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Background & Aims: The combination of atezolizumab and bevacizumab (atezo+bev) is the current standard of care for advanced hepatocellular carcinoma (HCC), providing a median overall survival (OS) of 19.2 months. Here, we aim to uncover the underlying cellular processes driving clinical benefit versus resistance to atezo+bev., Methods: We harnessed the power of single-cell RNA sequencing in advanced HCC to derive gene expression signatures recapitulating 21 cell phenotypes. These signatures were applied to 422 RNA-sequencing samples of advanced HCC patients treated with atezo+bev (n=317) versus atezolizumab (n=47) or sorafenib (n=58) as comparators., Results: We unveiled two distinct patterns of response to atezo+bev. First, an immune-mediated response characterized by the combined presence of CD8+ T effector cells and pro-inflammatory CXCL10+ macrophages, representing an immune rich microenvironment. Second, a non-immune, angiogenesis-related response distinguishable by a reduced expression of the VEGF co-receptor neuropilin-1 (NRP1), a biomarker that specifically predicts improved OS upon atezo+bev vs sorafenib (p = 0.039). Primary resistance was associated with an enrichment of immunosuppressive myeloid populations, namely CD14+ monocytes and TREM2+ macrophages, and Notch pathway activation. Based on these mechanistic insights we define "Immune-competent" and "Angiogenesis-driven" molecular subgroups, each associated with a significantly longer OS with atezo+bev versus sorafenib (p of interaction = 0.027), and a "Resistant" subset., Conclusion: Our study unveils two distinct molecular subsets of clinical benefit to atezolizumab plus bevacizumab in advanced HCC ("Immune-competent" and "Angiogenesis-driven") as well as the main traits of primary resistance to this therapy, thus providing a molecular framework to stratify patients based on clinical outcome and guiding potential strategies to overcome resistance., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: Therapeutic plasma exchange (PEX) has emerged as a potential treatment option for patients with acute liver failure (ALF). The effect of PEX on survival outcomes outside of clinical trials is not yet well established. In this study we aimed to evaluate the real-world use and outcomes of PEX for the treatment of ALF., Methods: This multicentre retrospective cohort study included consecutive patients with ALF admitted to all seven tertiary liver transplant centres in the UK between June 2013 and December 2021. Changes in clinical variables following PEX treatment were assessed, while overall survival and transplant-free survival up to hospital discharge in patients receiving PEX were compared to those receiving standard medical therapy Propensity score matching was performed to control for intergroup covariates and selection bias., Results: We included 378 patients with ALF (median [IQR] age 36 (28-48), 64% [n = 242] female) of whom 120 received PEX. There was a significant improvement in most clinical variables following PEX, including median dose of noradrenaline (reduction from 0.35 μg/kg/min [0.19-0.70 μg/kg/min] to 0.16 μg/kg/min [0.08-0.49], p = 0.001). There was no significant difference between PEX and standard medical therapy groups in overall survival (51.4% vs. 62.6%, respectively, p = 0.12) or transplant-free survival (42.6% vs. 53.1%, p = 0.24)., Conclusion: PEX is now frequently used in the management of patients with ALF in the UK. It is associated with significant improvement in haemodynamic parameters but not survival benefit., Impact and Implications: Therapeutic plasma exchange is frequently used in the management of patients with acute liver failure in the UK. This real-world study demonstrates significant improvement in haemodynamic status but has not confirmed the survival benefit seen in previous published literature. These results should help guide the future use of plasma exchange in this patient population., Competing Interests: Conflicts of interest RJ: has research collaborations with Yaqrit and Takeda. He is the inventor of OPA which has been patented by University College London and licensed to Mallinckrodt Pharma. He is also a founder of Yaqrit limited, a spin out company from University College London. He has also co-founded Hepyx Ltd. and Cyberliver Ltd. WB: personal fees from San Biotech and Flagship pioneering. SM has received fees from Norgine pharmaceuticals. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2024 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Patient-reported outcomes (PROs), such as health-related quality of life (HRQL), are important outcome measures for patients with chronic liver diseases (CLDs). Presence of cirrhosis and advanced liver disease have been associated with worsened HRQL and fatigue. On the other hand, some patients with earlier stages of CLD also experience fatigue, causing PRO impairment. Treatment for some CLDs may improve HRQL and, sometimes, levels of fatigue. We aimed to provide an in-depth expert review of concepts related to fatigue and HRQL in patients with primary biliary cholangitis, hepatitis C virus and MASLD (metabolic dysfunction-associated steatotic liver disease). A panel of experts in fatigue and CLD reviewed and discussed the literature and collaborated to provide this expert review of fatigue in CLD. Herein, we review and report on the complexity of fatigue, highlighting that it is comprised of peripheral (neuromuscular failure, often in conjunction with submaximal cardiorespiratory function) and central (central nervous system dysfunction) causes. Fatigue and HRQL are measured using validated self-report instruments. Additionally, fatigue can be measured through objective tests (e.g. grip strength). Fatigue has deleterious effects on HRQL and one's ability to be physically active and socially engaged but does not always correlate with CLD severity. Treatments for hepatitis C virus and MASLD can improve levels of fatigue and HRQL, but current treatments for primary biliary cholangitis do not seem to affect levels of fatigue. We conclude that obtaining PRO data, including on HRQL and fatigue, is essential for determining the comprehensive burden of CLD and its potential treatments., (Copyright © 2024 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Recurrent primary biliary cholangitis (rPBC) develops in approximately 30% of patients and negatively impacts graft and overall patient survival after liver transplantation (LT). There is a lack of data regarding the response rate to ursodeoxycholic acid (UDCA) in rPBC. We evaluated a large, international, multi-center cohort to assess the performance of PBC scores in predicting the risk of graft and overall survival after LT in patients with rPBC., Methods: A total of 332 patients with rPBC after LT were evaluated from 28 centers across Europe, North and South America. The median age at the time of rPBC was 58.0 years [IQR 53.2-62.6], and 298 patients (90%) were female. The biochemical response was measured with serum levels of alkaline phosphatase (ALP) and bilirubin, and Paris-2, GLOBE and UK-PBC scores at 1 year after UDCA initiation., Results: During a median follow-up of 8.7 years [IQR 4.3-12.9] after rPBC diagnosis, 52 patients (16%) had graft loss and 103 (31%) died. After 1 year of UDCA initiation the histological stage at rPBC (hazard ratio [HR] 3.97, 95% CI 1.36-11.55, p = 0.01), use of prednisone (HR 3.18, 95% CI 1.04-9.73, p = 0.04), ALP xULN (HR 1.59, 95% CI 1.26-2.01, p <0.001), Paris-2 criteria (HR 4.14, 95% CI 1.57-10.92, p = 0.004), GLOBE score (HR 2.82, 95% CI 1.71-4.66, p <0.001), and the UK-PBC score (HR 1.06, 95% CI 1.03-1.09, p <0.001) were associated with graft survival in the multivariate analysis. Similar results were observed for overall survival., Conclusion: Patients with rPBC and disease activity, as indicated by standard PBC risk scores, have impaired outcomes, supporting efforts to treat recurrent disease in similar ways to pre-transplant PBC., Impact and Implications: One in three people who undergo liver transplantation for primary biliary cholangitis develop recurrent disease in their new liver. Patients with recurrent primary biliary cholangitis and incomplete response to ursodeoxycholic acid, according to conventional prognostic scores, have worse clinical outcomes, with higher risk of graft loss and mortality in similar ways to the disease before liver transplantation. Our results supportsupport efforts to treat recurrent disease in similar ways to pre-transplant primary biliary cholangitis., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Strategies to reduce liver biopsy (LB) screen failures through better patient selection are needed for clinical trials. Standard fibrosis biomarkers were not derived to detect "at-risk" metabolic dysfunction-associated steatohepatitis (MASH; MASH with metabolic dysfunction-associated steatotic liver disease score ≥4 and fibrosis stage ≥2). We compared the performance of screening pathways that incorporate NIS2+™, an optimized version of the blood-based NIS4® technology designed to identify at-risk MASH, with those incorporating fibrosis (FIB)-4 within the RESOLVE-IT clinical trial (NCT02704403), aiming for optimized selection of patients for LB., Methods: A retrospective simulation analysis was conducted in the RESOLVE-IT screening pathway (RSP) cohort. LB failure rate (LBFR), number of patients needed to screen, and overall cost estimations of different pathways were calculated for a range of NIS2+™ and FIB-4 cut-offs and compared with those of the RSP, which relied on investigators' local practices. An analysis of potential recruitment bias based on histology, sex, age, or comorbidities was performed., Results: The analysis cohort included 1,929 patients, 765 (40%) with at-risk MASH. The NIS2+™ pathway resulted in a significantly lower LBFR (39%) compared with the FIB-4 pathway (58%) or the RSP (60%) when using cost-optimized cut-offs (NIS2+™, 0.53; FIB-4, 0.58). For every 1,000 inclusions, NIS2+™ significantly reduced unnecessary LBs (632 vs. 1,522; -58%) and screening costs (US$12.7 million vs. US$15.0 million) vs. the RSP, while the number of patients needed to screen increased moderately (3,220 to 4,033). NIS2+™ alone is better than FIB-4 alone or combined with FIB-4., Conclusions: This analysis demonstrated that patient selection for LB using NIS2+™ significantly reduced unnecessary biopsies and screening costs, which could greatly improve the feasibility of MASH clinical trials., Impact and Implications: Simple and accurate non-invasive strategies to optimize the selection of patients who should be referred for liver biopsy for inclusion in MASH clinical trials is critical to reduce the high liver biopsy failure rates. While the use of the Fibrosis-4 index alone did not lead to a significant improvement of the screening process, selecting patients using NIS2+™, a recently developed optimization of the NIS4® technology for the detection of at-risk MASH, showed improved performance by simultaneously reducing liver biopsy failure rates and the overall cost of the trial, while maintaining the number of patients needed to screen at a manageable level and not generating any bias in included patients' characteristics. This makes NIS2+™ an accurate and reliable screening tool that could improve the recruitment of patients in future MASH clinical trials, and would lead to increased patient comfort and security, ensuring timely and cost-efficient trial completion., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

The principal limitations of the terms NAFLD and NASH are the reliance on exclusionary confounder terms and the use of potentially stigmatising language. This study set out to determine if content experts and patient advocates were in favour of a change in nomenclature and/or definition. A modified Delphi process was led by three large pan-national liver associations. The consensus was defined a priori as a supermajority (67%) vote. An independent committee of experts external to the nomenclature process made the final recommendation on the acronym and its diagnostic criteria. A total of 236 panellists from 56 countries participated in 4 online surveys and 2 hybrid meetings. Response rates across the 4 survey rounds were 87%, 83%, 83%, and 78%, respectively. Seventy-four percent of respondents felt that the current nomenclature was sufficiently flawed to consider a name change. The terms "nonalcoholic" and "fatty" were felt to be stigmatising by 61% and 66% of respondents, respectively. Steatotic liver disease was chosen as an overarching term to encompass the various aetiologies of steatosis. The term steatohepatitis was felt to be an important pathophysiological concept that should be retained. The name chosen to replace NAFLD was metabolic dysfunction-associated steatotic liver disease (MASLD). There was consensus to change the definition to include the presence of at least 1 of 5 cardiometabolic risk factors. Those with no metabolic parameters and no known cause were deemed to have cryptogenic steatotic liver disease. A new category, outside pure metabolic dysfunction-associated steatotic liver disease, termed metabolic and alcohol related/associated liver disease (MetALD), was selected to describe those with metabolic dysfunction-associated steatotic liver disease, who consume greater amounts of alcohol per week (140-350 g/wk and 210-420 g/wk for females and males, respectively). The new nomenclature and diagnostic criteria are widely supported and non-stigmatising, and can improve awareness and patient identification., (Copyright © 2023 Mary E. Rinella, Jeffrey V. Lazarus, Vlad Ratziu, Sven M. Francque, Arun J. Sanyal, Fasiha Kanwal, Diana Romero, Manal F. Abdelmalek, Quentin M. Anstee, Juan Pablo Arab, Marco Arrese, Ramon Bataller, Ulrich Beuers, Jerome Boursier, Elisabetta Bugianesi, Christopher Byrne, Graciela E. Castro Narro, Abhijit Chowdhury, Helena Cortez-Pinto, Donna Cryer, Kenneth Cusi, Mohamed El-Kassas, Samuel Klein, Wayne Eskridge, Jiangao Fan, Samer Gawrieh, Cynthia D. Guy, Stephen A. Harrison, Seung Up Kim, Bart Koot, Marko Korenjak, Kris Kowdley, Florence Lacaille, Rohit Loomba, Robert Mitchell-Thain, Timothy R. Morgan, Elisabeth Powell, Michael Roden, Manuel Romero-Gómez, Marcelo Silva, Shivaram Prasad Singh, Silvia C. Sookoian, C. Wendy Spearman, Dina Tiniakos, Luca Valenti, Miriam B. Vos, Vincent Wai-Sun Wong, Stavra Xanthakos, Yusuf Yilmaz, Zobair Younossi, Ansley Hobbs, Marcela Villota-Rivas, Philip N. Newsome, NAFLD Nomenclature consensus group. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Obeticholic acid (OCA) is a first-in-class farnesoid X receptor agonist and antifibrotic agent in development for the treatment of pre-cirrhotic liver fibrosis due to non-alcoholic steatohepatitis (NASH). We aimed to validate the original 18-month liver biopsy analysis from the phase III REGENERATE trial of OCA for the treatment of NASH with a consensus panel analysis, provide additional histology data in a larger population, and evaluate safety from >8,000 total patient-years' exposure with nearly 1,000 participants receiving study drug for >4 years., Methods: Digitized whole-slide images were evaluated independently by panels of three pathologists using the NASH Clinical Research Network scoring system. Primary endpoints were (1) ≥1 stage improvement in fibrosis with no worsening of NASH or (2) NASH resolution with no worsening of fibrosis. Safety was assessed by laboratory values and adverse events., Results: Prespecified efficacy analyses included 931 participants. The proportion of participants achieving a ≥1 stage improvement in fibrosis with no worsening of NASH was 22.4% for OCA 25 mg vs. 9.6% for placebo (p <0.0001). More participants receiving OCA 25 mg vs. placebo achieved NASH resolution with no worsening of fibrosis (6.5% vs. 3.5%, respectively; p = 0.093). Histology data in a larger population of 1,607 participants supported these results. Safety data included 2,477 participants. The incidence of treatment-emergent adverse events (TEAEs), serious TEAEs, and deaths was not substantively different across treatment groups. Pruritus was the most common TEAE. Rates of adjudicated hepatic, renal, and cardiovascular events were low and similar across treatment groups., Conclusions: These results confirm the antifibrotic effect of OCA 25 mg. OCA was generally well tolerated over long-term dosing. These data support a positive benefit:risk profile in patients with pre-cirrhotic liver fibrosis due to NASH., Impact and Implications: Patients with non-alcoholic steatohepatitis (NASH) often have liver scarring (fibrosis), which causes an increased risk of liver-related illness and death. Preventing progression of fibrosis to cirrhosis or reversing fibrosis are the main goals of drug development for NASH. In this clinical trial of obeticholic acid (OCA) in patients with NASH (REGENERATE), we reaffirmed our previous results demonstrating that OCA was superior to placebo in improving fibrosis using a more rigorous consensus panel analysis of liver biopsies taken at month 18. We also showed that OCA treatment resulted in dose-dependent reductions of serum liver biochemistries and liver stiffness measurements compared with placebo, even in participants in whom histologic fibrosis did not change at 18 months, providing evidence that the benefit of OCA extends beyond what is captured by the ordinal NASH CRN scoring system. OCA was well tolerated with a favorable safety profile supporting a positive benefit: risk profile in patients with pre-cirrhotic liver fibrosis due to NASH., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Background & Aims: NIS4® is a blood-based non-invasive test designed to effectively rule in/rule out at-risk non-alcoholic steatohepatitis (NASH), defined as non-alcoholic fatty liver disease activity score ≥4 and significant fibrosis (stage ≥2), among patients with metabolic risk factors. Robustness of non-invasive test scores across characteristics of interest including age, type 2 diabetes mellitus, and sex, and optimised analytical aspects are critical for large-scale implementation in clinical practice. We developed and validated NIS2+™, an optimisation of NIS4®, specifically designed to improve score robustness., Methods: A well-balanced training cohort (n = 198) included patients from the GOLDEN-505 trial. The validation (n = 684) and test (n = 2,035) cohorts included patients from the RESOLVE-IT trial. Well-matched subgroups were created to avoid potential confounding effects during modelling and analysis of score robustness. Models were trained using logistic regressions for at-risk NASH detection and compared using Bayesian information criteria. Performance of NIS2+™ was compared with that of NIS4®, Fibrosis-4, and alanine aminotransferase using area under the receiver operating characteristic curve, and robustness was analysed through score distribution., Results: Using the training cohort to compare all combinations of NIS4® biomarkers, NIS2 (miR-34a-5p, YKL-40) was identified as the best combination of parameters. To correct for the sex effect on miR-34a-5p (validation cohort), sex and sex ∗ miR-34a-5p parameters were added, creating NIS2+™. In the test cohort, NIS2+™ exhibited a statistically higher area under the receiver operating characteristic curve (0.813) vs. NIS4® (0.792; p = 0.0002), Fibrosis-4 (0.653; p <0.0001), and alanine aminotransferase (0.699; p <0.0001). NIS2+™ scores were not affected by age, sex, BMI, or type 2 diabetes mellitus status, providing robust clinical performances irrespective of patient characteristics., Conclusion: NIS2+™ constitutes a robust optimisation of NIS4® technology for the detection of at-risk NASH., Impact and Implications: The development of non-invasive tests for accurate, large-scale detection of patients with at-risk non-alcoholic steatohepatitis (NASH; defined as NASH with non-alcoholic fatty liver disease activity score ≥4 and fibrosis stage ≥2) - who are at higher risk for disease progression and for developing liver-related life-threatening outcomes - is critical for identifying this patient population in the clinical setting and improving the screening process of NASH clinical trials. We report the development and validation of NIS2+™, a diagnostic test designed as an optimisation of NIS4® technology, a blood-based panel currently used to detect at-risk NASH in patients with metabolic risk factors. NIS2+™ showed improved performance for the detection of at-risk NASH compared with NIS4® and other non-invasive liver tests that was not impacted by patients' characteristics of interest, such as age, sex, type 2 diabetes mellitus, BMI, dyslipidaemia, and hypertension. This makes NIS2+™ a robust and reliable tool for the diagnosis of at-risk NASH among patients with metabolic risk factors, and an effective candidate for large-scale implementation in clinical practice and clinical trials., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: We conducted an individual patient data meta-analysis to establish stiffness cut-off values for magnetic resonance elastography (MRE) in staging liver fibrosis and to assess potential confounding factors., Methods: A systematic review of the literature identified studies reporting MRE data in patients with NAFLD. Data were obtained from the corresponding authors. The pooled diagnostic cut-off value for the various fibrosis stages was determined in a two-stage meta-analysis. Multilevel modelling methods were used to analyse potential confounding factors influencing the diagnostic accuracy of MRE in staging liver fibrosis., Results: Eight independent cohorts comprising 798 patients were included in the meta-analysis. The area under the receiver operating characteristic curve (AUROC) for MRE in detecting significant fibrosis was 0.92 (sensitivity, 79%; specificity, 89%). For advanced fibrosis, the AUROC was 0.92 (sensitivity, 87%; specificity, 88%). For cirrhosis, the AUROC was 0.94 (sensitivity, 88%, specificity, 89%). Cut-offs were defined to explore concordance between MRE and histopathology: ≥F2, 3.14 kPa (pretest probability, 39.4%); ≥F3, 3.53 kPa (pretest probability, 24.1%); and F4, 4.45 kPa (pretest probability, 8.7%). In generalized linear mixed model analysis, histological steatohepatitis with higher inflammatory activity (odds ratio 2.448, 95% CI 1.180-5.079, p <0.05) and high gamma-glutamyl transferase (GGT) concentration (>120U/L) (odds ratio 3.388, 95% CI 1.577-7.278, p <0.01] were significantly associated with elevated liver stiffness, and thus affecting accuracy in staging early fibrosis (F0-F1). Steatosis, as measured by magnetic resonance imaging proton density fat fraction, and body mass index(BMI) were not confounders., Conclusions: MRE has excellent diagnostic performance for significant, advanced fibrosis and cirrhosis in patients with NAFLD. Elevated inflammatory activity and GGT level may lead to overestimation of early liver fibrosis, but anthropometric measures such as BMI or the degree of steatosis do not., Impact and Implications: This individual patient data meta-analysis of eight international cohorts, including 798 patients, demonstrated that MRE achieves excellent diagnostic accuracy for significant, advanced fibrosis and cirrhosis in patients with NAFLD. Cut-off values (significant fibrosis, 3.14 kPa; advanced fibrosis, 3.53 kPa; and cirrhosis, 4.45 kPa) were established. Elevated inflammatory activity and gamma-glutamyltransferase level may affect the diagnostic accuracy of MRE, leading to overestimation of liver fibrosis in early stages. We observed no impact of diabetes, obesity, or any other metabolic disorder on the diagnostic accuracy of MRE., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: The progression of non-alcoholic steatohepatitis (NASH) to fibrosis and hepatocellular carcinoma (HCC) is aggravated by auto-aggressive T cells. The gut-liver axis contributes to NASH, but the mechanisms involved and the consequences for NASH-induced fibrosis and liver cancer remain unknown. We investigated the role of gastrointestinal B cells in the development of NASH, fibrosis and NASH-induced HCC., Methods: C57BL/6J wild-type (WT), B cell-deficient and different immunoglobulin-deficient or transgenic mice were fed distinct NASH-inducing diets or standard chow for 6 or 12 months, whereafter NASH, fibrosis, and NASH-induced HCC were assessed and analysed. Specific pathogen-free/germ-free WT and μMT mice (containing B cells only in the gastrointestinal tract) were fed a choline-deficient high-fat diet, and treated with an anti-CD20 antibody, whereafter NASH and fibrosis were assessed. Tissue biopsy samples from patients with simple steatosis, NASH and cirrhosis were analysed to correlate the secretion of immunoglobulins to clinicopathological features. Flow cytometry, immunohistochemistry and single-cell RNA-sequencing analysis were performed in liver and gastrointestinal tissue to characterise immune cells in mice and humans., Results: Activated intestinal B cells were increased in mouse and human NASH samples and licensed metabolic T-cell activation to induce NASH independently of antigen specificity and gut microbiota. Genetic or therapeutic depletion of systemic or gastrointestinal B cells prevented or reverted NASH and liver fibrosis. IgA secretion was necessary for fibrosis induction by activating CD11b+CCR2+F4/80+CD11c-FCGR1+ hepatic myeloid cells through an IgA-FcR signalling axis. Similarly, patients with NASH had increased numbers of activated intestinal B cells; additionally, we observed a positive correlation between IgA levels and activated FcRg+ hepatic myeloid cells, as well the extent of liver fibrosis., Conclusions: Intestinal B cells and the IgA-FcR signalling axis represent potential therapeutic targets for the treatment of NASH., Impact and Implications: There is currently no effective treatment for non-alcoholic steatohepatitis (NASH), which is associated with a substantial healthcare burden and is a growing risk factor for hepatocellular carcinoma (HCC). We have previously shown that NASH is an auto-aggressive condition aggravated, amongst others, by T cells. Therefore, we hypothesized that B cells might have a role in disease induction and progression. Our present work highlights that B cells have a dual role in NASH pathogenesis, being implicated in the activation of auto-aggressive T cells and the development of fibrosis via activation of monocyte-derived macrophages by secreted immunoglobulins (e.g., IgA). Furthermore, we show that the absence of B cells prevented HCC development. B cell-intrinsic signalling pathways, secreted immunoglobulins, and interactions of B cells with other immune cells are potential targets for combinatorial NASH therapies against inflammation and fibrosis., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Biomarkers have the potential to accelerate drug development, as early indicators of improved clinical response, to improve patient safety, and for personalised medicine. However, few have been approved through the biomarker qualification pathways of the regulatory agencies. This paper outlines how biomarkers can accelerate drug development, and reviews the lessons learned by the EU IMI2-funded LITMUS consortium, which has had several interactions with regulatory agencies in both the US and EU regarding biomarker qualification in patients with non-alcoholic fatty liver disease and non-alcoholic steatohepatitis. Sharing knowledge of such interactions with the scientific community is of paramount importance to increase the chances of qualification of relevant biomarkers that may accelerate drug development, and thereby help patients, across disease indications. A qualified biomarker enables a decision to be made that all understand and support in a common framework., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Non-alcoholic fatty liver disease is a condition that affects 25% of the population. Non-alcoholic steatohepatitis (NASH) is a progressive form of the disease that can lead to severe complications such as cirrhosis and hepatocellular carcinoma. Despite its high prevalence, no drugs are currently approved for the treatment of NASH. The drug development pipeline in NASH is very active, yet most assets do not progress to phase III trials and those that do reach phase III often fail to achieve the endpoints necessary for approval by regulatory agencies. Amongst other reasons, the methodological and operational features of traditional clinical trials in NASH might impede optimal drug development. In this regard, platform trials might be an attractive complement or alternative to conventional clinical trials. Platform trials use a master protocol which enables evaluation of multiple investigational medicinal products concurrently or sequentially with a single, shared control arm. Through Bayesian interim analyses, these trials allow for early exit of drugs from the trial based on success or futility, while providing participants better chances of receiving active compounds through adaptive randomisation. Overall, platform trials represent an alternative for patients, pharmaceutical companies, and clinicians in the quest to accelerate the approval of pharmacologic treatments for NASH., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: Liver stiffness measurement (LSM) by vibration-controlled transient elastography (VCTE) has been shown to predict outcomes of patients with primary biliary cholangitis (PBC) in small-size studies. We aimed to validate the prognostic value of LSM in a large cohort study., Methods: We performed an international, multicentre, retrospective follow-up study of 3,985 patients with PBC seen at 23 centres in 12 countries. Eligibility criteria included at least 1 reliable LSM by VCTE and a follow-up ≥ 1 year. Independent derivation (n = 2,740) and validation (n = 568) cohorts were built. The primary endpoint was time to poor clinical outcomes defined as liver-related complications, liver transplantation, or death. Hazard ratios (HRs) with CIs were determined using a time-dependent multivariable Cox regression analysis., Results: LSM was independently associated with poor clinical outcomes in the derivation (5,324 LSMs, mean follow-up 5.0 ± 3.1 years) and validation (1,470 LSMs, mean follow-up 5.0 ± 2.8 years) cohorts: adjusted HRs (95% CI) per additional kPa were 1.040 (1.026-1.054) and 1.042 (1.029-1.056), respectively (p <0.0001 for both). Adjusted C-statistics (95% CI) at baseline were 0.83 (0.79-0.87) and 0.92 (0.89-0.95), respectively. Between 5 and 30 kPa, the log-HR increased as a monotonic function of LSM. The predictive value of LSM was stable in time. LSM improved the prognostic ability of biochemical response criteria, fibrosis scores, and prognostic scores. The 8 kPa and 15 kPa cut-offs optimally separated low-, medium-, and high-risk groups. Forty percent of patients were at medium to high risk according to LSM., Conclusions: LSM by VCTE is a major, independent, validated predictor of PBC outcome. Its value as a surrogate endpoint for clinical benefit in PBC should be considered., Lay Summary: Primary biliary cholangitis (PBC) is a chronic autoimmune disease, wherein the body's immune system mistakenly attacks the bile ducts. PBC progresses gradually, so surrogate markers (markers that predict clinically relevant outcomes like the need for a transplant or death long before the event occurs) are often needed to expedite the drug development and approval process. Herein, we show that liver stiffness measurement is a strong predictor of clinical outcomes and could be a useful surrogate endpoint in PBC trials., Competing Interests: Conflict of interest Dr. Corpechot reports receiving grants from Arrow and Intercept France, consulting fees from Intercept France, Inventiva Pharma, Cymabay and Genkyotex, and fees for teaching from Intercept France and GlaxoSmithKline France; Dr. Chazouillères, receiving grant support from Aptalis, fees for teaching from Mayoly Spindler, consulting fees from Genfit, and fees for teaching and consulting fees from Intercept; Dr. Schramm, receiving lecture fees from Falk Pharma; Dr. Dumortier, receiving consulting and teaching fees from Intercept France; Dr. Parés, receiving grant funding, speaking fees, and advisory board fees from Intercept, advisory board fees and speaking fees from Novartis, and speaking fees from CymaBay and Inova Diagnostics; Dr. Bruns reports receiving advisory board fees from Intercept, Grifols and Sobi and receiving speaking fees from Falk Foundation, Abbvie, CSL Behring, Intercept, Merck and Gilead. No other potential conflict of interest relevant to this article was reported. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Inflammation, particularly that mediated by bacterial components translocating from the gut to the liver and binding to toll-like receptors (TLRs), is central to cholestatic liver injury. The triggering receptor expressed on myeloid cells-2 (TREM-2) inhibits TLR-mediated signaling and exerts a protective role in hepatocellular injury and carcinogenesis. This study aims to evaluate the role of TREM-2 in cholestasis., Methods: TREM-2 expression was analyzed in the livers of patients with primary biliary cholangitis (PBC) or primary sclerosing cholangitis (PSC), and in mouse models of cholestasis. Wild-type (WT) and Trem-2 deficient (Trem-2 -/- ) mice were subjected to experimental cholestasis and gut sterilization. Primary cultured Kupffer cells were incubated with lipopolysaccharide and/or ursodeoxycholic acid (UDCA) and inflammatory responses were analyzed., Results: TREM-2 expression was upregulated in the livers of patients with PBC or PSC, and in murine models of cholestasis. Compared to WT, the response to bile duct ligation (BDL)-induced obstructive cholestasis or alpha-naphtylisothiocyanate (ANIT)-induced cholestasis was exacerbated in Trem-2 -/- mice. This was characterized by enhanced necroptotic cell death, inflammatory responses and biliary expansion. Antibiotic treatment partially abrogated the effects observed in Trem-2 -/- mice after BDL. Experimental overexpression of TREM-2 in the liver of WT mice downregulated ANIT-induced IL-33 expression and neutrophil recruitment. UDCA regulated Trem-1 and Trem-2 expression in primary cultured mouse Kupffer cells and dampened inflammatory gene transcription via a TREM-2-dependent mechanism., Conclusions: TREM-2 acts as a negative regulator of inflammation during cholestasis, representing a novel potential therapeutic target., Lay Summary: Cholestasis (the reduction or cessation of bile flow) causes liver injury. This injury is exacerbated when gut-derived bacterial components interact with receptors (specifically Toll-like receptors or TLRs) on liver-resident immune cells, promoting inflammation. Herein, we show that the anti-inflammatory receptor TREM-2 dampens TLR-mediated signaling and hence protects against cholestasis-induced liver injury. Thus, TREM-2 could be a potential therapeutic target in cholestasis., Competing Interests: Conflict of interest The authors declare no conflicts of interest that pertain to this work. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: Non-alcoholic fatty liver disease (NAFLD) is the leading cause of liver disorders and has a strong heritable component. The aim of this study was to identify new loci that contribute to severe NAFLD by examining rare variants., Methods: We performed whole-exome sequencing in individuals with NAFLD and advanced fibrosis or hepatocellular carcinoma (n = 301) and examined the enrichment of likely pathogenic rare variants vs. the general population. This was followed by validation at the gene level., Results: In patients with severe NAFLD, we observed an enrichment of the p.P426L variant (rs143545741 C>T; odds ratio [OR] 5.26, 95% CI 2.1-12.6; p = 0.003) of autophagy-related 7 (ATG7), which we characterized as a loss-of-function, vs. the general population, and an enrichment in rare variants affecting the catalytic domain (OR 13.9; 95% CI 1.9-612; p = 0.002). In the UK Biobank cohort, loss-of-function ATG7 variants increased the risk of cirrhosis and hepatocellular carcinoma (OR 3.30; 95% CI 1.1-7.5 and OR 12.30, 95% CI 2.6-36, respectively; p <0.001 for both). The low-frequency loss-of-function p.V471A variant (rs36117895 T>C) was also associated with severe NAFLD in the clinical cohort (OR 1.7; 95% CI 1.2-2.5; p = 0.003), predisposed to hepatocellular ballooning (p = 0.007) evolving to fibrosis in the Liver biopsy cohort (n = 2,268), and was associated with liver injury in the UK Biobank (aspartate aminotransferase levels, p <0.001), with a larger effect in severely obese individuals in whom it was linked to hepatocellular carcinoma (p = 0.009). ATG7 protein localized to periportal hepatocytes, particularly in the presence of ballooning. In the Liver Transcriptomic cohort (n = 125), ATG7 expression correlated with suppression of the TNFα pathway, which was conversely upregulated in p.V471A carriers., Conclusions: We identified rare and low-frequency ATG7 loss-of-function variants that promote NAFLD progression by impairing autophagy and facilitating ballooning and inflammation., Lay Summary: We found that rare mutations in a gene called autophagy-related 7 (ATG7) increase the risk of developing severe liver disease in individuals with dysmetabolism. These mutations cause an alteration in protein function and impairment of self-renewal of cellular content, leading to liver damage and inflammation., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest relevant to the present study. LV has received speaking fees from MSD, Gilead, AlfaSigma and AbbVie, served as a consultant for Gilead, Pfizer, AstraZeneca, Novo Nordisk, Intercept, Diatech Pharmacogenetics and Ionis Pharmaceuticals, and received research grants from Gilead. SR has served as a consultant for AstraZeneca, Celgene, Sanofi, Amgen, Akcea Therapeutics, Camp4, AMbys, Medacorp and Pfizer in the past 5 years, and received research grants from AstraZeneca, Sanofi and Amgen. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Background & Aims: We examined the efficacy and safety of seladelpar, a selective peroxisome proliferator-activated receptor-delta agonist, in adults with primary biliary cholangitis (PBC) at risk of disease progression (alkaline phosphatase [ALP] ≥1.67xupper limit of normal [ULN]) who were receiving or intolerant to ursodeoxycholic acid., Methods: In this 52-week, phase II, dose-ranging, open-label study, patients were randomized (1:1) to seladelpar 5 mg/day (n = 53) or 10 mg/day (n = 55) or assigned to 2 mg/day (n = 11; United Kingdom sites after interim analysis) for 12 weeks. Doses could then be uptitrated to 10 mg/day. The primary efficacy endpoint was ALP change from baseline to Week 8., Results: Mean baseline ALP was 300, 345, and 295 U/L in the 2 mg, 5 mg, and 10 mg cohorts, respectively. Twenty-one percent of patients had cirrhosis, 71% had pruritus. At Week 8, mean ± standard error ALP reductions from baseline were 26 ± 2.8%, 33 ± 2.6%, and 41 ± 1.8% in the 2 mg (n = 11), 5 mg (n = 49), and 10 mg (n = 52) cohorts (all p ≤0.005), respectively. Responses were maintained or improved at Week 52, after dose escalation in 91% and 80% of the 2 mg and 5 mg cohorts, respectively. At Week 52, composite response (ALP <1.67xULN, ≥15% ALP decrease, and normal total bilirubin) rates were 64%, 53%, and 67%, and ALP normalization rates were 9%, 13%, and 33% in the 2 mg, 5 mg, and 10 mg cohorts, respectively. Pruritus visual analog scale score was decreased in the 5 mg and 10 mg cohorts. There were no treatment-related serious adverse events, and 4 patients discontinued due to adverse events., Conclusions: Seladelpar demonstrated robust, dose-dependent, clinically significant, and durable improvements in biochemical markers of cholestasis and inflammation in patients with PBC at risk of disease progression. Seladelpar appeared safe and well tolerated and was not associated with any increase in pruritus., Gov Number: NCT02955602 CLINICALTRIALSREGISTER., Eu Number: 2016-002996-91 LAY SUMMARY: Current treatment options for patients living with primary biliary cholangitis (PBC) are not optimal due to inadequate effectiveness or undesirable side effects. Patients with PBC who took seladelpar, a new treatment being developed for PBC, at increasing doses (2, 5, or 10 mg/day) for 1 year had clinically significant, dose-dependent improvements in key liver tests. Treatment appeared safe and was not associated with any worsening in patient self-reported itch scores., Competing Interests: Conflict of interest Christopher Bowlus has received grants from Arena Pharmaceuticals, Cara Therapeutics, Genfit, Genkyotex, and Novartis and grants and personal fees from CymaBay Therapeutics, Eli Lilly, Gilead, GlaxoSmithKline, and Intercept. Michael Galambos has received consulting and investigator fees from CymaBay Therapeutics. Richard Aspinall owns stock in CymaBay Therapeutics. Gideon M. Hirschfield has received personal fees from CymaBay Therapeutics, Genfit, GlaxoSmithKline, HighTide, Intercept Pharma, Mirum, and Pliant. David E. Jones has received personal fees from Abbott and Falk and personal fees and a grant from Intercept. Stuart C. Gordon has received personal fees and advisory board compensation from CymaBay Therapeutics and research support was paid to his institution by AbbVie, Brigham and Women’s Hospital, CymaBay Therapeutics, DURECT, Eiger, Genfit, Gilead, GlaxoSmithKline, Intercept, Merck, Pliant, Shire, and Viking. Stephen A. Harrison has received personal fees from AgonAB, Altimmune, Boston Pharmaceuticals, B Riley FBR, Canfite Biopharma, Echosens North America, Fibronostics, Foresite Labs, Fortress Biotech, GNS Healthcare, Histoindex Pte, Inipharm, Ionis, Kowa Research Institute, Medpace, Microba Pty, Nutrasource, Piper Sandler & Co, Prometic Pharma Smt, Ridgeline Therapeutics, Sonic Incytes Medical Corporation, and Terns; grants from Cirius Therapeutics, Galmed Research & Development, Hepion Pharmaceuticals, and Pathai; personal fees and grants from Akero Therapeutics, Axcella Health, Civi Biopharma, CymaBay Therapeutics, Enyo Pharma S.A., Galectin Therapeutics, Genfit, Gilead Sciences, High Tide, Intercept, Madrigal Pharmaceuticals, Metracrine, NGM Biopharmaceuticals, Northsea Therapeutics B.V., Novartis, Novo Nordisk, Poxel, Sagimet Biosciences, and Viking Therapeutics; advisory board compensation from 89Bio, Akero Therapeutics, Altimmune, Arrowhead Pharmaceuticals, Axcella Health, Civi Biopharma, CymaBay Therapeutics, Echosens North America, Foresite Labs, Galectin Therapeutics, Galmed Research & Development, Genfit, Gilead, Hepion Pharmaceuticals, High Tide, Histoindex Pte, Indalo Therapeutics, Intercept, Madrigal Pharmaceuticals, Medpace, Metacrine, NGM Biopharmaceuticals, Northsea Therapeutics, Novartis, Novo Nordisk, Poxel, Prometic Pharma Smt, Ridgeline Therapeutics, Sagimet Biosciences, Terns, and Theratechnologies; and owns stock in Akero Therapeutics, Cirius Therapeutics, Galectin Therapeutics, Genfit, Hepion Pharmaceuticals, Histoindex Pte, Metacrine, NGM Biopharmaceuticals, Northsea Therapeutics B.V., and Pathai. Andreas E. Kremer has received personal fees from AbbVie, Bayer, CymaBay Therapeutics, Eisai, Eli Lilly, Escient, Falk, FMC, Gilead, GlaxoSmithKline, Mirum, MSD, Myr, Newbridge, Novartis, and Zambon and personal fees and a grant from Intercept. Marlyn J. Mayo has received consulting and investigator fees from CymaBay Therapeutics. Paul J. Thuluvath has received a grant from CymaBay Therapeutics. Cynthia Levy has received grants from Genkyotex, Gilead, High Tide, Intercept, Novartis, and Zydus and grants and personal fees from Cara Therapeutics, CymaBay Therapeutics, Genfit, and GlaxoSmithKline. Mark Swain has received grants from CymaBay Therapeutics and Genfit and a grant and personal fees from Intercept. Yvonne Dörffel, Guy Neff, David Sheridan, Carmen Stanca, Christoph Berg, and Aparna Goel have nothing to disclose. Mitchell L. Shiffman has received grants from CymaBay Therapeutics, High Tide, and Genfit and grants and personal fees from Intercept. John M. Vierling has received grants from Gilead, GlaxoSmithKline, and Intercept; personal fees from Intercept and Novartis; and personal fees and grants from CymaBay Therapeutics. Pol Boudes is a former employee of CymaBay Therapeutics who owns stock in CymaBay Therapeutics and holds a patent broadly relevant to this work. Alexandra Steinberg is a former employee of CymaBay Therapeutics who owns stock in CymaBay Therapeutics. Yun-Jung Choi is an employee of and owns stock in CymaBay Therapeutics. Charles McWherter is an employee of and owns stock in CymaBay Therapeutics and holds patents broadly relevant to this work. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

HCV hijacks many host metabolic processes in an effort to aid viral replication. The resulting hepatic metabolic dysfunction underpins many of the hepatic and extrahepatic manifestations of chronic hepatitis C (CHC). However, the natural history of CHC is also substantially influenced by the host metabolic status: obesity, insulin resistance and hepatic steatosis are major determinants of CHC progression toward hepatocellular carcinoma (HCC). Direct-acting antivirals (DAAs) have transformed the treatment and natural history of CHC. While DAA therapy effectively eradicates the virus, the long-lasting overlapping metabolic disease can persist, especially in the presence of obesity, increasing the risk of liver disease progression. This review covers the mechanisms by which HCV tunes hepatic and systemic metabolism, highlighting how systemic metabolic disturbance, lipotoxicity and chronic inflammation favour disease progression and a precancerous niche. We also highlight the therapeutic implications of sustained metabolic dysfunction following sustained virologic response as well as considerations for patients who develop HCC on the background of metabolic dysfunction., Competing Interests: Conflicts of interest D.A.M. is a director of Fibrofind limited. J.L. and D.A.M. are shareholders in Fibrofind limited. The other authors report no conflicts of interest. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Over the last two decades, there have been tremendous advances in the non-invasive diagnosis and risk stratification of chronic liver diseases (CLDs). Non-invasive approaches are based on the quantification of biomarkers in serum samples or on the measurement of liver stiffness, using either ultrasound- or magnetic resonance-based elastography techniques. The fibrosis-4 index (non-patented) and enhanced liver fibrosis test (patented) are the most widely adopted serum markers, whereas vibration-controlled transient elastography is the most widely adopted elastography technique. In this review, we discuss the role of non-invasive tests in the current era, as well as their accuracy and how their use in clinical practice has changed the practice of hepatology, including identification of early cirrhosis in patients with risk factors for CLD, diagnosis of portal hypertension, establishing prognosis in compensated cirrhosis, guiding antiviral treatment, and screening for fibrosis and cirrhosis in primary care., Competing Interests: Conflicts of interest Quentin Anstee is Coordinator of the EU IMI-2 LITMUS consortium, which is funded by the EU Horizon 2020 programme and EFPIA. This multi-stakeholder consortium includes industry partners. He reports research Grant Funding: Allergan/Tobira, AstraZeneca, Boehringer-Ingelheim, GlaxoSmithKline, Glympse Bio, Intercept, Novartis Pharma AG, Pfizer Ltd. Consultancy: 89Bio, Abbvie/Allergan, Akero, Altimentiv, Altimmune, AstraZeneca, Axcella, Blade, BMS, BNN Cardio, Boehringer-Ingelheim, Cirius, CymaBay, EcoR1, E3Bio, Eli Lilly & Company Ltd., Galmed, Genentech, Genfit SA, Gilead, Grunthal, HistoIndex, Indalo, Intercept Pharma Europe Ltd., Inventiva, IQVIA, Janssen, Johnson & Johnson, Madrigal, MedImmune, Medpace, Merck, Metacrine, NGMBio, North Sea Therapeutics, Novartis, Novo Nordisk A/S, PathAI, Pfizer Ltd., Poxel, ProSciento, Raptor Pharma, Roche, Servier, Shionogi, Terns, The Medicines Company, Viking Therapeutics. Speaker: Abbott Laboratories, Allergan/Tobira, BMS, Clinical Care Options, Falk, Fishawack, Genfit SA, Gilead, Integritas Communications, Kenes, Medscape. Royalties: Elsevier Ltd. Laurent Castera has received consultancy fees from Alexion, Echosens, Gilead, Intercept, MSD, Novo Nordisk, and Pfizer, and lecture fees from Abbvie, Echosens, Gilead, Intercept and Novo Nordisk. Rohit Loomba serves as a consultant to Aardvark Therapeutics, Altimmune, Anylam/Regeneron, Amgen, Ar Pharmaceuticals, AstraZeneca, Bristol-Myer Squibb, CohBar, Eli Lilly, Galmed, Gilead, Glympse bio, Hightide, Inipharma, Intercept, Inventiva, Ionis, Janssen Inc., Madrigal, Metacrine, Inc., NGM Biopharmaceuticals, Novartis, Novo Nordisk, Merck, Pfizer, Sagimet, Theratechnologies, 89 bio, Terns Pharmaceuticals and Viking Therapeutics. In addition his institutions received research grants from Ar Pharmaceuticals, Astrazeneca, Boehringer-Ingelheim, Bristol-Myers Squibb, Eli Lilly, Galectin Therapeutics, Galmed Pharmaceuticals, Gilead, Intercept, Hanmi, Intercept, Inventiva, Ionis, Janssen, Madrigal Pharmaceuticals, Merck, NGM Biopharmaceuticals, Novo Nordisk, Merck, Pfizer, Sonic Incytes and Terns Pharmaceuticals. Co-founder of LipoNexus Inc. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Histologically assessed hepatocyte ballooning is a key feature discriminating non-alcoholic steatohepatitis (NASH) from steatosis (NAFL). Reliable identification underpins patient inclusion in clinical trials and serves as a key regulatory-approved surrogate endpoint for drug efficacy. High inter/intra-observer variation in ballooning measured using the NASH CRN semi-quantitative score has been reported yet no actionable solutions have been proposed., Methods: A focused evaluation of hepatocyte ballooning recognition was conducted. Digitized slides were evaluated by 9 internationally recognized expert liver pathologists on 2 separate occasions: each pathologist independently marked every ballooned hepatocyte and later provided an overall non-NASH NAFL/NASH assessment. Interobserver variation was assessed and a 'concordance atlas' of ballooned hepatocytes generated to train second harmonic generation/two-photon excitation fluorescence imaging-based artificial intelligence (AI)., Results: The Fleiss kappa statistic for overall interobserver agreement for presence/absence of ballooning was 0.197 (95% CI 0.094-0.300), rising to 0.362 (0.258-0.465) with a ≥5-cell threshold. However, the intraclass correlation coefficient for consistency was higher (0.718 [0.511-0.900]), indicating 'moderate' agreement on ballooning burden. 133 ballooned cells were identified using a ≥5/9 majority to train AI ballooning detection (AI-pathologist pairwise concordance 19-42%, comparable to inter-pathologist pairwise concordance of between 8-75%). AI quantified change in ballooned cell burden in response to therapy in a separate slide set., Conclusions: The substantial divergence in hepatocyte ballooning identified amongst expert hepatopathologists suggests that ballooning is a spectrum, too subjective for its presence or complete absence to be unequivocally determined as a trial endpoint. A concordance atlas may be used to train AI assistive technologies to reproducibly quantify ballooned hepatocytes that standardize assessment of therapeutic efficacy. This atlas serves as a reference standard for ongoing work to refine how ballooning is classified by both pathologists and AI., Lay Summary: For the first time, we show that, even amongst expert hepatopathologists, there is poor agreement regarding the number of ballooned hepatocytes seen on the same digitized histology images. This has important implications as the presence of ballooning is needed to establish the diagnosis of non-alcoholic steatohepatitis (NASH), and its unequivocal absence is one of the key requirements to show 'NASH resolution' to support drug efficacy in clinical trials. Artificial intelligence-based approaches may provide a more reliable way to assess the range of injury recorded as "hepatocyte ballooning"., Competing Interests: Conflicts of interest AJS is president of Sanyal Bio. He reports consultancy fees from Merck, Regeneron, Alnylam, Genentech, Amgen, Pfizer, Novo Nordisk, Eli Lilly, Boehringer Ingelhiem, Inventiva, Madrigal, Malinckrodt, Salix, Genfit, Hemoshear, Histoindex, Siemens, Gilead, NGM, Terns, Rivus, Endiva, 89Bio, Akero, Blade, Novartis, Axcella, Intercept; grant funding from Intercept, Gilead, Bristol Myers Squibb, Merck, Pfizer, Novo Nordisk, Astra Zeneca, Boehringer Ingelhiem, Eli Lilly, Viking, Madrigal, Akero, Hanmi; stock/stock options from Genfit, Tiziana, Indalo, Exhalenz, Hemoshear; royalties from Elsevier. AW reports fees from Clinnovate. BNT reports consultancy fees from Histoindex. CDG reports consultancy fees from HistoIndex, NGMBio, CymaBay and Madrigal. CDG has or has had consulting agreements with the following companies: HistoIndex, NGM, CymaBay, Madrigal and 89Bio. DGT reports consultancy for Intercept Pharmaceuticals Inc, Allergan plc, Verily Life Sciences LLC, Cirius Therapeutics Inc, Alimentiv Inc, Clinnovate Health UK Ltd, ICON Clinical Research Ltd and grants from Histoindex Pte Ltd. DT is an employee of, and holds stock in, Histoindex. EC and RY are employees of Histoindex. EMB has been on Advisory Board for Pfizer Ltd. And served as a consultant for Arrowhead, CymaBay, Intercept Pharmaceuticals, Medpace, Perspectum Diagnostics and Histoindex. Paid slide evaluation for CymaBay, Medpace and Perspectum Diagnostics. MR reports consultancy fees from Alnylam, Amgen, AMRA, BMS, Boehringer Ingelheim, Centara, Coherus, Enanta, Galecto, Intercept Pharmaceuticals, Madrigal, NGM, Biopharmaceuticals, Novo Nordisk, Pfizer, Fractyl, Gelesis, Siemens, Thetis, Terns, Rivus, 3vBio (Sagimet), 89Bio and Novartis, Immuron, Merck, Taiwan J. QMA is coordinator of the IMI2 LITMUS consortium, which is funded by the EU Horizon 2020 programme and EFPIA. This multi-stakeholder consortium includes industry partners. He reports research grant funding from Allergan/Tobira, AstraZeneca, GlaxoSmithKline, Glympse Bio, Novartis Pharma AG, Pfizer Ltd., Vertex; consultancy on behalf of Newcastle University for 89Bio, Allergan/Tobira, Altimmune, AstraZeneca, Axcella, Blade, BMS, BNN Cardio, Cirius, CymaBay, EcoR1, E3Bio, Eli Lilly & Company, Galmed, Genentech, Genfit, Gilead, Grunthal, HistoIndex, Indalo, Intercept, Inventiva, IQVIA, Janssen, Madrigal, MedImmune, Medpace, Metacrine, NGMBio, North Sea Therapeutics, Novartis, Novo Nordisk A/S, PathAI, Pfizer Ltd., Poxel, ProSciento, Raptor Pharma, Roche, Servier, Terns, The Medicines Company, Viking Therapeutics; and speaker fees from Abbott Laboratories, Allergan/Tobira, BMS, Clinical Care Options, Falk, Fishawack, Genfit SA, Gilead, Integritas Communications, Kenes, MedScape. SAH reports grants from Akero, Axcella, Cirius, CiVi, CymaBay, Enyo, Galectin, Galmed, Genfit, Gilead Sciences, Hepion, Hightide, Intercept, Madrigal, Metacrine, NGM, Northsea, Novartis, Novo Nordisk, Poxel, Sagimet, Viking; consultancy fees from AgomAB, Akero, Alentis, Alimentiv, Altimmune, Axcella, Boston Pharma, B Riley, BVF Partners, Canfite, CiVi, Corcept, CymaBay, Echosens, Enyo, Fibronostics, Foresite, Fortress, Galectin, Genfit, Gilead, GNS, Hepion, Hightide, HistoIndex, Inipharm, Intercept, Ionis, Kowa, Madrigal, Metacrine, Microba, NGM, NorthSea, Novartis, Novo Nordisk, Nutrasource, Piper Sandler, Poxel, Prometic Pharma, Sagimet, Sonic Incytes, Terns, Viking; and stock holdings or stock options in Akero, Chronwell, Cirius, Galectin, Genfit, Hepion, HistoIndex, Metacrine, NGM, NorthSea, PathAI, Sonic Incytes. VR reports consultancy fees from Novo-Nordisk, Intercept, NGM, Theratechnologies, Galmed, Madrigal, Hanmi, Astra-Zeneca. VT reports research grants from Gilead sciences and consulting fees from Novo-Nordisk, Intercept, NGM, Theratechnologies, Galmed, Madrigal, Hanmi, Astra-Zeneca. The other authors report no relevant conflicts of interest. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2022. Published by Elsevier B.V.)

Background & Aims: Obesity-associated inflammation is a key player in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). However, the role of macrophage scavenger receptor 1 (MSR1, CD204) remains incompletely understood., Methods: A total of 170 NAFLD liver biopsies were processed for transcriptomic analysis and correlated with clinicopathological features. Msr1 -/- and wild-type mice were subjected to a 16-week high-fat and high-cholesterol diet. Mice and ex vivo human liver slices were treated with a monoclonal antibody against MSR1. Genetic susceptibility was assessed using genome-wide association study data from 1,483 patients with NAFLD and 430,101 participants of the UK Biobank., Results: MSR1 expression was associated with the occurrence of hepatic lipid-laden foamy macrophages and correlated with the degree of steatosis and steatohepatitis in patients with NAFLD. Mice lacking Msr1 were protected against diet-induced metabolic disorder, showing fewer hepatic foamy macrophages, less hepatic inflammation, improved dyslipidaemia and glucose tolerance, and altered hepatic lipid metabolism. Upon induction by saturated fatty acids, MSR1 induced a pro-inflammatory response via the JNK signalling pathway. In vitro blockade of the receptor prevented the accumulation of lipids in primary macrophages which inhibited the switch towards a pro-inflammatory phenotype and the release of cytokines such as TNF-ɑ. Targeting MSR1 using monoclonal antibody therapy in an obesity-associated NAFLD mouse model and human liver slices resulted in the prevention of foamy macrophage formation and inflammation. Moreover, we identified that rs41505344, a polymorphism in the upstream transcriptional region of MSR1, was associated with altered serum triglycerides and aspartate aminotransferase levels in a cohort of over 400,000 patients., Conclusions: Taken together, our data suggest that MSR1 plays a critical role in lipid-induced inflammation and could thus be a potential therapeutic target for the treatment of NAFLD., Lay Summary: Non-alcoholic fatty liver disease (NAFLD) is a chronic disease primarily caused by excessive consumption of fat and sugar combined with a lack of exercise or a sedentary lifestyle. Herein, we show that the macrophage scavenger receptor MSR1, an innate immune receptor, mediates lipid uptake and accumulation in Kupffer cells, resulting in liver inflammation and thereby promoting the progression of NAFLD in humans and mice., Competing Interests: Conflict of interest The authors have no potential conflicts (financial, professional or personal) directly relevant to the manuscript. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Non-alcoholic fatty liver disease (NAFLD) is a highly prevalent, yet largely underappreciated liver condition which is closely associated with obesity and metabolic disease. Despite affecting an estimated 1 in 4 adults globally, NAFLD is largely absent on national and global health agendas., Methods: We collected data from 102 countries, accounting for 86% of the world population, on NAFLD policies, guidelines, civil society engagement, clinical management, and epidemiologic data. A preparedness index was developed by coding questions into 6 domains (policies, guidelines, civil awareness, epidemiology and data, NAFLD detection, and NAFLD care management) and categorising the responses as high, medium, and low; a multiple correspondence analysis was then applied., Results: The highest scoring countries were India (42.7) and the United Kingdom (40.0), with 32 countries (31%) scoring zero out of 100. For 5 of the domains a minority of countries were categorised as high-level while the majority were categorised as low-level. No country had a national or sub-national strategy for NAFLD and <2% of the different strategies for related conditions included any mention of NAFLD. National NAFLD clinical guidelines were present in only 32 countries., Conclusions: Although NAFLD is a pressing public health problem, no country was found to be well prepared to address it. There is a pressing need for strategies to address NAFLD at national and global levels., Lay Summary: Around a third of the countries scored a zero on the NAFLD policy preparedness index, with no country scoring over 50/100. Although NAFLD is a pressing public health problem, a comprehensive public health response is lacking in all 102 countries. Policies and strategies to address NAFLD at the national and global levels are urgently needed., Competing Interests: Conflict of interest HEM, MV-R, and AP report receiving consulting fees from the EASL International Liver Foundation related to this work. The EASL International Liver Foundation received grants from Intercept, Bristol-Myers-Squibb Company, Genfit, and MSD to support this study. The funders played no role in the conceptualisation, design, data collection, analysis, decision to publish, or preparation of the manuscript. All other authors declare no relevant completing interests. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: Non-alcoholic steatohepatitis (NASH) is a chronic, progressive fibrotic liver disease that can lead to cirrhosis. While liver biopsy is considered the reference standard for the histologic diagnosis of NASH and staging of fibrosis, its use in clinical practice is limited. Non-invasive tests (NITs) are increasingly being used to identify and stage liver fibrosis in patients with NASH, and several can assess liver-related outcomes. We report changes in various NITs in patients treated with obeticholic acid (OCA) or placebo in the phase III REGENERATE study., Methods: Patients with NASH and fibrosis stage F2 or F3 (n = 931) were randomized (1:1:1) to receive placebo, OCA 10 mg, or OCA 25 mg once daily. Various NITs based on clinical chemistry and/or imaging were evaluated at baseline and throughout the study., Results: Rapid, sustained reductions from baseline in alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutamyltransferase levels, as well as in Fibrosis-4 (FIB-4), FibroTest, FibroMeter, and FibroScan-AST scores were observed in OCA-treated vs. placebo-treated patients. Reduction in liver stiffness by vibration-controlled transient elastography was observed in the OCA 25 mg group vs. the placebo group at Month 18. NIT changes were associated with shifts in histologic fibrosis stage. The greatest improvements were observed in patients with ≥1-stage fibrosis improvement; however, improvements in ALT, AST, FIB-4, and FibroTest were also observed in OCA-treated patients whose histologic fibrosis remained stable., Conclusions: Based on the REGENERATE Month 18 interim analysis, rapid and sustained improvements in various NITs were observed with OCA treatment. Dynamic changes in selected NITs separated histologic responders from non-responders. These results suggest that NITs may be useful in assessing histologic response to OCA therapy. CLINICALTRIALS., Gov Number: NCT02548351 LAY SUMMARY: Non-alcoholic steatohepatitis (NASH) is a chronic, progressive liver disease that can lead to cirrhosis. To diagnose and assess liver fibrosis (scarring) in patients with NASH, non-invasive tests (NITs) are increasingly being used rather than liver biopsy, which is invasive, expensive, and can be risky. In the REGENERATE study, which is evaluating the effects of obeticholic acid vs. placebo in patients with NASH, various NITs were also evaluated. This analysis shows that improvements in levels of certain blood components, as well as favorable results of ultrasound imaging and proprietary tests of liver function, were associated with improvements in liver fibrosis after treatment with obeticholic acid, suggesting that NITs may be useful alternatives to liver biopsy in assessing NASH patients' response to therapy., Competing Interests: Conflict of interest MR was a consultant over the past 36 months for Alnylam, Amgen, AMRA, BMS, Boehringer Ingelheim, Centara, Coherus, Enanta, Galecto, Intercept Pharmaceuticals, Inc., Madrigal, NGM Biopharmaceuticals, Novo Nordisk, Pfizer, Fractyl, Gelesis, Siemens, Thetis, Terns, Rivus, 3vbio (Sagimet), 89Bio, and Novartis. She currently has no active consulting contracts. J-FD reports advisory committee participation for AbbVie, Bayer, Bristol-Myers Squibb, Falk, Genfit, Genkyotex, Gilead Sciences, HepaRegeniX, Intercept Pharmaceuticals, Inc., Lilly, Merck, Novartis; speaking and teaching fees from Bayer, Bristol-Myers Squibb, Intercept Pharmaceuticals, Inc., Genfit, Gilead Sciences, Novartis, Roche. QMA reports support from Intercept Pharmaceuticals, Inc. in relation to the present study; research funds as Coordinator of the EU IMI-2 LITMUS consortium, which is funded by the EU Horizon 2020 programme and EFPIA. This multi-stakeholder consortium includes industry partners, in addition, Allergan/Tobira, AstraZeneca, GlaxoSmithKline, Glympse Bio, Novartis Pharma AG, Pfizer Ltd; royalties or licenses from Elsevier Ltd; consulting fees from 89Bio, Allergan/Tobira, Altimmune, AstraZeneca, Axcella, Blade, BMS, BNN Cardio, Cirius, CymaBay, EcoR1, E3Bio, Eli Lilly & Company Ltd., Galmed, Genentech, Genfit SA, Gilead, Grunthal, HistoIndex, Indalo, Intercept Pharma Europe Ltd., Inventiva, IQVIA, Janssen, Madrigal, MedImmune, Medpace, Metacrine, NGMBio, North Sea Therapeutics, Novartis, Novo Nordisk A/S, PathAI, Pfizer Ltd., Poxel, ProSciento, Raptor Pharma, Roche, Servier, Terns, The Medicines Company, Viking Therapeutics; honoraria from Abbott Laboratories, Allergan/Tobira, BMS, Clinical Care Options, Falk, Fishawack, Genfit SA, Gilead, Integritas Communications, Kenes, Medscape; member of advisory board for Medpace (NorthSea Therapeutics). ZG has no conflicts to report. ZY reports consulting fees and research grants from Gilead Sciences, Intercept Pharmaceuticals, Inc., BMS, Novo Nordisk, Viking, Terns, Siemens, Quest, AbbVie, Madrigal, Merck, and Novartis. SAH reports consulting/advisory, stock ownership/equity from Akero, Galectin, Genfit, Madrigal, Metacrine, Histoindex and NGM Bio; grants from Axcella, Cirius, CymaBay, Galectin, Galmed, Genfit, Gilead, HighTide, Madrigal, NGM, Novartis, Novo Nordisk, Pfizer, and Second Genome; consulting/advisory for Alentis, Axcella, Bristol-Meyers Squibb, CiVi, CLDF, Corcept, Echosens, , Gilead, HighTide, Medpace, Novartis, Novo Nordisk, Perspectum, Prometric, Viking, Hepagene, Hepion, and Terns. RL reports research grants from Intercept Pharmaceuticals, Inc., Boehringer -Ingelheim, Bristol-Myers Squibb, Eli Lilly, Galectin Therapeutics, Galmed Pharmaceuticals, Genfit, Gilead, Inventiva, Janssen, Madrigal Pharmaceuticals, Merck, NGM Biopharmaceuticals, Pfizer, Siemens; consulting fees from Anylam/Regeneron, Amgen, Ar Pharmaceuticals, AstraZeneca, Bristol-Myers Squibb, CohBar, Eli Lilly, Galmed, Gilead, Glympse Bio, Inipharm, Intercept, Ionis, Janssen Inc., Madrigal, Metacrine, Inc., NGM Biopharmaceuticals, Novartis, Novo Nordisk, Pfizer, Sagimet, 89Bio, Viking Therapeutics; advisory committee participation for Intercept Pharmaceuticals, Inc.; co-founder of Liponexus, Inc. AJS is President of Sanyal Biotechnology; he reports stock options in Sanyal Bio, Exhalenz, Akarna, GenFit, Hemoshear, Durect, Indalo, Tiziana, Rivus; paid consultant to 89Bio, Albireo, Amgen, Ardelyx, Boehringer Ingelheim, Bird Rock, Bristol-Myers, Conatus, Covance, Echosens-Sandhill, ENYO, Genentech, General Electric, Genfit, Gilead, Hemoshear, HistoIndex, Inventiva, Janssen, Lilly, Madrigal, Mallinckrodt, Merck, NGM Bio, Nimbus, NorthSea, Novartis, Novo Nordisk, Owl, PathAI, Perspectum, Pfizer, Poxel, Prosciento, Regeneron, Rivus, Roche, Salix, Sanofi, Second Genome, Servier, Siemens, Takeda, Terns, Tiziana, Zydus; unpaid consultant to Immuron, Intercept Pharmaceuticals, Inc., Galectin, Sequana, Fractyl; Durect, Indalo, Allergan, Chemomab, Affimune, Teva, BASF, AMRA, Perspectum, Biocellvia; advisory board member for Immuron; royalties from Elsevier and UptoDate; grant support from Gilead, Mallinckrodt, Salix, Novartis, Galectin, Bristol-Myers, Sequana; receipt of equipment from Echosens-Sandhill; ongoing research collaboration with Echosens-Sandhill, Owl, Second Genome, Siemens; his institution has received grant support from Conatus, Gilead, Mallinckrodt, Boehringer Ingelheim, Novartis, Bristol-Myers, Merck, Lilly, Novo Nordisk, Fractyl, Madrigal, Inventiva, Covance; his institution has received fees from AstraZeneca. MB reports employment at and stock ownership in Intercept Pharmaceuticals, Inc. at the time of the study. AT reports employment at and stock ownership in Intercept Pharmaceuticals, Inc. MN reports employment at and stock ownership in Intercept Pharmaceuticals, Inc. at the time of the study. RS reports employment at and stock ownership in Intercept Pharmaceuticals, Inc. at the time of the study. TG reports employment at and stock ownership in Intercept Pharmaceuticals, Inc. at the time of the study. AV reports employment at Intercept Pharmaceuticals, Inc. at the time of the study. VR reports consulting fees from Galmed, Genfit, Madrigal, NGM, Bristol-Myers Squibb, Boehringer Ingelheim, Theratechnologies, Terns; advisory committee participation for Intercept Pharmaceuticals, Inc. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021. Published by Elsevier B.V.)

Background & Aims: Only a minority of excess alcohol drinkers develop cirrhosis. We developed and evaluated risk stratification scores to identify those at highest risk., Methods: Three cohorts (GenomALC-1: n = 1,690, GenomALC-2: n = 3,037, UK Biobank: relevant n = 6,898) with a history of heavy alcohol consumption (≥80 g/day (men), ≥50 g/day (women), for ≥10 years) were included. Cases were participants with alcohol-related cirrhosis. Controls had a history of similar alcohol consumption but no evidence of liver disease. Risk scores were computed from up to 8 genetic loci identified previously as associated with alcohol-related cirrhosis and 3 clinical risk factors. Score performance for the stratification of alcohol-related cirrhosis risk was assessed and compared across the alcohol-related liver disease spectrum, including hepatocellular carcinoma (HCC)., Results: A combination of 3 single nucleotide polymorphisms (SNPs) (PNPLA3:rs738409, SUGP1-TM6SF2:rs10401969, HSD17B13:rs6834314) and diabetes status best discriminated cirrhosis risk. The odds ratios (ORs) and (95% CIs) between the lowest (Q1) and highest (Q5) score quintiles of the 3-SNP score, based on independent allelic effect size estimates, were 5.99 (4.18-8.60) (GenomALC-1), 2.81 (2.03-3.89) (GenomALC-2), and 3.10 (2.32-4.14) (UK Biobank). Patients with diabetes and high risk scores had ORs of 14.7 (7.69-28.1) (GenomALC-1) and 17.1 (11.3-25.7) (UK Biobank) compared to those without diabetes and with low risk scores. Patients with cirrhosis and HCC had significantly higher mean risk scores than patients with cirrhosis alone (0.76 ± 0.06 vs. 0.61 ± 0.02, p = 0.007). Score performance was not significantly enhanced by information on additional genetic risk variants, body mass index or coffee consumption., Conclusions: A risk score based on 3 genetic risk variants and diabetes status enables the stratification of heavy drinkers based on their risk of cirrhosis, allowing for the provision of earlier preventative interventions., Lay Summary: Excessive chronic drinking leads to cirrhosis in some people, but so far there is no way to identify those at high risk of developing this debilitating disease. We developed a genetic risk score that can identify patients at high risk. The risk of cirrhosis is increased >10-fold with just two risk factors - diabetes and a high genetic risk score. Risk assessment using this test could enable the early and personalised management of this disease in high-risk patients., Competing Interests: Conflict of interest NPC has a number of consulting agreements with and research grants from the pharmaceutical industry but they are not significantly or directly related to this paper. MP receives research funding from various organisations including the MRC and NIHR. He has also received partnership funding for the following: MRC Clinical Pharmacology Training Scheme (co-funded by MRC and Roche, UCB, Eli Lilly and Novartis); a PhD studentship jointly funded by EPSRC and Astra Zeneca; and grant funding from Vistagen Therapeutics. He has also unrestricted educational grant support for the UK Pharmacogenetics and Stratified Medicine Network from Bristol-Myers Squibb and UCB. He has developed an HLA genotyping panel with MC Diagnostics, but does not benefit financially from this. He is part of the IMI Consortium ARDAT (www.ardat.org). None of these funding sources were deployed in the undertaking of this study. TRM has conducted clinical research with AbbVie, Genfit, Gilead, and Merck but none of these are related to this manuscript. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 European Association for the Study of the Liver. All rights reserved.)

Background and Aims: Vibration-controlled transient elastography (VCTE), point shear wave elastography (pSWE), 2-dimensional shear wave elastography (2DSWE), magnetic resonance elastography (MRE), and magnetic resonance imaging (MRI) have been proposed as non-invasive tests for patients with non-alcoholic fatty liver disease (NAFLD). This study evaluated their diagnostic accuracy for liver fibrosis and non-alcoholic steatohepatitis (NASH)., Methods: PubMED/MEDLINE, EMBASE and the Cochrane Library were searched for studies examining the diagnostic accuracy of these index tests, against histology as the reference standard, in adult patients with NAFLD. Two authors independently screened and assessed methodological quality of studies and extracted data. Summary estimates of sensitivity, specificity and area under the curve (sAUC) were calculated for fibrosis stages and NASH, using a random effects bivariate logit-normal model., Results: We included 82 studies (14,609 patients). Meta-analysis for diagnosing fibrosis stages was possible in 53 VCTE, 11 MRE, 12 pSWE and 4 2DSWE studies, and for diagnosing NASH in 4 MRE studies. sAUC for diagnosis of significant fibrosis were: 0.83 for VCTE, 0.91 for MRE, 0.86 for pSWE and 0.75 for 2DSWE. sAUC for diagnosis of advanced fibrosis were: 0.85 for VCTE, 0.92 for MRE, 0.89 for pSWE and 0.72 for 2DSWE. sAUC for diagnosis of cirrhosis were: 0.89 for VCTE, 0.90 for MRE, 0.90 for pSWE and 0.88 for 2DSWE. MRE had sAUC of 0.83 for diagnosis of NASH. Three (4%) studies reported intention-to-diagnose analyses and 15 (18%) studies reported diagnostic accuracy against pre-specified cut-offs., Conclusions: When elastography index tests are acquired successfully, they have acceptable diagnostic accuracy for advanced fibrosis and cirrhosis. The potential clinical impact of these index tests cannot be assessed fully as intention-to-diagnose analyses and validation of pre-specified thresholds are lacking., Lay Summary: Non-invasive tests that measure liver stiffness or use magnetic resonance imaging (MRI) have been suggested as alternatives to liver biopsy for assessing the severity of liver scarring (fibrosis) and fatty inflammation (steatohepatitis) in patients with non-alcoholic fatty liver disease (NAFLD). In this study, we summarise the results of previously published studies on how accurately these non-invasive tests can diagnose liver fibrosis and inflammation, using liver biopsy as the reference. We found that some techniques that measure liver stiffness had a good performance for the diagnosis of severe liver scarring., Competing Interests: Conflict of interest EAS, FEM, MHZ, YV, JAL, CKL, LAJY, NP, CHL, GPA and PMB have nothing to declare. ANAJ, SN and MP are shareholders of Perspectum Ltd. MRG reports grant funding from Gilead Sciences and Intercept; consultancy from Allergan, Gilead Science, Intercept, Medimmune, Shionogi, ProSciento, Kaleido, Siemens, Abbvie, Novo Nordisk, Genfit and Zydus. MJB and TAT are shareholders and employees of Pfizer. QMA is coordinator of the IMI2 LITMUS consortium and he reports research grant funding from Abbvie, Allergan/Tobira, AstraZeneca, GlaxoSmithKline, Glympse Bio, Novartis Pharma AG, Pfizer Ltd., Vertex; consultancy on behalf of Newcastle University for Abbott Laboratories, Acuitas Medical, Allergan/Tobira, Blade, BNN Cardio, Cirius, CymaBay, EcoR1, E3Bio, Eli Lilly & Company Ltd., Galmed, Genfit SA, Gilead, Grunthal, HistoIndex, Indalo, Imperial Innovations, Intercept Pharma Europe Ltd., Inventiva, IQVIA, Janssen, Kenes, Madrigal, MedImmune, Metacrine, NewGene, NGMBio, North Sea Therapeutics, Novartis, Novo Nordisk A/S, Pfizer Ltd., Poxel, ProSciento, Raptor Pharma, Servier, Viking Therapeutics; and speaker fees from Abbott Laboratories, Allergan/Tobira, BMS, Clinical Care Options, Falk, Fishawack, Genfit SA, Gilead, Integritas Communications, MedScape. SAH has research grants from Akero, Axcella, Cirius, CiVi Biopharma, Cymabay, Galectin, Galmed, Genfit, Gilead Sciences, Hepion Pharmaceuticals, Hightide Therapeutics, Intercept, Madrigal, Metacrine, NGM Bio, Northsea Therapeutics, Novartis, Novo Nordisk, Poxel, Sagimet, Viking. He has received consulting fees from Akero, Altimmune, Alentis, Arrowhead, Axcella, Canfite, Cirius, CiVi, Cymabay, Echosens, Enyo, Fibronostics, Foresite Labs, Fortress Biotech, Galectin, Genfit, Gilead Sciences, Hepion, HIghtide, HistoIndex, Intercept, Kowa, Madrigal, Metacrine, NGM, Northsea, Novartis, Novo Nordisk, Poxel, Prometic, Ridgeline, Sagimet, Terns, and Viking. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background & Aims: Non-invasive scoring systems (NSS) are used to identify patients with non-alcoholic fatty liver disease (NAFLD) who are at risk of advanced fibrosis, but their reliability in predicting long-term outcomes for hepatic/extrahepatic complications or death and their concordance in cross-sectional and longitudinal risk stratification remain uncertain., Methods: The most common NSS (NFS, FIB-4, BARD, APRI) and the Hepamet fibrosis score (HFS) were assessed in 1,173 European patients with NAFLD from tertiary centres. Performance for fibrosis risk stratification and for the prediction of long-term hepatic/extrahepatic events, hepatocarcinoma (HCC) and overall mortality were evaluated in terms of AUC and Harrell's c-index. For longitudinal data, NSS-based Cox proportional hazard models were trained on the whole cohort with repeated 5-fold cross-validation, sampling for testing from the 607 patients with all NSS available., Results: Cross-sectional analysis revealed HFS as the best performer for the identification of significant (F0-1 vs. F2-4, AUC = 0.758) and advanced (F0-2 vs. F3-4, AUC = 0.805) fibrosis, while NFS and FIB-4 showed the best performance for detecting histological cirrhosis (range AUCs 0.85-0.88). Considering longitudinal data (follow-up between 62 and 110 months), NFS and FIB-4 were the best at predicting liver-related events (c-indices>0.7), NFS for HCC (c-index = 0.9 on average), and FIB-4 and HFS for overall mortality (c-indices >0.8). All NSS showed limited performance (c-indices <0.7) for extrahepatic events., Conclusions: Overall, NFS, HFS and FIB-4 outperformed APRI and BARD for both cross-sectional identification of fibrosis and prediction of long-term outcomes, confirming that they are useful tools for the clinical management of patients with NAFLD at increased risk of fibrosis and liver-related complications or death., Lay Summary: Non-invasive scoring systems are increasingly being used in patients with non-alcoholic fatty liver disease to identify those at risk of advanced fibrosis and hence clinical complications. Herein, we compared various non-invasive scoring systems and identified those that were best at identifying risk, as well as those that were best for the prediction of long-term outcomes, such as liver-related events, liver cancer and death., Competing Interests: Conflict of interest The authors declare no conflicts of interest that pertain to this work. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background and Aims: Non-alcoholic steatohepatitis (NASH)-related hepatocellular carcinoma (HCC) is increasing globally, but its molecular features are not well defined. We aimed to identify unique molecular traits characterising NASH-HCC compared to other HCC aetiologies., Methods: We collected 80 NASH-HCC and 125 NASH samples from 5 institutions. Expression array (n = 53 NASH-HCC; n = 74 NASH) and whole exome sequencing (n = 52 NASH-HCC) data were compared to HCCs of other aetiologies (n = 184). Three NASH-HCC mouse models were analysed by RNA-seq/expression-array (n = 20). Activin A receptor type 2A (ACVR2A) was silenced in HCC cells and proliferation assessed by colorimetric and colony formation assays., Results: Mutational profiling of NASH-HCC tumours revealed TERT promoter (56%), CTNNB1 (28%), TP53 (18%) and ACVR2A (10%) as the most frequently mutated genes. ACVR2A mutation rates were higher in NASH-HCC than in other HCC aetiologies (10% vs. 3%, p <0.05). In vitro, ACVR2A silencing prompted a significant increase in cell proliferation in HCC cells. We identified a novel mutational signature (MutSig-NASH-HCC) significantly associated with NASH-HCC (16% vs. 2% in viral/alcohol-HCC, p = 0.03). Tumour mutational burden was higher in non-cirrhotic than in cirrhotic NASH-HCCs (1.45 vs. 0.94 mutations/megabase; p <0.0017). Compared to other aetiologies of HCC, NASH-HCCs were enriched in bile and fatty acid signalling, oxidative stress and inflammation, and presented a higher fraction of Wnt/TGF-β proliferation subclass tumours (42% vs. 26%, p = 0.01) and a lower prevalence of the CTNNB1 subclass. Compared to other aetiologies, NASH-HCC showed a significantly higher prevalence of an immunosuppressive cancer field. In 3 murine models of NASH-HCC, key features of human NASH-HCC were preserved., Conclusions: NASH-HCCs display unique molecular features including higher rates of ACVR2A mutations and the presence of a newly identified mutational signature., Lay Summary: The prevalence of hepatocellular carcinoma (HCC) associated with non-alcoholic steatohepatitis (NASH) is increasing globally, but its molecular traits are not well characterised. In this study, we uncovered higher rates of ACVR2A mutations (10%) - a potential tumour suppressor - and the presence of a novel mutational signature that characterises NASH-related HCC., Competing Interests: Conflict of interest J.M.L. receives research support from Bayer HealthCare Pharmaceuticals, Eisai Inc, Bristol-Myers Squibb, Boehringer-Ingelheim and Ipsen, and consulting fees from Eli Lilly, Bayer HealthCare Pharmaceuticals, Bristol-Myers Squibb, Eisai Inc, Celsion Corporation, Exelixis, Merck, Ipsen, Genentech, Roche, Glycotest, Leerink Swann LLC, Fortress Biotech, Nucleix, Can-Fite Biopharma, Sirtex, Mina Alpha Ltd and AstraZeneca. S.L.F consults for the following companies: 89 Bio, Amgen, Axcella Health, Blade Therapeutics, Bristol Myers Squibb, Can-Fite Biopharma, ChemomAb, Escient Pharmaceuticals, Forbion, Foresite laboratories, Galmed, Gordian Biotechnology, Glycotest, Glympse Bio, Hepgene, In sitro, Morphic Therapeutics, North Sea Therapeutics, Novartis, Ono Pharmaceuticals, Pfizer Pharmaceuticals, Scholar Rock Surrozen. He has stock options in the following companies: Blade Therapeutics, Escient, Galectin, Galmed, Genfit, Glympse, Hepgene, Lifemax, Metacrine, Morphic Therapeutics, Nimbus, North Sea Therapeutics, Scholar Rock, Surrozen. He receives research support from Morphic Therapeutics, Novo Nordisk, and Galmed, and has an SBIR grant with Abalone Bio. A.L. is a consultant for Neuwave and Histosonics. C.P.M.S.O. consults for Bayer, Novartis, Novonordisk, Allergan, Pfizer, Roche and Zambon. P.S. is receiving research grants from BMS, Roche, Incyte, Chugai and consulting fees from BMS, MSD, Incyte, Janssen, Roche, AstraZeneca and Amgen. H.W., P.T., and A.V.U. are or were salaried employees of Sema4 at the time of the study. H.W., P.T., and A.V.U. hold Sema4 stock options. B.M. received consultancy fees from Bayer-Shering Pharma, and speaker fees from Eisai and MSD. The rest of authors have nothing to disclose. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: The SALVE Histopathology Group (SHG) developed and validated a grading and staging system for the clinical and full histological spectrum of alcohol-related liver disease (ALD) and evaluated its prognostic utility in a multinational cohort of 445 patients., Methods: SALVE grade was described by semiquantitative scores for steatosis, activity (hepatocellular injury and lobular neutrophils) and cholestasis. The histological diagnosis of steatohepatitis due to ALD (histological ASH, hASH) was based on the presence of hepatocellular ballooning and lobular neutrophils. Fibrosis staging was adapted from the Clinical Research Network staging system for non-alcoholic fatty liver disease and the Laennec staging system and reflects the pattern and extent of ALD fibrosis. There are 7 SALVE fibrosis stages (SFS) ranging from no fibrosis to severe cirrhosis., Results: Interobserver κ-value for each grading and staging parameter was >0.6. In the whole study cohort, long-term outcome was associated with activity grade and cholestasis, as well as cirrhosis with very broad septa (severe cirrhosis) (p <0.001 for all parameters). In decompensated ALD, adverse short-term outcome was associated with activity grade, hASH and cholestasis (p = 0.038, 0.012 and 0.001, respectively), whereas in compensated ALD, hASH and severe fibrosis/cirrhosis were associated with decompensation-free survival (p = 0.011 and 0.001, respectively). On multivariable analysis, severe cirrhosis emerged as an independent histological predictor of long-term survival in the whole study cohort. Severe cirrhosis and hASH were identified as independent predictors of short-term survival in decompensated ALD, and also as independent predictors of decompensation-free survival in compensated ALD., Conclusion: The SALVE grading and staging system is a reproducible and prognostically relevant method for the histological assessment of disease activity and fibrosis in ALD., Lay Summary: Patients with alcohol-related liver disease (ALD) may undergo liver biopsy to assess disease severity. We developed a system to classify ALD under the microscope by grading ALD activity and staging the extent of liver scarring. We validated the prognostic performance of this system in 445 patients from 4 European centers., Competing Interests: Conflicts of interest The authors declare no conflicts of interest that pertain to this work. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Backgrounds & Aims: Primary biliary cholangitis (PBC) is a chronic liver disease in which autoimmune destruction of the small intrahepatic bile ducts eventually leads to cirrhosis. Many patients have inadequate response to licensed medications, motivating the search for novel therapies. Previous genome-wide association studies (GWAS) and meta-analyses (GWMA) of PBC have identified numerous risk loci for this condition, providing insight into its aetiology. We undertook the largest GWMA of PBC to date, aiming to identify additional risk loci and prioritise candidate genes for in silico drug efficacy screening., Methods: We combined new and existing genotype data for 10,516 cases and 20,772 controls from 5 European and 2 East Asian cohorts., Results: We identified 56 genome-wide significant loci (20 novel) including 46 in European, 13 in Asian, and 41 in combined cohorts; and a 57 th genome-wide significant locus (also novel) in conditional analysis of the European cohorts. Candidate genes at newly identified loci include FCRL3, INAVA, PRDM1, IRF7, CCR6, CD226, and IL12RB1, which each play key roles in immunity. Pathway analysis reiterated the likely importance of pattern recognition receptor and TNF signalling, JAK-STAT signalling, and differentiation of T helper (T H )1 and T H 17 cells in the pathogenesis of this disease. Drug efficacy screening identified several medications predicted to be therapeutic in PBC, some of which are well-established in the treatment of other autoimmune disorders., Conclusions: This study has identified additional risk loci for PBC, provided a hierarchy of agents that could be trialled in this condition, and emphasised the value of genetic and genomic approaches to drug discovery in complex disorders., Lay Summary: Primary biliary cholangitis (PBC) is a chronic liver disease that eventually leads to cirrhosis. In this study, we analysed genetic information from 10,516 people with PBC and 20,772 healthy individuals recruited in Canada, China, Italy, Japan, the UK, or the USA. We identified several genetic regions associated with PBC. Each of these regions contains several genes. For each region, we used diverse sources of evidence to help us choose the gene most likely to be involved in causing PBC. We used these 'candidate genes' to help us identify medications that are currently used for treatment of other conditions, which might also be useful for treatment of PBC., Competing Interests: Conflicts of interest GMH has consulted and/or been a speaker for Intercept, Genfit, Cymabay, GSK, and Falk. RNS and GFM have each received research funding from Intercept Pharmaceuticals. HJC, JJF, KU, RD, YA, YH, MK, NN, S-SK, OG, YK, MN, KT, RT, YS, ZL, BDJ, EJA, AG, MC, RA, AC, MdA, AB, JH, MARF, DS, DEJ, SF, AS, VLM, KNL, CIA, MFS, PI, KAS, XM and MN report no conflicts of interest. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021. Published by Elsevier B.V.)

Background & Aims: Patients with primary biliary cholangitis (PBC) who have an incomplete response to ursodeoxycholic acid remain at risk of disease progression. We investigated the safety and efficacy of elafibranor, a dual PPARα/δ agonist, in patients with PBC., Methods: This 12-week, double-blind phase II trial enrolled 45 adults with PBC who had incomplete response to ursodeoxycholic acid (alkaline phosphatase levels ≥1.67-fold the upper limit of normal (ULN). Patients were randomly assigned to elafibranor 80 mg, elafibranor 120 mg or placebo. The primary endpoint was the relative change of ALP at 12 weeks (NCT03124108)., Results: At 12 weeks, ALP was reduced by -48.3±14.8% in the elafibranor 80 mg group (p <0.001 vs. placebo) and by -40.6±17.4% in the elafibranor 120 mg group (p <0.001) compared to a +3.2±14.8% increase in the placebo group. The composite endpoint of ALP ≤1.67-fold the ULN, decrease of ALP >15% and total bilirubin below the ULN was achieved in 67% patients in the elafibranor 80 mg group and 79% patients in the elafibranor 120 mg group, vs. 6.7% patients in the placebo group. Levels of gamma-glutamyltransferase decreased by 37.0±25.5% in the elafibranor 80 mg group (p <0.001) and 40.0±24.1% in the elafibranor 120 mg group (p <0.01) compared to no change (+0.2±26.0%) in the placebo group. Levels of disease markers such as IgM, 5'-nucleotidase or high-sensitivity C-reactive protein were likewise reduced by elafibranor. Pruritus was not induced or exacerbated by elafibranor and patients with pruritus at baseline reported less pruritic symptoms at the end of treatment. All possibly drug-related non-serious adverse events were mild to moderate., Conclusion: In this randomized phase II trial, elafibranor was generally safe and well tolerated and significantly reduced levels of ALP, composite endpoints of bilirubin and ALP, as well as other markers of disease activity in patients with PBC and an incomplete response to ursodeoxycholic acid., Lay Summary: Patients with primary biliary cholangitis (a rare chronic liver disease) that do not respond to standard therapy remain at risk of disease progression toward cirrhosis and impaired quality of life. Elafibranor is a nuclear receptor agonist that we tested in a randomized clinical trial over 12 weeks. It successfully decreased levels of disease activity markers, including alkaline phosphatase. Thus, this study is the foundation for a larger prospective study that will determine the efficacy and safety of this drug as a second-line therapy., Clinical Trial Registration Number: Clinical Trials.gov NCT03124108., Competing Interests: Conflict of interest JMS reports consultancy: BMS, Boehringer Ingelheim, Echosens, Galmed, Genfit, Gilead Sciences, Intercept Pharmaceuticals, Madrigal, Nordic Bioscience, Novartis, Pfizer, Roche, Sanofi, Zydus. Research Funding: Gilead Sciences. AP has received grant funding, personal fees, and advisory board fees from Intercept Pharmaceuticals; advisory board fees and fees for teaching from Novartis; and personal fees from CymaBay Therapeutics and Inova Diagnostics. KVK: serves as consultant to or an advisory boards for Conatus, CymaBay, Gilead, Intercept, La Jolla, Merck and Novartis. He receives research support from Genfit, Gilead, High Tide, Intercept, NGM Biopharma and Novartis and serves as a speaker for Abbvie, Gilead Sciences and Intercept. MAH: Consultancy for Roche, Novartis, Falk and Intercept. SC received research support from Genfit, Gilead and Zydus. DP: nothing to disclose. AB: nothing to disclose. GMH has consulted for Intercept, Genfit, Novartis, GSK, Cymabay and Gilead. CL reports research grants: Gilead, Intercept, CymaBay, Genfit, Genkyotex, Enanta, GSK, Novartis, NGM, High Tide, Durect, Alnylam, Zydus, Cara Therapeutics, Target PharmaSolutions; Consulting fees/Advisory boards: CymaBay, GSK, Shire, Pliant, Target PharmaSolutions, Flashlight Therapeutics, Cara Therapeutics; Royalties: Up-to-date; other: Editorial board Liver Transplantation. JV Research Grants: Allergan, Arena, CymaBay, Enanta, Genkyotex, Intercept, Lilly, NGM Pharmaceuticals, Novartis, TaiwanJ, Scientific Advisor: Arena, BioIncept, Blade, CymaBay, Enanta, Genkyotex, Glaxo-Smith-Kline, Intercept, Lilly, Novartis, TaiwanJ, Authorship: Up-to-Date Immunosuppression in Liver Transplantation; AASLD Writing Committee AIH Guidance In Press, 2019. DJ reports consultancy and grant funding from Intercept and Consultancy from Novartis. AT has nothing to disclose. BS is consultant and president of the SAB of Genfit SA. SM was a former Genfit employee and has currently no COI. RH, DM and PB are Genfit employees. VL reports consultancy for Genfit., (Copyright © 2021 European Association for the Study of the Liver. All rights reserved.)

Background & Aims: Hepatocellular carcinoma (HCC) risk stratification in individuals with dysmetabolism is a major unmet need. Genetic predisposition contributes to non-alcoholic fatty liver disease (NAFLD). We aimed to exploit robust polygenic risk scores (PRS) that can be evaluated in the clinic to gain insight into the causal relationship between NAFLD and HCC, and to improve HCC risk stratification., Methods: We examined at-risk individuals (NAFLD cohort, n = 2,566; 226 with HCC; and a replication cohort of 427 German patients with NAFLD) and the general population (UK Biobank [UKBB] cohort, n = 364,048; 202 with HCC). Variants in PNPLA3-TM6SF2-GCKR-MBOAT7 were combined in a hepatic fat PRS (PRS-HFC), and then adjusted for HSD17B13 (PRS-5)., Results: In the NAFLD cohort, the adjusted impact of genetic risk variants on HCC was proportional to the predisposition to fatty liver (p = 0.002) with some heterogeneity in the effect. PRS predicted HCC more robustly than single variants (p <10 -13 ). The association between PRS and HCC was mainly mediated through severe fibrosis, but was independent of fibrosis in clinically relevant subgroups, and was also observed in those without severe fibrosis (p <0.05). In the UKBB cohort, PRS predicted HCC independently of classical risk factors and cirrhosis (p <10 -7 ). In the NAFLD cohort, we identified high PRS cut-offs (≥0.532/0.495 for PRS-HFC/PRS-5) that in the UKBB cohort detected HCC with ~90% specificity but limited sensitivity; PRS predicted HCC both in individuals with (p <10 -5 ) and without cirrhosis (p <0.05)., Conclusions: Our results are consistent with a causal relationship between hepatic fat and HCC. PRS improved the accuracy of HCC detection and may help stratify HCC risk in individuals with dysmetabolism, including those without severe liver fibrosis. Further studies are needed to validate our findings., Lay Summary: By analyzing variations in genes that contribute to fatty liver disease, we developed two risk scores to help predict liver cancer in individuals with obesity-related metabolic complications. These risk scores can be easily tested in the clinic. We showed that the risk scores helped to identify the risk of liver cancer both in high-risk individuals and in the general population., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest relevant to the present study. SR has served as a consultant for AstraZeneca, Celgene, Sanofi, Amgen, Akcea Therapeutics, Camp4, AMbys, Medacorp and Pfizer in the past 5 years, and received research grants from AstraZeneca, Sanofi and Amgen. LV has received speaking fees from MSD, Gilead, AlfaSigma and AbbVie, served as a consultant for Gilead, Pfizer, Astra Zeneca, Novo Nordisk, Intercept, Diatech Pharmacogenetics and Ionis Pharmaceuticals, and received research grants from Gilead. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2020 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: A common genetic variant near MBOAT7 (rs641738C>T) has been previously associated with hepatic fat and advanced histology in NAFLD; however, these findings have not been consistently replicated in the literature. We aimed to establish whether rs641738C>T is a risk factor across the spectrum of NAFLD and to characterise its role in the regulation of related metabolic phenotypes through a meta-analysis., Methods: We performed a meta-analysis of studies with data on the association between rs641738C>T genotype and liver fat, NAFLD histology, and serum alanine aminotransferase (ALT), lipids or insulin. These included directly genotyped studies and population-level data from genome-wide association studies (GWAS). We performed a random effects meta-analysis using recessive, additive and dominant genetic models., Results: Data from 1,066,175 participants (9,688 with liver biopsies) across 42 studies were included in the meta-analysis. rs641738C>T was associated with higher liver fat on CT/MRI (+0.03 standard deviations [95% CI 0.02-0.05], p z  = 4.8×10 -5 ) and diagnosis of NAFLD (odds ratio [OR] 1.17 [95% CI 1.05-1.3], p z  = 0.003) in Caucasian adults. The variant was also positively associated with presence of advanced fibrosis (OR 1.22 [95% CI 1.03-1.45], p z  = 0.021) in Caucasian adults using a recessive model of inheritance (CC + CT vs. TT). Meta-analysis of data from previous GWAS found the variant to be associated with higher ALT (p z  = 0.002) and lower serum triglycerides (p z  = 1.5×10 -4 ). rs641738C>T was not associated with fasting insulin and no effect was observed in children with NAFLD., Conclusions: Our study validates rs641738C>T near MBOAT7 as a risk factor for the presence and severity of NAFLD in individuals of European descent., Lay Summary: Fatty liver disease is a common condition where fat builds up in the liver, which can cause liver inflammation and scarring (including 'cirrhosis'). It is closely linked to obesity and diabetes, but some genes are also thought to be important. We did this study to see whether one specific change ('variant') in one gene ('MBOAT7') was linked to fatty liver disease. We took data from over 40 published studies and found that this variant near MBOAT7 is linked to more severe fatty liver disease. This means that drugs designed to work on MBOAT7 could be useful for treating fatty liver disease., Competing Interests: Conflict of interest C.E. reports receiving personal fees from Navitor Pharma and Novartis. The other authors declare no conflicts of interest that pertain to this work. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2020 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background & Aims: Genetic factors associated with non-alcoholic fatty liver disease (NAFLD) remain incompletely understood. To date, most genome-wide association studies (GWASs) have adopted radiologically assessed hepatic triglyceride content as the reference phenotype and so cannot address steatohepatitis or fibrosis. We describe a GWAS encompassing the full spectrum of histologically characterised NAFLD., Methods: The GWAS involved 1,483 European NAFLD cases and 17,781 genetically matched controls. A replication cohort of 559 NAFLD cases and 945 controls was genotyped to confirm signals showing genome-wide or close to genome-wide significance., Results: Case-control analysis identified signals showing p values ≤5 × 10 -8 at 4 locations (chromosome [chr] 2 GCKR/C2ORF16; chr4 HSD17B13; chr19 TM6SF2; chr22 PNPLA3) together with 2 other signals with p <1 × 10 -7 (chr1 near LEPR and chr8 near IDO2/TC1). Case-only analysis of quantitative traits showed that the PNPLA3 signal (rs738409) had genome-wide significance for steatosis, fibrosis and NAFLD activity score and a new signal (PYGO1 rs62021874) had close to genome-wide significance for steatosis (p = 8.2 × 10 -8 ). Subgroup case-control analysis for NASH confirmed the PNPLA3 signal. The chr1 LEPR single nucleotide polymorphism also showed genome-wide significance for this phenotype. Considering the subgroup with advanced fibrosis (≥F3), the signals on chr2, chr19 and chr22 maintained their genome-wide significance. Except for GCKR/C2ORF16, the genome-wide significance signals were replicated., Conclusions: This study confirms PNPLA3 as a risk factor for the full histological spectrum of NAFLD at genome-wide significance levels, with important contributions from TM6SF2 and HSD17B13. PYGO1 is a novel steatosis modifier, suggesting that Wnt signalling pathways may be relevant in NAFLD pathogenesis., Lay Summary: Non-alcoholic fatty liver disease is a common disease where excessive fat accumulates in the liver and may result in cirrhosis. To understand who is at risk of developing this disease and suffering liver damage, we undertook a genetic study to compare the genetic profiles of people suffering from fatty liver disease with genetic profiles seen in the general population. We found that particular sequences in 4 different areas of the human genome were seen at different frequencies in the fatty liver disease cases. These sequences may help predict an individual's risk of developing advanced disease. Some genes where these sequences are located may also be good targets for future drug treatments., Competing Interests: Conflict of interest Quentin Anstee reports grants from European Commission during the conduct of the study; other from Acuitas Medical, grants, personal fees and other from Allergan/Tobira, other from E3Bio, other from Eli Lilly & Company Ltd, other from Galmed, grants, personal fees and other from Genfit SA, personal fees and other from Gilead, other from Grunthal, other from Imperial Innovations, grants and other from Intercept Pharma Europe Ltd, other from Inventiva, other from Janssen, personal fees from Kenes, other from MedImmune, other from NewGene, grants and other from Pfizer Ltd, other from Raptor Pharma, grants from GlaxoSmithKline, grants and other from Novartis Pharma AG, grants from AbbVie, personal fees from BMS, grants from GSK, other from NGMBio, other from Madrigal, other from Servier, outside the submitted work; Dina Tiniakos reports consultation fees from Intercept Pharmaceuticals Inc, Allergan, Cirius Therapeutics and an educational grant from Histoindex Pte Ltd; Guruprasad P. Aithal reports institutional consultancy income outside the scope of this study from GSK and Pfizer; Michael Allison reports consultancy/advisory with MedImmune/Astra Zeneca, E3Bio, honoraria from Intercept, Grant support from GSK, Takeda; Jean-Francois Dufour reports advisory committees with AbbVie, Bayer, BMS, Falk, Genfit, Genkyotex, Gilead Science, HepaRegenix, Intercept, Lilly, Merck, Novartis and speaking and teaching with Bayer, BMS, Intercept, Genfit, Gilead Science, Novartis; Pietro Invernizzi reports grants from Intercept, Gilead and Bruschettini; Mattias Ekstedt reports personal fees from AbbVie, AstraZeneca, Albireo, Diapharma, Gilead and non-financial support from Echosens (through LITMUS IMI project); Karine Clement has no personal honoraria but has consultancy and scientific collaboration activity for LNC therapeutics, Confotherapeutics and Danone Research; Jörn M. Schattenberg reports grants from Gilead and Boehringer Ingelheim and fees from Gilead, Boehringer Ingelheim, Galmed, Genfit, Intercept, Novartis, Pfizer and AbbVie outside the submitted work. All other authors report no conflicts of interest. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2020 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)