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Abstract: Mini-module aging tests with differently interconnected heterojunction solar cells having industrially viable copper metallization are presented. The plating process comprises 3 steps: firstly, screen printing of a seed-grid layout using a copper-based paste, followed by deposition of a dielectric layer over the entire wafer surface, and finally, selective copper electrodeposition on grid positions. Modules with Smartwire interconnection, fabricated with M6 half-cells, are stable in extended TC and PID tests. DH degradation is at 5% after 2700 h (glass-glass modules without edge sealing). Shingle modules, realized in collaboration with CEA INES and AMAT, exhibit notably higher fill factor compared to reference modules with screen-printed silver paste. This improvement is attributed to the superior line conductivity achieved with plated copper. TC stability of shingle modules is very good, whereas after 2000 h damp-heat aging more than 2% loss in fill factor is observed. Using pattern-transfer-printing technology narrow, high aspect-ratio lines have been obtained: with a seed-grid of pure copper paste, reinforced with electrodeposited copper. Line dimensions and line resistance as well as first cell results are presented.

This work aims at the full recovery of efficiency losses induced by shingling double-side poly-Si/SiOx passivated contacts crystalline silicon solar cells. It focuses on thermally-activated Aluminium Oxide (AlOx) layers elaborated by thermal Atomic Layer Deposition (ALD) to passivate the edges of shingled cells cut by using the innovative “45° tilt squaring approach”. The whole procedure featuring high-temperature AlOx annealing led to very low cut-related performance losses. Indeed, the efficiency and FF of the passivated shingled cells surpassed the values obtained for the as-cut shingles by 0.5%abs and 2.6%abs, respectively. Approaches for further improvements are also discussed, particularly to overcome the short-circuit current density decrease observed for passivated shingles.

Nickel ferrite (NiFe2O4) was synthesized by mixing stoichiometric amounts of α-Fe2O3 and NiO using mechanical milling and heat treatments at high temperatures. The physical characterization of the samples was carried out using X-ray diffraction, infrared and Raman spectroscopies, Mössbauer spectrometry, magnetization measurements, scanning electron microscopy and energy dispersive X-ray spectroscopy. We found that NiFe2O4 production increases from 81 to 100 wt. % with increasing temperature. Additionally, the lattice parameter and the saturation magnetization increase with increasing temperature. On the other hand, Mössbauer spectrometry showed that there is a decrease in the subspectral areas ratio for Fe3+ cations at tetrahedral (A) and octahedral [B] sites, AA/AB, with the increase of the temperature. In the SEM micrographs it was observed that the samples consisted of particles with irregular shapes and micrometric sizes. From IR spectra, the intensity of the 411 cm-1 band (vibrations at octahedral sites) increases relative to the intensity of the 599 cm-1 band (vibrations at tetrahedral sites) with increasing temperature. From the results obtained in the magnetization curves, it was possible to confirm the synthesis of NiFe2O4. As the heat treatment temperature increases, hysteresis loops with S-type geometric forms were obtained. All the results suggest that a defective spinel NiFe2O4 is formed at 1000 °C, and that as the temperature increases, the defects gradually disappear. Neither cation reordering phenomena nor possible evaporation of chemical elements were the dominant effects to account for the results. The results can be explained if it is assumed that [B]-sites cation vacancies are gradually filled with cations as the temperature of the heat treatment increases.

The observed increase in emission of greenhouse gases, with attendant effects on global warming, have raised interests in identifying sources and sinks of carbon in the environment. Terrestrial carbon (C) sequestration involves capture of atmospheric C through photosynthesis and storage in biota, soil and wetlands. Particularly, wetland systems function primarily as long-term reservoirs for atmospheric carbon dioxide (CO2) and as sources of atmospheric methane (CH4). The objective of this study was to evaluate the patterns of carbon reservoirs in two high-altitude wetlands in the central Andean mountain of Colombia. Carbon cycle in both systems is related mainly with the plant biomass dynamics from the littoral zone. Thus, total organic carbon concentrate an average up to 329 kg of N ha-1 and 125 kg of P ha-1 every year vs only 17 kg N ha-1 and 6 kg P ha-1 in the water column of the limnetic zone in the wetland, evidencing spatial differences in carbon concentrations for these types of ecosystems. Results revealed that these systems participate in the balance and sequestration of carbon in the Colombian Andes.

The control of bovine tuberculosis and atypical mycobacterioses in cattle in developing countries is important but difficult because of the existence of wildlife reservoirs. In cattle farms in Tanzania, mycobacteria were detected in 7.3% of 645 small mammals and in cow's milk. The cattle farms were divided into “reacting” and “nonreacting” farms, based on tuberculin tests, and more mycobacteria were present in insectivores collected in reacting farms as compared to nonreacting farms. More mycobacteria were also present in insectivores as compared to rodents. All mycobacteria detected by culture and PCR in the small mammals were atypical mycobacteria. Analysis of the presence of mycobacteria in relation to the reactor status of the cattle farms does not exclude transmission between small mammals and cattle but indicates that transmission to cattle from another source of infection is more likely. However, because of the high prevalence of mycobacteria in some small mammal species, these infected animals can pose a risk to humans, especially in areas with a high HIV-prevalence as is the case in Tanzania.

Abstract Background Quantifying the improvements in lower limb or quadriceps muscle mass following resistance training (RT), is an important outcome measure in COPD. Ultrasound is a portable, radiation free imaging technique that can measure the size of superficial muscles belonging to the quadriceps group such as the rectus femoris, but has not been previously used in COPD patients following RT. We compared the responsiveness of ultrasound derived measures of quadriceps mass against dual energy x-ray absorptiometry (DEXA), in patients with COPD and healthy controls following a programme of high intensity knee extensor RT. Methods Portable ultrasound was used to assess the size of the dominant quadriceps in 45 COPD patients and 19 healthy controls-before, during, and after 8 weeks of bilateral high intensity isokinetic knee extensor RT. Scanning was performed at the mid-thigh region, and 2 indices of quadriceps mass were measured-rectus femoris cross-sectional area (RFcsa) and quadriceps muscle thickness (Qt). Thigh lean mass (Tdexa) was determined by DEXA. Results Training resulted in a significant increase in Tdexa, RFcsa and Qt in COPD patients [5.7%, 21.8%, 12.1% respectively] and healthy controls [5.4%, 19.5%, 10.9 respectively]. The effect size for the changes in RFcsa (COPD= 0.77; Healthy=0.83) and Qt (COPD=0.36; Healthy=0.78) were greater than the changes in Tdexa (COPD=0.19; Healthy=0.26) following RT. Conclusions Serial ultrasound measurements of the quadriceps can detect changes in muscle mass in response to RT in COPD. The technique has good reproducibility, and may be more sensitive to changes in muscle mass when compared to DEXA. Trial registration http://www.controlled-trials.com (Identifier: ISRCTN22764439)

Abstract Background The floral dip method of transformation by immersion of inflorescences in a suspension of Agrobacterium is the method of choice for Arabidopsis transformation. The presence of a marker, usually antibiotic- or herbicide-resistance, allows identification of transformed seedlings from untransformed seedlings. Seedling selection is a lengthy process which does not always lead to easily identifiable transformants. Selection for kanamycin-, phosphinothricin- and hygromycin B-resistance commonly takes 7–10 d and high seedling density and fungal contamination may result in failure to recover transformants. Results A method for identifying transformed seedlings in as little as 3.25 d has been developed. Arabidopsis T1 seeds obtained after floral dip transformation are plated on 1% agar containing MS medium and kanamycin, phosphinothricin or hygromycin B, as appropriate. After a 2-d stratification period, seeds are subjected to a regime of 4–6 h light, 48 h dark and 24 h light (3.25 d). Kanamycin-resistant and phosphinothricin-resistant seedlings are easily distinguished from non-resistant seedlings by green expanded cotyledons whereas non-resistant seedlings have pale unexpanded cotyledons. Seedlings grown on hygromycin B differ from those grown on kanamycin and phosphinothricin as both resistant and non-resistant seedlings are green. However, hygromycin B-resistant seedlings are easily identified as they have long hypocotyls (0.8–1.0 cm) whereas non-resistant seedlings have short hypocotyls (0.2–0.4 cm). Conclusion The method presented here is an improvement on current selection methods as it allows quicker identification of transformed seedlings: transformed seedlings are easily discernable from non-transformants in as little as 3.25 d in comparison to the 7–10 d required for selection using current protocols.

Metabolic dysfunction-associated steatohepatitis (MASH) is a major cause of liver-related morbidity and mortality, yet treatment options are limited. Manual scoring of liver biopsies, currently the gold standard for clinical trial enrollment and endpoint assessment, suffers from high reader variability. This study represents the most comprehensive multisite analytical and clinical validation of an artificial intelligence (AI)-based pathology system, AI-based measurement of metabolic dysfunction-associated steatohepatitis (AIM-MASH), to assist pathologists in MASH trial histology scoring. AIM-MASH demonstrated high repeatability and reproducibility compared to manual scoring. AIM-MASH-assisted reads by expert MASH pathologists were superior to unassisted reads in accurately assessing inflammation, ballooning, MAS ≥ 4 with ≥1 in each score category and MASH resolution, while maintaining non-inferiority in steatosis and fibrosis assessment. These findings suggest that AIM-MASH could mitigate reader variability, providing a more reliable assessment of therapeutics in MASH clinical trials., (© 2024. The Author(s).)

Background & Aims: Metabolic dysfunction-associated steatohepatitis (MASH) and fibrotic MASH are significant health challenges. This multi-national study aimed to validate the acMASH index (including serum creatinine and aspartate aminotransferase concentrations) for MASH diagnosis and develop a new index (acFibroMASH) for non-invasively identifying fibrotic MASH and exploring its predictive value for liver-related events (LREs)., Methods: We analyzed data from 3004 individuals with biopsy-proven metabolic dysfunction-associated fatty liver disease (MAFLD) across 29 Chinese and 9 international cohorts to validate the acMASH index and develop the acFibroMASH index. Additionally, we utilized the independent external data from a multi-national cohort of 9034 patients with MAFLD to examine associations between the acFibroMASH index and the risk of LREs., Results: In the pooled global cohort, the acMASH index identified MASH with an area under the receiver operating characteristic curve (AUROC) of 0.802 (95% confidence interval [CI], 0.786-0.818). The acFibroMASH index (including the acMASH index plus liver stiffness measurement) accurately identified fibrotic MASH with an AUROC of 0.808 in the derivation cohort and 0.800 in the validation cohort. Notably, the AUROC for the acFibroMASH index was 0.835 (95% CI, 0.786-0.882), superior to that of the FAST score at 0.750 (95% CI, 0.693-0.800; P < .01) in predicting the 5-year risk of LREs. Patients with acFibroMASH >0.39 had a higher risk of LREs than those with acFibroMASH <0.15 (adjusted hazard ratio, 11.23; 95% CI, 3.98-31.66)., Conclusions: This multi-ethnic study validates the acMASH index as a reliable, noninvasive test for identifying MASH. The newly proposed acFibroMASH index is a reliable test for identifying fibrotic MASH and predicting the risk of LREs., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

The intersection of cardiovascular disease, metabolic disorders and chronic kidney disease represents a complex clinical picture challenging healthcare systems worldwide. Metabolic-dysfunction-associated steatotic liver disease (MASLD) often manifests sequentially or concomitantly with these diseases, and may share underlying mechanisms and risk factors. Growing evidence suggests that new therapies could have benefits across these diseases, but trial sponsors and investigators tend to be reluctant to include patients with comorbidities-particularly liver diseases-in clinical trials. In this Perspective, we call for inclusion of patients with MASLD and measurement of liver outcomes in cardio-kidney-metabolic trials, when data suggest mechanistically plausible benefits and liver and cardiovascular safety. We discuss the implications of this new paradigm for clinical trial design and considerations for regulatory approval. Finally, we outline the challenges to implementing such an approach and provide recommendations for future clinical trial conduct., (© 2024. Springer Nature America, Inc.)

Metabolic dysfunction-associated steatotic liver disease (MASLD) affects 1 in 3-4 adult individuals and can progress to metabolic dysfunction-associated steatohepatitis (MASH) and cirrhosis. Insulin resistance plays a central role in MASLD/MASH pathophysiology with higher rates of MASLD (2 in 3) and MASH with fibrosis (1 in 5) in adults with obesity and diabetes. This review summarizes the role of glucagon-like peptide-1 receptor agonists in treating MASLD/MASH. Although not approved by the Food and Drug Administration for the treatment of MASLD, this class of medication is available to treat obesity and type 2 diabetes and has been shown to reverse steatohepatitis, reduce cardiovascular risk, and is safe to use across the spectrum of MASLD with or without fibrosis., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Metabolic dysfunction-associated steatotic liver disease affects 1 in 4 people in the United States and western Europe, with an important proportion developing metabolic dysfunction-associated steatohepatitis (MASH), the progressive subtype of metabolic dysfunction-associated steatotic liver disease. Cirrhosis caused by MASH is a leading indication for liver transplantation and the most common cause of hepatocellular carcinoma. Hitherto, there have been no specific pharmacotherapies for MASH. The recent conditional approval by the Food and Drug Administration of resmetirom for the treatment of moderate or advanced MASH presents a much-anticipated therapeutic option for patients with noncirrhotic advanced MASH. Specifically, the intended population for resmetirom are patients with MASH and fibrosis stages 2 or 3. The approval of resmetirom also presents important challenges, including how to noninvasively identify patients with fibrosis stages 2-3, and how to exclude patients with more advanced disease who should not be treated until further data emerge on the use of resmetirom in this population. Herein we consider the available literature with regard to identifying the intended population for treatment with resmetirom and in proposing criteria for stopping treatment., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background & Aims: In the American Gastroenterological Association/American Association for the Study of Liver Diseases (AGA/AASLD) Clinical Care Pathway, Fibrosis-4 index (FIB-4) is used to stratify patients at risk for metabolic dysfunction-associated steatotic liver disease (MASLD) as low-, indeterminate-, or high-risk for developing advanced liver fibrosis. We assessed the performance of FIB-4 in a general population., Methods: Using the 2017 to 2020 National Health and Nutrition Examination Surveys dataset, we selected subjects ≥18 years who had FibroScan data. We followed AGA/AASLD guidelines to identify subjects with characteristics that place them at risk for MASLD-associated liver fibrosis. Other causes of liver disease were excluded. Our final cohort had 3741 subjects. We then categorized these subjects based on recommended FIB-4 cutoffs. FibroScan liver stiffness measurement (LSM) served as the outcome measurement., Results: Among the 2776 subjects (74.2%) classified as low risk by FIB-4, 277 subjects (10%) were not classified at low risk by LSM, and 75 subjects (2.7%) were classified as high risk by LSM. Among the 86 subjects classified as high risk by FIB-4, 68 subjects (79.1%) were not at high risk by LSM, and 54 subjects (62.8%) were at low risk by LSM. Subjects misclassified by FIB-4 as low risk were older; had a higher body mass index, waist circumference, glycohemoglobin A1c level, alanine transaminase, aspartate transaminase, diastolic blood pressure, controlled attenuation parameter score, white blood cell count, alkaline phosphatase, and fasting glucose level; but had lower high-density lipoprotein, and albumin level (all P < .05). Misclassified subjects were also more likely to have prediabetes/diabetes., Conclusion: Using FIB-4 in the AGA/AASLD guidelines to risk-stratify subjects at risk for MASLD-associated fibrosis results in many subjects being misclassified into the low- and high-risk categories. Therefore, it may be worthwhile considering caution in interpretation and/or alternative strategies., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background & Aims: In phase 2 studies, efruxifermin, an Fc-FGF21 analog, significantly reduced steatohepatitis and fibrosis in patients with non-alcoholic steatohepatitis, now called metabolic dysfunction-associated steatohepatitis (MASH), for which there is no approved treatment. Type 2 diabetes (T2D) and obesity are prevalent among patients with MASH and increasingly treated with glucagon-like peptide-1 receptor agonists (GLP-1RAs). This study evaluated the safety and efficacy of efruxifermin in patients with MASH, fibrosis, and T2D taking a GLP-1RA., Methods: Cohort D was a double-blind, placebo-controlled, phase 2b study in adults with T2D and MASH with fibrosis (F1-F3) on stable GLP-1RA therapy randomized (2:1) to receive efruxifermin 50 mg or placebo, once weekly for 12 weeks. The primary endpoint was safety and tolerability of efruxifermin added to a stable dose of GLP-1RA. Secondary endpoints included changes in hepatic fat fraction (HFF), markers of liver injury and fibrosis, and metabolic parameters., Results: Adults (N = 31) with T2D and MASH fibrosis (F1-F3) on a stable GLP-1RA (semaglutide, 48.4%; dulaglutide, 45.2%; liraglutide, 6.5%) received efruxifermin 50 mg (n = 21) or placebo (n = 10) for 12 weeks. The addition of efruxifermin to a GLP-1RA appeared safe and well-tolerated. The most frequent efruxifermin-related adverse events were mild to moderate gastrointestinal events. One patient receiving efruxifermin discontinued due to nausea, and another withdrew consent. There were no treatment-related serious adverse events. After 12 weeks, efruxifermin reduced HFF by 65% (P < .0001 vs placebo) compared with a 10% reduction for placebo (GLP-1RA alone). Efruxifermin also improved noninvasive markers of liver injury, fibrosis, glucose, and lipid metabolism while maintaining GLP-1RA-mediated weight loss., Conclusions: The tolerability profile of efruxifermin added to GLP-1RA appeared comparable to that of either drug alone, while also significantly reducing HFF and noninvasive markers of fibrosis in patients with MASH and T2D. Liver health in patients already on a GLP-1RA may be further improved by addition of efruxifermin., Clinicaltrials: gov, Number: NCT05039450., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Oat crown rust, caused by Puccinia coronata Corda f. sp. avenae Eriks. (Pca), is a major biotic impediment to global oat production. Crown rust resistance has been described in oat diploid species A. strigosa accession PI 258731 and resistance from this accession has been successfully introgressed into hexaploid A. sativa germplasm. The current study focuses on 1) mapping the location of QTL containing resistance and evaluating the number of quantitative trait loci (QTL) conditioning resistance in PI 258731; 2) understanding the relationship between the original genomic location in A. strigosa and the location of the introgression in the A. sativa genome; 3) identifying molecular markers tightly linked with PI 258731 resistance loci that could be used for marker assisted selection and detection of this resistance in diverse A. strigosa accessions. To achieve this, A. strigosa accessions, PI 258731 and PI 573582 were crossed to produce 168 F5:6 recombinant inbred lines (RILs) through single seed descent. Parents and RILs were genotyped with the 6K Illumina SNP array which generated 168 segregating SNPs. Seedling reactions to two isolates of Pca (races TTTG, QTRG) were conditioned by two genes (0.6 cM apart) in this population. Linkage mapping placed these two resistant loci to 7.7 (QTRG) to 8 (TTTG) cM region on LG7. Field reaction data was used for QTL analysis and the results of interval mapping (MIM) revealed a major QTL (QPc.FD-AS-AA4) for field resistance. SNP marker assays were developed and tested in 125 diverse A. strigosa accessions that were rated for crown rust resistance in Baton Rouge, LA and Gainesville, FL and as seedlings against races TTTG and QTRG. Our data proposed SNP marker GMI&#95;ES17&#95;c6425&#95;188 as a candidate for use in marker-assisted selection, in addition to the marker GMI&#95;ES02&#95;c37788&#95;255 suggested by Rine's group, which provides an additional tool in facilitating the utilization of this gene in oat breeding programs., Competing Interests: The authors have declared that no competing interests exist, (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Background & Aims: Robust performance of non-invasive tests (NITs) across ages is critical to assess liver disease among patients with metabolic dysfunction-associated liver disease (MASLD). We evaluated the impact of age on the performance of NIS2+™ vs. other NITs., Methods: An analysis cohort (N = 1,926) with biopsy-proven MASLD was selected among individuals screened for the phase III RESOLVE-IT clinical trial and divided into ≤45, 46-55, 56-64, and ≥65 years groups. To avoid potential confounding effects, a well-balanced cohort (n = 708; n = 177/age group) was obtained by applying a propensity score-matching algorithm to the analysis cohort. Baseline values of biomarkers and NITs were compared across age groups using one-way ANOVA, and the impact of age and histology were compared through three-way ANOVA. The impact of age on NIT performance for the detection of at-risk metabolic dysfunction-associated steatohepatitis (MASH; MASLD activity score [MAS] ≥4 and fibrosis stage [F] ≥2) was also evaluated., Results: Age did not affect the distributions of NIS2+™ and APRI (aspartate aminotransferase-to-platelet ratio index), but significantly ( p <0.0001) impacted those of NFS (NAFLD fibrosis score), FIB-4 (Fibrosis-4 index), and Enhanced Liver Fibrosis (ELF™) score. NIS2+™ was the only NIT on which fibrosis and MAS exerted a moderate to large effect. While the impact of fibrosis on APRI was moderate, that of MAS was low. The impact of age on FIB-4 and NFS was larger than that of fibrosis. NIS2+™ exhibited the highest AUROC values for detecting at-risk MASH across age groups, with stable performances irrespective of cut-offs., Conclusions: NIS2+™ was not significantly impacted by age and was sensitive to both fibrosis and MAS grade, demonstrating a robust performance to rule in/out at-risk MASH with fixed cut-offs., Impact and Implications: While metabolic dysfunction-associated steatotic liver disease (MASLD) can affect individuals of all ages, patient age could represent an important confounding factor when interpreting non-invasive test (NIT) results, highlighting the need for reliable and efficient NITs that are not impacted by age and that could be interpreted with fixed cut-offs, irrespective of patient age. We report the impact of age on different well-established NITs - among those tested, only two panels, NIS2+™ and APRI, were not impacted by age and can be used and interpreted independently of patient age. NIS2+™ was also sensitive to both fibrosis and MAS, further confirming its efficiency for the detection of the composite endpoint of at-risk MASH and its potential as a valuable candidate for large-scale implementation in clinical practice and clinical trials., Competing Interests: QMA: research support from LITMUS (Liver Investigation: Testing Marker Utility in Steatohepatitis) consortium funded by the Innovative Medicines Initiative Program of the European Union under Grant Agreement 777377 (this multistakeholder consortium includes industry partners and received funding from EFPIA); grants or contracts from AstraZeneca, Boehringer Ingelheim, and Intercept Pharmaceuticals; royalties or licenses from Elsevier Ltd.; consulting fees (on behalf of Newcastle University) from Alimentiv, Akero Therapeutics, Inc., AstraZeneca, Axcella Health, Inc., 89bio, Inc., Boehringer Ingelheim, Bristol Myers Squibb, Galmed Pharmaceuticals, Genfit S.A., Genentech, Gilead Sciences Inc., GlaxoSmithKline, Hanmi, HistoIndex Pte Ltd., Intercept Pharmaceuticals, Inventiva, Ionis, IQVIA, Janssen, Madrigal Pharmaceuticals, Medpace, Merck, NGM Biopharmaceuticals, Novartis Pharmaceuticals, Novo Nordisk, PathAI, Pfizer, Prosciento, Poxel S.A., Resolution Therapeutics, Roche, Ridgeline Therapeutics, RTI, Shionogi, and Terns Pharmaceuticals; payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events from Fishawack, Integritas Communications, Kenes, Novo Nordisk, Madrigal Pharmaceuticals, Medscape, and Springer Healthcare; served on advisory boards or data safety monitoring boards (on behalf of Newcastle University) for Medpace (NorthSea Therapeutics B.V., DSMB). BS: consulting fees from Genfit S.A. MAC: Labcorp employee. RL: grants/funding support from the National Center for Advancing Translational Sciences, the National Institute of Diabetes and Digestive and Kidney Diseases, the National Heart, Lung, and Blood Institute, Arrowhead Pharmaceuticals, AstraZeneca, Boehringer Ingelheim, Bristol Myers Squibb, Eli Lilly, Galectin Therapeutics, Inc., Galmed Pharmaceuticals, Gilead Sciences, Inc., Hanmi, Intercept Pharmaceuticals, Inventiva, Ionis, Janssen, Madrigal Pharmaceuticals, Merck, NGM Biopharmaceuticals, Novo Nordisk, Pfizer, Sonic Incytes, and Terns Pharmaceuticals; consulting fees from Aardvark Therapeutics, Altimmune, Anylam/Regeneron, Amgen, Arrowhead Pharmaceuticals, AstraZeneca, Bristol Myers Squibb, CohBar, Inc., Eli Lilly, Galmed Pharmaceuticals, Gilead Sciences, Inc., Glympse Bio, HighTide Therapeutics, Ini Pharma, Intercept Pharmaceuticals, Intercept Pharmaceuticals, Ionis, Janssen, Madrigal Pharmaceuticals, Metacrine, NGM Biopharmaceuticals, Novartis Pharmaceuticals, Novo Nordisk, Merck, Pfizer, Sagimet Biosciences, Theratechnologies, Inc., 89bio, Inc., Terns Pharmaceuticals, and Viking Therapeutics; other financial interests: LipoNexus, Inc. (co-founder). SAH: grants or contracts from Akero Therapeutics, Alentis Therapeutics, Altimmune, B. Riley FBR, ChronWell, Corcept Therapeutics, Echosens, Axcella Health, Cirius Therapeutics, CiVi Biopharma, Cymabay Therapeutics, Inc., Enyo Pharma S.A., Galectin Therapeutics, Inc., Galmed Research & Development, Ltd., Genfit S.A., Gilead Sciences, Inc., Hepion Pharmaceuticals, Inc., Hepta Bio, HighTide Therapeutics, Inc, HistoIndex, Intercept Pharmaceuticals, Ionis, Madrigal Pharmaceuticals, Medpace, NGM Biopharmaceuticals, Inc., NeuroBo, NorthSea Therapeutics B.V., Novartis Pharmaceuticals, Novo Nordisk, Path AI, Perspectum, Poxel S.A., Sagimet Biosciences, Sonic Incytes, Terns Pharmaceuticals, and Viking Therapeutics; stock or stock options for Akero Therapeutics, ChronWell, Cirius Therapeutics, Galectin Therapeutics, Inc., Genfit S.A., Hepion Pharmaceuticals, Inc., HistoIndex Pte Ltd., Metacrine, NGM Biopharmaceuticals, Inc., NorthSea Therapeutics B.V. VR: grants or contracts from Intercept Pharmaceuticals, and Gilead Sciences, Inc.; consulting fees from Boehringer Ingelheim, Novo Nordisk, Poxel S.A., Enyo Pharma S.A., Madrigal Pharmaceuticals, Terns Pharmaceuticals, Intercept Pharmaceuticals, NGM Biopharmaceuticals Inc., and Pfizer. AJS: stock options in Genfit S.A., Tiziana, Indalo, Durect, Inversago, and Galmed Pharmaceuticals; consultant to AstraZeneca, Salix, Tobira, Takeda, Jannsen, Gilead Sciences, Inc., Terns Pharmaceuticals, Merck, Madrigal Pharmaceuticals, NGM Biopharmaceuticals, Inc., Sagimet Biosciences, Valeant, Boehringer Ingelheim, Bristol Myers Squibb, Eli Lilly, Hemoshear, Novartis Pharmaceuticals, Inventiva, Enyo, Akero Therapeutics, 89bio, Inc., Novo Nordisk, Pfizer, Amgen, Genentech, Regeneron, Alnylam, Hanmi, LG Chem, Histoindex, Thera Technologies, Intercept Pharmaceuticals, Target-RWE, Surrozen, Zydus, Path AI, Exhalenz, and Genfit S.A.; his institution has received grant support from Gilead Sciences, Inc., Salix, Tobira, Bristol Myers Squibb, Pfizer, Intercept Pharmaceuticals, Merck, AstraZeneca, Mallinckrodt, and Novartis Pharmaceuticals; royalties received from Elsevier and UpToDate. JM, YH, AC, ZM, CR, and DWH: stock or stock options from Genfit S.A. and serve as Genfit S.A. employees. Please refer to the accompanying ICMJE disclosure forms for further details., (© 2024 The Author(s).)

Background: Causality assessment of suspected drug-induced liver injury (DILI) during metabolic dysfunction-associated steatohepatitis (MASH) clinical trials can be challenging, and liver biopsies are not routinely performed as part of this evaluation. While the field is moving away from liver biopsy as a diagnostic and prognostic tool, information not identified by non-invasive testing may be provided on histology., Aim: To address the appropriate utilisation of liver biopsy as part of DILI causality assessment in this setting., Methods: From 2020 to 2022, the Liver Forum convened a series of webinars on issues pertaining to liver biopsy during MASH trials. The Histology Working Group was formed to generate a series of consensus documents addressing these challenges. This manuscript focuses on liver biopsy as part of DILI causality assessment., Results: Expert opinion, guidance and recommendations on the role of liver biopsy as part of causality assessment of suspected DILI occurring during clinical trials for a drug(s) being developed for MASH are provided. Lessons learned from prior MASH programs are reviewed and gaps identified., Conclusions: Although there are no pathognomonic features, histologic evaluation of suspected DILI during MASH clinical trials may alter patient management, define the pattern and severity of injury, detect findings that favour a diagnosis of DILI versus MASH progression, identify prognostic features, characterise the clinicopathological phenotype of DILI, and/or define lesions that influence decisions about trial discontinuation and further development of the drug., (© 2023 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.)

Background: Non-alcoholic steatohepatitis (NASH) is a progressive form of non-alcoholic fatty liver disease (NAFLD) associated with steatosis, hepatocellular injury, inflammation and fibrosis. In a Phase 2 trial in adults with NASH (NCT02912260), resmetirom, an orally administered, liver-targeted thyroid hormone receptor-β selective agonist, significantly reduced hepatic fat (via imaging) and resolved NASH without worsening fibrosis (via liver biopsy) in a significant number of patients compared with placebo., Aims: To present the design of the Phase 3 MAESTRO clinical programme evaluating resmetirom for treatment of NASH (MAESTRO-NAFLD-1 [NCT04197479], MAESTRO-NAFLD-OLE [NCT04951219], MAESTRO-NASH [NCT03900429], MAESTRO-NASH-OUTCOMES [NCT05500222])., Methods: MAESTRO-NASH is a pivotal serial biopsy trial in up to 2000 adults with biopsy-confirmed at-risk NASH. Patients are randomised to a once-daily oral placebo, 80 mg resmetirom, or 100 mg resmetirom. Liver biopsies are conducted at screening, week 52 and month 54. MAESTRO-NAFLD-1 is a 52-week safety trial in ~1400 adults with NAFLD/presumed NASH (based on non-invasive testing); ~700 patients from MAESTRO-NAFLD-1 are enrolled in MAESTRO-NAFLD-OLE, a 52-week active treatment extension to further evaluate safety. MAESTRO-NASH-OUTCOMES is enrolling 700 adults with well-compensated NASH cirrhosis to evaluate the potential for resmetirom to slow progression to hepatic decompensation events. Non-invasive tests (biomarkers, imaging) are assessed longitudinally throughout, in addition to validated patient-reported outcomes., Conclusion: The MAESTRO clinical programme was designed in conjunction with regulatory authorities to support approval of resmetirom for treatment of NASH. The surrogate endpoints, based on week 52 liver biopsy, serum biomarkers and imaging, are confirmed by long-term clinical liver-related outcomes in MAESTRO-NASH (month 54) and MAESTRO-NASH-OUTCOMES (time to event)., (© 2023 Madrigal Pharmaceuticals. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.)

Background: Seladelpar is a potent and selective peroxisome proliferator-activated receptor-δ agonist that targets multiple cell types involved in primary biliary cholangitis (PBC), leading to anti-cholestatic, anti-inflammatory and anti-pruritic effects., Aims: To evaluate the long-term safety and efficacy of seladelpar in patients with PBC., Methods: In an open-label, international, long-term extension study, patients with PBC completing seladelpar lead-in studies continued treatment. Seladelpar was taken orally once daily at doses of 5 or 10 mg with dose adjustment permitted for safety or tolerability. The primary analysis was for safety and the secondary efficacy analysis examined biochemical markers of cholestasis and liver injury. The study was terminated early due to the unexpected histological findings in a concurrent study for non-alcoholic steatohepatitis, which were subsequently found to predate treatment. Safety and efficacy data were analysed through 2 years., Results: There were no serious treatment-related adverse events observed among 106 patients treated with seladelpar for up to 2 years. There were four discontinuations for safety, one possibly related to seladelpar. Among 53 patients who completed 2 years of seladelpar, response rates increased from years 1 to 2 for the composite endpoint (alkaline phosphatase [ALP] <1.67 × ULN, ≥15% decrease in ALP, and total bilirubin ≤ULN) and ALP normalisation from 66% to 79% and from 26% to 42%, respectively. In those with elevated bilirubin at baseline, 43% achieved normalisation at year 2., Conclusions: Seladelpar was safe, and markedly improved biochemical markers of cholestasis and liver injury in patients with PBC. These effects were maintained or improved throughout the second year., Clinicaltrials: gov: NCT03301506; Clinicaltrialsregister.eu: 2017-003910-16., (© 2023 CymaBay Therapeutics, Inc and The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.)

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 favor 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 panelists 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. 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 nonstigmatising, and can improve awareness and patient identification., Competing Interests: Declaration of interests Manal F. Abdelmalek consults, advises, and received grants from Bristol Myers Squibb, Hanmi, Intercept, Inventiva, and Madrigal. She consults and advises 89Bio, Merck, NGM Bio, Novo Nordisk, Sonic Incytes, and Theratechnologies. She is on the speakers’ bureau for the Chronic Liver Disease Foundation, Clinical Care Options, Fishawack, Medscape, and Terra Firma. She received grants from Allergan, Boehringer Ingelheim, Celgene, Durect, Enanta, Enyo, Galmed, Genentech, Gilead, Novo Nordisk, Poxel, Target NASH, and Viking. Quentin M. Anstee, on behalf of Newcastle University, consults for Alimentiv, Akero, AstraZeneca, Axcella, 89Bio, Boehringer Ingelheim, Bristol Myers Squibb, Galmed, GENFIT, Genentech, Gilead, GSK, Hanmi, HistoIndex, Intercept, Inventiva, Ionis, IQVIA, Janssen, Madrigal, Medpace, Merck, NGM Bio, Novartis, Novo Nordisk, PathAI, Pfizer, Pharmanest, Prosciento, Poxel, RTI, Resolution Therapeutics, Ridgeline Therapeutics, Roche, Shionogi, and Terns. He is on the speakers’ bureau for Fishawack, Integritas Communications, Kenes, Novo Nordisk, Madrigal, Medscape, and Springer Healthcare. He received grants from AstraZeneca, Boehringer Ingelheim, and Intercept. He holds intellectual property rights with Elsevier, Ltd. Ramon Bataller is on the speakers’ bureau for Abbvie and Gilead. Ulrich Beuers consults for CSL Behring. He is on the speakers’ bureau for Abacus and Zambon. Elisabetta Bugianesi advises Boehringer Ingelheim, MSD, and Novo Nordisk. Helena Cortez-Pinto consults and received grants from Novo Nordisk and Roche. She received grants from Eisai, Gilead, GMP-Orphan, and Intercept. Kenneth Cusi Consults for Aligos, Arrowhead, AstraZeneca, Boehringer Ingelheim, Bristol Myers Squibb, Covance, Lilly, Madrigal, Myovant, Novo Nordisk, Prosciento, Sagimet, and Siemens. He received grants from Echosens, Inventiva, LabCorp, Nordic Biosciences, and Target NASH. Sven M. Francque consults and received grants from Astellas, Falk, GENFIT, Gilead, Glympse Bio, Janssen, Inventiva, Merck, Pfizer, and Roche. He consults for AbbVie., Actelion, Aelin Therapeutics, Allergan, AstraZeneca, Bayer, Boehringer Ingelheim, Bristol Myers Squibb, Coherus, CSL Behring, Echosens, Eisai, ENYO, Galapagos, Galmed, Genetech, Intercept, Julius Clinical, Madrigal, Medimmune, NGM Bio, Novartis, Novo Nordisk, and Promethera. Samer Gawrieh consults for Pfizer and TransMedics. He received grants from LiverIncytes, Viking, and Zydus. Manuel Romero-Gómez advises and received grants from Novo Nordisk and Siemens. He advises AbbVie., Alpha-sigma, Allergan, AstraZeneca, Axcella, Boehringer Ingelheim, Bristol Myers Squibb, Gilead, Intercept, Inventia, Kaleido, MSD, Pfizer, Prosciento, Rubió, Shionogi, Sobi, and Zydus. He received grants from Echosens and Theratechnologies. Cynthia D. Guy consults for 89Bio, CymaBay, HistoIndex, Madrigal, and NGM. Stephen Harrison consults, advises, is involved with trials, received grants, and owns stock in Akero, Galectin, GENFIT, Hepion, and NGM Bio. He consults, advises, is involved with trials, and received grants from Axcella, Gilead, Intercept, Madrigal, and Poxel. He consults, advises, received grants, and owns stock in NorthSea Therapeutics. He consults, advises, and is involved with trials for Terns. He consults, advises, and received grants from HighTide, Novartis, Novo Nordisk, and Sagimet. He consults, advises, and owns stock in HistoIndex, Metacrine, and Sonic Incytes. He consults, received grants, and owns stock in Cirius. He consults, is involved with trials, and received grants from ENYO and Viking. He is involved with trials and received grants from Genentech. He consults and is involved with trials for Ionis. He consults and received grants from CiVi, CymaBay, Galmed, and Pfizer. He consults and owns stock in Hepta Bio. He consults and advises for Altimmune, Echosens North America, Foresite Labs, and Medpace. He advises and owns stock in ChronWell. He consults for AgomAb, Alentis, Aligos Therapeutics, Alimentiv, Blade, Bluejay, Boston Pharmaceuticals, Boxer Capital, CanFite BioPharma, the Chronic Liver Disease Foundation (CLDF), CohBar, Corcept, Fibronostics, Fortress Biotech, Galecto, Gelesis, GSK, GNS Healthcare, GRI Bio, Hepagene, Indalo, Inipharm, Innovate Biopharmaceuticals, Kowa Research Institute, Merck, MGGM, NeuroBo, Nutrasource, Perspectum, Piper Sandler, Prometic (now Liminal BioSciences), Ridgeline Therapeutics, Silverback, and Zafgen (now Larimar). He advises Arrowhead BVF Partners, Humana, and Pathai. He received grants from Bristol Myers Squibb, Conatus, Immuron, and Second Genome. Samuel Klein advises Alnylam, Altimmune, and Merck. Kris V. Kowdley advises, is on the speakers’ bureau, and received grants from Gilead and Intercept. He advises, received grants, and owns stock in Inipharm. He advises and received grants from 89bio, CymaBay, GENFIT, Ipsen, Madrigal, Mirum, NGM Bio, Pfizer, Pliant, and Zeds. He advises Enact, HighTide, and Protagonist. He is on the speakers’ bureau for AbbVie. He received grants from Boston Pharmaceuticals, Corcept, GSK, Hanmi, Janssen, Novo Nordisk, Terns, and Viking. Jeffrey V. Lazarus consults for Novavax. He received grants from AbbVie, Gilead, MSD, and Roche Diagnostics. Rohit Loomba consults and received grants from Arrowhead, AstraZeneca, Bristol Myers Squibb, Galmed, Gilead, Intercept, Inventiva, Ionis, Janssen, Lilly, Madrigal, Merck, NGM Bio, Novo Nordisk, Pfizer, and Terns. He consults and owns stock in 89Bio and Sagimet. Consults for Altimmune, Anylam, Amgen, CohBar, Glympse Bio, HighTide, Inipharm, Metacrine, Novartis, Regeneron, Theratechnologies, and Viking. He received grants from Boehringer Ingelheim, Galectin Therapeutics, Hanmi, and Sonic Incytes. He cofounded and owns stock in LipoNexus. Phillip N. Newsome consults, advises, is on the speakers’ bureau, and received grants from Novo Nordisk. He consults and advises Boehringer Ingelheim, Bristol Myers Squibb, Gilead, GSK, Intercept, Madrigal, Pfizer, Poxel, and Sun Pharma. He is on the speakers’ bureau for AiCME. Elizabeth E. Powell advises and received grants from Novo Nordisk. Vlad Ratziu consults and received grants from Intercept. He consults for Boehringer Ingelheim, Eny, Madrigal, NorthSea, Novo Nordisk, E. Poxel, and Sagimet. He received grants from Gilead. Mary E. Rinella consults for Boehringer Ingelheim, CytoDyn, GSK, Novo Nordisk, HistoIndex, Intercept, Madrigal, NGM Bio, and Sonic Incytes. Michael Roden consults and received grants from Boehringer Ingelheim and Novo Nordisk. He consults for Lilly. He is on the speakers’ bureau for AstraZeneca. Arun J. Sanyal consults and advises Avant Santé and AstraZeneca. He consults and received grants from Akero, Bristol Myers Squibb, Intercept, Lilly, Madrigal, and Novo Nordisk. He consults and owns stock in Rivus. He consults for AGED Diagnostics, Albireo, Alnylam, Altimmune, Boehringer Ingelhiem, 89Bio, Echosense, Genentech, Gilead, GSK, HistoIndex, Malinckrodt, Merck, NGM Bio, Novartis, PathAI, Pfizer, Poxel, Regeneron, Salix, Siemens, Surrozen, Takeda, Terns, and Zydus. He owns stock in Durect, Exhalenz, GENFIT, Indalo, Inversago, and Tiziana. He received royalties from Elsevier and Wolters Kluwer. Marcelo Silva consults, advises, and received grants from Zydus. He received grants from Inventiva and MSD. Dina Tiniakos consults for Clinnovate Health, ICON, Ionis, Inventiva, Merck, and Verily. Luca Valenti consults and received grants from Gilead. He consults for AstraZeneca, Boehringer Ingelheim, MSD, Novo Nordisk, Pfizer, and Resalis Therapeutics. Miriam B. Vos consults and advises Thiogenesis. She consults and received grants from Target Real World Evidence. She consults and owns stock in Intercept. She consults for Albireo, Boehringer Ingelheim, Lilly, Novo Nordisk, and Takeda. She received grants from Bristol Myers Squibb, Quest, and Sonic Incytes. Vincent Wai-Sun Wong consults and received grants from Gilead. He consults for AbbVie, Boehringer Ingelheim, Echosens, Intercept, Inventiva, Novo Nordisk, Pfizer, Sagimet, and TARGET PharmaSolutions. He owns stock in Illuminatio Medical Technology. Yusuf Yilmaz consults for Zydus. He advises Novo Nordisk. He is on the speakers’ bureau for Echosens. Zobair Younossi consults for Bristol Myers Squibb, Gilead, Intercept, Madrigal, Merck, Novartis, Novo Nordisk, Quest, Siemens, and Terns. The remaining authors have no conflicts to report., (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 D. 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 (senior), NAFLD Nomenclature consensus group. Published by Elsevier España, S.L.U. All rights reserved.)

Background & Aims: Pegbelfermin is a polyethylene glycol-conjugated analog of human fibroblast growth factor 21, a nonmitogenic hormone that regulates energy metabolism. This phase 2b study evaluated 48-week pegbelfermin treatment in patients with nonalcoholic steatohepatitis (NASH) with compensated cirrhosis., Methods: FALCON 2 (NCT03486912) was a randomized (1:1:1:1), double-blind, placebo-controlled study. Eligible adults had biopsy-confirmed NASH and stage 4 fibrosis. Pegbelfermin (10, 20, or 40 mg) or placebo was injected subcutaneously once weekly. The primary endpoint was 1 or more stages of improvement in the NASH Clinical Research Network fibrosis score without NASH worsening at week 48; pegbelfermin dose response was assessed using a Cochran-Armitage trend test across proportions (1-sided α = .05). Additional endpoints included histologic and noninvasive measures of steatosis, fibrosis, and liver injury/inflammation., Results: Overall, 155 patients were randomized, and 154 patients received treatment. At week 48, 24% to 28% of the pegbelfermin arms had primary endpoint responses vs 31% of the placebo arm (P = .361). Nonalcoholic fatty liver disease activity score improvements were more frequent with pegbelfermin vs placebo and were driven primarily by reduced lobular inflammation. Numerically higher proportions of the pegbelfermin arms had liver stiffness (magnetic resonance elastography) and steatosis (magnetic resonance imaging-proton density fat fraction) improvements vs placebo; these differences were not statistically significant. Mean N-terminal type III collagen propeptide, alanine aminotransferase, and aspartate aminotransferase values were numerically lower in the 20- and/or 40-mg pegbelfermin arms compared with placebo. Serious adverse events were more frequent with pegbelfermin vs placebo, although none were treatment related. One patient (40-mg pegbelfermin) discontinued treatment because of a treatment-emergent adverse event (worsening ascites)., Conclusions: FALCON 2 did not meet its primary endpoint of 1 or more stages of improvement in the NASH Clinical Research Network fibrosis without NASH worsening assessed via biopsy. Pegbelfermin generally was well tolerated in this advanced NASH population., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Pegbelfermin is a polyethlene glycol-conjugated analog of human fibroblast growth factor 21, a nonmitogenic hormone that regulates energy metabolism. This phase 2b study evaluated 48-week pegbelfermin treatment in patients with nonalcoholic steatohepatitis (NASH) and stage 3 (bridging) fibrosis., Methods: The FALCON 1 study (NCT03486899) was a multicenter, randomized (1:1:1:1), double-blind, placebo-controlled study. Patients with biopsy-confirmed NASH and stage 3 fibrosis (N = 197) received weekly subcutaneous pegbelfermin (10, 20, or 40 mg) or placebo injections for 48 weeks. The week 24 primary endpoint was a ≥1-point decrease in fibrosis score without NASH worsening or NASH improvement without fibrosis worsening; pegbelfermin dose response was assessed using a Cochran-Armitage trend test across proportions (1-sided α = 0.05). Secondary/exploratory endpoints included histological and noninvasive measures of steatosis, fibrosis, and liver injury/inflammation., Results: At week 24, the primary endpoint was met by 14% (placebo) vs 24%-31% (pegbelfermin arms); statistical significance was not reached due to lack of pegbelfermin dose response (P = .134). At weeks 24 and 48, more patients who received pegbelfermin had ≥30% relative reductions in hepatic fat fraction (magnetic resonance imaging-proton density fat fraction) vs placebo, although no differences reached statistical significance. In the pegbelfermin arms, improvements in liver fibrosis (magnetic resonance elastography and N-terminal type III collagen propeptide) and liver injury/inflammation (alanine aminotransferase, aspartate aminotransferase) were observed vs placebo. Adverse events occurred at similar frequencies across arms. No treatment-related serious adverse events were observed., Conclusions: The FALCON 1 study did not meet its primary endpoint; a ≥1-point decrease in fibrosis score without NASH worsening or NASH improvement without fibrosis worsening assessed via biopsy. Pegbelfermin was generally well tolerated during 48 weeks of treatment., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Liver histopathologic assessment is the accepted surrogate endpoint in NASH trials; however, the scoring of NASH Clinical Research Network (CRN) histologic parameters is limited by intraobserver and interobserver variability. We designed a consensus panel approach to minimize variability when using this scoring system. We assessed agreement between readers, estimated linear weighted kappas between 2 panels, compared them with published pairwise kappa estimates, and addressed how agreement or disagreement might impact the precision and validity of the surrogate efficacy endpoint in NASH trials., Methods: Two panels, each comprising 3 liver fellowship-trained pathologists who underwent NASH histology training, independently evaluated scanned whole slide images, scoring fibrosis, inflammation, hepatocyte ballooning, and steatosis from baseline and month 18 biopsies for 100 patients from the precirrhotic NASH study REGENERATE. The consensus score for each parameter was defined as agreement by ≥2 pathologists. If consensus was not reached, all 3 pathologists read the slide jointly to achieve a consensus score., Results: Between the 2 panels, the consensus was 97%-99% for steatosis, 91%-93% for fibrosis, 88%-92% for hepatocyte ballooning, and 84%-91% for inflammation. Linear weighted kappa scores between panels were similar to published NASH CRN values., Conclusions: A panel of 3 trained pathologists independently scoring 4 NASH CRN histology parameters produced high consensus rates. Interpanel kappa values were comparable to NASH CRN metrics, supporting the accuracy and reproducibility of this method. The high concordance for fibrosis scoring was reassuring, as fibrosis is predictive of liver-specific outcomes and all-cause mortality., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Association for the Study of Liver Diseases.)

S100 proteins are a subfamily of EF-hand calcium-binding proteins found primarily in vertebrate animals. They are distinguished by binding of transition metals and functioning in both the intracellular and extracellular milieu. S100A7 functions in the protection of the skin and mucous membranes and is a biomarker in inflammatory skin disease. A recent study of Neisseria gonorrhoeae infection revealed that human but not murine S100A7 could be used to evade host nutritional immunity. To understand the molecular basis for this difference, we carried out a comparative analysis of the physical and structural properties of human and murine S100A7. The X-ray crystal structure of Ca 2+ -loaded mouse S100A7 (mS100A7) was determined to 1.69 Å resolution, and Ca 2+ -induced conformational changes were assessed by NMR. Unlike human S100A7 (hS100A7), which exhibits conformational changes in response to binding of Ca 2+ , no significant changes in mS100A7 were detected. Dynamic light scattering, circular dichroism, and a competition chelator assay were used to compare the Zn 2+ affinity and the effects of ion binding on mS100A7 versus hS100A7. Alignment of their sequences revealed a substantial difference in the C-terminal region, which is an important mediator of protein-protein interactions, suggesting a rationale for the specificity of N. gonorrhoeae for hS100A7. These data, along with more detailed analysis of S100A7 sequence conservation across different species, support the proposal that, although hS100A7 is highly conserved in many mammals, the murine protein is a distinct ortholog. Our results highlight the potential limitations of using mouse models for studying bacterial infections in humans., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Nonalcoholic steatohepatitis (NASH) is a progressive liver disease with no approved treatment. MAESTRO-NAFLD-1 was a 52-week randomized, double-blind, placebo-controlled phase 3 trial evaluating the safety of resmetirom in adults with nonalcoholic fatty liver disease and presumed NASH. Patients were randomized to three double-blind arms (100 mg resmetirom (n = 325), 80 mg resmetirom (n = 327) or placebo (n = 320)) or open-label 100 mg resmetirom (n = 171). The primary end point was incidence of treatment-emergent adverse events (TEAEs) over 52 weeks and key secondary end points were LDL-C, apoB, triglycerides (over 24 weeks), hepatic fat (over 16 and 52 weeks) and liver stiffness (over 52 weeks). Resmetirom was safe and well tolerated. TEAEs occurred in 86.5% (open-label 100 mg resmetirom), 86.1% (100 mg resmetirom), 88.4% (80 mg resmetirom) and 81.8% (placebo) of patients. TEAEs in excess of placebo included diarrhea and nausea at the initiation of treatment. Key secondary end points included least square means difference from placebo at 80 mg, 100 mg resmetirom: LDL-C (-11.1%, -12.6%), apoB (-15.6%, -18.0%), triglycerides (-15.4%, -20.4%), 16-week hepatic fat (-34.9%, -38.6%), (P < 0.0001) and liver stiffness (-1.02, -1.70) and 52-week hepatic fat (-28.8, -33.9). These findings demonstrate resmetirom was safe and well tolerated in adults with presumed NASH, supporting a role for further clinical development. (ClinicalTrials.gov identifier NCT04197479 )., (© 2023. The Author(s).)

Background: An approved therapy for nonalcoholic steatohepatitis (NASH) and fibrosis remains a major unmet medical need., Aim: To investigate the histological and metabolic benefits of pegozafermin, a glycoPEGylated FGF21 analogue, in subjects with biopsy-confirmed NASH., Methods: This proof-of-concept, open-label, single-cohort study, part 2 of a phase 1b/2a clinical trial, was conducted at 16 centres in the United States. Adults (age 21-75 years) with NASH (stage 2 or 3 fibrosis, NAS≥4) and magnetic resonance imaging proton density fat fraction (MRI-PDFF) ≥8% received subcutaneous pegozafermin 27 mg once weekly for 20 weeks. Primary outcomes were improvements in liver histology, and safety and tolerability., Results: Of 20 enrolled subjects, 19 completed the study. Twelve subjects (63%) met the primary endpoint of ≥2-point improvement in NAFLD activity score with ≥1-point improvement in ballooning or lobular inflammation and no worsening of fibrosis. Improvement of fibrosis without worsening of NASH was observed in 26% of subjects, and NASH resolution without worsening of fibrosis in 32%. Least-squares mean relative change from baseline in MRI-PDFF was -64.7% (95% CI: -71.7, -57.7; p < 0.0001). Significant improvements from baseline were also seen in serum aminotransferases, noninvasive fibrosis tests, serum lipids, glycaemic control and body weight. Adverse events (AEs) were reported in 18 subjects (90%). The most frequently reported AEs were mild/moderate nausea and diarrhoea. There were no serious AEs, discontinuations due to AEs, or deaths., Conclusions: Pegozafermin treatment for 20 weeks had beneficial effects on hepatic and metabolic parameters and was well tolerated in subjects with NASH., Clinicaltrials: gov: NCT04048135., (© 2023 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.)

Background & Aims: The MRI-AST (MAST) score accurately identifies patients with at-risk nonalcoholic steatohepatitis, defined as nonalcoholic steatohepatitis with nonalcoholic fatty liver disease activity score ≥4 and fibrosis stage ≥2 at highest risk for disease progression. It is important to determine the robustness of the MAST score in predicting major adverse liver outcomes (MALO), hepatocellular carcinoma (HCC), liver transplant, and death., Methods: This retrospective analysis included patients with nonalcoholic fatty liver disease from a tertiary care center who underwent magnetic resonance imaging proton density fat fraction, magnetic resonance elastography, and laboratory testing within 6 months from 2013 to 2022. Other causes of chronic liver disease were excluded. Hazard ratios between logit MAST and MALO (ascites, hepatic encephalopathy, or bleeding esophageal varices), liver transplant, HCC, or liver-related death were computed using a Cox proportional hazards regression model. We computed the hazard ratio of MALO or death associated with MAST scores 0.165-0.242 and 0.242-1.000, using MAST scores 0.000-0.165 as the reference group., Results: Among 346 total patients, average age was 58.8 years with 52.9% females and 34.4% with type 2 diabetes. Average alanine aminotransferase was 50.7 IU/L (24.3-60.0 IU/L), aspartate aminotransferase was 38.05 IU/L (22.00-41.00 IU/L), platelets were 242.9 × 10 9 /L (193.8-290.0 × 10 9 /L), proton density fat fraction was 12.90% (5.90%-18.22%), and liver stiffness on magnetic resonance elastography was 2.75 kPa (2.07-2.90 kPa). Median follow-up was 29.5 months. Fourteen had adverse outcomes, including 10 MALO, 1 HCC, 1 liver transplant, and 2 liver-related deaths. The Cox regression of MAST versus adverse event rate had a hazard ratio of 2.01 (95% confidence interval, 1.59-2.54; P < .0001) for each 1 logit unit increases in MAST. The corresponding Harrell concordance statistic (C statistic) was 0.919 (95% confidence interval, 0.865-0.953). The MAST score ranges of 0.165-0.242 and 0.242-1.0, respectively, had adverse event rate hazard ratio of 7.75 (1.40-42.9; P = .0189) and 22.11 (6.59-74.2; P < .0000) relative to MAST 0-0.165., Conclusions: The MAST score noninvasively identifies at-risk nonalcoholic steatohepatitis and accurately predicts MALO, HCC, liver transplant, and liver-related death., (Copyright © 2023 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Nonalcoholic fatty liver disease consists of a spectrum starting from nonalcoholic fatty liver disease that may progress to nonalcoholic steatohepatitis (NASH), which can lead to fibrosis, cirrhosis, hepatocellular carcinoma, or even liver failure. The prevalence of NASH has increased in parallel with the rising rate of obesity and type 2 diabetes. Given the high prevalence and deadly complications of NASH, there have been significant efforts to develop effective treatments. Phase 2A studies have assessed various mechanisms of action across the spectrum of the disease, while phase 3 studies have focused mainly on NASH and fibrosis stage 2 and higher, as these patients have a higher risk of disease morbidity and mortality. The primary efficacy endpoints also vary, by using noninvasive tests in early-phase trials while relying on liver histological endpoints in phase 3 studies as required by regulatory agencies. Despite initial disappointment due to the failure of several drugs, recent phase 2 and 3 studies have shown promising results, with the first Food and Drug Administration-approved drug for NASH expected to be approved in 2023. In this review, we discuss the various drugs under development for NASH, their mechanisms of action, and the results of their clinical trials. We also highlight the potential challenges in developing pharmacological therapies for NASH., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

A protometabolic approach to the origins of life assumes that the conserved biochemistry of metabolism has direct continuity with prebiotic chemistry. One of the most important amino acids in modern biology is aspartic acid, serving as a nodal metabolite for the synthesis of many other essential biomolecules. Aspartate's prebiotic synthesis is complicated by the instability of its precursor, oxaloacetate. In this paper, we show that the use of the biologically relevant cofactor pyridoxamine, supported by metal ion catalysis, is sufficiently fast to offset oxaloacetate's degradation. Cu 2+ -catalysed transamination of oxaloacetate by pyridoxamine achieves around a 5% yield within 1 h, and can operate across a broad range of pH, temperature, and pressure. In addition, the synthesis of the downstream product β-alanine may also take place in the same reaction system at very low yields, directly mimicking an archaeal synthesis route. Amino group transfer supported by pyridoxal is shown to take place from aspartate to alanine, but the reverse reaction (alanine to aspartate) shows a poor yield. Overall, our results show that the nodal metabolite aspartate and related amino acids can indeed be synthesised via protometabolic pathways that foreshadow modern metabolism in the presence of the simple cofactor pyridoxamine and metal ions.

Laboratory models are critical to basic and translational microbiology research. Models serve multiple purposes, from providing tractable systems to study cell biology to allowing the investigation of inaccessible clinical and environmental ecosystems. Although there is a recognized need for improved model systems, there is a gap in rational approaches to accomplish this goal. We recently developed a framework for assessing the accuracy of microbial models by quantifying how closely each gene is expressed in the natural environment and in various models. The accuracy of the model is defined as the percentage of genes that are similarly expressed in the natural environment and the model. Here, we leverage this framework to develop and validate two generalizable approaches for improving model accuracy, and as proof of concept, we apply these approaches to improve models of Pseudomonas aeruginosa infecting the cystic fibrosis (CF) lung. First, we identify two models, an in vitro synthetic CF sputum medium model (SCFM2) and an epithelial cell model, that accurately recapitulate different gene sets. By combining these models, we developed the epithelial cell-SCFM2 model which improves the accuracy of over 500 genes. Second, to improve the accuracy of specific genes, we mined publicly available transcriptome data, which identified zinc limitation as a cue present in the CF lung and absent in SCFM2. Induction of zinc limitation in SCFM2 resulted in accurate expression of 90% of P. aeruginosa genes. These approaches provide generalizable, quantitative frameworks for microbiological model improvement that can be applied to any system of interest.

Background and Aims: We explored 2 novel scores, Agile 3+ and 4, to identify advanced fibrosis (≥F3) and cirrhosis (F4), respectively, in NAFLD and compared their diagnostic performances to liver stiffness measurement (LSM) by vibration-controlled transient elastography and fibrosis-4 index (FIB-4) (for Agile 3+)., Approach and Results: This multicenter study included 548 NAFLD patients with laboratory testing, liver biopsy, and vibration-controlled transient elastography within 6 months. Agile 3+ and 4 were applied and compared with FIB-4 or LSM alone. Goodness of fit was evaluated using a calibration plot and discrimination using area under the receiver operating curve. Area under the receiver operating curves was compared using the Delong test. Dual cutoff approaches were applied to rule out and rule in ≥F3 and F4. Median (interquartile range) age was 58 (15) years. Median body mass index was 33.3 (8.5) kg/m2. Fifty-three percent had type 2 diabetes, 20% had F3, and 26% had F4. Agile 3+ demonstrated an area under the receiver operating curve of 0.85 (0.81; 0.88) similar to that of LSM [0.83 (0.79; 0.86), p=0.142] but significantly higher than that of FIB-4 [0.77 (0.73; 0.81), p<0.0001). Agile 4's area under the receiver operating curve [0.85 (0.81; 0.88)] was similar to that of LSM [0.85 (0.81; 0.88), p=0.065). However, the percentage of patients with indeterminate results was significantly lower with Agile scores compared with FIB-4 and LSM (Agile 3+: 14% vs. FIB-4: 31% vs. LSM: 13%, p<0.001; Agile 4: 23% vs. LSM: 38%, p<0.001)., Conclusions: Agile 3+ and 4 are novel vibration-controlled transient elastography-based noninvasive scores that increase accuracy in the identification of advanced fibrosis and cirrhosis respectively and are ideal for clinical use due to a lower percentage of indeterminant outputs compared with FIB-4 or LSM alone., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Association for the Study of Liver Diseases.)

The genetic code conceals a 'code within the codons', which hints at biophysical interactions between amino acids and their cognate nucleotides. Yet, research over decades has failed to corroborate systematic biophysical interactions across the code. Using molecular dynamics simulations and NMR, we have analysed interactions between the 20 standard proteinogenic amino acids and 4 RNA mononucleotides in 3 charge states. Our simulations show that 50% of amino acids bind best with their anticodonic middle base in the -1 charge state common to the backbone of RNA, while 95% of amino acids interact most strongly with at least 1 of their codonic or anticodonic bases. Preference for the cognate anticodonic middle base was greater than 99% of randomised assignments. We verify a selection of our results using NMR, and highlight challenges with both techniques for interrogating large numbers of weak interactions. Finally, we extend our simulations to a range of amino acids and dinucleotides, and corroborate similar preferences for cognate nucleotides. Despite some discrepancies between the predicted patterns and those observed in biology, the existence of weak stereochemical interactions means that random RNA sequences could template non-random peptides. This offers a compelling explanation for the emergence of genetic information in biology.

Oat crown rust caused by Puccinia coronata f. sp. avenae P. Syd. & Syd (Pca) is a major constraint to oat (Avena sativa L.) production in many parts of the globe. The objectives of this study were to locate Pc96 on the oat consensus map and to develop SNP markers linked to Pc96 for use in marker-assisted selection. SNP loci linked to the crown rust resistance gene Pc96 were identified by linkage analysis and PACE assays were developed for marker-assisted selection in breeding programs. Pc96 is a race-specific crown rust resistance gene originating from cultivated oat that has been deployed in North American oat breeding programs. Pc96 was mapped in a recombinant inbred line population (n = 122) developed from a cross between the oat crown rust differential known to carry Pc96 and the differential line carrying Pc54. A single resistance locus was identified on chromosome 7D between 48.3 and 91.2 cM. The resistance locus and linked SNPs were validated in two additional biparental populations, Ajay × Pc96 (F2:3, n = 139) and Pc96 × Kasztan (F2:3, n = 168). Based on all populations, the most probable location of the oat crown rust resistance gene Pc96 on the oat consensus map was on chromosome 7D approximately at 87.3 cM. In the Ajay × Pc96 population, a second unlinked resistance gene was contributed by the Pc96 differential line, which mapped to chromosome 6C at 75.5 cM. A haplotype of nine linked SNPs predicted the absence of Pc96 in a diverse group of 144 oat germplasm. SNPs that are closely linked to the Pc96 gene may be beneficial as PCR-based molecular markers in marker-assisted selection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Nonalcoholic fatty liver disease (NAFLD) and its more severe form, nonalcoholic steatohepatitis (NASH), represent a growing worldwide epidemic and a high unmet medical need, as no licensed drugs have been approved thus far. Currently, histopathological assessment of liver biopsies is mandatory as a primary endpoint for conditional drug approval. This requirement represents one of the main challenges in the field, as there is substantial variability in this invasive histopathological assessment, which leads to dramatically high screen-failure rates in clinical trials. Over the past decades, several non-invasive tests have been developed to correlate with liver histology and, eventually, outcomes to assess disease severity and longitudinal changes non-invasively. However, further data are needed to ensure their endorsement by regulatory authorities as alternatives to histological endpoints in phase 3 trials. This Review describes the challenges of drug development in NAFLD-NASH trials and potential mitigating strategies to move the field forward., (© 2023. Springer Nature America, Inc.)

Background and Aims: In cirrhotic nonalcoholic steatohepatitis (NASH) clinical trials, primary efficacy endpoints have been hepatic venous pressure gradient (HVPG), liver histology and clinical liver outcomes. Important histologic features, such as septa thickness, nodules features and fibrosis area have not been included in the histologic assessment and may have important clinical relevance. We assessed these features with a machine learning (ML) model., Methods: NASH patients with compensated cirrhosis and HVPG ≥6 mm Hg (n = 143) from the Belapectin phase 2b trial were studied. Liver biopsies, HVPG measurements and upper endoscopies were performed at baseline and at end of treatment (EOT). A second harmonic generation/two-photon excitation fluorescence provided an automated quantitative assessment of septa, nodules and fibrosis (SNOF). We created ML scores and tested their association with HVPG, clinically significant HVPG (≥10 mm Hg) and the presence of varices (SNOF-V)., Results: We derived 448 histologic variables (243 related to septa, 21 related to nodules and 184 related to fibrosis). The SNOF score (≥11.78) reliably distinguished CSPH at baseline and in the validation cohort (baseline + EOT) [AUC = 0.85 and 0.74, respectively]. The SNOF-V score (≥0.57) distinguished the presence of varices at baseline and in the same validation cohort [AUC = 0.86 and 0.73, respectively]. Finally, the SNOF-C score differentiated those who had >20% change in HVPG against those who did not, with an AUROC of 0.89., Conclusion: The ML algorithm accurately predicted HVPG, CSPH, the development of varices and HVPG changes in patients with NASH cirrhosis. The use of ML histology model in NASH cirrhosis trials may improve the assessment of key outcome changes., (© 2023 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.)

Background & Aims: The effect of race on routinely available noninvasive tests of fibrosis is incompletely understood. This study evaluated the performance of noninvasive tests among white and Asian patients in the STELLAR trials (NCT03053050 and NCT03053063), which evaluated selonsertib in patients with advanced (F3-F4) fibrosis due to nonalcoholic steatohepatitis (NASH)., Methods: Baseline liver biopsies were centrally read using the NASH Clinical Research Network system, and 4 noninvasive tests (Nonalcoholic fatty liver disease fibrosis score [NFS], Fibrosis-4 index [FIB-4], Enhanced Liver Fibrosis test [ELF], and liver stiffness by vibration-controlled transient elastography) were measured. The performance of these tests to discriminate advanced fibrosis was evaluated using areas under the receiver operating characteristics curves with 5-fold cross-validation repeated 100 times., Results: Among 3207 patients screened with evaluable liver histology, 2281 were whites and 762 were Asians. Seventy-two percent of whites and 67% of Asians had advanced fibrosis. The areas under the receiver operating characteristics curves of the noninvasive tests for advanced fibrosis were similar in whites and Asians: 0.73 and 0.75 for NFS, 0.78 and 0.80 for FIB-4, 0.79 and 0.81 for ELF, and 0.80 and 0.83 for liver stiffness, respectively. At the published cutoffs, the tests had similar sensitivities and specificities in the 2 groups. However, the sensitivities of NFS, FIB-4, and ELF were low in both white and Asian patients younger than 40 years., Conclusions: In the global phase III STELLAR trials, the diagnostic performance of routinely available noninvasive tests for the detection of advanced fibrosis due to NASH was acceptable and similar between white and Asian patients., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Rencofilstat (RCF) demonstrated antifibrotic effects in preclinical models and was safe and well tolerated in Phase 1 studies. The aim of this Phase 2a study was safety, tolerability, pharmacokinetics, and exploration of efficacy biomarkers in subjects with nonalcoholic steatohepatitis (NASH). This Phase 2a, multicenter, single-blind, placebo-controlled study randomized 49 presumed F2/F3 subjects to RCF 75 mg once daily (QD), RCF 225 mg QD, or placebo for 28 days. Primary safety and tolerability endpoints were explored using descriptive statistics with post hoc analyses comparing active to placebo groups. Pharmacokinetics were evaluated using population pharmacokinetics methods. Efficacy was explored using biomarkers, transcriptomics, and lipidomics. RCF was safe and well tolerated, with no safety signals identified. The most frequently reported treatment-emergent adverse events were constipation, diarrhea, back pain, dizziness, and headache. No clinically significant changes in laboratory parameters were observed, and RCF pharmacokinetics were unchanged in subjects with NASH. Alanine transaminase (ALT) reduction was greater in active subjects than in placebo groups. Nonparametric analysis suggested that ALT reductions were statistically different in the 225-mg cohort compared with matching placebo: -16.3 ± 25.5% versus -0.7 ± 13.4%, respectively. ProC3 and C6M reduction was statistically significant in groups having baseline ProC3 > 15.0 ng/ml. RCF was safe and well tolerated after 28 days in subjects with presumed F2/F3 NASH. Presence of NASH did not alter its pharmacokinetics. Reductions in ALT, ProC3, and C6M suggest direct antifibrotic effects with longer treatment duration. Reductions in key collagen genes support a mechanism of action via suppression and/or regression of collagen deposition. Conclusion: These results support advancement of rencofilstat into a larger and longer Phase 2b study., (© 2022 The Authors. Hepatology Communications published by Wiley Periodicals LLC on behalf of American Association for the Study of Liver Diseases.)

The universal core of metabolism could have emerged from thermodynamically favoured prebiotic pathways at the origin of life. Starting with H 2 and CO 2 , the synthesis of amino acids and mixed fatty acids, which self-assemble into protocells, is favoured under warm anoxic conditions. Here, we address whether it is possible for protocells to evolve greater metabolic complexity, through positive feedbacks involving nucleotide catalysis. Using mathematical simulations to model metabolic heredity in protocells, based on branch points in protometabolic flux, we show that nucleotide catalysis can indeed promote protocell growth. This outcome only occurs when nucleotides directly catalyse CO 2 fixation. Strong nucleotide catalysis of other pathways (e.g. fatty acids and amino acids) generally unbalances metabolism and slows down protocell growth, and when there is competition between catalytic functions cell growth collapses. Autocatalysis of nucleotide synthesis can promote growth but only if nucleotides also catalyse CO 2 fixation; autocatalysis alone leads to the accumulation of nucleotides at the expense of CO 2 fixation and protocell growth rate. Our findings offer a new framework for the emergence of greater metabolic complexity, in which nucleotides catalyse broad-spectrum processes such as CO 2 fixation, hydrogenation and phosphorylation important to the emergence of genetic heredity at the origin of life.

The origin of the genetic code is an abiding mystery in biology. Hints of a 'code within the codons' suggest biophysical interactions, but these patterns have resisted interpretation. Here, we present a new framework, grounded in the autotrophic growth of protocells from CO 2 and H 2 . Recent work suggests that the universal core of metabolism recapitulates a thermodynamically favoured protometabolism right up to nucleotide synthesis. Considering the genetic code in relation to an extended protometabolism allows us to predict most codon assignments. We show that the first letter of the codon corresponds to the distance from CO 2 fixation, with amino acids encoded by the purines (G followed by A) being closest to CO 2 fixation. These associations suggest a purine-rich early metabolism with a restricted pool of amino acids. The second position of the anticodon corresponds to the hydrophobicity of the amino acid encoded. We combine multiple measures of hydrophobicity to show that this correlation holds strongly for early amino acids but is weaker for later species. Finally, we demonstrate that redundancy at the third position is not randomly distributed around the code: non-redundant amino acids can be assigned based on size, specifically length. We attribute this to additional stereochemical interactions at the anticodon. These rules imply an iterative expansion of the genetic code over time with codon assignments depending on both distance from CO 2 and biophysical interactions between nucleotide sequences and amino acids. In this way the earliest RNA polymers could produce non-random peptide sequences with selectable functions in autotrophic protocells., Competing Interests: Declaration of competing interest The authors 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 © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)