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Metabolic dysfunction-associated steatotic liver disease (MASLD) is common worldwide. Genes and proteins contributing to drug disposition may show altered expression as MASLD progresses. To assess this further, we undertook transcriptomic and proteomic analysis of 137 pharmacogenes in liver biopsies from a large MASLD cohort. We performed sequencing on RNA from 216 liver biopsies (206 MASLD and 10 controls). Untargeted mass spectrometry proteomics was performed on a 103 biopsy subgroup. Selected RNA sequencing signals were replicated with an additional 187 biopsies. Comparison of advanced MASLD (fibrosis score 3/4) with milder disease (fibrosis score 0-2) by RNA sequencing showed significant alterations in expression of certain phase I, phase II and ABC transporters. For cytochromes P450, CYP2C19 showed the most significant decreased expression (30 % of that in mild disease) but significant decreased expression of other CYPs (including CYP2C8 and CYP2E1) also occurred. CYP2C19 also showed a significant decrease comparing the inflammatory form of MASLD (MASH) with non-MASH biopsies. Findings for CYP2C19 were confirmed in the replication cohort. Proteomics on the original discovery cohort confirmed decreased levels of several CYPs as MASLD advanced but this decrease was greatest for CYP2C19 where levels fell to 40 % control. This decrease may result in decreased CYP2C19 activity that could be problematic for prescription of drugs activated or metabolized by CYP2C19 as MASLD advances. More limited decreases for other P450s suggest fewer issues with non-CYP2C19 drug substrates. Negative correlations at RNA level between CYP2C19 and several cytokine genes provided initial insights into the mechanism underlying decreased expression., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ann Daly and Quentin Anstee reports financial support was provided by European Union. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Non-alcoholic fatty liver disease (NAFLD) is the liver manifestation of the metabolic syndrome with global prevalence reaching epidemic levels. Despite the high disease burden in the population only a small proportion of those with NAFLD will develop progressive liver disease, for which there is currently no approved pharmacotherapy. Identifying those who are at risk of progressive NAFLD currently requires a liver biopsy which is problematic. Firstly, liver biopsy is invasive and therefore not appropriate for use in a condition like NAFLD that affects a large proportion of the population. Secondly, biopsy is limited by sampling and observer dependent variability which can lead to misclassification of disease severity. Non-invasive biomarkers are therefore needed to replace liver biopsy in the assessment of NAFLD. Our study addresses this unmet need. The LITMUS Imaging Study is a prospectively recruited multi-centre cohort study evaluating magnetic resonance imaging and elastography, and ultrasound elastography against liver histology as the reference standard. Imaging biomarkers and biopsy are acquired within a 100-day window. The study employs standardised processes for imaging data collection and analysis as well as a real time central monitoring and quality control process for all the data submitted for analysis. It is anticipated that the high-quality data generated from this study will underpin changes in clinical practice for the benefit of people with NAFLD. Study Registration: clinicaltrials.gov: NCT05479721., Competing Interests: Declaration of Competing Interest FEM, SA, KW, JC, MA, EMG, SF, MRG, JC, IFL, RA, RSG, JMP, JB, VR, MW, SP, MA, RF, LM, ET, ME, PL, ATH, GP, HYJ, KP and PMB declare no conflicts of interest. MP is a shareholder in Perspectum Ltd. EMT is a shareholder in Perspectum Ltd. SA is currently employed by Boehringer Ingelheim (but was not during his participation in the project). GPA has served as a consultant and an advisory board member for Pfizer Inc., Inventiva Pharma, GlaxoSmithKline and KaNDy Therapeutics; he has been a consultant to BerGenBio ASA, Median Technologies, FRACTYL, Amryt Pharmaceuticals and AstraZeneca; and has given presentations on behalf of Roche Diagnostics and Medscape all through the University of Nottingham contract. CA declares the following potential conflicts of interest: Hologic: Support of study and expert; Guerbet: Member of board, PI of study; Siemens: Expert. EB has served as a consultant or advisory board member for Boehringer Ingelheim, Gilead Sciences, Intercept, Merck, Novo Nordisk, Pfizer, ProSciento; and a speaker for Gilead Sciences, Intercept, Merck, Novo Nordisk, Pfizer. She has also received a research grant from Gilead Sciences for fatty liver research. AG served as a speaker and consultant for AbbVie, Alexion, AstraZeneca, Bayer, BMS, CSL Behring, Eisai, Falk, Gilead, Heel, Intercept, Ipsen, Merz, MSD, Novartis, Pfizer, Roche, Sanofi-Aventis, Sequana; received research funding from Intercept, Falk, Novartis. JMS reports consultancy for BMS, Boehringer Ingelheim, Echosens, Genfit, Gilead Sciences, Intercept Pharmaceuticals, Madrigal, Novartis, Pfizer, Roche, Sanofi; received research funding from Gilead Sciences and was on the speaker's bureau for Falk Foundation MSD Sharp & Dohme GmbH. MJS and PH are employed by Antaros Medical AB, Mölndal, Sweden. ES is employed by Perspectum Ltd., Oxford, UK. RB is a shareholder and employed (CEO) by Perspectum Ltd., Oxford, UK. KP and MK are employed by Resoundant Inc., Rochester, MN, USA. RLE and the Mayo Clinic have intellectual property rights and a financial interest in magnetic resonance elastography technology. AT is employed by ADVANZPHARMA, Capital House, 1st Floor, 85 King William Street, London, EC4N 7BL, United Kingdom. HC is employed by Boehringer Ingelheim Pharma GmbH & Co. MM is employed by Novartis AG, Basel, Switzerland. CY is employed by Pfizer Inc., Lake Mary, FL, USA. TT was employed by Pfizer at the time of her involvement with the project. QMA is coordinator of the EU IMI-2 LITMUS consortium, which is funded by the EU Horizon 2020 programme and EFPIA. This multistakeholder consortium includes industry partners. QMA has received research grant funding from AstraZeneca, Boehringer Ingelheim, and Intercept Pharmaceuticals, Inc.; has served as a consultant on behalf of Newcastle University for Alimentiv, Akero, AstraZeneca, Axcella, 89bio, Boehringer Ingelheim, Bristol Myers Squibb, Galmed, Genfit, Genentech, Gilead, GSK, Hanmi, HistoIndex, Intercept Pharmaceuticals, Inc., Inventiva, Ionis, IQVIA, Janssen, Madrigal, Medpace, Merck, NGM Bio, Novartis, Novo Nordisk, PathAI, Pfizer, Poxel, Resolution Therapeutics, Roche, Ridgeline Therapeutics, RTI, Shionogi, and Terns; has served as a speaker for Fishawack, Integritas Communications, Kenes, Novo Nordisk, Madrigal, Medscape, and Springer Healthcare; and receives royalties from Elsevier Ltd. SN is a shareholder in Perspectum Ltd. SAH has research grants from Akero, Altimmune, Axcella-Cirius, CiVi Biopharma, Cymabay, Galectin, 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, Echosens, Enyo, Foresite Labs, Galectin, Genfit, Gilead Sciences, Hepion, HIghtide, HistoIndex, Intercept, Kowa, Madrigal, Metacrine, NeuroBo, NGM, Northsea, Novartis, Novo Nordisk, Poxel, Perspectum, Sagimet, Terns, and Viking., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: The reference standard for detecting non-alcoholic steatohepatitis (NASH) and staging fibrosis-liver biopsy-is invasive and resource intensive. Non-invasive biomarkers are urgently needed, but few studies have compared these biomarkers in a single cohort. As part of the Liver Investigation: Testing Marker Utility in Steatohepatitis (LITMUS) project, we aimed to evaluate the diagnostic accuracy of 17 biomarkers and multimarker scores in detecting NASH and clinically significant fibrosis in patients with non-alcoholic fatty liver disease (NAFLD) and identify their optimal cutoffs as screening tests in clinical trial recruitment., Methods: This was a comparative diagnostic accuracy study in people with biopsy-confirmed NAFLD from 13 countries across Europe, recruited between Jan 6, 2010, and Dec 29, 2017, from the LITMUS metacohort of the prospective European NAFLD Registry. Adults (aged ≥18 years) with paired liver biopsy and serum samples were eligible; those with excessive alcohol consumption or evidence of other chronic liver diseases were excluded. The diagnostic accuracy of the biomarkers was expressed as the area under the receiver operating characteristic curve (AUC) with liver histology as the reference standard and compared with the Fibrosis-4 index for liver fibrosis (FIB-4) in the same subgroup. Target conditions were the presence of NASH with clinically significant fibrosis (ie, at-risk NASH; NAFLD Activity Score ≥4 and F≥2) or the presence of advanced fibrosis (F≥3), analysed in all participants with complete data. We identified thres holds for each biomarker for reducing the number of biopsy-based screen failures when recruiting people with both NASH and clinically significant fibrosis for future trials., Findings: Of 1430 participants with NAFLD in the LITMUS metacohort with serum samples, 966 (403 women and 563 men) were included after all exclusion criteria had been applied. 335 (35%) of 966 participants had biopsy-confirmed NASH and clinically significant fibrosis and 271 (28%) had advanced fibrosis. For people with NASH and clinically significant fibrosis, no single biomarker or multimarker score significantly reached the predefined AUC 0·80 acceptability threshold (AUCs ranging from 0·61 [95% CI 0·54-0·67] for FibroScan controlled attenuation parameter to 0·81 [0·75-0·86] for SomaSignal), with accuracy mostly similar to FIB-4. Regarding detection of advanced fibrosis, SomaSignal (AUC 0·90 [95% CI 0·86-0·94]), ADAPT (0·85 [0·81-0·89]), and FibroScan liver stiffness measurement (0·83 [0·80-0·86]) reached acceptable accuracy. With 11 of 17 markers, histological screen failure rates could be reduced to 33% in trials if only people who were marker positive had a biopsy for evaluating eligibility. The best screening performance for NASH and clinically significant fibrosis was observed for SomaSignal (number needed to test [NNT] to find one true positive was four [95% CI 4-5]), then ADAPT (six [5-7]), MACK-3 (seven [6-8]), and PRO-C3 (nine [7-11])., Interpretation: None of the single markers or multimarker scores achieved the predefined acceptable AUC for replacing biopsy in detecting people with both NASH and clinically significant fibrosis. However, several biomarkers could be applied in a prescreening strategy in clinical trial recruitment. The performance of promising markers will be further evaluated in the ongoing prospective LITMUS study cohort., Funding: The Innovative Medicines Initiative 2 Joint Undertaking., Competing Interests: Declaration of interests QMA is the coordinator of the Innovative Medicines Initiative 2 (IMI2) LITMUS consortium funded by the IMI2 Program of the EU (777377) and is a Newcastle National Institute for Health and Care Research Biomedical Research Centre investigator. The multistakeholder LITMUS consortium includes industry partners and received funding from the European Federation of Pharmaceutical Industries and Associations. QMA also declares research grant funding from AbbVie, AstraZeneca, Boehringer Ingelheim, Glympse Bio, Intercept, Novartis, and Pfizer; is a consultant on behalf of Newcastle University to Alimentiv, Akero, AstraZeneca, Axcella, 89Bio, Boehringer Ingelheim, Bristol-Myers Squibb, Galmed, Genfit, Genentech, Gilead, GlaxoSmithKline, Hanmi, HistoIndex, Intercept, Inventiva, Ionis, IQVIA, Janssen, Madrigal, Medpace, Merck, NGMBio, Novartis, Novo Nordisk, PathAI, Pfizer, Poxel, Resolution Therapeutics, Roche, Ridgeline Therapeutics, RTI International, Shionogi, and Terns; has received speaker fees from Fishawack, Integritas Communications, Kenes, Novo Nordisk, Madrigal, Medscape, and Springer Healthcare; is on the North Sea Therapeutics Data Safety and Monitoring Committee at Medpace on behalf of Newcastle University; and has received royalties from Elsevier. JB declares research grant funding from Echosens, Intercept, Inventiva, and Siemens; consulting fees from Echosens, Intercept, and Siemens; speaker fees from Gilead, Intercept, Lilly, and Siemens; and is a member of advisory boards of Bristol-Myers Squibb, Echosens, Intercept, Novo Nordisk, and Merck Sharp & Dohme. DT declares grants from and contracts with Histoindex; consulting fees from Alimentiv, Clinnovate Health UK, Allergan, Cirius Therapeutics, Intercept Pharmaceuticals, ICON Clinical Research, and Ionis Pharmaceuticals; and was the past President of the European Society of Pathology (unpaid). PB is the owner and Director of Liverpat. AG declares support from the IMI2 LITMUS project and research grants from Falk and Intercept; consulting fees from AbbVie, Alexion, Bayer, Bristol-Meyers Squibb, Eisai, Gilead, Intercept, Ipsen, Merck Sharp & Dohme, Novartis, Pfizer, Roche, Sanofi-Aventis, and Sequana; research fees from AbbVie, Alexion, Bristol-Meyers Squibb, Burger Stein, CSL Behring, Falk, Gilead, Intercept, Mert, Merck Sharp & Dohme, Novartis, Roche, and Sequana; travel and meeting support from Intercept, Gilead, AbbVie, and Falk; and is a member of an advisory board of Falk. SF has a Senior Clinical Investigator fellowship from the Research Foundation Flanders (1802154N); declares grants to his institution (University of Antwerp, Belgium) from Genfit, Gilead Sciences, Roche, and Bristol-Meyers Squibb; has been a consultant to Roche, Intercept, Gilead Sciences, Echosens, Allergan, Genentech, AbbVie, Novo Nordisk, Bayer, Novartis, Bristol-Meyers Squibb, AstraZeneca, Boehringer Ingelheim, Galmed, Merck Sharp & Dohme, Promethera, Janssen Pharmaceutica, Coherus, Actelion Madrigal, Astellas, Julius Clinical, Genfit, NGM Bio, and Inventiva; and has been lecturer for AbbVie, Allergan, Bayer, Genfit, and Gilead Sciences. MA declares research grants from GlaxoSmithKline, Takeda, and AstraZeneca and consulting fees from Intercept and AstraZeneca. GP declares support from AbbVie, Gilead Sciences, Janssen, and Takeda; consulting fees from AbbVie, Amgen, Astellas, Astra Zeneca, Gilead Sciences, GlaxoSmithKline, Janssen, Ipsen, Merck Sharp & Dohme, Novartis, Novo Nordisk, and Roche; speaker fees from AbbVie, Gilead Sciences, GlaxoSmithKline, Ipsen, Merck Sharp & Dohme, Novartis, Novo Nordisk, Roche, and Takeda; and travel support from AbbVie, Gilead Sciences, Ipsen, Janssen, and Novo Nordisk. HC-P declares support from the IMI2 LITMUS project; consulting fees from Intercept, Novo Nordisk, and Orphalan; and speaker fees from Eisai and Roche Portugal. J-FD is a member of the advisory committees of Alentis, AstraZeneca, Bayer, Bristol-Myers Squibb, Enyo, Esai, Falk, Genfit, Gilead Sciences, Intercept, Inventiva, Ipsen, Lilly, Madrigal, Merck, Novartis, Novo Nordisk, and Roche; declares travel and meeting support from Gilead and speaking and teaching fees from Bayer, Bristol-Myers Squibb, Intercept, Gilead Sciences, Novartis, and Roche; has received fees from the journal GUT as an Associate Editor and Uptodate; and is President of the Swiss NASH Foundation and the Swiss Foundation against Liver Cancer. DJL declares support from the IMI2 LITMUS project and Nordic Bioscience and has stock in Nordic Bioscience. SAH declares grants from Akero Therapeutics, Axcella Health, Cirius Therapeutics, CiVi Biopharma, Cymabay, Enyo Pharma, Galectin, Galmed Research and Development, Genfit, Gilead Sciences, Hepion Pharmaceuticals, Hightide Therapeutics, Intercept Pharmaceuticals, Madrigal Pharmaceuticals, Metacrine, NGM Bio, Northsea Therapeutics, Novartis Pharmaceuticals, Novo Nordisk, Poxel, Sagimet Biosciences, and Viking Therapeutics; consulting and speaker fees from AgomAB, Akero Therapeutics, Alentis Therapeutics, Alimentiv, Altimmune, Axcella Health, Boston Pharmaceuticals, Riley FBR, BVF Partners, Cohbar, Canfite, Corcept Therapeutics, Cymabay, Echosens North America, Enyo Pharma, Fibronostics, Foresite Labs, Fortress Biotech, Galectin Therapeutics, Genfit, Genius Group, Hepion Pharmaceuticals, Hightide Therapeutics, HistoIndex, Inipharm, Intercept Pharmaceuticals, Ionis, Kowa Research Institute, Madrigal Pharmaceuticals, Medpace, Metacrine, Microba, NGM Biopharmaceuticals, Northsea Therapeutics, Novo Nordisk, Nutrasource, Perspectum Diagnostics, Piper, Sandler, Poxel, Prometic Pharma, Ridgeline, Sagimet Biosciences, Sonic Incytes, Medical, Terns, and Viking Therapeutics; is a member of advisory boards of 89 Bio, Akero Therapeutics, Altimmune, Arrowhead, Axcella Health, Chronwell, CiVi, Cymabay, Echosens North America, Foresite Labs, Galectin Therapeutics, Galmed Research and Development, Genfit, Gilead Sciences, Hepion Pharmaceuticals, Hightide Therapeutics, HistoIndex, Indalo, Intercept Pharmaceuticals, Madrigal Pharmaceuticals, Medpace, Metacrine, NGM Bio, Northsea Therapeutics, Novartis Pharmaceuticals, Novo Nordisk, PathAI, Poxel, Prometic Pharma, Ridgeline, Sagimet Biosciences, Sonic Incytes Medical, Terns, and Theratechnologies; has stock in Akero Therapeutics, Chronwell, Cirius Therapeutics, Galectin Therapeutics, Genfit, Hepion Pharmaceuticals, HistoIndex, Metacrine, NGM Bio, Northsea Therapeutics, Sonic Incytes, and Medical Corp; and declares consultancy services to Akero Therapeutics, Axcella Health, Cirius Therapeutics, CiVi Biopharma, Cymabay, Enyo Pharma, Galectin Therapeutics, Galmed Research and Development, Genfit, Gilead Sciences, Hepion Pharmaceuticals, Hightide Therapeutics, Intercept Pharmaceuticals, Madrigal Pharmaceuticals, Metacrine, NGM Bio, Northsea Therapeutics, Novartis Pharmaceuticals, Novo Nordisk, Poxel, Sagimet Biosciences, and Viking Therapeutics. YC is an employee of Eli Lilly and the company and hold stocks and shares in Eli Lilly. JFC declares support from Oxford National Institute for Health and Care Research Biomedical Research Centre; consulting fees from Intercept and Alnylam; speaker fees from AstraZeneca; and is a member of an advisory board of Intercept. MP declares support from the IMI2 LITMUS project and grants from Cancer Research UK (C30358/A29725); has a patent for medical imaging (WO2015155521A1); and is a shareholder in Perspectum. HY-J declares support from the IMI2 LITMUS project; grants from Academy of Finland, Sigrid Juselius Foundation, and EVO foundation; consulting fees from Boehringer Ingelheim; speaker fees from Novo Nordisk and GlaxoSmithKline; and is a member of advisory boards of Merck Sharp & Dohme, Eli Lilly, and Hamni. JC declares grants and research support from Gilead Sciences, AbbVie, and Intercept Pharmaceuticals; consultant fees from Gilead Sciences, AbbVie, Merck Sharp & Dohme, Shionogi, Intercept Pharmaceuticals, Janssen Pharmaceuticals, Celgene, and Alexion; speaker support from Gilead Sciences, AbbVie, Shionogi, Janssen Pharmaceuticals, and Celgene; meeting and travel support from Gilead Sciences, AbbVie, and Janssen Pharmaceuticals; and is President of the Spanish Society of Digestive Pathology. MK declares support from the IMI2 LITMUS project and Nordic Bioscience. He is a Nordic Bioscience stock share holder. RO is an employee of SomaLogic; declares support from the IMI2 LITMUS project (SomaLogic funded the SomaScan proteomic assay results as a contribution to LITMUS); is on the patent application of SomaLogic for NASH biomarkers; and has stock in SomaLogic. RT is an employee of and holds stock in AstraZeneca, Sweden. KD is an employee of Eli Lilly and the company and hold stocks and shares in Eli Lilly. CY is an employee and stock and share holder of Pfizer. CB is a former employee of Novartis Pharma and declares financial support from Novartis Pharma; has pending patents for treatment of NASH (Novartis); and has stock in Novartis Pharma and Merck. ME is a member of the advisory board of AMRA Medical. GPA declares support from the IMI2 LITMUS project and consulting fees from Clinicpace, Servier Pharmaceuticals, GlaxoSmithKline, NuCana, AstraZeneca, and Benevolent AI. JMS declares grants from Gilead Sciences, Boehringer Ingelheim, Nordic Bioscience, and Siemens Healthcare; consulting fees from Apollo Endosurgery, Albireo Pharma, Bayer, Bristol-Meyers Squibb, Boehringer Ingelheim, Echosens, Genfit, Gilead Sciences, GlaxoSmithKline, Heel, Intercept Pharmaceuticals, Ipsen, Inventiva Pharma, Julius Clinical, Madrigal, Merck Sharp & Dohme, Nordic Bioscience, Novartis, Novo Nordisk, Pfizer, Roche, Sanofi, Shinogi, Siemens Healthcare, and Summit Clinical Research; speaker fees from Apollo Endosurgery, MedPublico, and Boehringer Ingelheim; and is a member of advisory boards of Novo Nordisk and Boehringer Ingelheim. EB declares consulting fees from Gilead, Novo Nordisk, Boehringer Ingelheim, and Intercept; speaker fees from Novo Nordisk, Intercept, and Merck Sharp & Dohme; and is a member of advisory boards of Novo Nordisk and Intercept. MR-G declares grants from Siemens, Gilead, and Intercept; consulting fees from AbbVie, Alpha-sigma, Allergen, AstraZeneca, Inventia, Kaleido, Novo Nordisk, Pfizer, Axcella, Bristol-Meyers Squibb, Boehringer Ingelheim, and Gilead; speaker fees from Inventia, Sobi, Novo Nordisk, Rubio, and Shionogi; meeting and travel support from Abbvie and Gilead; and is a member of an advisory board of Galmed. VR declares grants from Gilead Sciences and Intercept Pharmaceuticals and consulting fees from GlaxoSmithKline, Galmed, Novo Nordisk, Prosciento, Terns, NorthSea Therapeutics, Enyo, Sagimet, NGM Pharmaceuticals, and Madrigal. All other authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background and Aims: Detecting NASH remains challenging, while at-risk NASH (steatohepatitis and F≥ 2) tends to progress and is of interest for drug development and clinical application. We developed prediction models by supervised machine learning techniques, with clinical data and biomarkers to stage and grade patients with NAFLD., Approach and Results: Learning data were collected in the Liver Investigation: Testing Marker Utility in Steatohepatitis metacohort (966 biopsy-proven NAFLD adults), staged and graded according to NASH CRN. Conditions of interest were the clinical trial definition of NASH (NAS ≥ 4;53%), at-risk NASH (NASH with F ≥ 2;35%), significant (F ≥ 2;47%), and advanced fibrosis (F ≥ 3;28%). Thirty-five predictors were included. Missing data were handled by multiple imputations. Data were randomly split into training/validation (75/25) sets. A gradient boosting machine was applied to develop 2 models for each condition: clinical versus extended (clinical and biomarkers). Two variants of the NASH and at-risk NASH models were constructed: direct and composite models.Clinical gradient boosting machine models for steatosis/inflammation/ballooning had AUCs of 0.94/0.79/0.72. There were no improvements when biomarkers were included. The direct NASH model produced AUCs (clinical/extended) of 0.61/0.65. The composite NASH model performed significantly better (0.71) for both variants. The composite at-risk NASH model had an AUC of 0.83 (clinical and extended), an improvement over the direct model. Significant fibrosis models had AUCs (clinical/extended) of 0.76/0.78. The extended advanced fibrosis model (0.86) performed significantly better than the clinical version (0.82)., Conclusions: Detection of NASH and at-risk NASH can be improved by constructing independent machine learning models for each component, using only clinical predictors. Adding biomarkers only improved the accuracy of fibrosis., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc.)

The mechanisms that drive nonalcoholic fatty liver disease (NAFLD) remain incompletely understood. This large multicenter study characterized the transcriptional changes that occur in liver tissue across the NAFLD spectrum as disease progresses to cirrhosis to identify potential circulating markers. We performed high-throughput RNA sequencing on a discovery cohort comprising histologically characterized NAFLD samples from 206 patients. Unsupervised clustering stratified NAFLD on the basis of disease activity and fibrosis stage with differences in age, aspartate aminotransferase (AST), type 2 diabetes mellitus, and carriage of PNPLA3 rs738409 , a genetic variant associated with NAFLD. Relative to early disease, we consistently identified 25 differentially expressed genes as fibrosing steatohepatitis progressed through stages F2 to F4. This 25-gene signature was independently validated by logistic modeling in a separate replication cohort ( n = 175), and an integrative analysis with publicly available single-cell RNA sequencing data elucidated the likely relative contribution of specific intrahepatic cell populations. Translating these findings to the protein level, SomaScan analysis in more than 300 NAFLD serum samples confirmed that circulating concentrations of proteins AKR1B10 and GDF15 were strongly associated with disease activity and fibrosis stage. Supporting the biological plausibility of these data, in vitro functional studies determined that endoplasmic reticulum stress up-regulated expression of AKR1B10 , GDF15 , and PDGFA , whereas GDF15 supplementation tempered the inflammatory response in macrophages upon lipid loading and lipopolysaccharide stimulation. This study provides insights into the pathophysiology of progressive fibrosing steatohepatitis, and proof of principle that transcriptomic changes represent potentially tractable and clinically relevant markers of disease progression., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Non-Alcoholic Fatty Liver Disease (NAFLD), a progressive liver disease that is closely associated with obesity, type 2 diabetes, hypertension and dyslipidaemia, represents an increasing global public health challenge. There is significant variability in the disease course: the majority exhibit only fat accumulation in the liver but a significant minority develop a necroinflammatory form of the disease (non-alcoholic steatohepatitis, NASH) that may progress to cirrhosis and hepatocellular carcinoma. At present our understanding of pathogenesis, disease natural history and long-term outcomes remain incomplete. There is a need for large, well characterised patient cohorts that may be used to address these knowledge gaps and to support the development of better biomarkers and novel therapies. The European NAFLD Registry is an international, prospectively recruited observational cohort study that aims to establish a large, highly-phenotyped patient cohort and linked bioresource. Here we describe the infrastructure, data management and monitoring plans, and the standard operating procedures implemented to ensure the timely and systematic collection of high-quality data and samples. Already recruiting subjects at secondary/tertiary care centres across Europe, the Registry is supporting the European Union IMI2-funded LITMUS 'Liver Investigation: Testing Marker Utility in Steatohepatitis' consortium, which is a major international effort to robustly validate biomarkers that diagnose, risk stratify and/or monitor NAFLD progression and liver fibrosis stage. The European NAFLD Registry has the demonstrable capacity to support research and biomarker development at scale and pace., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)