largest genetically defined cohort of FIC1 deficiency patients to date. This multicenter, combined retrospective and prospective study included 130 patients with compound heterozygous or homozygous predicted pathogenic ATP8B1 variants. Patients were categorized according to the number of PPTMs (i.e., splice site, frameshift due to deletion or insertion, nonsense, duplication); FIC1-A (n=67; no PPTM), FIC1-B (n=29; one PPTM) or FIC1-C (n=34; two PPTMs). Survival analysis showed an overall native liver survival (NLS) of 44% at age 18y. NLS was comparable between FIC1-A, FIC1-B, and FIC1-C (%NLS at age 10y: 67%, 41%, and 59%, respectively; P=0.12), despite FIC1-C undergoing SBD less often (%SBD at age 10y: 65%, 57%, and 45%, respectively; P=0.03). sBAs at presentation were negatively associated with NLS (NLS at age 10y; sBAs, University of Colorado; Baylor College of Medicine; Children’s Hospital of Philadelphia; Children’s Hospital of Pittsburgh; St Louis Children’s Hospital; Riley Hospital for Children; Seattle Children’s Hospital; M.D./Ph.D. Scholarship from the University of Groningen; European Society for Pediatric Gastroenterology, Hepatology and Nutrition (ESPGHAN) Networking Grant 2019; ChilDReN National Institutes of Health Grants; Ann & Robert H. Lurie Children’s Hospital; Albireo and Mirum Pharmaceuticals

Objectives: The aim of the study was to determine the frequency and natural history of infantile idiopathic cholestasis (IC) in a large, prospective, multicenter cohort of infants. Methods: We studied 94 cholestatic infants enrolled up to 6 months of age in the NIDDK ChiLDReN (Childhood Liver Disease Research Network) “PROBE” protocol with a final diagnosis of IC; they were followed up to 30 months of age. Results: Male sex (66/94; 70%), preterm birth (22/90 with data; 24% born at < 37 weeks’ gestational age), and low birth weight (25/89; 28% born at 1 mg/dL and/or ALT > 35 U/L; n = 7), and exited healthy (resolved disease per study site report but without documented biochemical resolution; n = 34). Biochemical resolution occurred at median of 9 months of age. GGT was

Alagille syndrome (ALGS) is an autosomal dominant disorder caused by pathogenic variants in JAG1 or NOTCH2, which encode fundamental components of the Notch signaling pathway. Clinical features span multiple organ systems including hepatic, cardiac, vascular, renal, skeletal, craniofacial, and ocular, and occur with variable phenotypic penetrance. Genotype–phenotype correlation studies have not yet shown associations between mutation type and clinical manifestations or severity, and it has been hypothesized that modifier genes may modulate the effects of JAG1 and NOTCH2 pathogenic variants. Medical management is supportive, focusing on clinical manifestations of disease, with liver transplant indicated for severe pruritus, liver synthetic dysfunction, portal hypertension, bone fractures, and/or growth failure. New therapeutic approaches are under investigation, including ileal bile acid transporter (IBAT) inhibitors and other approaches that may involve targeted interventions to augment the Notch signaling pathway in involved tissues.

Alagille syndrome is an autosomal dominant disease with a known molecular etiology of dysfunctional Notch signaling caused primarily by pathogenic variants in JAGGED1 (JAG1), but also by variants in NOTCH2. The majority of JAG1 variants result in loss of function, however disease has also been attributed to lesser understood missense variants. Conversely, the majority of NOTCH2 variants are missense, though fewer of these variants have been described. In addition, there is a small group of patients with a clear clinical phenotype in the absence of a pathogenic variant. Here, we catalog our single‐center study, which includes 401 probands and 111 affected family members amassed over a 27‐year period, to provide updated mutation frequencies in JAG1 and NOTCH2 as well as functional validation of nine missense variants. Combining our cohort of 86 novel JAG1 and three novel NOTCH2 variants with previously published data (totaling 713 variants), we present the most comprehensive pathogenic variant overview for Alagille syndrome. Using this data set, we developed new guidance to help with the classification of JAG1 missense variants. Finally, we report clinically consistent cases for which a molecular etiology has not been identified and discuss the potential for next generation sequencing methodologies in novel variant discovery.

Biliary atresia is a progressive fibroinflammatory cholangiopathy of infancy that is associated with activation of innate and adaptive immune responses targeting bile ducts. A recently completed multicenter phase I/IIA trial of intravenous immunoglobulin in biliary atresia did not improve serum total bilirubin levels at 90 days after hepatoportoenterostomy or survival with the native liver at 1 year. A mechanistic aim of this trial was to determine if the peripheral blood immunophenotype was associated with clinical outcomes. Flow cytometry of peripheral blood cell markers (natural killer [NK], macrophage subsets, T‐ and B‐cell subsets, regulatory T cells), neutrophils, and activation markers (clusters of differentiation [CD]38, CD69, CD86, human leukocyte antigen‐DR isotype [HLA‐DR]) was performed on 29 patients with biliary atresia at baseline and at 60, 90, 180, and 360 days after hepatoportoenterostomy. Plasma cytokines and neutrophil products were also measured. Spearman correlations of change of an immune marker from baseline to day 90 with change in serum bilirubin revealed that an increase in total bilirubin correlated with 1) increased percentage of HLA‐DR+CD38+ NK cells and expression of NK cell activation markers CD69 and HLA‐DR, 2) decreased percentage of regulatory T cells, and 3) increased interleukin (IL)‐8 and associated neutrophil products (elastase and neutrophil extracellular traps). Cox modeling revealed that the change from baseline to day 60 of the percentage of HLA‐DR+CD38+ NK cells and plasma IL‐8 levels was associated with an increased risk of transplant or death by day 360. Conclusion: Poor outcomes in biliary atresia correlated with higher peripheral blood NK cells and IL‐8 and lower regulatory T cells. Future studies should include immunotherapies targeting these pathways in order to protect the biliary tree from ongoing damage.

Biliary atresia (BA) is the most common cause of end-stage liver disease in children and the primary indication for pediatric liver transplantation, yet underlying etiologies remain unknown. Approximately 10% of infants affected by BA exhibit various laterality defects (heterotaxy) including splenic abnormalities and complex cardiac malformations-a distinctive subgroup commonly referred to as the biliary atresia splenic malformation (BASM) syndrome. We hypothesized that genetic factors linking laterality features with the etiopathogenesis of BA in BASM patients could be identified through whole-exome sequencing (WES) of an affected cohort. DNA specimens from 67 BASM subjects, including 58 patient-parent trios, from the National Institute of Diabetes and Digestive and Kidney Diseases-supported Childhood Liver Disease Research Network (ChiLDReN) underwent WES. Candidate gene variants derived from a prespecified set of 2,016 genes associated with ciliary dysgenesis and/or dysfunction or cholestasis were prioritized according to pathogenicity, population frequency, and mode of inheritance. Five BASM subjects harbored rare and potentially deleterious biallelic variants in polycystic kidney disease 1 like 1 (PKD1L1), a gene associated with ciliary calcium signaling and embryonic laterality determination in fish, mice, and humans. Heterozygous PKD1L1 variants were found in 3 additional subjects. Immunohistochemical analysis of liver from the one BASM subject available revealed decreased PKD1L1 expression in bile duct epithelium when compared to normal livers and livers affected by other noncholestatic diseases. Conclusion: WES identified biallelic and heterozygous PKD1L1 variants of interest in 8 BASM subjects from the ChiLDReN data set; the dual roles for PKD1L1 in laterality determination and ciliary function suggest that PKD1L1 is a biologically plausible, cholangiocyte-expressed candidate gene for the BASM syndrome.

Osteopenia and bone fractures are significant causes of morbidity in children with cholestatic liver disease. Dual-energy X-ray absorptiometry (DXA) analysis was performed in children with intrahepatic cholestatic diseases who were enrolled in the Longitudinal Study of Genetic Causes of Intrahepatic Cholestasis in the Childhood Liver Disease Research Network. DXA was performed on participants aged >5 years (with native liver) diagnosed with bile acid synthetic disorder (BASD), alpha-1 antitrypsin deficiency (A1AT), chronic intrahepatic cholestasis (CIC), and Alagille syndrome (ALGS). Weight, height, and body mass index Z scores were lowest in CIC and ALGS. Total bilirubin (TB) and serum bile acids (SBA) were highest in ALGS. Bone mineral density (BMD) and bone mineral content (BMC) Z scores were significantly lower in CIC and ALGS than in BASD and A1AT (P < 0.001). After anthropometric adjustment, bone deficits persisted in CIC but were no longer noted in ALGS. In ALGS, height-adjusted and weight-adjusted subtotal BMD and BMC Z scores were negatively correlated with TB (P < 0.001) and SBA (P = 0.02). Mean height-adjusted and weight-adjusted subtotal BMC Z scores were lower in ALGS participants with a history of bone fractures. DXA measures did not correlate significantly with biliary diversion status. Conclusion: CIC patients had significant bone deficits that persisted after adjustment for height and weight and generally did not correlate with degree of cholestasis. In ALGS, low BMD and BMC reference Z scores were explained by poor growth. Anthropometrically adjusted DXA measures in ALGS correlate with markers of cholestasis and bone fracture history. Reduced bone density in this population is multifactorial and related to growth, degree of cholestasis, fracture vulnerability, and contribution of underlying genetic etiology.

OBJECTIVES To advance our understanding of monogenic forms of intrahepatic cholestasis. METHODS Analyses included participants with pathogenic biallelic mutations in adenosine triphosphate (ATP)-binding cassette subfamily B member 11 (ABCB11) (bile salt export pump; BSEP) or adenosine triphosphatase (ATPase) phospholipid transporting 8B1 (ATP8B1) (familial intrahepatic cholestasis; FIC1), or those with monoallelic or biallelic mutations in adenosine triphosphate (ATP)-binding cassette subfamily B member 4 (ABCB4) (multidrug resistance; MDR3), prospectively enrolled in the Longitudinal Study of Genetic Causes of Intrahepatic Cholestasis (LOGIC; NCT00571272) between November 2007 and December 2013. Summary statistics were calculated to describe baseline demographics, history, anthropometrics, laboratory values, and mutation data. RESULTS Ninety-eight participants with FIC1 (n = 26), BSEP (n = 53, including 8 with biallelic truncating mutations [severe] and 10 with p.E297G or p.D482G [mild]), or MDR3 (n = 19, including four monoallelic) deficiency were analyzed. Thirty-five had a surgical interruption of the enterohepatic circulation (sEHC), including 10 who underwent liver transplant (LT) after sEHC. Onset of symptoms occurred by age 2 years in most with FIC1 and BSEP deficiency, but was later and more variable for MDR3. Pruritus was nearly universal in FIC1 and BSEP deficiency. In participants with native liver, failure to thrive was common in FIC1 deficiency, high ALT was common in BSEP deficiency, and thrombocytopenia was common in MDR3 deficiency. sEHC was successful after more than 1 year in 7 of 19 participants with FIC1 and BSEP deficiency. History of LT was most common in BSEP deficiency. Of 102 mutations identified, 43 were not previously reported. CONCLUSIONS In this cohort, BSEP deficiency appears to be correlated with a more severe disease course. Genotype-phenotype correlations in these diseases are not straightforward and will require the study of larger cohorts.

Background and aims Many children with primary sclerosing cholangitis (PSC) receive oral vancomycin therapy (OVT) or ursodeoxycholic acid (UDCA). There is a paucity of data on whether these medications improve outcomes. Approach and results We analyzed retrospective data from the Pediatric PSC Consortium. Children treated with OVT were matched 1:1:1 to those treated with UDCA or managed with observation (no treatment) based on the closest propensity score, ensuring similar baseline characteristics. Two hundred sixty-four patients (88 each with OVT, UDCA, or observation) had matching propensity scores and were similar in demographics, phenotype, immunosuppression, baseline biochemistry, and hepatic fibrosis. After 1 year in an intention-to-treat analysis, all outcome metrics were similar regardless of treatment group. In OVT, UDCA, and untreated groups, respectively: Gamma-glutamyltransferase normalized in 53%, 49%, and 52% (P = not significant [NS]), liver fibrosis stage was improved in 20%, 13%, and 18% and worsened in 11%, 29%, and 18% (P = NS), and the 5-year probability of liver transplant listing was 21%, 10%, and 12% (P = NS). Favorable outcome was associated with having a mild phenotype of PSC and minimal hepatic fibrosis. Conclusions We presented the largest-ever description of outcomes on OVT in PSC and compared them to carefully matched patients on UDCA or no therapy. Neither OVT nor UDCA showed improvement in outcomes compared to a strategy of observation. Patients progressed to end-stage liver disease at similar rates. Spontaneous normalization of biochemistry is common in children receiving no therapy, particularly in the majority of children with a mild phenotype and an early stage of disease. Placebo-controlled treatment trials are needed to identify effective treatments for pediatric PSC.

Background and Aims: Disease progression in children with primary sclerosing cholangitis (PSC) is variable. Prognostic and risk-stratification tools exist for adult-onset PSC, but not for children. We aimed to create a tool that accounts for the biochemical and phenotypic features and early disease stage of pediatric PSC. Approach and Results: We used retrospective data from the Pediatric PSC Consortium. The training cohort contained 1,012 patients from 40 centers. We generated a multivariate risk index (Sclerosing Cholangitis Outcomes in Pediatrics [SCOPE] index) that contained total bilirubin, albumin, platelet count, gamma glutamyltransferase, and cholangiography to predict a primary outcome of liver transplantation or death (TD) and a broader secondary outcome that included portal hypertensive, biliary, and cancer complications termed hepatobiliary complications (HBCs). The model stratified patients as low, medium, or high risk based on progression to TD at rates of

TRMU is a nuclear gene crucial for mitochondrial DNA translation by encoding tRNA 5-methylaminomethyl-2-thiouridylate methyltransferase, which thiolates mitochondrial tRNA. Biallelic pathogenic variants in TRMU are associated with transient infantile liver failure. Other less common presentations such as Leigh syndrome, myopathy, and cardiomyopathy have been reported. Recent studies suggested that provision of exogenous L-cysteine or N-acetylcysteine may ameliorate the effects of disease-causing variants and improve the natural history of the disease. Here, we report six infants with biallelic TRMU variants, including four previously unpublished patients, all treated with exogenous cysteine. We highlight the first report of an affected patient undergoing orthotopic liver transplantation, the long-term effects of cysteine supplementation, and the ability of the initial presentation to mimic multiple inborn errors of metabolism. We propose that TRMU deficiency should be suspected in all children presenting with persistent lactic acidosis and hypoglycemia, and that combined N-acetylcysteine and L-cysteine supplementation should be considered prior to molecular diagnosis, as this is a low-risk approach that may increase survival and mitigate the severity of the disease course.

OBJECTIVES: Most patients with primary sclerosing cholangitis (PSC) also have inflammatory bowel disease (IBD). The liver and colon express MAdCAM-1, a target of lymphocyte homing integrins. Vedolizumab (VDZ) is an α4β7 integrin antibody used to treat IBD. We investigated liver outcomes in children with PSC-IBD treated with VDZ. METHODS: Patients were identified within the Pediatric PSC Consortium, a multicenter research registry. Retrospective demographic, phenotypic, biochemical, radiological, histopathologic and IBD data for up to 1 year of VDZ therapy were collected. Liver biochemical and IBD responses were defined as: a 75% or greater reduction in initial γ-glutamyltransferase (GGT), or a GGT that fell to

Alagille syndrome (ALGS) is an autosomal dominant disorder attributed to mutations in the Notch signaling pathway. Children with ALGS are at increased risk for fragility fracture of unknown etiology. Our objective was to characterize bone mass, geometry, and microarchitecture in children with ALGS. This was a cross-sectional study of 10 children (9 females), ages 8-18 years, with a clinical diagnosis of ALGS. Bone density was assessed via DXA (Hologic Discovery A) at several skeletal regions. Tibia trabecular and cortical bone was assessed via pQCT (Stratec XCT 2000) at the distal 3% and 38% sites, respectively. Tibia bone microarchitecture was assessed via HR-pQCT (Scanco XtremeCT II) at an ultradistal site located at 4% of tibia length and a cortical site at 30% of tibia length. Z-scores were calculated for DXA and pQCT measures. In the absence of XtremeCT II HR-pQCT reference data, these outcome measures were descriptively compared to a sample of healthy children ages 5-20 years (n = 247). Anthropometrics and labs were also collected. Based on one-sample t-tests, mean Z-scores for height and weight (both p .05), were significantly less than zero. DXA bone Z-scores were not significantly different from zero, but were highly variable. For pQCT bone measures, Z-scores for total bone mineral content at the distal 3% site and cortical bone mineral content, cortical area, and cortical thickness at the distal 38% site were significantly less than zero (all p .05). There was good correspondence between pQCT measures of cortical thickness Z-scores and DXA Z-scores for aBMD at the whole body less head, 1/3 radius, and femoral neck (all p .05). Compared to healthy children, those with ALGS generally had lower trabecular number and greater trabecular separation despite having greater trabecular thickness (measured via HR-pQCT). Bilirubin and bile acids, markers of hepatic cholestasis, were associated with poorer bone measures. For example, greater bilirubin was associated with lower trabecular number (Spearman's rho [ρ] = -0.82, p = .023) and greater trabecular separation (ρ = 0.82, p = .023) measured via HR-pQCT, and greater bile acids were associated with lower cortical area measured via pQCT (ρ = -0.78, p = .041) and lower serum insulin-like growth factor-1 (ρ = -0.86, p = .002). In summary, deficits in cortical bone size and trabecular bone microarchitecture were evident in children with ALGS. Further investigation is needed to understand the factors contributing to these skeletal inadequacies, and the manner in which these deficits contribute to increased fracture risk.

Alagille syndrome (ALGS) is a multisystem autosomal dominant developmental disorder caused predominantly by pathogenic variants in JAGGED1 (JAG1), and also by pathogenic variants in NOTCH2 in a much smaller number of individuals. Clinical presentation is highly variable and includes liver, heart, eye, skeleton, and facial abnormalities, with a subset of individuals also presenting with kidney, vascular, and central nervous system phenotypes. Hepatocellular carcinoma (HCC) is a rare complication of ALGS, though little is known about its incidence or etiology among affected individuals. Previous reports have identified HCC occurrence in both pediatric and adult cases of ALGS. We present a case report of HCC in a 58‐year‐old woman with a pathogenic JAG1 variant and no overt hepatic features of ALGS. Through a comprehensive literature review, we compile all reported pediatric and adult cases, and further highlight one previously reported case of HCC onset in an adult ALGS patient without any hepatic disease features, similar to our own described patient. Our case report and literature review suggest that ALGS‐causing variants could confer risk for developing HCC regardless of phenotypic severity and highlight a need for a cancer screening protocol that would enable early detection and treatment in this at‐risk population.