Whole-exome sequencing reveals novel cancer genes and actionable targets in biliary tract cancers in primary sclerosing cholangitis.

BACKGROUND: People with primary sclerosing cholangitis (PSC) have a 20% lifetime risk of biliary tract cancer (BTC). Using whole-exome sequencing, we characterized genomic alterations in tissue samples from BTC with underlying PSC. METHODS: We extracted DNA from formalin-fixed, paraffin-embedded tumor and paired nontumor tissue from 52 resection or biopsy specimens from patients with PSC and BTC and performed whole-exome sequencing. Following copy number analysis, variant calling, and filtering, putative PSC-BTC-associated genes were assessed by pathway analyses and annotated to targeted cancer therapies. RESULTS: We identified 53 candidate cancer genes with a total of 123 nonsynonymous alterations passing filtering thresholds in 2 or more samples. Of the identified genes, 19% had not previously been implicated in BTC, including CNGA3, KRT28, and EFCAB5. Another subset comprised genes previously implicated in hepato-pancreato-biliary cancer, such as ARID2, ELF3, and PTPRD. Finally, we identified a subset of genes implicated in a wide range of cancers such as the tumor suppressor genes TP53, CDKN2A, SMAD4, and RNF43 and the oncogenes KRAS, ERBB2, and BRAF. Focal copy number variations were found in 51.9% of the samples. Alterations in potential actionable genes, including ERBB2, MDM2, and FGFR3 were identified and alterations in the RTK/RAS (p = 0.036), TP53 (p = 0.04), and PI3K (p = 0.043) pathways were significantly associated with reduced overall survival. CONCLUSIONS: In this exome-wide characterization of PSC-associated BTC, we delineated both PSC-specific and universal cancer genes. Our findings provide opportunities for a better understanding of the development of BTC in PSC and could be used as a platform to develop personalized treatment approaches.

Comparing Survival of Perihilar Cholangiocarcinoma After R1 Resection Versus Palliative Chemotherapy for Unresected Localized Disease.

BACKGROUND: Resection of perihilar cholangiocarcinoma (pCCA) is a complex procedure with a high risk of postoperative mortality and early disease recurrence. The objective of this study was to compare patient characteristics and overall survival (OS) between pCCA patients who underwent an R1 resection and patients with localized pCCA who received palliative systemic chemotherapy. METHODS: Patients with a diagnosis of pCCA between 1997-2021 were identified from the European Network for the Study of Cholangiocarcinoma (ENS-CCA) registry. pCCA patients who underwent an R1 resection were compared with patients with localized pCCA (i.e., nonmetastatic) who were ineligible for surgical resection and received palliative systemic chemotherapy. The primary outcome was OS. RESULTS: Overall, 146 patients in the R1 resection group and 92 patients in the palliative chemotherapy group were included. The palliative chemotherapy group more often underwent biliary drainage (95% vs. 66%, p < 0.001) and had more vascular encasement on imaging (70% vs. 49%, p = 0.012) and CA 19.9 was more frequently >200 IU/L (64 vs. 45%, p = 0.046). Median OS was comparable between both groups (17.1 vs. 16 months, p = 0.06). Overall survival at 5 years after diagnosis was 20.0% with R1 resection and 2.2% with chemotherapy. Type of treatment (i.e., R1 resection or palliative chemotherapy) was not an independent predictor of OS (hazard ratio 0.76, 95% confidence interval 0.55-1.07). CONCLUSIONS: Palliative systemic chemotherapy should be considered instead of resection in patients with a high risk of both R1 resection and postoperative mortality.

Intergenic risk variant rs56258221 skews the fate of naive CD4+ T cells via miR4464-BACH2 interplay in primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is an immune-mediated liver disease of unknown pathogenesis, with a high risk to develop cirrhosis and malignancies. Functional dysregulation of T cells and association with genetic polymorphisms in T cell-related genes were previously reported for PSC. Here, we genotyped a representative PSC cohort for several disease-associated risk loci and identified rs56258221 (BACH2/MIR4464) to correlate with not only the peripheral blood T cell immunophenotype but also the functional capacities of naive CD4+ T (CD4+ TN) cells in people with PSC. Mechanistically, rs56258221 leads to an increased expression of miR4464, in turn causing attenuated translation of BACH2, a major gatekeeper of T cell quiescence. Thereby, the fate of CD4+ TN is skewed toward polarization into pro-inflammatory subsets. Clinically, people with PSC carrying rs56258221 show signs of accelerated disease progression. The data presented here highlight the importance of assigning functional outcomes to disease-associated genetic polymorphisms as potential drivers of diseases.

Whole blood RNA sequencing identifies transcriptional differences between primary sclerosing cholangitis and ulcerative colitis.

Background & Aims: Genetic and microbiome studies across patients with primary sclerosing cholangitis (PSC) and ulcerative colitis (UC) have indicated that UC in PSC is a separate disease entity to primary UC, but expression studies for PSC are lacking. Methods: We conducted whole blood RNA sequencing experiments for 495 patients with UC, 220 patients with PSC (including 177 with UC), and 320 healthy controls from Germany and Norway. Differential expression analyses, gene ontology and coexpression analyses and random forest machine learning were performed to identify genes, ontologies and transcriptional features that discriminate diagnoses. Results: The blood transcriptome in UC and PSC is dominated by neutrophil activation genes (e.g. S100A12). In UC, but not in PSC (neither PSC alone nor patients with an additional diagnosis of UC [PSC/UC]), ribosomal, mitochondrial, and energy metabolism genes are upregulated in conjunction with antibody transcript expression (MZB1, IGJ). In PSC, there is an increase in modules related to apoptosis and expression of genes of interferon-I-related ontologies. Random forest analysis could poorly discriminate PSC alone from PSC/UC (AUROC 0.56), but could discriminate PSC, UC, and controls with high accuracy (AUROC UC vs. controls 0.95, PSC vs. controls 0.88, UC vs. PSC 0.986). The main coexpression modules relevant for distinguishing PSC, UC, and controls are enriched in neutrophil degranulation and antibody production genes. Conclusions: Supported by machine learning results, PSC and UC appear to be separate entities on a molecular level, while PSC/UC and PSC are indistinguishable. Impact and implications: Clinical and genetic studies suggest that the colitis-like symptoms in primary sclerosing cholangitis (PSC) represent a different disease entity from primary ulcerative colitis (UC). The present study supports this assumption with transcriptomic data from whole blood and describes notable differences in gene expression between primary UC and PSC, providing insights into the still unclear pathophysiology of both diseases. These findings are of interest to scientists seeking to decipher the molecular pathophysiology of both diseases and provide evidence that a redefinition of the PSC-UC phenotype should be considered. The study practically supports future molecular research by providing a large transcriptomic whole blood reference cohort.

Osteopontin characterizes bile duct-associated macrophages and correlates with liver fibrosis severity in primary sclerosing cholangitis.

BACKGROUND AND AIMS: Primary sclerosing cholangitis (PSC) is an immune-mediated cholestatic liver disease for which pharmacological treatment options are currently unavailable. PSC is strongly associated with colitis and a disruption of the gut-liver axis, and macrophages are involved in the pathogenesis of PSC. However, how gut-liver interactions and specific macrophage populations contribute to PSC is incompletely understood. APPROACH AND RESULTS: We investigated the impact of cholestasis and colitis on the hepatic and colonic microenvironment, and performed an in-depth characterization of hepatic macrophage dynamics and function in models of concomitant cholangitis and colitis. Cholestasis-induced fibrosis was characterized by depletion of resident KCs, and enrichment of monocytes and monocyte-derived macrophages (MoMFs) in the liver. These MoMFs highly express triggering-receptor-expressed-on-myeloid-cells-2 ( Trem2 ) and osteopontin ( Spp1 ), markers assigned to hepatic bile duct-associated macrophages, and were enriched around the portal triad, which was confirmed in human PSC. Colitis induced monocyte/macrophage infiltration in the gut and liver, and enhanced cholestasis-induced MoMF- Trem2 and Spp1 upregulation, yet did not exacerbate liver fibrosis. Bone marrow chimeras showed that knockout of Spp1 in infiltrated MoMFs exacerbates inflammation in vivo and in vitro , while monoclonal antibody-mediated neutralization of SPP1 conferred protection in experimental PSC. In human PSC patients, serum osteopontin levels are elevated compared to control, and significantly increased in advanced stage PSC and might serve as a prognostic biomarker for liver transplant-free survival. CONCLUSIONS: Our data shed light on gut-liver axis perturbations and macrophage dynamics and function in PSC and highlight SPP1/OPN as a prognostic marker and future therapeutic target in PSC.

Primær skleroserende kolangitt.

Primary sclerosing cholangitis is a severe liver disease and a leading cause of liver transplantation in Scandinavia. This clinical review article examines recently revised recommendations on diagnosis, follow-up and treatment of patients with this disease. Treatment of symptoms, assessment of fibrosis and monitoring for the development of cancer in the liver and bowel are central.

A critical spotlight on the paradigms of FFPE-DNA sequencing.

In the late 19th century, formalin fixation with paraffin-embedding (FFPE) of tissues was developed as a fixation and conservation method and is still used to this day in routine clinical and pathological practice. The implementation of state-of-the-art nucleic acid sequencing technologies has sparked much interest for using historical FFPE samples stored in biobanks as they hold promise in extracting new information from these valuable samples. However, formalin fixation chemically modifies DNA, which potentially leads to incorrect sequences or misinterpretations in downstream processing and data analysis. Many publications have concentrated on one type of DNA damage, but few have addressed the complete spectrum of FFPE-DNA damage. Here, we review mitigation strategies in (I) pre-analytical sample quality control, (II) DNA repair treatments, (III) analytical sample preparation and (IV) bioinformatic analysis of FFPE-DNA. We then provide recommendations that are tested and illustrated with DNA from 13-year-old liver specimens, one FFPE preserved and one fresh frozen, applying target-enriched sequencing. Thus, we show how DNA damage can be compensated, even when using low quantities (50 ng) of fragmented FFPE-DNA (DNA integrity number 2.0) that cannot be amplified well (Q129 bp/Q41 bp = 5%). Finally, we provide a checklist called 'ERROR-FFPE-DNA' that summarises recommendations for the minimal information in publications required for assessing fitness-for-purpose and inter-study comparison when using FFPE samples.

Clinical and biochemical impact of vitamin B6 deficiency in primary sclerosing cholangitis before and after liver transplantation.

BACKGROUND AND AIMS: We previously demonstrated that people with primary sclerosing cholangitis (PSC) had reduced gut microbial capacity to produce active vitamin B6 (pyridoxal 5'-phosphate [PLP]), which corresponded to lower circulating PLP levels and poor outcomes. Here, we define the extent and biochemical and clinical impact of vitamin B6 deficiency in people with PSC from several centers before and after liver transplantation (LT). METHODS: We used targeted liquid chromatography-tandem mass spectrometry to measure B6 vitamers and B6-related metabolic changes in blood from geographically distinct cross-sectional cohorts totaling 373 people with PSC and 100 healthy controls to expand on our earlier findings. Furthermore, we included a longitudinal PSC cohort (n = 158) sampled prior to and serially after LT, and cohorts of people with inflammatory bowel disease (IBD) without PSC (n = 51) or with primary biliary cholangitis (PBC) (n = 100), as disease controls. We used Cox regression to measure the added value of PLP to predict outcomes before and after LT. RESULTS: In different cohorts, 17-38% of people with PSC had PLP levels below the biochemical definition of a vitamin B6 deficiency. The deficiency was more pronounced in PSC than in IBD without PSC and PBC. Reduced PLP was associated with dysregulation of PLP-dependent pathways. The low B6 status largely persisted after LT. Low PLP independently predicted reduced LT-free survival in both non-transplanted people with PSC and in transplant recipients with recurrent disease. CONCLUSIONS: Low vitamin B6 status with associated metabolic dysregulation is a persistent feature of PSC. PLP was a strong prognostic biomarker for LT-free survival both in PSC and recurrent disease. Our findings suggest that vitamin B6 deficiency modifies the disease and provides a rationale for assessing B6 status and testing supplementation. IMPACT AND IMPLICATIONS: We previously found that people with PSC had reduced gut microbial potential to produce essential nutrients. Across several cohorts, we find that the majority of people with PSC are either vitamin B6 deficient or have a marginal deficiency, which remains prevalent even after liver transplantation. Low vitamin B6 levels strongly associate with reduced liver transplantation-free survival as well as deficits in biochemical pathways dependent on vitamin B6, suggesting that the deficiency has a clinical impact on the disease. The results provide a rationale for measuring vitamin B6 and to investigate whether vitamin B6 supplementation or modification of the gut microbial community can help improve outcomes for people with PSC.

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.

Multitrait genome-wide analyses identify new susceptibility loci and candidate drugs to primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is a rare autoimmune bile duct disease that is strongly associated with immune-mediated disorders. In this study, we implemented multitrait joint analyses to genome-wide association summary statistics of PSC and numerous clinical and epidemiological traits to estimate the genetic contribution of each trait and genetic correlations between traits and to identify new lead PSC risk-associated loci. We identified seven new loci that have not been previously reported and one new independent lead variant in the previously reported locus. Functional annotation and fine-mapping nominated several potential susceptibility genes such as MANBA and IRF5. Network-based in silico drug efficacy screening provided candidate agents for further study of pharmacological effect in PSC.

Environmental chemicals and endogenous metabolites in bile of USA and Norway patients with primary sclerosing cholangitis.

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 < 0.20) and 3143 metabolic features (FDR < 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 < .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.

Algebraic topology-based machine learning using MRI predicts outcomes in primary sclerosing cholangitis.

BACKGROUND: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease that can lead to cirrhosis and hepatic decompensation. However, predicting future outcomes in patients with PSC is challenging. Our aim was to extract magnetic resonance imaging (MRI) features that predict the development of hepatic decompensation by applying algebraic topology-based machine learning (ML). METHODS: We conducted a retrospective multicenter study among adults with large duct PSC who underwent MRI. A topological data analysis-inspired nonlinear framework was used to predict the risk of hepatic decompensation, which was motivated by algebraic topology theory-based ML. The topological representations (persistence images) were employed as input for classification to predict who developed early hepatic decompensation within one year after their baseline MRI. RESULTS: We reviewed 590 patients; 298 were excluded due to poor image quality or inadequate liver coverage, leaving 292 potentially eligible subjects, of which 169 subjects were included in the study. We trained our model using contrast-enhanced delayed phase T1-weighted images on a single center derivation cohort consisting of 54 patients (hepatic decompensation, n = 21; no hepatic decompensation, n = 33) and a multicenter independent validation cohort of 115 individuals (hepatic decompensation, n = 31; no hepatic decompensation, n = 84). When our model was applied in the independent validation cohort, it remained predictive of early hepatic decompensation (area under the receiver operating characteristic curve = 0.84). CONCLUSIONS: Algebraic topology-based ML is a methodological approach that can predict outcomes in patients with PSC and has the potential for application in other chronic liver diseases.

Survey uncovering variations in the management of primary sclerosing cholangitis across Europe.

Background & Aims: Data on the management of primary sclerosing cholangitis (PSC) in European expert centres are sparse. In this study, a PSC group from the ERN RARE-LIVER surveyed European hepatologists to uncover differences in real-life clinical practices. Methods: In April 2020 a survey questionnaire was sent to members of the International PSC Study Group and ERN RARE-LIVER. Participants were asked about the size of their PSC cohort, use of medical treatments including ursodeoxycholic acid (UDCA) and surveillance for cholangiocarcinoma, gallbladder polyps and inflammatory bowel disease (IBD). Data were presented descriptively. Results: Eighty-two of 278 members responded. Fifty percent of physicians prescribed UDCA routinely to all their patients with PSC, whereas 12% never prescribed UDCA. UDCA was used for one or more indications including: alkaline phosphatase >1.5x the upper limit of normal, severe PSC changes, pruritus, PSC-IBD or patient demand. Few physicians offered other medical treatments than UDCA. The use of medical treatments was generally comparable in small (<99 patients) and large (≥99 patients) cohorts, as well as for adult and paediatric physicians. Most physicians routinely screened for cholangiocarcinoma and the most frequent modalities used were MRI and ultrasound. At detection of a gallbladder polyp of 6 mm, 46% of physicians recommended repeated ultrasound after 3-6 months, whereas 44% of physicians recommended immediate cholecystectomy. In patients with PSC without IBD at PSC diagnosis, 68% of physicians repeated colonoscopy within 3-5 years whereas 27% referred only patients who developed symptoms of IBD. Conclusion: Substantial variations in treatment and monitoring of European patients with PSC were discovered. Harmonisation of strategies is desirable to enable improved interpretation of outcome data and to optimise clinical patient care. Lay summary: In this study, we explored how different centres in Europe manage primary sclerosing cholangitis (PSC), a rare inflammatory disease of the bile ducts. We collected information through a questionnaire sent to specialist physicians who were part of a European network for rare liver diseases. We found several differences in how patients with PSC were monitored and treated. This includes differences in surveillance for bile duct cancer, gallbladder polyps and inflammatory bowel disease. By pointing out these differences, we hope that management of PSC will be standardized, which could aid clinical research and benefit patients.

Impact of Positive Lymph Nodes and Resection Margin Status on the Overall Survival of Patients with Resected Perihilar Cholangiocarcinoma: The ENSCCA Registry.

Background: Lymph node metastasis and positive resection margins have been reported to be major determinants of overall survival (OS) and poor recurrence-free survival (RFS) for patients who underwent resection for perihilar cholangiocarcinoma (pCCA). However, the prognostic value of positive lymph nodes independently from resection margin status on OS has not been evaluated. Methods: From the European Cholangiocarcinoma (ENSCCA) registry, patients who underwent resection for pCCA between 1994 and 2021 were included in this retrospective cohort study. The primary outcome was OS stratified for resection margin and lymph node status. The secondary outcome was recurrence-free survival. Results: A total of 325 patients from 11 different centers and six European countries were included. Of these, 194 (59.7%) patients had negative resection margins. In 113 (34.8%) patients, positive lymph nodes were found. Lymph node status, histological grade, and ECOG performance status were independent prognostic factors for survival. The median OS for N0R0, N0R1, N+R0, and N+R1 was 38, 30, 18, and 12 months, respectively (p < 0.001). Conclusion: These data indicate that in the presence of positive regional lymph nodes, resection margin status does not determine OS or RFS in patients with pCCA. Achieving negative margins in patients with positive nodes should not come at the expense of more extensive surgery and associated higher mortality.

Portal fibroblasts with mesenchymal stem cell features form a reservoir of proliferative myofibroblasts in liver fibrosis.

BACKGROUND AND AIMS: In liver fibrosis, myofibroblasts derive from HSCs and as yet undefined mesenchymal cells. We aimed to identify portal mesenchymal progenitors of myofibroblasts. APPROACH AND RESULTS: Portal mesenchymal cells were isolated from mouse bilio-vascular tree and analyzed by single-cell RNA-sequencing. Thereby, we uncovered the landscape of portal mesenchymal cells in homeostatic mouse liver. Trajectory analysis enabled inferring a small cell population further defined by surface markers used to isolate it. This population consisted of portal fibroblasts with mesenchymal stem cell features (PMSCs), i.e., high clonogenicity and trilineage differentiation potential, that generated proliferative myofibroblasts, contrasting with nonproliferative HSC-derived myofibroblasts (-MF). Using bulk RNA-sequencing, we built oligogene signatures of the two cell populations that remained discriminant across myofibroblastic differentiation. SLIT2, a prototypical gene of PMSC/PMSC-MF signature, mediated profibrotic and angiogenic effects of these cells, which conditioned medium promoted HSC survival and endothelial cell tubulogenesis. Using PMSC/PMSC-MF 7-gene signature and slit guidance ligand 2 fluorescent in situ hybridization, we showed that PMSCs display a perivascular portal distribution in homeostatic liver and largely expand with fibrosis progression, contributing to the myofibroblast populations that form fibrotic septa, preferentially along neovessels, in murine and human liver disorders, irrespective of etiology. We also unraveled a 6-gene expression signature of HSCs/HSC-MFs that did not vary in these disorders, consistent with their low proliferation rate. CONCLUSIONS: PMSCs form a small reservoir of expansive myofibroblasts, which, in interaction with neovessels and HSC-MFs that mainly arise through differentiation from a preexisting pool, underlie the formation of fibrotic septa in all types of liver diseases.

Cholangiocarcinoma landscape in Europe: Diagnostic, prognostic and therapeutic insights from the ENSCCA Registry.

BACKGROUND & AIMS: Cholangiocarcinoma (CCA) is a rare and heterogeneous biliary cancer, whose incidence and related mortality is increasing. This study investigates the clinical course of CCA and subtypes (intrahepatic [iCCA], perihilar [pCCA], and distal [dCCA]) in a pan-European cohort. METHODS: The ENSCCA Registry is a multicenter observational study. Patients were included if they had a histologically proven diagnosis of CCA between 2010-2019. Demographic, histomorphological, biochemical, and clinical studies were performed. RESULTS: Overall, 2,234 patients were enrolled (male/female=1.29). iCCA (n = 1,243) was associated with overweight/obesity and chronic liver diseases involving cirrhosis and/or viral hepatitis; pCCA (n = 592) with primary sclerosing cholangitis; and dCCA (n = 399) with choledocholithiasis. At diagnosis, 42.2% of patients had local disease, 29.4% locally advanced disease (LAD), and 28.4% metastatic disease (MD). Serum CEA and CA19-9 showed low diagnostic sensitivity, but their concomitant elevation was associated with increased risk of presenting with LAD (odds ratio 2.16; 95% CI 1.43-3.27) or MD (odds ratio 5.88; 95% CI 3.69-9.25). Patients undergoing resection (50.3%) had the best outcomes, particularly with negative-resection margin (R0) (median overall survival [mOS] = 45.1 months); however, margin involvement (R1) (hazard ratio 1.92; 95% CI 1.53-2.41; mOS = 24.7 months) and lymph node invasion (hazard ratio 2.13; 95% CI 1.55-2.94; mOS = 23.3 months) compromised prognosis. Among patients with unresectable disease (49.6%), the mOS was 10.6 months for those receiving active palliative therapies, mostly chemotherapy (26.2%), and 4.0 months for those receiving best supportive care (20.6%). iCCAs were associated with worse outcomes than p/dCCAs. ECOG performance status, MD and CA19-9 were independent prognostic factors. CONCLUSION: CCA is frequently diagnosed at an advanced stage, a proportion of patients fail to receive cancer-specific therapies, and prognosis remains dismal. Identification of preventable risk factors and implementation of surveillance in high-risk populations are required to decrease cancer-related mortality. LAY SUMMARY: This is, to date, the largest international (pan-European: 26 hospitals and 11 countries) observational study, in which the course of cholangiocarcinoma has been investigated, comparing the 3 subtypes based on the latest International Classification of Diseases 11th Edition (ICD-11) (i.e., intrahepatic [2C12], perihilar [2C18], or distal [2C15] affected bile ducts), which come into effect in 2022. General and tumor-type specific features at diagnosis, risk factors, biomarker accuracy, as well as patient management and outcomes, are presented and compared, outlining the current clinical state of cholangiocarcinoma in Europe.

Risk of hepato-pancreato-biliary cancer is increased by primary sclerosing cholangitis in patients with inflammatory bowel disease: A population-based cohort study.

BACKGROUND: There is continued uncertainty regarding the risks of hepato-pancreato-biliary cancers in patients with inflammatory bowel disease (IBD) with or without concomitant primary sclerosing cholangitis (PSC). OBJECTIVE: To give updated estimates on risk of hepato-pancreato-biliary cancers in patients with IBD, including pancreatic cancer, hepatocellular carcinoma, gall bladder cancer, and intra - and extrahepatic cholangiocarcinoma. METHODS: In a population-based cohort study, we included all patients diagnosed with IBD in Norway and Sweden from 1987 to 2016. The cohort comprised of 141,960 patients, identified through hospital databases and the National Patient Register. Participants were followed through linkage to national cancer, cause of death, and population registries. We calculated absolute risk and standardized incidence ratios (SIRs) of hepato-pancreato-biliary cancers by PSC and other clinical characteristics. RESULTS: Of the 141,960 IBD patients, 3.2% were diagnosed with PSC. During a median follow-up of 10.0 years, we identified 443 biliary tract cancers (SIR 5.2, 95% confidence interval [CI] 4.8-5.7), 161 hepatocellular carcinomas (SIR 2.4, 95% CI 2.0-2.7) and 282 pancreatic cancers (SIR 1.3, 95% CI 1.2-1.5). The relative risks were considerably higher in PSC-IBD patients, with SIR of 140 (95% CI 123-159) for biliary tract, 38.6 (95% CI 29.2-50.0) for hepatocellular, and 9.0 (95% CI 6.3-12.6) for pancreatic cancer. The SIRs were still slightly increased in non-PSC-IBD patients, compared to the general population. For biliary tract cancer, the cumulative probability at 25 years was 15.6% in PSC-IBD patients, and 0.4% in non-PSC-IBD patients. CONCLUSIONS: The dramatically increased risks of hepato-pancreato-biliary cancers in PSC-IBD patients support periodic surveillance for these malignancies. While much lower, the excess relative risks in non-PSC-IBD patients were not trivial compared to non-IBD related risk factors.

Cross-tissue transcriptome-wide association studies identify susceptibility genes shared between schizophrenia and inflammatory bowel disease.

Genetic correlations and an increased incidence of psychiatric disorders in inflammatory-bowel disease have been reported, but shared molecular mechanisms are unknown. We performed cross-tissue and multiple-gene conditioned transcriptome-wide association studies for 23 tissues of the gut-brain-axis using genome-wide association studies data sets (total 180,592 patients) for Crohn's disease, ulcerative colitis, primary sclerosing cholangitis, schizophrenia, bipolar disorder, major depressive disorder and attention-deficit/hyperactivity disorder. We identified NR5A2, SATB2, and PPP3CA (encoding a target for calcineurin inhibitors in refractory ulcerative colitis) as shared susceptibility genes with transcriptome-wide significance both for Crohn's disease, ulcerative colitis and schizophrenia, largely explaining fine-mapped association signals at nearby genome-wide association study susceptibility loci. Analysis of bulk and single-cell RNA-sequencing data showed that PPP3CA expression was strongest in neurons and in enteroendocrine and Paneth-like cells of the ileum, colon, and rectum, indicating a possible link to the gut-brain-axis. PPP3CA together with three further suggestive loci can be linked to calcineurin-related signaling pathways such as NFAT activation or Wnt.