A Teufel

Genome-wide association study of primary sclerosing cholangitis identifies new risk loci and quantifies the genetic relationship with inflammatory bowel disease

Primary sclerosing cholangitis (PSC) is a rare progressive disorder leading to bile duct destruction; ∼75% of patients have comorbid inflammatory bowel disease (IBD). We undertook the largest genome-wide association study of PSC (4,796 cases and 19,955 population controls) and identified four new genome-wide significant loci. The most associated SNP at one locus affects splicing and expression of UBASH3A, with the protective allele (C) predicted to cause nonstop-mediated mRNA decay and lower expression of UBASH3A. Further analyses based on common variants suggested that the genome-wide genetic correlation (rG) between PSC and ulcerative colitis (UC) (rG = 0.29) was significantly greater than that between PSC and Crohns disease (CD) (rG = 0.04) (P = 2.55 × 10−15). UC and CD were genetically more similar to each other (rG = 0.56) than either was to PSC (P < 1.0 × 10−15). Our study represents a substantial advance in understanding of the genetics of PSC.

Smad7 regulates compensatory hepatocyte proliferation in damaged mouse liver and positively relates to better clinical outcome in human hepatocellular carcinoma.

Transforming growth factor β (TGF-β) is cytostatic towards damage-induced compensatory hepatocyte proliferation. This function is frequently lost during hepatocarcinogenesis, thereby switching the TGF-β role from tumour suppressor to tumour promoter. In the present study, we investigate Smad7 overexpression as a pathophysiological mechanism for cytostatic TGF-β inhibition in liver damage and hepatocellular carcinoma (HCC). Transgenic hepatocyte-specific Smad7 overexpression in damaged liver of fumarylacetoacetate hydrolase (FAH)-deficient mice increased compensatory proliferation of hepatocytes. Similarly, modulation of Smad7 expression changed the sensitivity of Huh7, FLC-4, HLE and HLF HCC cell lines for cytostatic TGF-β effects. In our cohort of 140 HCC patients, Smad7 transcripts were elevated in 41.4% of HCC samples as compared with adjacent tissue, with significant positive correlation to tumour size, whereas low Smad7 expression levels were significantly associated with worse clinical outcome. Univariate and multivariate analyses indicate Smad7 levels as an independent predictor for overall (P<0.001) and disease-free survival (P=0.0123). Delineating a mechanism for Smad7 transcriptional regulation in HCC, we identified cold-shock Y-box protein-1 (YB-1), a multifunctional transcription factor. YB-1 RNAi reduced TGF-β-induced and endogenous Smad7 expression in Huh7 and FLC-4 cells respectively. YB-1 and Smad7 mRNA expression levels correlated positively (P<0.0001). Furthermore, nuclear co-localization of Smad7 and YB-1 proteins was present in cancer cells of those patients. In summary, the present study provides a YB-1/Smad7-mediated mechanism that interferes with anti-proliferative/tumour-suppressive TGF-β actions in a subgroup of HCC cells that may facilitate aspects of tumour progression.

Benefit of adjuvant chemotherapy in patients with T4 UICC II colon cancer.

BackgroundColorectal cancer is the third most common cancer and a major cause of morbidity and mortality worldwide. Adjuvant chemotherapy is considered the standard of care in patients with UICC stage III colon cancer after R0 resection. Adjuvant therapy was not shown to be beneficial in patients with UICC stage II colon cancer. However, there is an ongoing discussion as to whether adjuvant chemotherapy may be beneficial for a subgroup of UICC II patients in a “high-risk situation” (such as T4).MethodsWe investigated a Bavarian population-based (2.1 million inhabitants) cohort of 1937 patients with UICC II CRC treated between 2002 and 2012 in regard of the benefit of adjuvant chemotherapy for large (T4) tumors. Patients older than 80xa0years of age were excluded. Of 1937 patients, 240 had a T4 tumor (12xa0%); 77 of all T4 patients received postoperative chemotherapy (33xa0%). Kaplan-Meier analysis and Cox regression models were used for survival analyses.ResultsPatients with a T4 tumor who received postoperative chemotherapy had a highly significant survival benefit in respect of overall survival (pu2009<u20090.001) and recurrence-free survival (pu2009=u20090.008). However, no difference was observed between oxaliplatin-containing and non-oxaliplatin-containing treatment regimens. G2 and G3 tumors were found to particularly benefit from adjuvant treatment. Chemotherapy, age at diagnosis, and tumor grading remained independent risk factors in the multivariate cox regression analysis.ConclusionOur retrospective study demonstrated the significant benefit of adjuvant chemotherapy in the T4 subgroup of patients with UICC II colon cancer. Our data suggest that adjuvant chemotherapy should be seriously considered in these patients.

CellMinerHCC: A microarray-based expression database for hepatocellular carcinoma cell lines

Therapeutic options for hepatocellular carcinoma (HCC) still remain limited. Development of gene targeted therapies is a promising option. A better understanding of the underlying molecular biology is gained in in vitro experiments. However, even with targeted manipulation of gene expression varying treatment responses were observed in diverse HCC cell lines. Therefore, information on gene expression profiles of various HCC cell lines may be crucial to experimental designs. To generate a publicly available database containing microarray expression profiles of diverse HCC cell lines.

Increased liver carcinogenesis and enrichment of stem cell properties in livers of Dickkopf 2 (Dkk2) deleted mice

Dkk2 a antagonist of the Wnt/β-catenin-signaling pathway was shown to be silenced in diverse cancers. More recent data indicate that Dkk family members may also possess functions independent of Wnt-signaling during carcinogenesis. The detailed biological function of Dkks and its relevance for liver cancer is unknown. We analyzed the effects of a genetic deletion of Dkk2 (Dkk2−/−) in a hepatocarcinogenesis model using DEN/Phenobarbital. Untreated Dkk2−/− animals, showed considerable atypia with variation of hepatocyte size and chromatin density. In livers of Dkk2−/− mice nodule formation was seen at 9 months of age with focal loss of trabecular architecture and atypical hepatocytes and after DEN induction Dkk2−/− mice developed significantly more liver tumors compared to controls. Whole transcriptome analysis of untreated Dkk2−/− liver tissue revealed a Dkk2-dependent genetic network involving Wnt/β-Catenin but also multiple additional oncogenic factors, such as e.g. Pdgf-b, Gdf-15 and Hnf4a. Dkk2−/− tumor cells showed a significant deregulation of stemness genes associated with enhanced colony forming properties. Integration of the Dkk2−/− signature into human data was strongly associated with patients survival. Dkk2 deletion results in alterations of liver morphology leading to an increased frequency of liver cancer. The associated genetic changes included factors not primarily related to Wnt/β-Catenin-signaling and correlated with the clinical outcome of HCC-patients.

Personalisierte Hepatologie − Gegenwärtige Konzepte, Entwicklungen und Erwartungen im Postgenomzeitalter

Promoted by the decoding of the human genome as part of the human genome project, individualised therapy approaches have become a realistic perspective for therapies that are more effective, less prone to side effects and economically reasonable. This also applies to chronic liver disease. With the aim not only to expand the current knowledge base through basic research on the underlying disease processes and treatment options but also to identify and characterise biomarkers, the creation of genetic fingerprints for individualised diagnosis, prognosis and treatment of patients takes its place in the centre of translational hepatology. For certain liver diseases personalised therapy approaches are already existent. Examples are the determination of viral genotypes, viral kinetics and genotyping of the IL28B polymorphism to optimise the treatment of chronic hepatitis C. The challenges of the next few years relate to the broadening of the knowledge base, the establishment of reliable and standardised technologies, and the development of intelligent bioinformatics strategies for data analysis and data integration. The following review not only summarises the current state of progress and possibilities of personalised medicine in hepatological diseases, but also explains the technical background of the limitations that currently hinder a consistent clinical implementation.

Genotype-phenotype analysis across 130 422 genetic variants identifies RSPO3 as the first genome-wide significant modifier gene in primary sclerosing cholangitis

Background n nUlcerative colitis (UC), a complex polygenic disorder, is one of the main subphenotypes of inflammatory bowel disease. A comprehensive dissection of the genetic etiology of UC needs to assess the contribution of rare genetic variants including copy number variations (CNVs) to disease risk. In this study, we performed a multi-step genome-wide case-control analysis to interrogate the presence of disease-relevant rare copy number variants. nMethods n nOne thousand one hundred twenty-one German UC patients and 1770 healthy controls were initially screened for rare deletions and duplications employing SNP-array data. Quantitative PCR and high density custom array-CGH were used for validation of identified CNVs and fine mapping. Two main follow-up panels consisted of an independent cohort of 451 cases and 1274 controls, in which CNVs were assayed through quantitative PCR, and a British cohort of 2396 cases versus 4886 controls with CNV genotypes based on array data. Additional sample sets were assessed for targeted and in silico replication. nResults n nTwenty-four rare copy number variants (14 deletions and 10 duplications), overrepresented in UC patients were identified in the initial screening panel. Follow-up of these CNV regions in four independent case-control series as well as an additional public in silico control group (totaling 4439 UC patients and 15,961 healthy controls) revealed three copy number variants enriched in UC patients; a 15.8 kb deletion upstream of ABCC4 and CLDN10 at13q32.1 (0.43 % cases, 0.11 % controls), a 119 kb duplication at 7p22.1, overlapping RNF216, ZNF815, OCM and CCZ1 (0.13 % cases, 0.01 % controls) and a 134 kb large duplication upstream of the KCNK9 gene at 8q24.3 (0.22 % carriers among cases, 0.03 % carriers among controls). The trend of association with UC was present after the P-values were corrected for combining data from different subpopulations. Break-point mapping of the deleted region suggested non-allelic homologous recombination as the mechanism underlying its formation. nConclusion n nOur study presents a pragmatic approach for effective rare CNV screening of SNP-array data sets and implicates the potential contribution of rare structural variants in the pathogenesis of UC.

HCC-Cluster-Analyse anhand genetischer Signalwege

In den letzten Jahren wurden viele Gene entdeckt, die in der Hepatokarzinogenese beteiligt sind. Im Vergleich zur Expression tausender Gene lasst sich das Risiko einer neoplastischen Erkrankung durch eine deutlich geringere Anzahl aktivierter, genetisch definierter Signalwege beschreiben. In microarray-basierten HCC-Subgruppen konnten wiederholt einzelne Gene oder kleinere Cluster von Genen identifiziert werden, welche mit einem unterschiedlichen Krankheitsverlauf assoziiert waren. Weiterhin sind mehrere genetische Signalwege bei der Entwicklung von HCCs von kritischer Bedeutung. Bisher ist wenig bekannt uber die Aktivierung der Signalwege in diesen genetisch unterschiedlichen HCC Subgruppen. Wir haben die Microarray basierten Genexpressionsdaten von 141 HCCs auf differentiell aktivierte Signalwege untersucht. In einem Cluster A, fur das zuvor ein signifikant kurzeres Uberleben beschrieben worden war, zeigten sich unter den charakteristischen Signalwegen zahlreiche mit der Karzinogenese assoziierte Signalwege, z.B. Wnt-Signalweg, „MAPK“, „TGFbeta“ und „mTOR“, aber auch die Cluster „Extracellular Region“ und „Focal Adhesion“. Am starksten uberreprasentiert waren „Ribosomal Proteins“, was auf eine essentielle Rolle dieses Clusters in der Hepatokarzinogenese hinweisen konnte. Im Gegensatz hierzu waren diese Signalwege im Cluster B, welches mit einem besseren Uberleben assoziiert war, nicht uberreprasentiert. Allerdings zeigte sich eine Anreicherung von „Complement and Coagulation“ und „PPAR“. Somit fand sich eine klare Unterscheidung von mindestens zwei HCC-Subtypen auf dem Boden einer differentiellen Aktivierung von genetischen Signalwegen. Im weiteren zeigte eine Clusterung mittels der Signalwege „Ribosomal Proteins“ und „Focal Adhesion“ sowie „Complement and Coagulation“ und „PPAR“ eine weitestgehend identische Auftrennung der Subgruppen, womit diesen Signalwegen eine Schlusselrolle in der Karzinogenese der genetisch differenten HCC-Subtypen zukommen konnte.

Identifizierung genetischer Netzwerke bei der PDGF-B vermittelten Leberfibrose

Fibrotische Veranderungen der Leber sind eine wichtige Vorstufe zu weit schwereren Lebererkrankungen wie Leberzirrhose oder Hepatozellularem Karzinom. Daher ist es wichtig, die zu Grunde liegenen genetischen Signalnetzwerke von profibrogenetischen Faktoren zu verstehen. Wir konnten vor kurzem die essentielle Rolle von PDGF-B im Rahmen der Leberfibrogenese im transgenen Mausmodell zeigen. Um die genetischen Zusammenhange bei der PDGF-B vermittelten Leberfibrogenese genauer zu untersuchen, wurden sowohl PDGF-B transgene Mause als auch Wildtypen mittels Microarrays analysiert. Verwedet wurden dazu Illumina MouseRef-8 BeadChips, welche das gesamte Mausgenom abdecken. Dabei zeigte sich, das die Uberexpression von PDGF-B die Expression von Genen beeinflusst, welche fur den Aufbau der extrazellularen Matrix, matrixmodellierenden Proteinen und deren Regulatoren, intrazellulare Komponenten des TGF-b Signalweges und dessen Zielgene von Bedeutung sind. Desweiteren werden durch PDGF-B auch Gene beeinflusst, die eine wichtige Rolle bei der Zellorganisation, Zelldifferenzierung und der Angiogenese spielen. Weiter Analysen mittels RealTime-PCR bestatigeten den signifikanten Einfluss von PDGF-B auf den TGF-b Signalweg und zeigte eine signifikant erhohte Expression von VEGF und b-catenin. Letzeres verknupft somit PDGF-B auch mit dem Wnt/b-catenin Signalweg. Histologische Untersuchungen der Leber zeigten desweiteren, dass in PDGF-B transgenen Tieren nicht nur die Expression von b-catenin erhoht ist, sondern dass es auch zu einer massiven Dislokalisation von b-catenin von der extrazellularen Matrix in das Zytoplasma und den Nucleus kommt. In Zusammenhang mit der erhohten Expression von VEGF und des VEGF-R1 weisen PDGF-B transgene Tiere auch vermehrt CD-31 positive Regionen auf.