Juran, Brian D.

ABSTRACT Background Primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC) are rare, chronic cholestatic liver diseases with heterogeneous progression and limited biomarkers of disease biology. Epigenetic age acceleration has been demonstrated in PSC and PBC, suggesting accelerated biological aging. DNA methylation–derived telomere length (DNAmTL) reflects replicative aging and cellular turnover, but its role in PSC and PBC remains unclear. We aimed to evaluate DNAmTL in PSC and PBC and its association with disease severity. Methods We analysed and compared DNAmTL and age‐adjusted DNAmTL among patients with PSC ( n = 474) versus PSC controls ( n = 484) and patients with PBC ( n = 291) versus PBC controls ( n = 291). Associations with clinical outcomes were assessed using nonparametric tests and multivariable logistic regression models. Correlations with primary sclerosing cholangitis Risk Estimate Tool (PREsTO) score and epigenetic age acceleration (AgeAccelGrim) were evaluated using linear regression. Results Patients with PSC demonstrated significantly shorter age‐adjusted DNAmTL compared with controls (median −0.036 vs. 0.016, p < 0.001). Patients with PBC also exhibited reduced DNAmTL relative to controls (median 6.814 vs. 6.885, p < 0.001). Within PSC, shorter DNAmTL was associated with cirrhosis (OR 0.21, 95% CI 0.06–0.69, p = 0.013) and advanced disease (OR 0.20, 95% CI 0.06–0.67, p = 0.009). Shorter DNAmTL was also observed in hepatic decompensation and hepatobiliary malignancy, including cholangiocarcinoma. DNAmTL inversely correlated with PReSTO score ( p < 0.001). DNAmTL was strongly inversely correlated with AgeAccelGrim in both PSC and PBC ( p < 0.001). Conclusion Cholestatic liver diseases are associated with shortened DNAmTL, which independently correlates with disease severity in PSC. Together with epigenetic age acceleration, these findings support a model of accelerated biological aging in PSC and PBC characterized by converging signatures of accelerated biological aging that may contribute to progressive biliary injury and fibrosis.

ABSTRACT Background Primary Biliary Cholangitis (PBC) is an autoimmune cholangiopathy with polygenic architecture and unknown aetiology. Evidence links Neanderthal‐derived genetic variants to autoimmune conditions; however, their contribution to PBC susceptibility remains unexplored. Objective We investigate whether archaic alleles influence PBC risk. Design Genotype data from four PBC cohorts were analysed: CANUK (4615 cases, 9233 controls), OLD IT (444 cases, 901 controls), NEW IT (255 cases, 579 controls) and MAYO (891 cases, 621 controls). Association testing with 235 592 Neanderthal Informative Markers (NIMs) was performed, adjusting for population stratification. Heritability was estimated using RHE‐mc and temporal haplotype dynamics were assessed using ancient DNA data from Europe and Asia. Results A Neanderthal‐derived variant in TNPO3 (rs12531711), previously reported by Cordell et al. showed strong association in CANUK ( p = 2.04 × 10 −26 ) and replicated across all validation cohorts. This variant functions as an eQTL, upregulating IRF5 and downregulating TNPO3 in blood. It is rare in African populations but common both in Europeans and Asians. Temporal analysis of the haplotype carrying rs12531711 revealed frequency increased substantially during the Bronze Age (5000–3000 BP), rising from 2.0% to 7.7% in Europe and 3.4% to 17.4% in Asia, before stabilizing. Despite this strong single‐variant association, the heritability explained by NIMs was not significantly different from that of non‐NIM variants. Conclusion A Neanderthal‐derived variant contributes to PBC risk and its frequency increased in the Bronze Age, possibly due to pathogen‐driven selection. Archaic introgression overall has limited influence on PBC heritability. Functional studies are needed to elucidate the role of rs12531711.

Background and Aims: Cholangiocarcinoma (CCA) is a rare but aggressive malignancy with poorly understood genetic susceptibility. To date, genome-wide association studies (GWAS) investigating germline variants associated with CCA risk remain limited. We aimed to identify genetic risk loci for CCA and its clinical subtypes through comprehensive GWAS and post-GWAS analyses. Approach and Results: We conducted a GWAS of 2366 CCA cases and 11,750 controls of European ancestry. Genome-wide significant loci ( p <5×10 −8 ) were identified and further examined through fine-mapping, functional annotation, and HLA imputation. Subgroup analyses were conducted by CCA subtypes and primary sclerosing cholangitis (PSC) status. Cross-trait linkage disequilibrium score regression and Mendelian randomization were employed to investigate the shared genetic architecture and potential causal relationships with a diverse range of traits. We identified 1 new genome-wide significant variant, rs535777 (OR=1.44), near HLA-DRB1/DQA1 associated with CCA, and 2 variants associated with extrahepatic CCA: rs116224263 (OR=0.17) in LINC02506 at 4p15.1 and rs6914950 (OR=1.63) near HLA-DRB1/DQB1 . Stratified analyses revealed rs2395184 (OR=3.51) near HLA-DRA/DRB5 associated with PSC-related CCA, and rs142674434 (OR=2.98) in THSD7A at 7p21.3 associated with non-PSC-related CCA. HLA imputation uncovered new amino acid residues associated with disease risk. Cross-trait analyses identified shared genetic signals between CCA and anthropometric, lipidemic, lifestyle, and medical traits. Mendelian randomization supported putative causal associations for 12 traits with CCA or its subtypes. Conclusions: Our large-scale GWAS highlights new genetic variants and HLA-linked mechanisms underlying CCA susceptibility. Integrating multi-step post-GWAS approaches enhances understanding of CCA pathogenesis and may facilitate the development of risk biomarkers for early detection and precision prevention strategies.

Background: The epigenome, the set of modifications to DNA and associated molecules that control gene expression, cellular identity, and function, plays a major role in mediating cellular responses to outside factors. Thus, evaluation of the epigenetic state can provide insights into cellular adaptions occurring over the course of disease. Methods: We performed epigenome-wide association studies of primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC) using the Illumina MethylationEPIC Bead Chip. Results: We found evidence of increased epigenetic age acceleration and differences in predicted immune cell composition in patients with PSC and PBC. Epigenetic profiles demonstrated differences in predicted protein levels including increased levels of tumor necrosis factor receptor superfamily member 1B in patients with cirrhotic compared to noncirrhotic PSC and PBC. Epigenome-wide association studies of PSC discovered strongly associated 5′-C-phosphate-G-3′ sites in genes including vacuole membrane protein 1 and SOCS3, and epigenome-wide association studies of PBC found strong 5′-C-phosphate-G-3′ associations in genes including NOD-like receptor family CARD domain containing 5, human leukocyte antigen-E, and PSMB8. Analyses identified disease-associated canonical pathways and upstream regulators involved with immune signaling and activation of macrophages and T-cells. A comparison of PSC and PBC data found relatively little overlap at the 5′-C-phosphate-G-3′ and gene levels with slightly more overlap at the level of pathways and upstream regulators. Conclusions: This study provides insights into methylation profiles of patients that support current concepts of disease mechanisms and provide novel data to inspire future research. Studies to corroborate our findings and expand into other -omics layers will be invaluable to further our understanding of these rare diseases with the goal to improve and individualize prognosis and treatment.

Abstract Primary sclerosing cholangitis (PSC) is a complex bile duct disorder. Its etiology is incompletely understood, but environmental chemicals likely contribute to risk. Patients with PSC have an altered bile metabolome, which may be influenced by environmental chemicals. This novel study utilized state-of-the-art high-resolution mass spectrometry (HRMS) with bile samples to provide the first characterization of environmental chemicals and metabolomics (collectively, the exposome) in PSC patients located in the United States of America (USA) (n = 24) and Norway (n = 30). First, environmental chemical- and metabolome-wide association studies were conducted to assess geographic-based similarities and differences in the bile of PSC patients. Nine environmental chemicals (false discovery rate, FDR &lt; 0.20) and 3143 metabolic features (FDR &lt; 0.05) differed by site. Next, pathway analysis was performed to identify metabolomic pathways that were similarly and differentially enriched by the site. Fifteen pathways were differentially enriched (P &lt; .05) in the categories of amino acid, glycan, carbohydrate, energy, and vitamin/cofactor metabolism. Finally, chemicals and pathways were integrated to derive exposure–effect correlation networks by site. These networks demonstrate the shared and differential chemical–metabolome associations by site and highlight important pathways that are likely relevant to PSC. The USA patients demonstrated higher environmental chemical bile content and increased associations between chemicals and metabolic pathways than those in Norway. Polychlorinated biphenyl (PCB)-118 and PCB-101 were identified as chemicals of interest for additional investigation in PSC given broad associations with metabolomic pathways in both the USA and Norway patients. Associated pathways include glycan degradation pathways, which play a key role in microbiome regulation and thus may be implicated in PSC pathophysiology.

Abstract Background: Primary sclerosing cholangitis (PSC) patients have a risk of developing cholangiocarcinoma (CCA). Establishing predictive models for CCA in PSC is important. Methods: In a large cohort of 1,459 PSC patients seen at Mayo Clinic (1993-2020), we quantified the impact of clinical/laboratory variables on CCA development using univariate and multivariate Cox models and predicted CCA using statistical and artificial intelligence approaches. We explored plasma bile acid (BA) levels’ predictive power of CCA (subset of 300 patients, BA cohort). Results: Eight significant risk factors (false discovery rate: 20%) were identified with univariate analysis; prolonged inflammatory bowel disease (IBD) was the most important one. IBD duration, PSC duration, and total bilirubin remained significant (p<0.05) with multivariate analysis. Clinical/laboratory variables predicted CCA with cross-validated C-indexes of 0.68–0.71 at different time points of disease, significantly better compared to commonly used PSC risk scores. Lower chenodeoxycholic acid, higher conjugated fraction of lithocholic acid and hyodeoxycholic acid, and higher ratio of cholic acid to chenodeoxycholic acid were predictive of CCA. BAs predicted CCA with a cross-validated C-index of 0.66 (std: 0.11, BA cohort), similar to clinical/laboratory variables (C-index=0.64, std: 0.11, BA cohort). Combining BAs with clinical/laboratory variables leads to the best average C-index of 0.67 (std: 0.13, BA cohort). Conclusions: In a large PSC cohort, we identified clinical and laboratory risk factors for CCA development and demonstrated the first artificial intelligence based predictive models that performed significantly better than common risk scores. More predictive data modalities are needed for clinical adoption of these models.

Progress in development of prognostic and therapeutic options for the rare cholestatic liver diseases, primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), is hampered by limited knowledge of their pathogeneses. In particular, the potential role of hepatotoxic and/or metabolism‐altering environmental chemicals in the pathogenesis of these diseases remains relatively unstudied. Moreover, the extent to which metabolic pathways are altered due to ongoing cholestasis and subsequent liver damage or possibly influenced by hepatotoxic chemicals is poorly understood. In this study, we applied a comprehensive exposomics‐metabolomics approach to uncover potential pathogenic contributors to PSC and PBC. We used untargeted high‐resolution mass spectrometry to characterize a wide range of exogenous chemicals and endogenous metabolites in plasma and tested them for association with disease. Exposome‐wide association studies (EWAS) identified environmental chemicals, including pesticides, additives and persistent pollutants, that were associated with PSC and/or PBC, suggesting potential roles for these compounds in disease pathogenesis. Metabolome‐wide association studies (MWAS) found disease‐associated alterations to amino acid, eicosanoid, lipid, co‐factor, nucleotide, mitochondrial and microbial metabolic pathways, many of which were shared between PSC and PBC. Notably, this analysis implicates a potential role of the 5‐lipoxygenase pathway in the pathogenesis of these diseases. Finally, EWAS × MWAS network analysis uncovered linkages between environmental agents and disrupted metabolic pathways that provide insight into potential mechanisms for PSC and PBC. Conclusion: This study establishes combined exposomics‐metabolomics as a generalizable approach to identify potentially pathogenic environmental agents and enumerate metabolic alterations that may impact PSC and PBC, providing a foundation for diagnostic and therapeutic strategies.