COBALT: A Confirmatory Trial of Obeticholic Acid in Primary Biliary Cholangitis With Placebo and External Controls.

OBJECTIVES: Obeticholic acid (OCA) treatment for primary biliary cholangitis (PBC) was conditionally approved in the phase 3 POISE trial. The COBALT confirmatory trial assessed whether clinical outcomes in PBC patients improve with OCA therapy. METHODS: Patients randomized to OCA (5-10 mg) were compared with placebo (randomized controlled trial [RCT]) or external control (EC). The primary composite endpoint was time to death, liver transplant, model for end-stage liver disease score ≥15, uncontrolled ascites, or hospitalization for hepatic decompensation. A prespecified propensity score-weighted EC group was derived from a US healthcare claims database. RESULTS: In the RCT, the primary endpoint occurred in 28.6% of OCA (n=168) and 28.9% of placebo patients (n=166; intent-to-treat [ITT] analysis hazard ratio [HR]=1.01, 95% CI=0.68-1.51), but functional unblinding and crossover to commercial therapy occurred, especially in the placebo arm. Correcting for these using inverse probability of censoring weighting (IPCW) and as-treated analyses shifted the HR to favor OCA. In the EC (n=1051), the weighted primary endpoint occurred in 10.1% of OCA and 21.5% of non-OCA patients (HR=0.39; 95% CI=0.22-0.69; P=0.001). No new safety signals were identified in the RCT. CONCLUSIONS: Functional unblinding and treatment crossover, particularly in the placebo arm, confounded the ITT estimate of outcomes associated with OCA in the RCT. Comparison with the real-world EC showed that OCA treatment significantly reduced the risk of negative clinical outcomes. These analyses demonstrate the value of EC data in confirmatory trials and suggest that treatment with OCA improves clinical outcomes in patients with PBC.

Efficacy and safety of palliative treatment in patients with autoimmune liver disease-associated hepatocellular carcinoma.

INTRODUCTION AND OBJECTIVES: Autoimmune liver diseases (AILD) are rare causes hepatocellular carcinoma (HCC). and data on the efficacy and tolerability of anti-tumour therapies are scarce. This pan-European study aimed to assess outcomes in AILD-HCC patients treated with tyrosine kinase inhibitors (TKIs) or transarterial chemoembolization (TACE) compared with patients with more common HCC etiologies, including viral, alcoholic or non-alcoholic fatty liver disease. MATERIALS AND METHODS: 107 patients with HCC-AILD (AIH:55; PBC:52) treated at 13 European centres between 1996 and 2020 were included. 65 received TACE and 28 received TKI therapy. 43 (66%) were female (median age 73 years) with HCC tumour stage BCLC A (34%), B (46%), C (9%) or D (11%). For each treatment type, propensity score matching was used to match AILD to non-AILD-HCC on a 1:1 basis, yielding in a final cohort of 130 TACE and 56 TKI patients for comparative analyses of median overall survival (mOS) and treatment tolerability. RESULTS: HCC-AILD patients showed comparable mOS to controls for both TACE (19.5 vs 22.1 months, p=0.9) and TKI (15.4 vs 15.1 months, p=0.5). Adverse events were less frequent in AILD-HCC patients than controls (33% vs 62%, p=0.003). For TKIs, there were no significant differences in adverse events (73% vs. 86%, p=0.2) or interruption rates (44% vs. 36%, p=0.7). CONCLUSIONS: In summary, this study demonstrates comparable mOS for AILD-HCC patients undergoing local and systemic treatments, with better tolerability than HCC of other causes. TKIs remain important therapeutic options for AILD-HCC patients, particularly given their exclusion from recent immunotherapy trials.

Quality of Life and Clinical Outcomes of Endosonography-Guided Biliary Drainage in Patients with Malignant Biliary Obstruction: A Single-Center, Prospective Analysis.

Introduction: Endosonography-guided biliary drainage (EUS-BD) serves as a rescue treatment modality for patients with malignant biliary obstruction when endoscopic retrograde cholangiopancreatography (ERCP) fails. Objectives: This study explores the effects of EUS-BD on liver function and quality of life (QoL). Patients and Methods: Patients with malignant biliary obstruction and failed ERCP were enrolled to undergo EUS-BD. QoL, including pruritus severity, was evaluated using EQ-5D-5L and PSS-10 questionnaires before and after EUS-BD. Serum bilirubin and liver function tests were measured on the procedure day, two days, and at least 14 days post-procedure. Results: During a 20-month study period, 1755 ERCPs were performed, with 595 for malignant cases. Of these, 49 underwent EUS-BD following failed ERCP, and 37 (54% women, age range 34-87 years) completed the 14-day follow-up. Pancreatic cancer was the most common (49%) condition, and the median hospital stay was 4 days. Serum bilirubin, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and gamma-glutamyl transferase significantly decreased 2 and 14 days after EUS-BD (all p < 0.001). Pruritus significantly improved, with an average reduction of 5.19 points on the PSS-10 scale two weeks post-procedure (p < 0.001). EUS-BD led to improvements in anxiety and depression according to the EQ-5D-5L (p = 0.013). Conversely, deteriorations were observed in the Mobility, Self-Care, and Usual Activities domains over time (all p < 0.05). Successful EUS-BD enabled the resumption of chemotherapy in 11 (30%) patients. The median post-procedure survival was 112 (range 27-1030) days. Conclusions: EUS-BD improves liver parameters and some aspects of life quality in patients with malignant biliary obstruction, thereby increasing their eligibility for optimal palliative care.

Modulation of miR-155-5p signalling via 5-ASA for the prevention of high microsatellite instability: an in vitro study using human epithelial cell lines.

5-aminosalicylic acid (5-ASA) is a first-line treatment for maintaining colitis remission. It is a highly effective, safe, and well-tolerated drug with anti-inflammatory and chemo-preventive properties. While patients with primary sclerosing cholangitis (PSC) with concomitant ulcerative colitis are treated with 5-ASA, the molecular mechanisms underlying the drug's chemo-preventive effects are not entirely understood. We previously reported that bile acids and lipopolysaccharide-induced miR-155 expression was associated with downregulating mismatch repair (MMR) proteins in CACO-2 cell lines. Therefore, in this investigation, a set of in vitro functional studies was performed to show the possible mechanisms behind the epigenetic relationship between miR-155 and 5-ASA's prevention of high microsatellite instability (MSI-H). In transient transfection with miR-155Mimic, which behaves like endogenous miRNA, we confirmed the relationships between miR-155 and its target MMR in three human intestinal epithelial cell lines: CACO-2, NCM460D and HT-29. We have shown, for the first time, that 5-ASA modulates MLH1, MSH2, MSH6 in miR-155 transfected cells. These findings underline that chemoprotective 5-ASA therapy can effectively attenuate the expression of miR-155 and potentially prevent a development of MSI-H in a subset of colorectal cancers associated with PSC.

Unmet needs in autoimmune hepatitis: Results of the prospective multicentre European Reference Network Registry (R-LIVER).

BACKGROUND AND AIMS: The European Reference Network on Hepatological Diseases (ERN RARE-LIVER) launched the prospective, multicentre, quality-controlled R-LIVER registry on rare liver diseases. The aim of this study was to assess the presentation and outcome of autoimmune hepatitis (AIH) after 1 year of treatment. METHODS: Data were prospectively collected at the time of diagnosis and after 6 and 12 months follow-up. Complete biochemical response (CBR) was defined as normalization of alanine aminotransferase (ALT) and immunoglobulin G (IgG) serum levels. RESULTS: A total of 231 patients from six European centres were included in the analysis. After 6 months of treatment 50% (106/212), and after 12 months 63% (131/210) of patients reached CBR with only 27% (56/211) achieving a steroid-free CBR within the first year. Overall, 16 different treatment regimens were administered. Change of treatment, mostly due to intolerance, occurred in 30.4% within the first 6 months. In multivariate analysis, younger age at diagnosis (odds ratio [OR] = 1.03 [95% confidence interval (CI) 1.01-1.05]; p = .007), severe fibrosis (OR .38 [95% .16-.89], p = .026) and change of treatment within the first 6 months (OR .40 [95% CI .2-.86]; p = .018) were associated with a lesser chance of ALT normalization at 12 months follow-up. CONCLUSION: The landscape of AIH treatment in Europe is highly heterogeneous, even between expert centres. The results from this first European multicentre prospective registry reveal several unmet needs, highlighted by the overall low rates of CBR and the frequent failure to withdraw corticosteroids.

Antimicrobial glycoprotein 2 (GP2) in gallstones, bile fluid and peribiliary glands of patients with primary sclerosing cholangitis.

BACKGROUND: Glycoprotein-2 (GP2) IgA is a predictor of disease severity in primary sclerosing cholangitis (PSC). We examined GP2's occurrence in the biliary tract, the site of inflammation. METHODS: GP2 was analyzed using ELISA, immunoblotting, mass spectrometry, and immunohistochemistry. The samples included: 20 bile and 30 serum samples from PSC patients, 23 bile and 11 serum samples from patients with gallstone disease (GD), 15 bile samples from healthy individuals undergoing liver-donation surgery (HILD), 20 extracts of gallstones (GE) obtained during cholecystectomy, and 101 blood-donor sera. RESULTS: Biliary GP2 concentrations were significantly higher in patients with PSC and GD than in HILD (p < 0.0001). Serum GP2 levels were similar in PSC and GD patients, and controls, but lower than in bile (p < 0.0001). GP2 was detected in all 20 GEs. Mass spectrometry identified GP2 in the bile of 2 randomly selected GD and 2 PSC patients, and in none of 2 HILD samples. GP2 was found in peribiliary glands in 8 out of 12 PSC patients, showing morphological changes in acinar cells, but not in GD-gallbladders. CONCLUSIONS: GP2 is present in bile of PSC and GD patients. It is synthesized in the peribiliary glands of PSC patients, supporting a pathogenic role for biliary GP2 in PSC.

Loss of mucosal tolerance to glycoprotein 2 isoform 1 is a potential novel diagnostic biomarker for cholangiocarcinoma.

BACKGROUND: Anti-glycoprotein 2 (anti-GP2) IgA and antineutrophil-cytoplasmic antibodies to proteinase 3 (PR3-ANCA) have been reported as predictive markers of cholangiocarcinoma (CCA) in patients with primary sclerosing cholangitis (PSC), but their prevalence in CCA patients without PSC remains unclear. METHODS: This study involved Asian discovery (n = 118) and European validation (n = 38) cohorts of CCA patients without PSC, alongside 49 Asian and 82 European pancreatic ductal adenocarcinoma (PDAC) patients, 21 with benign pancreatic neoplasms (BPN) and 45 with hepatocellular carcinoma (HCC), and 157 healthy controls (HC) from Asia and Europe. We analyzed the prevalence of PR3-ANCA, IgA and IgG against GP21 and GP24, and the CA19-9 levels. RESULTS: Anti-GP21 IgA was the most prevalent in both CCA cohorts (discovery: 55.1 %; validation: 42.1 %) and significantly higher than in other groups except PDAC (all p < 0.05). It demonstrated the best diagnostic performance in distinguishing CCA from disease controls and HC, outperforming tumor markers. No significant correlation was found between anti-GP21 IgA levels and CA19-9 levels. CONCLUSION: Our findings show that anti-GP21 IgA revealing the loss of mucosal tolerance is a potential novel diagnostic biomarker for CCA.

Previous hepatitis E virus infection is associated with increased liver stiffness in patients with autoimmune hepatitis.

INTRODUCTION: Autoimmune hepatitis (AIH) is a chronic, progressive liver disease that, in most cases, may require lifelong immunosuppression. Hepatitis E virus (HEV) is a leading cause of acute, typically self­limited hepatitis worldwide, although immunocompromised patients may develop chronic hepatitis. OBJECTIVES: We aimed to evaluate the impact of HEV seropositivity on the clinical course of AIH. PATIENTS AND METHODS: The study involved a group of 374 adult patients with AIH (68% women; median [interquartile range] age, 34 [18-83] years; 38% with liver cirrhosis). Serum HEV immunoglobulin (Ig) G and IgM antibodies were measured by enzyme­linked immunosorbent assay, liver fibrosis was assessed by liver stiffness measurement (LSM), and liver cirrhosis was confirmed with liver histology or LSM. RESULTS: Fifty­five patients (15%) with AIH were HEV IgG­positive. These patients were older (P <0.001), had higher body mass index, and higher value of LSM (both P <0.05). In a multivariable model including the levels of alanine aminotransferase and IgG, the HEV seropositive status was associated with an increased risk of advanced liver fibrosis with odds ratio of 3.69 (95% CI, 1.26-10.77; P = 0.02), as reflected by liver stiffness equal to or above 10.5 kPa. HEV IgG seropositivity was, however, not linked with the type of treatment or worse AIH outcome. Seroprevalence of HEV in the patients with AIH was lower than in the general population of Polish blood donors (43%). CONCLUSIONS: Patients with AIH and HEV IgG­positive status seem to be at risk of more advanced liver fibrosis. However, the overall seroprevalence of HEV IgG is lower in patients with AIH than in blood donors in Poland.

The ABCG8 polymorphism increases the risk of gallbladder cancer in the general population and gallstones in obese patients from Poland.

BACKGROUND: Gallstone disease (GD) is common but remains asymptomatic in most cases. However, gallstones can lead to complications like choledocholithiasis or gallbladder cancer. In this study, we analyse the common genetic risk factor for GD, the p.D19H variant in the sterol transporter ABCG8, in Polish patients with gallstones and gallbladder cancer. METHODS: Three adult cohorts were prospectively recruited: 65 patients with gallbladder cancer, 170 obese individuals scheduled for bariatric surgery and 72 patients who underwent endoscopic retrograde cholangiopancreatography due to recurrent choledocholithiasis. The control cohort consisted of 172 gallstone-free adults. The ABCG8 p.D19H (rs11887534) polymorphism was genotyped using TaqMan assays. RESULTS: The minor allele frequency (MAF) of the ABCG8 p.D19H polymorphism was significantly (p = .02) higher among cases with either gallstones or gallbladder cancer (MAF = 8.4%) as compared to controls (MAF = 4.0%). The highest frequency of the risk allele was detected in patients with gallbladder cancer (18.5%) and obese patients with GD (17.5%), followed by individuals with choledocholithiasis (13.9%). Notably, the p.19H variant was associated with an increased risk of developing gallbladder cancer (OR 2.76, 95% CI 1.16-6.54, p = .01) and an increased risk of GD in obese individuals scheduled for bariatric surgery (OR = 2.70, 95% CI 1.05-6.49, p = .03), but did not significantly affect the risk of choledocholithiasis. CONCLUSIONS: The ABCG8 p.D19H common risk variant increases the risk of developing gallbladder cancer in Central Europeans and enhances the risk of gallstones in the obese. Carriers of the p.D19H variant might benefit from personalized preventive strategies, particularly regarding gallbladder cancer.

Oxidative stress in metabolic dysfunction-associated steatotic liver disease (MASLD): How does the animal model resemble human disease?

Despite decades of research, the pathogenesis of metabolic dysfunction-associated steatotic liver disease (MASLD) is still not completely understood. Based on the evidence from preclinical models, one of the factors proposed as a main driver of disease development is oxidative stress. This study aimed to search for the resemblance between the profiles of oxidative stress and antioxidant defense in the animal model of MASLD and the group of MASLD patients. C57BL/6J mice were fed with the Western diet for up to 24 weeks and served as the animal model of MASLD. The antioxidant profile of mice hepatic tissue was determined by liquid chromatography-MS3 spectrometry (LC-MS/MS). The human cohort consisted of 20 patients, who underwent bariatric surgery, and 6 controls. Based on histological analysis, 4 bariatric patients did not have liver steatosis and as such were also classified as controls. Total antioxidant activity was measured in sera and liver biopsy samples. The hepatic levels of antioxidant enzymes and oxidative damage were determined by Western Blot. The levels of antioxidant enzymes were significantly altered in the hepatic tissue of mice with MASLD. In contrast, there were no significant changes in the antioxidant profile of hepatic tissue of MASLD patients, except for the decreased level of carbonylated proteins. Decreased protein carbonylation together with significant correlations between the thioredoxin system and parameters describing metabolic health suggest alterations in the thiol-redox signaling. Altogether, these data show that even though the phenotype of mice closely resembles human MASLD, the animal-to-human translation of cellular and molecular processes such as oxidative stress may be more challenging.

Increased type-I interferon level is associated with liver damage and fibrosis in primary sclerosing cholangitis.

BACKGROUND: The level of type-I interferons (IFNs) in primary sclerosing cholangitis (PSC) was investigated to evaluate its association with disease activity and progression. METHODS: Bioactive type-I IFNs were evaluated in a murine model of PSC and human patients' sera using a cell-based reporter assay and ELISA techniques. In total, 57 healthy participants, 71 PSC, and 38 patients with primary biliary cholangitis were enrolled in this study. RESULTS: Bioactive type-I IFNs were elevated in the liver and serum of multidrug resistance protein 2-deficient animals and showed a correlation with the presence of CD45+ immune cells and serum alanine transaminase levels. Concordantly, bioactive type-I IFNs were elevated in the sera of patients with PSC as compared to healthy controls (sensitivity of 84.51%, specificity of 63.16%, and AUROC value of 0.8267). Bioactive IFNs highly correlated with alkaline phosphatase (r=0.4179, p<0.001), alanine transaminase (r=0.4704, p<0.0001), and gamma-glutamyl transpeptidase activities (r=0.6629, p<0.0001) but not with serum bilirubin. In addition, patients with PSC with advanced fibrosis demonstrated significantly higher type-I IFN values. Among the type-I IFN subtypes IFNα, ß and IFNω could be detected in patients with PSC with IFNω showing the highest concentration among the subtypes and being the most abundant among patients with PSC. CONCLUSIONS: The selectively elevated bioactive type-I IFNs specifically the dominating IFNω could suggest a novel inflammatory pathway that might also have a hitherto unrecognized role in the pathomechanism of PSC.

Maternal liver-related symptoms during pregnancy in primary sclerosing cholangitis.

Background & Aims: Although worsening liver-related symptoms during pregnancy can occur in primary sclerosing cholangitis (PSC), there are insufficient data to effectively counsel patients on their pre-conception risk and no clear recommendations on monitoring and management during pregnancy. We aimed to describe maternal liver-related symptoms in pregnancy, both before and after PSC diagnosis, and explore factors associated with worsening symptoms and liver-related outcomes. Methods: We conducted a multicentre retrospective observational study of females with PSC and known pregnancy with live birth, via the International PSC Study Group. We included 450 patients from 12 European centres. Data included clinical variables, liver-related symptoms (pruritus and/or cholangitis) during pregnancy, and liver biochemistry. A composite primary endpoint of transplant-free survival from time of PSC diagnosis was used. Results: There were 266 pregnancies in 178 patients following PSC diagnosis. Worsening liver-related symptoms were reported in 66/228 (28.9%) pregnancies; they had a reduced transplant-free survival (p = 0.03), which retained significance on multivariate analysis (hazard ratio 3.02, 95% CI 1.24-7.35; p = 0.02).Abnormal biochemistry and/or liver-related symptoms (pruritus and/or cholangitis) were noted during pregnancy before PSC diagnosis in 21/167 (12.6%) patients. They had a reduced transplant-free survival from pregnancy (p = 0.01), which did not retain significance in a multivariable model (hazard ratio 1.10, 95% CI 0.43-2.85; p = 0.84). Conclusions: Liver-related symptoms are frequently encountered during pregnancies before the diagnosis of PSC, and pregnancy may expose the pre-clinical phase of PSC in some patients. Worsening liver-related symptoms were seen in a third of our cohort with known PSC during pregnancy; and this subgroup had a poorer prognosis, which may be related to more advanced liver disease at time of pregnancy and/or a more severe disease phenotype. Impact and implications: Patients with PSC can develop worsening of their liver-related symptoms during pregnancy; however, risk factors for this and the long-term implications are not known. We identified that there is a significant risk of these symptoms in pregnancy, both before and after PSC has been diagnosed, particularly in patients with elevated alkaline phosphatase. Furthermore, our findings suggest that worsening symptoms during pregnancy may be associated with adverse long-term clinical outcomes of liver transplantation and death in patients with known PSC. This may be related to the presence of more advanced liver disease at time of pregnancy. This information can be used to counsel patients with PSC before conception and identify patients who need close follow-up after delivery.

Effect of Low Testosterone Levels on the Expression of Proliferator-Activated Receptor Alpha in Female Patients with Primary Biliary Cholangitis.

Sex-dependent patterns in chronic immune-mediated cholangiopathies, like primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC), remain poorly understood. Peroxisome proliferator-activated receptor alpha (PPAR-α), expressed in immune cells, plays a key role in innate defence. In this study, the relationship between PPAR-α expression in peripheral blood mononuclear cells (PBMCs), serum androgen levels, IFNγ production, and sex-dependent tendencies during the development of PBC and PSC was investigated. We confirmed that normal cholangiocytes respond to PPAR-α and inhibit the lipopolysaccharide-induced expression of IL-6, IL-1b, and TNFα. Compared with PSC patients, PPAR-α was downregulated, while IFNγ was upregulated, in the PBMCs of PBC patients. When the analysis was conducted on females only, there was no difference in PPAR-α, but IFNγ was elevated in females with PBC compared with those with PSC. Serum testosterone concentrations in females with PBC were below the normal range (regardless of age) and correlated positively with PPAR-α and negatively with IFNγ. While PPAR-α has been reported to be a target of miR-155 and miR-21, no correlations with these microRNAs were observed in the PBMCs. However, a positive correlation between miR-21 and IFNγ was observed. Our results showed suppressed PPAR-α expression accompanied by reduced testosterone levels in women with PBC, which should elicit interest in the role of testosterone in PBC development.

The Role of hsa-miR-125b-5p Interaction with S1P/Ceramide Axis in the Potential Development of Inflammation-Associated Colon Cancer in Primary Sclerosing Cholangitis.

Primary sclerosing cholangitis (PSC) is characterised by the co-occurrence of inflammatory bowel diseases, particularly ulcerative colitis (UC). We investigated how the interaction of miR-125b with the sphingosine-1-phosphate (S1P)/ceramide axis may predispose patients with PSC, PSC/UC, and UC to carcinogenesis in the ascending and sigmoid colons. The overexpression of miR-125b was accompanied by the upregulation of S1P, ceramide synthases, ceramide kinases, and the downregulation of AT-rich interaction domain 2 in the ascending colon of PSC/UC, which contributed to the progression of high microsatellite instability (MSI-H) colorectal carcinoma. We also showed that the overexpression of sphingosine kinase 2 (SPHK2) and the genes involved in the glycolytic pathway in the sigmoid colon of UC led to the upregulation of Interleukin 17 (IL-17). In vitro stimulation of human intestinal epithelial cells (Caco-2, HT-29, and NCM460D) with lipopolysaccharide suppressed miR-125b and increased proinflammatory cytokines, whereas the induction of miR-125b activity by either a miR-125b mimetic or lithocholic acid resulted in the inhibition of miR-125b targets. In summary, miR-125b overexpression was associated with an imbalance in the S1P/ceramide axis that can lead to MSI-H cancer progression in PSC/UC. Furthermore, SPHK2 overexpression and a change in the cellular metabolic flux are important players in inflammation-associated colon cancer in UC.

Health-related quality of life and symptoms in autoimmune liver diseases.

Assessment of Health-Related Quality of Life (HRQoL) has emerged as an important tool in the evaluation of both the well-being of patients and the results of their clinical management. Over the years, a large number of questionnaires focusing on various aspects of quality of life have been developed. They are frequently divided into generic questionnaires, which can be used under various conditions, disease-specific and symptom-specific questionnaires. Autoimmune liver diseases, such as autoimmune hepatitis, primary sclerosing cholangitis, or primary biliary cirrhosis, comprise a group of rare liver conditions (i.e. affecting fewer than 5 in 10,000 people in the general population). Unfortunately, HRQoL has not been well-studied in this group of patients. In this review, we comprehensively summarize the data available in the literature on HRQoL in these conditions, emphasizing the important role that quality of life plays in the successful management of such patients.

Liquid biopsy-based protein biomarkers for risk prediction, early diagnosis, and prognostication of cholangiocarcinoma.

BACKGROUND & AIMS: Cholangiocarcinoma (CCA), heterogeneous biliary tumours with dismal prognosis, lacks accurate early diagnostic methods especially important for individuals at high-risk (i.e. those with primary sclerosing cholangitis [PSC]). Here, we searched for protein biomarkers in serum extracellular vesicles (EVs). METHODS: EVs from patients with isolated PSC (n = 45), concomitant PSC-CCA (n = 44), PSC who developed CCA during follow-up (PSC to CCA; n = 25), CCAs from non-PSC aetiology (n = 56), and hepatocellular carcinoma (n = 34) and healthy individuals (n = 56) were characterised by mass spectrometry. Diagnostic biomarkers for PSC-CCA, non-PSC CCA, or CCAs regardless of aetiology (Pan-CCAs) were defined and validated by ELISA. Their expression was evaluated in CCA tumours at a single-cell level. Prognostic EV biomarkers for CCA were investigated. RESULTS: High-throughput proteomics of EVs identified diagnostic biomarkers for PSC-CCA, non-PSC CCA, or Pan-CCA, and for the differential diagnosis of intrahepatic CCA and hepatocellular carcinoma, which were cross-validated by ELISA using total serum. Machine learning-based algorithms disclosed CRP/FIBRINOGEN/FRIL for the diagnosis of PSC-CCA (local disease [LD]) vs. isolated PSC (AUC = 0.947; odds ratio [OR] =36.9) and, combined with carbohydrate antigen 19-9, overpowers carbohydrate antigen 19-9 alone. CRP/PIGR/VWF allowed the diagnosis of LD non-PSC CCAs vs. healthy individuals (AUC = 0.992; OR = 387.5). It is noteworthy that CRP/FRIL accurately diagnosed LD Pan-CCA (AUC = 0.941; OR = 89.4). Levels of CRP/FIBRINOGEN/FRIL/PIGR showed predictive capacity for CCA development in PSC before clinical evidence of malignancy. Multi-organ transcriptomic analysis revealed that serum EV biomarkers were mostly expressed in hepatobiliary tissues, and single-cell RNA sequencing and immunofluorescence analysis of CCA tumours showed their presence mainly in malignant cholangiocytes. Multivariable analysis unveiled EV prognostic biomarkers, with COMP/GNAI2/CFAI and ACTN1/MYCT1/PF4V associated negatively and positively with patients' survival, respectively. CONCLUSIONS: Serum EVs contain protein biomarkers for the prediction, early diagnosis, and prognostication of CCA that are detectable using total serum, representing a tumour cell-derived liquid biopsy tool for personalised medicine. IMPACT AND IMPLICATIONS: The accuracy of current imaging tests and circulating tumour biomarkers for cholangiocarcinoma (CCA) diagnosis is far from satisfactory. Most CCAs are considered sporadic, although up to 20% of patients with primary sclerosing cholangitis (PSC) develop CCA during their lifetime, constituting a major cause of PSC-related death. This international study has proposed protein-based and aetiology-related logistic models with predictive, diagnostic, or prognostic capacities by combining two to four circulating protein biomarkers, moving a step forward into personalised medicine. These novel liquid biopsy tools may allow the (i) easy and non-invasive diagnosis of sporadic CCAs, (ii) identification of patients with PSC with higher risk for CCA development, (iii) establishment of cost-effective surveillance programmes for the early detection of CCA in high-risk populations (e.g. PSC), and (iv) prognostic stratification of patients with CCA, which, altogether, may increase the number of cases eligible for potentially curative options or to receive more successful treatments, decreasing CCA-related mortality.

Improving quality of life in patients with rare autoimmune liver diseases by structured peer-delivered support (Q.RARE.LI): study protocol for a transnational effectiveness-implementation hybrid trial.

BACKGROUND: Psychosocial support is a crucial component of adequate rare disease care, but to date psychosocial support needs of this patient population are insufficiently met. Within Q.RARE.LI, we strive to evaluate the effectiveness of a structured, transdiagnostic, and location-independent psychosocial support intervention in routine care of patients with rare autoimmune liver diseases in five countries and prepare its implementation. METHODS: Within an effectiveness-implementation hybrid trial, we aim to a) investigate the effectiveness of the intervention in routine care in five diverse healthcare systems and b) assess implementation outcomes, examine and prepare the implementation context, and develop country-specific implementation strategies. To assess effectiveness, we will include N = 240 patients with rare autoimmune liver diseases. Within a two-armed randomized controlled trial (allocation ratio 1:1), we will compare structured and peer-delivered psychosocial support in addition to care-as-usual (CAU) with CAU alone. Outcomes will be assessed via electronic database entry prior to intervention, directly after, and at a three-month follow-up. Our primary effectiveness outcome will be mental health-related quality of life at post-assessment. Secondary outcomes include depression and anxiety severity, perceived social support, helplessness, and disease acceptance. Implementation outcomes will be assessed within a mixed-methods process evaluation. In a quantitative cross-sectional survey, we will examine perceived acceptability and feasibility in patients, peer-counselors, and healthcare providers involved in delivery of the intervention. In qualitative focus groups, we will analyze the implementation context and determine barriers and facilitators for implementation with different stakeholders (patients and/or representatives, peer-counselors, healthcare providers, health insurers). Based on these results, we will derive country-specific implementation strategies and develop a concrete implementation plan for each country. DISCUSSION: The intervention is expected to help patients adjust to their disease and improve their mental quality of life. The transdiagnostic and location-independent program has the potential to reach patients for psychosocial support who are usually hard to reach. By preparing the implementation in five countries, the project can help to make low-threshold psychosocial support available to many patients with rare diseases and improve comprehensive healthcare for an often neglected group. TRIAL REGISTRATION: ISRCTN15030282.

Liver phenotypes in PCOS: Analysis of exogenous and inherited risk factors for liver injury in two European cohorts.

BACKGROUND & AIMS: Fatty liver disease (FLD) is common in women with polycystic ovary syndrome (PCOS). Here, we use non-invasive tests to quantify liver injury in women with PCOS and analyse whether FLD-associated genetic variants contribute to liver phenotypes in PCOS. METHODS: Prospectively, we recruited women with PCOS and controls at two university centres in Germany and Poland. Alcohol abuse was regarded as an exclusion criterion. Genotyping of variants associated with FLD was performed using TaqMan assays. Liver stiffness measurements (LSM), controlled attenuation parameters (CAP) and non-invasive HSI, FLI, FIB-4 scores were determined to assess hepatic steatosis and fibrosis. RESULTS: A total of 42 German (age range 18-53 years) and 143 Polish (age range 18-40 years) women with PCOS, as well as 245 German and 289 Polish controls were recruited. In contrast to Polish patients, Germans were older, presented with more severe metabolic profiles and had significantly higher LSM (median 5.9 kPa vs. 3.8 kPa). In the German cohort, carriers of the PNPLA3 p.I148M risk variant had an increased LSM (p = .01). In the Polish cohort, the minor MTARC1 allele was linked with significantly lower serum aminotransferases activities, whereas the HSD17B13 polymorphism was associated with lower concentrations of 17-OH progesterone, total testosterone, and androstenedione (all p < .05). CONCLUSIONS: FLD is common in women with PCOS. Its extent is modulated by both genetic and metabolic risk factors. Genotyping of variants associated with FLD might help to stratify the risk of liver disease progression in women suffering from PCOS.

Variants of autoimmune liver diseases: how to diagnose and treat them?

Autoimmune liver diseases (AILDs), such as autoimmune hepatitis (AIH), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC), are classified as rare diseases, but their incidence is increasing. In this review, we present the characteristics of AILDs in adults, and mainly focus on their variants in terms of diagnosis and management. The classic AILDs have been well defined in clinical guidelines, but a proportion of patients with a single AILD tend to show features of other AILDs. In these cases, AIH­PSC or AIH­PBC variants should be suspected, prompting evaluation in experienced centers. These variants are more representative of clinical categories rather than pathological diagnoses, and the leading component of the disease determines its treatment. However, treating these patients is challenging, even for experienced clinicians. Progression to end­stage liver disease is, unfortunately, not a rare course, despite combined and second­line therapies, particularly for AIH­PSC variants. Thus, studies based on prospective registers are necessary to elaborate upon widely accepted guidelines, to offer better care to these patients, and to improve their prognosis.