Immune checkpoints and liver resection after neoadjuvant chemotherapy including bevacizumab in patients with microsatellite-stable colorectal liver metastases.

BACKGROUND: The clinical value of immune checkpoint expression as prognostic biomarker in bevacizumab-pretreated patients with resected microsatellite-stable (MMS) colorectal liver metastases is unclear and was retrospectively investigated in this study. METHODS: Expression analyses of IDO-1, PD-L1, and CTLA-4 were performed by immunohistochemistry in resected bevacizumab-pretreated colorectal liver metastases. Association of immune checkpoint expression in tumor cells and immune cells with response and clinical outcome was investigated. Expression profiles were compared with those of patients with anti-EGFR-targeted therapy and lung metastases, respectively. RESULTS: One hundred thirty-six patients with MMS disease were investigated (79 (58.1%) male/57 (41.9%) female, median age 62.9 years (range 31.0-80.4)). High expression of IDO-1 in immune cells was associated with longer OS (not reached versus 44.8 months, HR 0.23 (95% CI 0.09, 0.55), P = 0.001). Low expression of CTLA-4 in tumor cells was associated with better histological response (26 major, 19 partial, 18 none versus 14 major, 23 partial, 30 none, P = 0.032). Expression profiles differed compared to patients with anti-EGFR-targeted therapy and patients with lung metastases. CONCLUSION: Immune checkpoint expression was associated with response and survival. IDO-1 may serve as a novel prognostic and/or predictive biomarker in patients with MMS colorectal liver metastases.

Immune phenotype and histopathological growth pattern in patients with colorectal liver metastases.

BACKGROUND: Patients with desmoplastic (angiogenic) histopathological growth pattern (HGP) colorectal liver metastases (CLM) might derive more benefit from bevacizumab-based chemotherapy than those with replacement (non-angiogenic) HGP. This study investigated the association of HGP with the immune phenotype (IP) and clinical outcome after liver resection. METHODS: CLM of patients treated with perioperative bevacizumab-based chemotherapy and liver resection were investigated. Association of HGP and IP with response, recurrence-free survival (RFS) and overall survival (OS) was investigated. RESULTS: One hundred and eighteen patients (M/F 66/52, median age 62.3 (31.0-80.4) years, median follow-up 32.2 (5.0-92.7) months) were enrolled. The inflamed IP was associated with the desmoplastic HGP. The desmoplastic HGP was associated with better radiological and histological response compared to the replacement HGP, respectively. The replacement HGP was associated with shorter RFS (8.7 versus 16.3 months, HR 2.60, P = 0.001) and OS (36.6 months versus not reached, HR 2.32, P = 0.027), respectively. The non-inflamed IP was associated with shorter RFS (10.8 versus 16.5 months, HR 1.85, P = 0.029). The HGP but not the IP remained significant in multivariable analysis for RFS. CONCLUSIONS: The desmoplastic HGP is associated with the inflamed IP and HGP may be a potential biomarker for adjuvant treatment that includes targeting the immune contexture.

The diagnostic efficacy of quantitative liver MR imaging with diffusion-weighted, SWI, and hepato-specific contrast-enhanced sequences in staging liver fibrosis--a multiparametric approach.

PURPOSE: To assess the diagnostic efficacy of multiparametric MRI using quantitative measurements of the apparent diffusion coefficient (ADC) of the liver parenchyma on diffusion-weighted imaging (DWI), signal intensity (SI) on susceptibility-weighted imaging (SWI), and gadoxetic acid-enhanced T1-weighted imaging during the hepatobiliary phase for the staging of liver fibrosis. MATERIALS AND METHODS: Seventy-seven patients underwent a 3T MRI examination, including DWI/SWI sequences and gadoxetic acid-enhanced T1-weighted MRI. Liver fibrosis according to liver biopsy was staged using the Metavir fibrosis score: F0 (n = 21, 27.3%); F1 (n = 7, 9.1%); F2 (n = 8, 10.4%); F3 (n = 12, 15.6%); and F4 (n = 29, 37.7%). SI of the liver was defined using region-of-interest measurements to calculate the ADC values, the relative enhancement (RE) in the hepatobiliary phase, and the liver-to-muscle ratio (LMR) measurements for SWI. RESULTS: The values of RE, LMR, and ADC measurements were statistically significantly different among the five fibrosis stages (p < 0.004). Combining the three parameters in a multiparametric approach, the AUC for detecting F1 stage or greater (≥ F1) was 94%, for F2 or greater (≥F2) was 95%, for F3 or greater (≥F3) was 90%, and for stage F4 was 93%. CONCLUSIONS: Multiparametric MRI is an efficient non-invasive diagnostic tool for the staging of liver fibrosis. KEY POINTS: • Multiparametric MRI has high accuracy in predicting moderate or greater liver fibrosis. • Relative enhancement post- gadoxetic acid is an independent predictor of liver fibrosis. • Liver SWI signal intensity and ADC values enhance the diagnostic ability.

Genetic variations in angiopoietin and pericyte pathways and clinical outcome in patients with resected colorectal liver metastases.

BACKGROUND: Genes involved in the angiopoietin and pericyte pathways may become escape mechanisms under antivascular endothelial growth factor (anti-VEGF) therapy. The authors investigated whether variations within genes in these pathways are associated with clinical outcome in patients with colorectal liver metastases who undergo liver resection and receive perioperative, bevacizumab-based chemotherapy. METHODS: Single nucleotide polymorphisms (SNPs) in 9 genes (angiopoietin-1 [ANGPT1]; ANGPT2; TEK tyrosine kinase, endothelial [TEK]; platelet-derived growth factor ß [PDGFB]; ß-type platelet-derived growth factor receptor [PDGFRB]; insulin-like growth factor 1 [IGF1]; transforming growth factor ß1 [TGFB1]; RalA binding protein 1 [RALBP1]; and regulator of G-protein signaling 5 [RGS5]) were analyzed in samples of genomic DNA from 149 patients and were evaluated for associations with clinical outcome. RESULTS: RALBP1 reference SNP 329007 (rs329007) A>G resulted in a significant difference in recurrence-free survival (A/A genotype, 14.0 months; A/G or G/G genotype, 9.2 months; hazard ratio [HR], 1.60; P = .024). PDGFB rs1800818 A>G was associated with 3-year overall survival rates (A/A genotype, 78%; A/G genotype, 69%; [HR 1.37]; G/G genotype, 53%; [HR 2.12]; P = .048). In multivariate analysis, RALBP1 rs329007 A>G remained significant (HR, 1.99; P = .002). PDGFB rs1800818 A>G and RALBP1 rs329007 A>G were correlated with radiologic response (A/A or A/G genotype, 86%; G/G genotype, 71% [P = .042]; A/A genotype, 78%; A/G or G/G genotype, 94% [P = .018], respectively). RALBP1 rs329007 A>G demonstrated significantly different rates of histologic response (A/A genotype: major histologic response, 35%; partial histologic response, 34%; no histologic response, 30%; A/G or G/G genotype: 46%, 13%, and 41%, respectively; P = .029). Recursive partitioning analysis revealed that ANGPT2 rs2442599 T>C and RALBP1 rs329007 A>G were the main SNPs that predicted histologic response and recurrence-free survival, whereas PDGFB rs1800818 A>G was the leading SNP that predicted overall survival. ANGPT2 rs2916702 C>T and rs2442631 G>A were significantly associated with the probability of achieving a cure. CONCLUSIONS: The current data suggest that variations in genes involved in the angiopoietin and pericyte pathways may be predictive and/or prognostic biomarkers in patients with resected colorectal liver metastases who receive bevacizumab-based chemotherapy.

Hepatic steatosis in Wilson disease--Role of copper and PNPLA3 mutations.

BACKGROUND & AIMS: The earliest characteristic alterations of the liver pathology in Wilson disease (WD) include steatosis, which is sometimes indistinguishable from non-alcoholic fatty liver disease (NAFLD). Steatosis in WD may reflect copper-induced mitochondrial dysfunction. A genetic polymorphism in rs738409, in the patatin-like phospholipase domain-containing 3 gene (PNPLA3), is strongly associated with appearance of in NAFLD. This study evaluated the role of PNPLA3 and hepatic copper content for development of steatosis in patients with WD. METHODS: Liver biopsies obtained at diagnosis and the PNPLA3 genotype were analyzed in 98 Caucasian patients with WD (male: 52 [53.1%]; mean age: 27.6 years [CI 95%: 24.8-30.4, range: 5.8-61.5]). Steatosis was graded as percentage of lipid containing hepatocytes by an expert hepatopathologist unaware of the results of genetic testing. RESULTS: Moderate/severe steatosis (>33% of hepatocytes) was observed in 28 patients (pediatric: n=13/26 [50.0%], adult: n=15/72 [20.8%]; p=0.01). Forty-six patients (46.9%; pediatric: n=7, adult: n=39; p=0.022) had cirrhosis. Multivariate logistic regression identified PNPLA3 G allele (OR: 2.469, CI 95%: 1.203-5.068; p=0.014) and pediatric age (OR: 4.348; 1.577-11.905; p=0.004) as independent variables associated with moderate/severe steatosis. In contrast, hepatic copper content did not impact on moderate/severe steatosis (OR: 1.000, CI 95%: 1.000-1.001; p=0.297). CONCLUSIONS: Steatosis is common in WD and the PNPLA3 G allele contributes to its pathogenesis. The role of hepatic copper concentration and ATP7B mutations in steatosis development deserve further investigations.

Morphologic and Molecular Features of Hepatocellular Adenoma with Gadoxetic Acid-enhanced MR Imaging.

PURPOSE: To evaluate the diagnostic performance of imaging features of gadoxetic acid-enhanced magnetic resonance (MR) imaging to differentiate among hepatocellular adenoma (HCA) subtypes by using the histopathologic results of the new immunophenotype and genotype classification and to correlate the enhancement pattern on the hepatobiliary phase (HBP) with the degrees of expression of organic anion transporting polypeptide (OATP1B1/3), multidrug resistance-associated protein 2 (MRP) (MRP2), and MRP 3 (MRP3) transporters. MATERIALS AND METHODS: This retrospective study was approved by the institutional review board, and the requirement for informed consent waived. MR imaging findings of 29 patients with 43 HCAs were assessed by two radiologists independently then compared with the histopathologic analysis as the standard of reference. Receiver operating characteristic curves and Spearman rank correlation coefficient were used to test the diagnostic performance of gadoxetic acid-enhanced MR imaging features, which included the retention or washout at HBP and degree of transporter expression. Interreader agreement was assessed by using the κ statistic with 95% confidence interval. RESULTS: The area under the curve for the diagnosis of inflammatory HCA was 0.79 (95% confidence interval: 0.64, 0.90); for the steatotic type, it was 0.90 (95% confidence interval: 0.77, 0.97); and for the ß-catenin type, it was 0.87 (95% confidence interval: 0.74, 0.95). There were no imaging features that showed a significant statistical correlation for the diagnosis of unclassified HCAs. On immunohistochemical staining, OATP1B1/3 expression was the main determinant for the retention, whereas MRP3 was the key determinant for washout of gadoxetic acid at HBP (P < .001). MRP2 appeared to have no role. CONCLUSION: Gadoxetic acid-enhanced MR imaging features may suggest the subtype of HCA. The degree of OATP1B1/3 and MRP3 expression correlated statistically with gadoxetic acid retention and washout, respectively, in the HBP.

New reliability criteria for transient elastography increase the number of accurate measurements for screening of cirrhosis and portal hypertension.

BACKGROUND & AIMS: Transient elastography (TE) can non-invasively diagnose cirrhosis and portal hypertension (PHT). New TE reliability criteria suggest classifying measurements as very reliable (IQR/M < 0.1), reliable (IQR<0.3 or >0.3, if TE < 7.1 kPa) and poorly reliable (IQR/M > 0.3, if TE > 7.1 kPa). Compare traditional (reliable: success rate >60% + IQR/M ≤ 0.30) and new TE quality criteria (accurate: very reliable + reliable) regarding their diagnostic accuracy for cirrhosis and PHT and to identify potential confounders (age, aetiology, necroinflammatory activity, steatosis, siderosis, cholestasis, aminotransferases) of TE performance. METHODS: Patients undergoing simultaneous measurements of TE, portal pressure (hepatic venous pressure gradient, HVPG) and liver biopsy were analysed. RESULTS: Among 226 patients (48.7 ± 13.1 years, 74.7% male, 75.7% viral aetiology, 57% F3/F4), traditional TE quality criteria identified 71.6% reliable measurements, while new criteria yielded in 83.2% accurate results. Reliable TE values according to both criteria significantly correlated with fibrosis stage (r = 0.648 vs. r = 0.636) and HVPG (r = 0.836 vs. r = 0.846). Diagnostic accuracy for cirrhosis (cut-off >14.5 kPa) was 76.5% (AUC: 0.863) and 75.0% (AUC: 0.852) for traditional and new TE criteria, respectively, while for predicting HVPG ≥ 10 mmHg (>16.1 kPa), the accuracies were 88.9% (AUC: 0.957) and 89.8% (AUC: 0.962). New TE criteria allowed a better discrimination of reliable and non-reliable results for prediction of fibrosis and CSPH. Only aetiology and aminotransferases were independent confounders of the correlation of TE and fibrosis stage, while no confounder affected the correlation of TE and HVPG. CONCLUSIONS: New reliability criteria for TE measurements increase the number of patients with accurate measurements without affecting diagnostic performance for detecting cirrhosis and portal hypertension. Aetiology of liver disease and aminotransferases should be considered when assessing liver fibrosis by TE.

Early recognition of possible pitfalls in histological diagnosis of celiac disease.

OBJECTIVE: A state-of-the-art assessment of duodenal biopsies by the pathologist in the diagnosis of celiac disease (CelD) is of highest importance. However, inaccurate characterization of specific features can lead to misdiagnosis. Our hypothesis is that a detailed histological report guarantees a good quality and helps in the primary diagnostic process by preventing false-positive diagnosis of CelD. MATERIAL AND METHODS: A total of 52 patients primarily diagnosed with CelD and suspicion of misdiagnosis were selected for this retrospective study. External histological reports of duodenal biopsies were obtained and later reassessed by an experienced in-house pathologist. For an objective evaluation a histology quality score (HQS) was created. Both pathological reports were compared and causes for a possible misdiagnosis were analyzed. Diagnostic systems by Catassi and Korponay-Szabo were compared with each other. RESULTS: The original diagnosis had been confirmed in 27% by our pathologist. The HQS was significantly lower (worse) in cases where the original diagnosis were dismissed than in confirmed CelD (p = 0.018). The new diagnostic approach by Catassi and Korponay-Szabo showed a sensitivity of 89% and 83%, respectively, a specificity of 97%, a positive predictive value of 94% and a negative predictive value of 94% and 92%, respectively. CONCLUSIONS: A low quality of the histological evaluation can lead to a high probability of misdiagnosing CelD. By applying the HQS the physician can estimate whether the report is fraught with uncertainty. Ideally the score minimizes false-positive results and prevents a delay of the correct diagnosis.

Noninvasive differentiation of simple steatosis and steatohepatitis by using gadoxetic acid-enhanced MR imaging in patients with nonalcoholic fatty liver disease: a proof-of-concept study.

PURPOSE: To determine whether gadoxetic acid-enhanced magnetic resonance (MR) imaging can be used to distinguish between simple steatosis and nonalcoholic steatohepatitis (NASH) in patients with nonalcoholic fatty liver disease (NAFLD), defined according to the steatosis activity and fibrosis (SAF) scoring system, which is based on the semiquantitative scoring of steatosis activity and liver fibrosis. MATERIALS AND METHODS: The local institutional review committee approved this study and waived written informed consent. This was a retrospective study of gadoxetic acid-enhanced 3-T MR imaging performed in 81 patients with NAFLD (45 men [56%]; mean age, 56 years; range, 25-78 years). The MR images were analyzed by using the relative enhancement (the ratio of signal intensities of the liver parenchyma before and 20 minutes after intravenous administration of gadoxetic acid). Univariate and multiple regression analyses were applied to identify variables associated with relative enhancement measurements. The ability of relative enhancement to allow differentiation between simple steatosis and NASH was assessed by using area under the receiver operating characteristic (ROC) curve analysis. RESULTS: Relative enhancement negatively correlated with the degree of lobular inflammation (r = -0.59, P < .0001), ballooning (r = -0.44, P < .0001), and fibrosis (r = -0.59, P ≤ .0001), but not with steatosis (r = -0.16, P = .15). Patients with NASH had a significantly lower relative liver enhancement (0.82 ± 0.22) than those with simple steatosis (1.39 ± 0.52) (P < .001). Relative enhancement measurements performed well in the differentiation between simple steatosis and NASH, with an area under the ROC curve of 0.85 (95% confidence interval: 0.75, 0.91) (cutoff = 1.24, sensitivity = 97%, specificity = 63%). CONCLUSION: Gadoxetic acid relative enhancement was significantly lower in patients with NASH than in patients with simple steatosis, but further prospective studies are warranted.

Susceptibility-weighted MR imaging in the grading of liver fibrosis: a feasibility study.

PURPOSE: To assess the feasiblity of magnetic resonance (MR) susceptibility-weighted (SW) imaging as a tool to evaluate liver fibrosis grades in patients with chronic liver diseases (CLD) utilizing signal intensity (SI) measurements, with histopathologic findings as the reference standard. MATERIALS AND METHODS: This retrospective study was approved by the local ethics committee. All subjects gave written informed consent. Eighty consecutive patients (mean age, 56.8 years), 60% of whom were male [n = 48] and 40% of whom were female [n = 32], with CLD due to various underlying causes and histopathologically proved liver fibrosis were included. Biopsies were evaluated for liver fibrosis and necroinflammatory activity (according to METAVIR scoring system), iron load, and steatosis. Two radiologists, blinded to the clinical data, assessed regions of interest in the liver and spinal muscle in consensus. Liver-to-muscle SI ratios were calculated and correlated to histopathologic findings and clinical data by using univariate and multivariate regression analysis. RESULTS: Liver-to-muscle SI ratio decreased in parallel with the increasing grade of liver fibrosis and correlated strongly with liver fibrosis (r = -0.81, P < .0001) and moderately with necroinflammatory activity (r = -0.52, P < .0001) and iron load (r = -0.37, P = .0002) but did not correlate with steatosis (r = -0.18, P = .11). In multiple regression analysis, liver fibrosis and iron load independently influenced SW imaging measurements, explaining 69% of the variance of liver-to-muscle SI ratio (R(2) = 0.69, P < .001). Liver-to-muscle SI ratio performed well in grading liver fibrosis, with an area under the receiver operating characteristic curve of 0.92 for scores of F2 or higher and 0.93 for score of F4 (liver cirrhosis). CONCLUSION: SW imaging is a feasible noninvasive tool to detect moderate and advanced liver fibrosis in CLD patients.

[Gastrointestinal stromal tumors: recommendations for diagnosis, treatment and aftercare in Austria]. / Klinische Empfehlungen. Gastrointestinale Stromatumore: Empfehlungen zu Diagnose, Behandlung und Nachsorge in Osterreich.

Diagnosis and treatment of gastrointestinal stromal tumors (GIST) requires an interdisciplinary treatment approach. This strategy should be reflected by the content of this article. Austrian representatives of 'GIST relevant' specialties authored this publication on a consensual base. This manuscript should be regarded as a guideline for 'GIST involved' colleagues in Austria.

FGF18 in colorectal tumour cells: autocrine and paracrine effects.

Fibroblast growth factors (FGFs) and their high-affinity receptors contribute to the autocrine growth stimulation in several human malignancies. Here, we describe that FGF18 expression is up-regulated in 34/38 colorectal tumours and is progressively enhanced during colon carcinogenesis reaching very high levels in carcinoma. Moreover, our data suggest that FGF18 affects both tumour cells and tumour microenvironment in a pro-tumorigenic and pro-metastatic way. Addition of recombinant FGF18 to the culture media of slowly growing colorectal tumour cell lines LT97 and Caco-2 stimulated proliferation. Phosphorylation of externally regulated kinase 1/2 and S6 was increased already 5 min after growth factor addition. SW480 cells, endogenously producing large amounts of FGF18, were not affected in this setting, but recombinant FGF18 supported tumour cell survival under conditions of serum starvation. Down-modulation of endogenous FGF18 production by small interference RNA (siRNA) significantly reduced clonogenicity of SW480 cells and restored sensitivity to exogenous FGF18. With respect to the tumour microenvironment, both recombinant and tumour-derived FGF18 stimulated growth of colon-associated fibroblasts at 0.1 ng/ml and migration at 10 ng/ml. In addition, recombinant FGF18 (10 ng/ml) induced tube formation in human umbilical vein endothelial cells. siRNA knock down demonstrated that tube-forming activity of colon cancer cell supernatants depended to a large part on tumour cell-derived FGF18. In summary, this study demonstrates that FGF18 is almost generally over-expressed in colon cancer and exerts pro-tumorigenic effects both in the epithelial and the stromal compartments by stimulating growth and survival of tumour cells, migration of fibroblasts and neovascularization. Together, these data strongly support an oncogenic role of FGF18 in colorectal cancer.

Very early viral kinetics on interferon treatment in chronic hepatitis C virus genotype 4 infection.

BACKGROUND: Interferon (IFN)-resistant hepatitis C virus strains limit efficacy of antiviral combination therapy in patients infected with genotypes 1 and 4. A single test dose of IFN was useful to identify non-responders to IFN-alpha2b/ribavirin (RBV) or likely non-responders to pegylated (PEG)-IFN-alpha2a/RBV therapy in genotype 1 patients. Our aim was to investigate this approach in genotype 4 patients. METHODS: Viral load was measured in 46 patients before and 24 h after 10 megaunits (MU) IFN-alpha2b, and before and during 2 weeks of daily 5 MU IFN-alpha2b administration. Thereafter, patients received 48 weeks combination therapy with either 180 microg PEG-IFN-alpha2a/week (n=33), 1.5 microg/kg PEG-IFN-alpha2b/week (n=7) or 5 MU IFN-alpha2b/2 days (n=6), along with 1-1.2g RBV/day. For prediction analysis the largest group (PEG-IFN-alpha2a) was evaluated only. RESULTS: Median 24 h log10 change after 10 MU IFN-alpha2b was 1.15 (range 0.08-2.48) and after 5 MU IFN-alpha2b was 0.81 (-0.12-2.22; P<0.0001). Log10 changes after 2 weeks on 5 MU IFN-alpha2b daily and 24 h after 10 MU were the best predictors of early virological response (defined by negativity of a standard qualitative PCR) to PEG-IFN-alpha2a/RBV combination therapy (area under curve [AUC]=0.97; P<0.001, receiver operating characteristics), 24 h log10 change after 10 MU was the best predictor of sustained virological response (SVR; AUC=0.91, P=0.001). CONCLUSION: As in genotype 1 patients, there is large variation in IFN responsiveness, including the presence of resistant strains, in genotype 4 patients. A 24 h log10 change after 10 MU IFN-alpha2b is an excellent predictor of SVR on PEG-IFNalpha2a/RBV combination therapy. This test may be useful to obtain homogeneous groups for clinical studies and could help in clinical decision making.

Development of a new PCR protocol for the detection of species and genotypes (strains) of Echinococcus in formalin-fixed, paraffin-embedded tissues.

The aim of our study was to establish a new PCR protocol for the detection and discrimination of Echinococcus granulosus complex on one hand and Echinococcus multilocularis in formalin-fixed, paraffin-embedded tissues (FFPTs) on the other. The target sequences for all PCRs are located on a 471bp segment of the mitochondrial ND1 gene, the fragment sizes of the amplification products are 295bp (for the sheep strain of E. granulosus), 204bp (for the pig strain of E. granulosus) and 252bp (for E. multilocularis), respectively. In total, 80 FFPTs from patients with histologically confirmed echinococcosis (76 with E. granulosus and four with E. multilocularis) operated on in Austrian hospitals between 1978 and 2005 were examined. In 68 (85%) samples, we were able to detect specific DNA fragments with our newly established PCR protocols. Thirty-eight (47.5%) of 80 clinical samples were identified as the G1 strain, 26 (32.5%) as the G5, 6 or 7 strains and four (5%) as E. multilocularis. The specificity of all three PCRs was 100%; for the discrimination between G6 and G7 strains, sequencing of an additional 234bp PCR fragment was necessary and showed that three out of 26 G5, 6 or 7 PCR-positive patients were infected with E. granulosus genotype G6 (the camel strain).

Microscopic biliary and perineural invasion and clinical outcome after neoadjuvant bevacizumab-based chemotherapy and liver resection in patients with colorectal liver metastases.

BACKGROUND: The value of microscopic biliary and perineural invasion as prognostic biomarkers in patients with resectable colorectal liver metastases (CLM) who undergo neoadjuvant chemotherapy and liver resection is still unclear. This retrospective study was performed to elucidate this issue. METHODS: Histologic slides of resected CLM of patients who underwent neoadjuvant bevacizumab-based chemotherapy and liver resection were investigated with respect to biliary and perineural invasion. Presence of invasion was correlated with radiologic and histologic response, recurrence-free survival (RFS) and overall survival (OS). RESULTS: One hundred forty-one patients were enrolled. There was a significant association between biliary and perineural invasion, respectively (P = 0.001). Moreover, both biliary and perineural invasion were associated with bilobar metastatic spread and higher number of metastases, while perineural invasion was also associated with a higher Fong score. No significant association was found with response. In univariable analysis, biliary and perineural invasion were associated with shorter RFS (median 10.1 vs. 13.5 months, HR 2.09, P = 0.010 and 7.6 vs. 14.0, HR 2.23, P = 0.001, respectively). Biliary invasion was also associated with shorter OS (median 32.8 months vs. not reached, HR 2.78, P = 0.010), however these results did not remain significant in multivariable analysis. CONCLUSIONS: In patients with resectable colorectal liver metastases undergoing neoadjuvant bevacizumab-based chemotherapy and liver resection, biliary and perineural invasion are associated with higher tumor load but may not be prognostic biomarkers.

• Pancho trial (p53-adapted neoadjuvant chemotherapy for resectable esophageal cancer) completed-mutation rate of the marker higher than expected.

BACKGROUND: In operable esophageal cancer patients, neoadjuvant therapy benefits only those who respond to the treatment. The • Pancho trial represents the first prospective randomized trial evaluating the relevance of the mark53 status for predicting the effect of two different neoadjuvant chemotherapies. METHOD: Biomarker analysis was conducted using the mark53 analysis. Calculation of patient number needed was based on a 60% rate of marker positivity, deduced from the results of a phase II pilot study. RESULTS: From 2007-2012, the • Pancho trial recruited 235 patients with operable esophageal cancer in Austria. A total of 181 patients were eligible and could be subjected to mark53 analysis and randomization. After randomizing 74 patients, the overall TP53 mutation rate was 79%. However, due to the high prevalence of marker positivity, the number of projected patients was increased to 181 patients in order to ensure a sufficient number of marker-negative patients. After completion of the trial, the overall TP53 mutation rate was 77.9%. CONCLUSION: Due to high medical need, the recruitment for the academic trial was excellent. Mark53 analysis clearly detected more mutations in the TP53 gene as compared to the cancer-specific p53 literature. Final analysis examining the interaction between the mark53 status and the effect of chemotherapies applied in the • Pancho trial is now awaited.

Low hepatic copper content and PNPLA3 polymorphism in non-alcoholic fatty liver disease in patients without metabolic syndrome.

INTRODUCTION: The pathogenesis of non-alcoholic fatty liver disease (NAFLD) is multifactorial including metabolic, genetic (e.g. PNPLA3 [patatin-like phospholipase domain-containing 3 gene]), viral factors and drugs. Besides, there is evidence for a role of copper deficiency. Aim of the study was to evaluate the role of hepatic copper content, PNPLA3 in NAFLD patients with and without metabolic syndrome (MetS). METHODS: One-hundred seventy-four NAFLD patients, who underwent liver biopsy for diagnostic work-up, were studied. Diagnosis of MetS was based on the WHO Clinical Criteria. Steatosis was semiquantified as percentage of fat containing hepatocytes and was graded according to Brunt. Histological features of non-alcoholic steatohepatitis (NASH) were assessed using the Bedossa classification. Hepatic copper content (in µg/g dry weight) was measured by flame atomic absorption spectroscopy. SNP rs738409 in PNPLA3 was investigated by RT-PCR. RESULTS: Mean hepatic copper content was 22.3 (19.6-25.1) µg/g. The mean percentage of histologically lipid containing hepatocytes was 42.2% (38.3-46.0) and correlated inversely with hepatic copper content (ρ=-0.358, P<0.001). By subgroup analysis this inverse correlation remained significant only in patients without MetS (OR: 0.959 [CI95%: 0.926-0.944], P=0.020). Presence of minor allele (G) of PNPLA3 was also associated with moderate/severe steatosis (≥33%) both in patients with (OR: 2.405 [CI95%: 1.220-4.744], P=0.011) and without MetS (OR: 2.481 [CI95%: 1.172-5.250], P=0.018), but was only associated with NASH (OR: 2.002 [CI95%: 1.062-3.772], P=0.032) and liver fibrosis (OR: 2.646 [CI95%: 1.299-5.389], P=0.007) in patients without MetS. CONCLUSION: Hepatic copper content and PNPLA3 mutations are associated with disease activity in NAFLD patients without MetS. Presence of MetS appears to mask the effects of hepatic copper and PNPLA3.

Genetic detection of lymph node micrometastases: a selection criterion for liver transplantation in patients with liver metastases after colorectal cancer.

BACKGROUND: Liver transplantation for nonresectable liver metastases from colorectal cancer was abandoned in 1994 on account of high recurrence rates. The aim of this study was to investigate whether the genetic detection of micrometastases in histologically negative lymph nodes of the primary colon cancer could be applied to select patients for liver transplantation. METHODS: We analyzed 21 patients with colorectal cancer who had undergone liver transplantation between 1983 and 1994 for liver metastases. Eleven patients were histologically lymph node negative at the time of surgery; ten patients with lymph node metastases served as control group. DNA sequencing was used to screen tumor material for p53 and K-ras mutations. Mutant allele-specific amplification (MASA) was then used to search for micrometastases in DNA from regional lymph nodes of the primary colorectal cancer. RESULTS: p53 and K-ras mutations were detected in 12 (57%) and 3 (14%) of 21 patients in the colorectal cancer, respectively. The mutations were confirmed in the corresponding liver metastases. Of 11 patients with histologically negative lymph nodes, nine were eligible for MASA due to presence of p53 or K-ras mutation. MASA revealed six of nine patients to be genetically positive for micrometastases. Three patients were both genetically and histologically negative. These three patients showed a significantly longer overall survival (P = 0.011) of 4, 5, and 20 years, respectively. CONCLUSIONS: We conclude that the genetic detection of micrometastases by MASA could be a powerful prognostic indicator for selecting patients with colorectal liver metastases who could benefit from liver transplantation.

Chronic Paracoccidioidomycosis with adrenal involvement mimicking tuberculosis - A case report from Austria.

Paracoccidioidomycosis is a systemic fungal infection caused by Paracoccidioides brasiliensis and endemic in certain areas of Central and South America. We report a case of a 62-year-old-man with a complex history of tuberculosis and imaging findings of a cerebral lesion and bilateral adrenal enlargement. Biopsy of adrenal gland revealed Paracoccidioides brasiliensis. This case highlights the importance of travel history for diagnosis of paracoccidioidomycosis in non-endemic areas and emphasizes the clinical and histopathological similarities with tuberculosis.

Prevalence of Liver Fibrosis and its Association with Non-invasive Fibrosis and Metabolic Markers in Morbidly Obese Patients with Vitamin D Deficiency.

BACKGROUND: Morbidly obese patients are at risk for non-alcoholic fatty liver disease (NAFLD) and vitamin D deficiency (VDD). Non-alcoholic steatohepatitis (NASH) is the progressive variant of NAFLD and can advance to fibrosis, cirrhosis, and liver cancer. We aimed to examine prevalence of liver fibrosis and its non-invasive predictors in bariatric patients with VDD (<75 nmol/l). METHODS: Baseline liver biopsy of a randomized controlled trial was performed in 46 patients with omega loop gastric bypass. Clinical, laboratory, and histological data were examined and tested with univariate and multivariable analysis. RESULTS: In total, 80 % were females, aged 42 (SD 13) years with BMI 44 (4) kg/m(2). Twenty-six percent had diabetes mellitus (DM) and 44 % metabolic syndrome (MeS). Seventy-two percent had NASH, 11 % simple steatosis, and 17 % normal liver. In total, 30 % demonstrated significant fibrosis (F ≥ 2) with 9 % of advanced (F3) and 4 % cirrhosis (F4). Increased stages of fibrosis were primarily associated with higher levels of HOMA2-insulin resistance (IR), procollagen type I propeptide (P1NP), lower osteocalcin, albumin-corrected calcium, parathyroid hormone, vitamin D, male sex, and higher age. Other independent risk factors for advanced fibrosis were MeS (OR = 9.3 [0.99-87.5], p = 0.052) and DM (OR = 12.8 [1.2-137.4], p = 0.035). The fibrosis FIB-4 index <10.62 and NAFLD fibrosis score <-26.93 had a negative predictive value of 100 and 96 %, respectively. CONCLUSIONS: Liver fibrosis is frequent in morbidly obese patients with concurrent DM and/or MeS. Increased serum levels of IR, P1NP, lower osteocalcin, and VDD are clinically relevant predictors of fibrosis. Consequently, we suggest that patients with preoperative presence of these markers are at increased risk for liver fibrosis and should be monitored closely.