Epigenetics & Chromatin

DNMT3A is a de novo DNA methyltransferase whose recruitment to chromatin regulates its function. Missense mutations within the chromatin-binding PWWP domain are associated with diverse human disorders, yet how mutations in the same domain produce distinct phenotypes remains unclear. Here we systematically characterise 19 clinically reported mutations in the PWWP domain of DNMT3A that are associated with Heyn-Sproul-Jackson syndrome (HESJAS), paraganglioma (PG) and clonal haematopoiesis (CH). We show that all PWWP-domain mutations associated with HESJAS abolished interaction with H3K36me2 modified nucleosomes, defining this as a consistent biochemical feature of HESJAS. In contrast, mutations from all disease classes differentially altered DNA binding of the PWWP domain, driven by alterations in the net charge of the domain. However, these effects are largely overcome by inclusion of the DNNMT3A1 N-terminal region, which is absent from its embryonic isoform, suggesting that PWWP mutations may differentially affect DNMT3A function through development. Changes in the thermal stability of the isolated PWWP domain mutants did not directly translate into altered stability of full-length DNMT3A1 in cells. We show that HESJAS mutations can affect the intramolecular interaction between the PWWP and adjacent ADD domain, an interaction proposed to contribute to the autoinhibitory function of the ADD domain. However, not all mutations behaved in the same way, suggesting that multiple factors govern the intramolecular autoinhibition of DNMT3A. Together, this study advances our understanding of the molecular mechanisms by which DNMT3A PWWP-domain mutations are mechanistically heterogeneous, providing a biochemical framework that contributes to distinct disease phenotypes.

Nuclear pore complexes (NPCs) are nuclear envelope (NE)-embedded protein assemblies that mediate nucleocytoplasmic exchange and interact with the genome, including binding of an NPC component Nup93 to Polycomb chromatin domains. Here, we investigated the in vivo relevance of this relationship in Drosophila, which unusually contains two distinct paralogs of Nup93. Interestingly, we identified a Nup93-2-specific tumorigenic phenotype in larval wings, where depletion of Nup93-2, but not Nup93-1, led to tumor-like overgrowth, reminiscent of Polycomb mutations. Consistently, our transcriptomic analysis revealed a wide-spread loss of gene silencing in Nup93-2-depleted wings, particularly in a Nup93-bound Polycomb domain spanning genes for activators of JAK/STAT signaling. Nup93 paralogs were not found to differ in their effect on NPC biogenesis but strikingly, showed differences in subnuclear localization patterns. While Nup93-1 co-localized exclusively with fully assembled NPCs, Nup93-2 exhibited only partial co-localization and was found at additional NE locations in a tissue-specific manner. Together, our results identify an in vivo silencing role of a Nup93 paralog and suggest that Nup93-2 may form a unique NE-associated complex that targets a subset of Polycomb domains containing growth-promoting genes.

THAP9 is a transposable element-derived gene which encodes a protein that is homologous to the active Drosophila P-element transposase (DmTNP). Both THAP9 and DmTNP possess a C-terminal domain (CTD) which is functionally uncharacterized. Sequence and structural analysis suggest that the THAP9-CTD has a novel fold which is only found in THAP9 homologs. To explore the evolutionary history and characteristics of this novel domain, exhaustive phylogenetic analysis (using MSA, structure prediction, MSTA-based clustering) was performed. THAP9-CTD homologs were more widely distributed throughout the animal kingdom in comparison to DmTNP-CTD homologs which were restricted to arthropods. Moreover, the THAP9-CTD homologs were more conserved, especially among mammals and birds and their average length increased in a class-specific manner. Comparison with the DmTNP-CTD homologs demonstrates that although their respective CTDs may have evolved independently, they both surprisingly share similar secondary structure elements consisting of three conserved helical regions made of hydrophobic residues that are predicted to make up a conserved core. The role of the respective CTDs were further investigated by creating truncation mutants lacking the CTD. Interestingly both THAP9 and DmTNP truncation mutants are still capable of DNA excision and integration suggesting that their respective CTDs are not essential for DNA transposition. Moreover, CTD truncation favours DNA integration in THAP9: this suggests that CTD acquisition during evolution may have led to THAP9 domestication as observed in other transposable element-derived genes like Rag1 and piggybac, which have similar terminal regulatory domains.

X-chromosome inactivation involves chromosome-wide gene silencing accompanied by extensive chromatin changes, as well the loss of topologically associating domains. Yet discrete regions of the inactive X chromosome retain activity within localised 3D domains, which contain active genes that variably escape from X inactivation. The transcription factor and architectural protein CTCF has been proposed to be implicated in escape by insulating escape domains or sustaining their topology via cohesin-mediated loop extrusion. Here, we test the role of CTCF and cohesin in escape using acute degron-mediated depletion of CTCF and RAD21 in neural progenitor cells with established escape profiles. Although CTCF occupancy correlates with escape status on the inactive X chromosome, its removal - together with loss of loop extrusion - does not disrupt escapee gene expression, or domain organization, nor does it result in spreading of silencing or activation of genes in cis. Rather, we show that facultative escape regions are self-sustaining compartments of active chromatin enriched in H3K27 acetylation and depleted in H3K27 methylation, with the magnitude of compartment strength scaling up with the degree of transcriptional activity on the inactive X chromosome. These active escapee compartments are propagated independently of CTCF and RAD21-dependent 3D architecture. Our findings identify chromatin compartmentalization as the primary feature of facultative escapee domains.

Transcription factor Ets2 coalesce with the NF-{kappa}B pathway to regulate gene expression in specific signaling contexts. IKK2/{beta}-mediated phosphorylation events critically regulate the NF-{kappa}B pathway. However, any link between Ets2 and IKK2 remains elusive. Here we report Ets2 as a direct substrate of IKK2. In-vitro kinase assays using deletion constructs, high resolution MS-MS and site directed mutagenesis identified S295 as a prominent phosphorylation site distal to the DNA binding domain, substitution of which to phosphor-mimetic Glutamate triggers further phosphorylation of Ets2. MD simulations clearly indicate conformational constriction of the otherwise disordered N-terminal region and inhibition of DNA binding activity upon phosphorylation, which was further confirmed by Electrophoretic mobility shift assays. Our results uncover a phosphoregulatory connection between Ets2 and IKK2.

BACH1 is a heme-regulated basic-leucine-zipper containing transcriptional repressor that binds its DNA recognition elements as a heterodimer with MAFK. Heme-binding is thought to be mediated by several Cys-Proline (CP) motifs and this results in dissociation of the heterodimer from DNA. The mechanism of heme-binding and heme-mediated DNA dissociation remains unresolved. We have used UV-visible spectroscopy, 2D-NMR and DNA-binding assays to explore both heme-binding and DNA dissociation of a minimal BACH1 construct containing 2 CP motifs (C492(CP5) and C646(CP6)) flanking the DNA-binding domain. We find that heme is able to bind to both CP motifs, but also to other non-CP cysteines and histidines in the construct. Using NMR spectroscopy, we identify a structured binding pocket in which heme interacts with both C646(CP6) and Cys621. However, DNA-binding assays show that C646(CP6) is not required for heme-mediated DNA dissociation of the BACH1:MAFK heterodimer. Using UV-visible spectroscopy we show that C492(CP5) also recruits heme with a second ligand, a conserved histidine, His559, in the BACH1 DNA-recognition helix. Mutation of C492(CP5) reduces but does not abolish heme-mediated dissociation from DNA. Our findings suggest a mechanism for heme-binding to BACH1 and heme-mediated dissociation from DNA.

Paternally-inherited epigenetic information can influence phenotype in offspring (1). Here, we identify a critical mechanistic contribution of KDM6A (UTX), an X-linked histone modifier and tumor suppressor, in regulating transmissible epigenetic information in mammalian sperm. Paternal loss of KDM6A increases cancer risk in genetically wild type offspring, but how Kdm6a knockout sperm transmit this effect at the molecular level is unknown (2). We find that KDM6A functions in spermatogenesis to promote methylation of histone H3 lysine 4 (H3K4) via selective interaction with the COMPASS complex methyltransferase KMT2C (MLL3). KMT2C and KDM6A are coordinately recruited to promoters of active genes in spermatogenic cells, contrasting with recruitment to intergenic enhancers in other cell types (3, 4). Loss of KDM6A disrupts H3K4 methylation at promoters of tumor suppressor genes in spermatogonia, and some of these defects persist in epididymal sperm and correspond to impaired expression in preimplantation embryos. These genes are also misregulated in normal and malignant hematopoietic tissue of genetically wild type offspring, indicating that impaired H3K4 methylation in KDM6A-deficient male germ cells may preferentially alter regulation of tumor suppressor gene networks in development across generations.

Tumor-promoting myofibroblastic carcinoma-associated fibroblasts (myCAFs) are induced by activation of transforming growth factor-{beta} signaling. However, the molecular basis of myCAF-specific transcriptional programs regulated by TGF-{beta} signaling remains poorly understood. Using a meta-analysis of single-cell RNA-seq data from 132 human breast tumor and non-tumor tissues, we show that myCAFs activate gene regulatory programs relevant to skeletal and cardiovascular development that are associated with poorer outcomes in breast cancer patients. Of note, distal-less homeobox 5 (DLX5), a master transcription factor for skeletal development, is activated in human breast myCAFs at both epigenetic and transcriptional levels. DLX5 expression is also initiated by TGF-{beta}1 treatment in human mammary fibroblasts. Immunoprecipitation and CUT&RUN assays using DLX5-expressing fibroblasts demonstrate that DLX5 interacts with Smad2/3/4 proteins, enabling their cooperative occupancy at shared genomic binding sites of target genes, thereby promoting canonical TGF-{beta} signaling and the myCAF state. DLX5-primed myCAFs also enhance paracrine TGF-{beta} signaling and neuropilin-2 expression to promote collective tumor invasion. Our findings indicate that DLX5 induces myCAF formation and promotes breast tumor progression in collaboration with canonical TGF-{beta} signaling.

Chemoradiotherapy (CRT) is the standard-of-care therapy for many solid malignancies, yet predictive biomarkers of treatment response remain limited. We identified a germline single nucleotide polymorphism (SNP) in an intrinsically disordered region of the lysine demethylase KDM3C/JMJD1C (p.S464T) that is associated with CRT outcomes in locally advanced rectal cancers (LARC) and head and neck squamous cell carcinoma (LA-HNSCC). In silico modeling with AlphaFold predicted S464T substitution influenced interaction between phosphorylated KDM3C and RNF8 FHA domain. In cellular models, conversion of S464 to T464 increased sensitivity to DNA-damaging agents. S464T substitution impaired damage-induced MDC1-RAP80 signaling and downstream RAP80-BRCA1 colocalization. SNP carrying cells impaired DNA repair causing genotoxic stress that is associated with increased cGAS-cGAMP innate immune signaling and increased apoptosis. Population analyses with the SNP highlighted an increase incidence of UV-induced skin and other cancers, linking inherited variation in the chromatin regulatory gene KDM3C to genome instability, cancer risk, and therapeutic vulnerability.

Developmental regression is a severe but poorly understood complication of several neurodevelopmental disorders. In Kleefstra syndrome (KLEFS1), caused by EHMT1 haploinsufficiency, regression often emerges during adolescence or early adulthood and is frequently preceded by marked sleep disturbance. Experimental work implicating EHMT1/G9a in metabolic regulation and stress responses raises the possibility that impaired metabolic resilience contributes to this vulnerability. Here, we aimed to investigate whether altered glucose metabolism is a feature of KLEFS1 and whether it relates to clinical variability, including regression. Through [18F]FDG-PET/CT, individuals with KLEFS1 who had experienced regression (n=4) exhibited a hypometabolic brain profile, whereas one individual who had not experienced regression showed globally elevated metabolic activity. In parallel, G9a mutant flies exhibited increased baseline metabolic rate and neuronal ATP levels together with sleep fragmentation resembling the clinical phenotype. Providing flies with oxidative stress to model KLEFS1 regression further exacerbated sleep disruption and was associated with a reduction in metabolic output. Importantly, adult high sugar feeding in flies prevented oxidative stress-induced worsening of sleep and maintained metabolic stability under challenge. Together, these findings suggest that regression in KLEFS1 and associated sleep disturbances are linked to underlying metabolic vulnerability and impaired maintenance of energy homeostasis under stress.

Canonical McrBC enzymes are nucleotide-powered, motor-driven endonucleases that bind and cleave modified bacteriophage DNA. Non-canonical McrBC homologs like LlaI and BsuMI are distinguished by a unique three-gene organization and the ability to target DNA site-specifically. Here, we report the atomic-resolution crystal structures of the DNA-binding module LlaI.R1 and AAA+ motor LlaI.R2 from the Lactococcus lactis LlaI anti-phage defense system. The crystallized LlaI.R2 hexamer traps two distinct active site conformations that correlate to different states of the nucleotide hydrolysis cycle and reveal that the organization of the critical catalytic machinery present in canonical McrB homologs is also conserved in non-canonical R2 proteins. Although canonical McrB homologs are strictly GTP-specific, we find that the R2 proteins from LlaI and BsuMI do not discriminate between different nucleotides, even when in complex with their respective R1 partners. Using mutagenesis, we define surfaces on the LlaI.R1 structure that are critical for DNA-binding and interaction with LlaI.R2. These observations support computational modelling of the assembled LlaI restriction system bound to DNA. Together, our data provide new insights into the evolution of nucleotide specificity in McrBC restriction complexes and the molecular mechanisms governing McrBC-catalyzed DNA translocation and cleavage.

Bulk telomere length measured from tumour sequencing is routinely interpreted as a property of the cancer cells. However, a tumour specimen is a mixture, and the patient who supplies it has a telomere length of their own. Here I re-analyse published pan-cancer telomere estimates and ask how much of a tumour's telomere length is patient-specific. A calibration step comes first. Whole-genome and low-pass estimates recover the known cross-sectional attrition of leukocyte telomeres with age, at 26.6 bp per year in blood normals, whereas whole-exome estimates do not. After adjustment for cancer type, sequencing centre and sex, the exome slope is minus 0.6 bp per year. In 684 blood-normal aliquots sequenced by both assays, the whole-genome estimate declines at 38.9 bp per year, whereas the exome estimate from the same DNA shows no detectable decline. The difference between assays is 41.5 bp per year, with P = 3 x 10^-10. Because exome data constitute 78.6% of the original resource, downstream analyses use only whole-genome and low-pass libraries. Within those data, tumour telomere length tracks the patient's matched-normal telomere length. The Spearman correlation is 0.395 in TCGA, with positive associations in 22 of 23 cancer types. This finding replicates in PCAWG using a different telomere estimator, with a correlation of 0.472 and positive associations in all 24 histologies examined. Adjustment for cancer type, sequencing centre and library type leaves a regression coefficient of 0.385. The association is also stable after adjustment for age, sex, tumour purity, leukocyte fraction, ploidy, sequencing coverage and continental ancestry, with coefficients ranging from 0.406 to 0.429. Pure normal-cell admixture is rejected as the sole explanation. Under a two-compartment mixture model, the coefficient for host telomere length is expected to equal 1 and the host-by-purity interaction to equal minus 1. These restrictions are jointly rejected with P = 0.001. Tumour purity, leukocyte fraction and age each explain only about 1 to 3% of within-cohort variance and do not alter the cross-cancer ranking. By contrast, the between-cohort coefficient is not directly interpretable. Its apparent near one-to-one relationship with tissue-associated telomere length depends strongly on which tissue supplies the matched-normal reference and on the statistical spread of that predictor, falling to 0.44 when organ-matched solid tissue is used. Bulk tumour telomere length is therefore a composite phenotype containing a replicated patient-specific component. Telomere biomarker studies should include matched-normal telomere length as a covariate rather than treating tumour telomere length as exclusively tumour-intrinsic.

The Mediator complex, a vital transcriptional coregulator in eukaryotes, partners with transcription factors to orchestrate gene transcription and in turn many developmental and physiological processes. Mediator subunit MED15 is required for the pre-natal development and post-natal maturation of pancreatic {beta}-cells in mice. However, whether MED15 plays a role in {beta}-cell function after initial development and throughout adulthood is unknown. To investigate the role of MED15 in {beta}-cells post-maturation, we induced a {beta}-cell specific Med15 knockout at six weeks age in male and female mice. This post-developmental Med15 ablation led to glucose intolerance and impaired insulin secretion. RNA-sequencing revealed downregulation of {beta}-cell maturation markers, indicating that MED15 is continuously required to maintain {beta}-cell functionality. Further, we implanted insulin pellets into Med15 knockout mice to lower blood glucose and used RNA-seq to validate that the transcriptional changes we observed are a direct consequence of Med15 loss and not an indirect effect of hyperglycemia arising in the knockout mice. In sum, our study shows that MED15 is continuously required after weaning to maintain functional {beta}-cell maturity.

Lysophosphatidic acid (LPA) is widely recognized as an extracellular lipid mediator; however, the functional significance of intracellularly produced LPA remains poorly understood. Here, we investigated the regulatory mechanism and functional role of a LPA-producing lysophospholipase D GDE4, also known as GDPD1, in prostate cancer cells. GDE4 expression is induced under ER stress conditions in a PERK-dependent manner and requires the transcription factor ATF3. Disruption of GDE4 expression resulted in altered intracellular levels of LPA and LPA precursor lysophosphatidylethanolamine, accompanied by reduced cell proliferation. RNA sequencing and subsequent validation identified a set of genes downregulated in GDE4-depleted cells. Pharmacological inhibition experiments indicated that peroxisome proliferator-activated receptor and {gamma} (PPAR and PPAR{gamma}) signaling pathways contribute to the regulation of these GDE4-dependent genes. Collectively, our findings suggest that GDE4-dependent lipid remodeling is associated with PPAR/{gamma}-mediated transcriptional regulation under ER stress conditions. These results provide a potential framework for understanding the link between intracellular lipid metabolism and stress-responsive gene regulation.

The X chromosome carries an unusually high density of immune genes and is a major contributor to sex differences in immune function and autoimmune diseases. In females, X-chromosome inactivation (XCI) has two major functional consequences: it shapes X-linked gene dosage through XCI escape and determines the cellular exposure of heterozygous X-linked variants through XCI skew. Yet because XCI creates a mosaic of cells expressing different parental X chromosomes, these properties have remained largely inaccessible in individual women, becoming measurable only where XCI is non-random or after aggregation across large cohorts. Consequently, how X-linked variation contributes to sex-biased immunity and differs between individual women has remained unresolved. Here we present scDaisyChain, a graph-based framework that reconstructs chromosome-scale X haplotypes directly from heterozygous SNPs and single-cell long-read transcriptomes. scDaisyChain achieves near-ground-truth accuracy in highly polymorphic mouse hybrids and shows strong concordance with orthogonal long-read whole-genome phasing in human samples. Applied to peripheral blood immune cells from healthy women, it reveals a lineage-specific escape program in which lymphoid cells escape XCI more broadly than monocytes, with corresponding gains in the inactive X chromatin accessibility and female-biased expression. Lineage-specific skew further alters the proportion of cells expressing each heterozygous X-linked variant, a property we term variant exposure. Predicted deleterious variants are preferentially found in low-exposure states, exemplified by a splice-altering TLR8 variant expressed in few cytotoxic T cells. In rheumatoid arthritis (RA), the monocyte compartment - which has the lowest escape in health - shows reproducible inactive X dysregulation converging on a trained-immunity programme linked to disease flare and synovial macrophage activation, with elevated escape of IL13RA1 and HDAC8. These findings establish lineage-specific escape, skew and variant exposure as quantifiable, patient-resolved determinants of sex-biased immune gene dosage and X-linked variant penetrance in health and autoimmune disease, resolving a dimension of female biology that has been previously inaccessible in individual donors.

Causal mediation analysis is widely used to identify biological pathways linking exposures to outcomes, but most methods assume homogeneous mediation effects across individuals. In high-dimensional omics settings, this assumption can mask important heterogeneity driven by demographic, genetic, or environmental factors. We propose the M-high-learner, a flexible framework for detecting heterogeneous mediation effects with high-dimensional mediators. The method identifies mediators with subgroup-specific indirect effects while distinguishing them from null or homogeneous signals and controlling the type I error rate. It is computationally efficient, scalable, and yields interpretable sub-types. Simulation studies show that the proposed approach achieves high power while maintaining accurate error control. Applications to the Framingham Heart Study and the Multi-Ethnic Study of Atherosclerosis reveal that the mediation role of gene expression in sexs effect on high-density lipoprotein varies across subgroups defined by body mass index and age. Our framework provides a practical tool for uncovering heterogeneous biological mechanisms in high-dimensional genomic studies. Author SummaryBiological processes linking risk factors to disease often differ across individuals, but many existing methods assume these processes are the same for everyone. This can hide important differences between groups. We developed a powerful method to identify when these pathways vary across subgroups using large-scale molecular data. Our approach detects differences in how intermediate biological factors contribute to outcomes in populations defined by characteristics such as age and body mass index. Applying our method to population studies, we found that some biological pathways operate differently across groups, suggesting that key mechanisms may be missed when differences are ignored. Our work provides a tool to better understand how disease-related processes vary across individuals, which may support more targeted and personalized approaches to health research.

Initiator-methionine excision is among the earliest protein modifications, yet its relationship to assembly geometry is unknown. Burial of the mature first residue was measured across 7,246 deposited human biological assemblies (22,291 chain-level observations; 1,191 proteins). Among 1,143 analyzable proteins, termini in MetAP-permissive penultimate-residue sequence classes were less often interface-engaged than termini in MetAP-nonpermissive classes (37.4% versus 47.4%; adjusted odds ratio 0.65, p = 7.2e-4). Curated processing annotations did not show a corresponding burial difference, and correlated residue properties preclude attributing the sequence-class association specifically to iMet removal. The analysis identified 264 interface-engaged MetAP-permissive candidates concentrated in cellular machines. In a fully recomputed conformer scan of deeply buried proteasome positions, modeled methionine accommodation was less favorable than at observed-methionine controls (median overlap -0.30 versus -1.12 angstrom, p = 0.0049), although most scoreable sites permitted a nonoverlapping placement. The census therefore reveals a graded structural constraint - not universal steric failure - and prioritizes complexes in which altered packing, assembly kinetics, lipidation or N-terminal methylation can be tested.

Elucidating the biological and molecular mechanisms that govern male fertility and mating behavior in mosquitoes is critical for optimizing genetic and sterile insect technique-based vector control strategies. Here, we examined age-related changes in male reproductive capacity in Anopheles culicifacies, using female egg output as an indirect indicator of male fertility. Our results demonstrated that male reproductive age follows a non-linear pattern of fertility. Morphometric analysis from emergence to day 13 post-eclosion revealed a strong correlation between seminal vesicle capacity and female fecundity, suggesting that age-dependent gonadal development directly influences reproductive potential. At the molecular level, we identified AcLITAF6 as a key regulator of male reproductive homeostasis. RNAi-mediated knockdown of AcLITAF6 impaired apoptosis-associated and phagocytic clearance, reduced sperm viability, and decreased female productive outcomes. Conclusively, we reveal a previously unrecognized role of LITAF in sperm quality control and male reproductive fitness, highlighting AcLITAF6 as a potential target for mosquito population suppression strategies.

Background: Prostate cancer enzalutamide treatment is approved at a standard dose of 160 mg daily. Concerns for real-world patients -- older and more fragile than those enrolled in clinical trials -- have prompted consideration of initiating treatment with lower doses, but the long-term efficacy of this approach remains unknown. We evaluate the long-term survival and longevity in patients treated with standard versus upfront low-dose enzalutamide. Methods: Retrospective analysis of 151 patients treated with enzalutamide (102 receiving 160 mg; 49 receiving [≤]80 mg) between 2014--2021 at the Centre Hospitalier Universitaire de Martinique, with complete follow-up through end of life (98.7% completeness of follow-up). Primary outcomes were overall survival (OS), progression-free survival (PFS), and longevity (attained age). Results: Doses [≤]80 mg were associated with longer median OS (36.3 vs. 20.7 months), improved restricted mean OS (difference of 0.7 years, p=0.05), and enhanced longevity (median 82.5 vs. 78.3 years, p=0.004). PSA response rate at 12 weeks was higher with lower-dose (71.4% vs. 48.8%, p=0.016). In multivariable models adjusted for prognostic factors, [≤]40 mg compared with 160 mg was non-inferior regarding OS (HR=0.61, 95% CI 0.36--1.06), superior regarding PFS (HR=0.59, 95% CI 0.35--0.99), and superior regarding longevity (HR=0.48, 95% CI 0.28--0.84). Bone metastasis, poor performance status, PSA response, time to PSA nadir, and disease duration were independent predictors of outcomes. A post-hoc analysis revealed a strong association between dose and physician-prescribing profiles, ranging from "endorse-lowest-dose" to "never-deviate-from-full-dose". Conclusions: Lower doses of enzalutamide were non-inferior to full-dose. Dose-adapted strategies warrant further investigation.

Background & Aims: Fatigue is a central symptom of chronic liver disease (CLD), substantially impacting health-related quality of life (HRQoL). This study aimed to further understand CLD symptomatology, including fatigue, and its impact on HRQoL from a patient perspective. Methods: Abbott Global Assessment of Patients unmet needs (aGAP) was a multinational, cross-sectional survey in adults with compensated CLD in China, India and Mexico, conducted between July and November 2024. Adult participants who self-reported that they had physician-diagnosed CLD and were experiencing fatigue completed a quantitative survey to assess symptom burden and included three HRQoL patient-reported outcome (PRO) questionnaires (Patient-Reported Outcomes Measurement Information System [PROMIS]-29+2, Work Productivity and Activity Impairment - Specific Health Problem version 2.0 [WPAI: SHP], Multidimensional Fatigue Inventory [MFI]). Results: Overall, 505 participants (China: 200; Mexico: 105; India: 200) completed the study. Participants reported that their CLD-related fatigue sometimes, often or always affected their self-esteem/confidence (45.1%) and ability to maintain or acquire new employment (38.6%). Most participants reported moderate (51.3%) or serious (26.9%) fatigue, with 33.5% experiencing fatigue every day or almost every day. Many participants felt their social life was negatively impacted by their fatigue (47.3%) and that there were related financial difficulties (53.9%). Use of validated PRO tools demonstrated severe fatigue (MFI: overall mean [SD] 13.9 [3.4] general fatigue and 13.4 [3.6] physical fatigue) as well as substantial levels of work and activity impairment (WPAI: SHP overall mean [SD] 53.0 [26.4]) and high levels of anxiety, pain interference, depression and sleep interference (PROMIS T-scores [≥]54). Conclusions: Fatigue has a substantial impact on HRQoL among adults with CLD across several countries, highlighting a global unmet need for targeted interventions to effectively identify and manage the condition.