Prevalence of anti-Hepatitis E antibodies and impact on disease severity in non-alcoholic fatty liver disease.

AIM: In most immune-competent individuals, hepatitis E (HEV) infections appear silent. It is unclear whether past HEV infections deteriorate disease severity in patients with non-alcoholic fatty liver disease (NAFLD). METHODS: Patients with biopsy-proven NAFLD and data on anti-HEV immunoglobulin M (HEV-IgM) and anti-HEV IgG antibodies (HEV-IgG) were included. The NAFLD activity score (NAS) was used to grade and stage all liver biopsy samples. The HEV-IgG prevalence was compared to a healthy cohort of 997 subjects. RESULTS: One hundred sixty-seven patients with NAFLD were included with the following characteristics: age, 50 ± 13 years; NAS ≤4, 89 (53.3%); NAS 5-8, 78 (46.7%); cirrhosis, 16 (9.6%). Two patients (1.2%) were HEV-IgM-positive, however HEV polymerase chain reaction remained negative and no signs of acute hepatitis were seen. Forty-four patients (26.3%) were HEV-IgG-positive and they were significantly older (55 ± 10 years vs. 48 ± 13 years, P < 0.001) and predominantly men (31 [70.5%] vs.13 [29.5%], P = 0.022). Distribution across NAS (P = 0.610) was not different. However, HEV-IgG-positive patients were significantly more often found with cirrhosis (8 [18.2%] vs. 8 [6.5%], P = 0.024) and liver stiffness values >10 kPa (14 [58.2%] vs. 29 [43.3%], P = 0.026). Multivariable analyses revealed age (odds ratio [OR], 1.054 [1.022-1.086]) and male sex (OR 2.77 [1.27-6.04]) associated with HEV-IgG positivity. Presence of diabetes (OR 3.86 [1.18-12.59]), higher aspartate aminotransferase levels (OR, 1.02 [1.006-1.033]), and HEV-IgG seropositivity (OR 3.52 [1.11-11.13]) were independently linked to cirrhosis. Finally, HEV-IgG positivity was not independently associated with NAFLD patients in a case-control study including healthy subjects. CONCLUSIONS: Prevalence of anti-HEV-IgG antibodies in patients with NAFLD is linked to age and male sex. Furthermore, previous HEV infection was an independent risk factor for cirrhosis. Whether this finding is causal or solely associative is unclear and should be elucidated in future studies.

Variants in ABCB4 (MDR3) across the spectrum of cholestatic liver diseases in adults.

The ATP binding cassette subfamily B member 4 (ABCB4) gene on chromosome 7 encodes the ABCB4 protein (alias multidrug resistance protein 3 [MDR3]), a P-glycoprotein in the canalicular membrane of the hepatocytes that acts as a translocator of phospholipids into bile. Several variants in ABCB4 have been shown to cause ABCB4 deficiency, accounting for a disease spectrum ranging from progressive familial cholestasis type 3 to less severe conditions like low phospholipid-associated cholelithiasis, intrahepatic cholestasis of pregnancy or drug-induced liver injury. Furthermore, whole genome sequencing has shown that ABCB4 variants are associated with an increased incidence of gallstone disease, gallbladder and bile duct carcinoma, liver cirrhosis or elevated liver function tests. Diagnosis of ABCB4 deficiency-related diseases is based on clinical presentation, serum biomarkers, imaging techniques, liver histology and genetic testing. Nevertheless, the clinical presentation can vary widely and clear genotype-phenotype correlations are currently lacking. Ursodeoxycholic acid is the most commonly used medical treatment, but its efficacy has yet to be proven in large controlled clinical studies. Future pharmacological options may include stimulation/restoration of residual function by chaperones (e.g. 4-phenyl butyric acid, curcumin) or induction of ABCB4 transcription by FXR (farnesoid X receptor) agonists or PPARα (peroxisome proliferator-activated receptor-α)-ligands/fibrates. Orthotopic liver transplantation remains the last and often only therapeutic option in cirrhotic patients with end-stage liver disease or patients with intractable pruritus.

Non-invasive diagnosis of cirrhosis and long-term disease monitoring by transient elastography in patients with Wilson disease.

BACKGROUND & AIMS: The value of liver stiffness measurement (LSM) by transient elastography (TE) for non-invasive fibrosis staging and disease monitoring has not been established in patients with Wilson disease (WD). METHODS: Liver stiffness measurement by TE and non-invasive fibrosis scores (APRI, FIB-4) were analysed from 188 WD patients with liver biopsy (LBX). Longitudinal LSM was performed in 128 (68.1%) patients. RESULTS: One hundred and eighty-eight patients (mean age: 35 ± 14 years, 54.8% women; 27.1% with histological cirrhosis) were studied. Forty-four[23.4%] patients were recently diagnosed with WD, while 144[76.6%] were previously diagnosed (>1 year between LBX and LSM). Overall, LSM (11.3 vs 6.1 kPa, P < .001), APRI (0.72 vs 0.38, P < .001) and FIB-4 (1.54 vs 0.89, P < .001) were higher in cirrhotic than in non-cirrhotic patients. This was even more pronounced in recently diagnosed patients (35.2 kPa vs 6.4 kPa, P < .001). Accuracy for diagnosing cirrhosis at an LSM cut-off ≥9.9 kPa was better in recently diagnosed (PPV: 74%, NPV: 100%) vs previously diagnosed (PPV: 53%, NPV: 82%) patients. Recently diagnosed patients had higher Area Under the Curve (AUC) for APRI (0.79 vs 0.61) and FIB-4 (0.84 vs 0.65) than previously diagnosed patients. At APRI <1.5 and FIB-4 <3.25 cirrhosis was ruled out with a specificity of 93% and 95% respectively. During a median follow-up of 46 (24-66) months, only 5.9% (5/85) of non-cirrhotic WD patients showed progression to cirrhotic LSM values, while 30.8% (4/13) of cirrhotic WD patients showed LSM suggestive of cirrhosis regression. CONCLUSION: TE-based LSM ≥9.9 kPa accurately identifies cirrhosis in WD patients. Next to TE-LSM <9.9 kPa, APRI <1.5 and FIB-4 <3.25 values assist to non-invasively rule out cirrhosis. LSM remains stable in most non-cirrhotic patients on WD therapy, while one-third of cirrhotic patients present clinically relevant decreases in LSM.

Chronic Hepatitis E is associated with cholangitis.

BACKGROUND AND AIMS: Sporadic hepatitis E is an emerging indigenous disease in Europe induced by genotype 3 of the virus. While the disease takes an acute self-limited course in immunocompetent individuals, under immunocompromised conditions chronic hepatitis E might develop. The histology of chronic hepatitis E has not been described in detail systematically. METHODS: Liver biopsies from 19 immunosuppressed patients with chronic hepatitis E were collected: 17 were organ transplant recipients, one had a CD4-deficiency and one had received steroid therapy because of ulcerative colitis. Biopsies were processed with standard stains. Evaluation of histologic activity and fibrosis was performed according to Ishak. Additionally, immunohistochemistry with antibodies directed against open reading frame 2 and 3 of the virus was performed and liver biopsies were tested for hepatitis E virus RNA. RESULTS: Biochemical data showed an increase in alanine transaminase, aspartate transaminase, gamma-glutamyl transferase and total bilirubin. Histopathology displayed typical features of chronic hepatitis with mild to moderate activity. The number of polymorphonuclear leucocytes was considerably increased and all patients had a florid cholangitis that presented as a destructive form in five of them. Hepatocytes and bile duct epithelia stained positive for hepatitis E virus by immunohistochemistry. CONCLUSIONS: Chronic hepatitis E in immunocompromised individuals runs a similar course as hepatitis B and C and shows similar histopathology. However, the presence of destructive cholangitis in some cases accompanied by an increased number of polymorphonuclear leucocytes is markedly different. Immunohistochemically the virus is present in bile duct epithelia, seemingly the cause for cholangitis.

EGFR in Tumor-Associated Myeloid Cells Promotes Development of Colorectal Cancer in Mice and Associates With Outcomes of Patients.

BACKGROUND & AIMS: Inhibitors of the epidermal growth factor receptor (EGFR) are the first-line therapy for patients with metastatic colorectal tumors without RAS mutations. However, EGFR inhibitors are ineffective in these patients, and tumor level of EGFR does not associate with response to therapy. We screened human colorectal tumors for EGFR-positive myeloid cells and investigated their association with patient outcome. We also performed studies in mice to evaluate how EGFR expression in tumor cells and myeloid cells contributes to development of colitis-associated cancer and ApcMin-dependent intestinal tumorigenesis. METHODS: We performed immunohistochemical and immunofluorescent analyses of 116 colorectal tumor biopsies to determine levels of EGFR in tumor and stroma; we also collected information on tumor stage and patient features and outcomes. We used the Mann-Whitney U and Kruskal-Wallis tests to correlate tumor levels of EGFR with tumor stage, and the Kaplan-Meier method to estimate patients' median survival time. We performed experiments in mice lacking EGFR in intestinal epithelial cells (Villin-Cre; Egfrf/f and Villin-CreERT2; Egfrf/f mice) or myeloid cells (LysM-Cre; Egfrf/f mice) on a mixed background. These mice were bred with ApcMin/+ mice; colitis-associated cancer and colitis were induced by administration of dextran sodium sulfate (DSS), with or without azoxymethane (AOM), respectively. Villin-CreERT2 was activated in developed tumors by administration of tamoxifen to mice. Littermates that expressed full-length EGFR were used as controls. Intestinal tissues were collected; severity of colitis, numbers and size of tumors, and intestinal barrier integrity were assessed by histologic, immunohistochemical, quantitative reverse transcription polymerase chain reaction, and flow cytometry analyses. RESULTS: We detected EGFR in myeloid cells in the stroma of human colorectal tumors; myeloid cell expression of EGFR associated with tumor metastasis and shorter patient survival time. Mice with deletion of EGFR from myeloid cells formed significantly fewer and smaller tumors than the respective EGFR-expressing controls in an ApcMin/+ background as well as after administration of AOM and DSS. Deletion of EGFR from intestinal epithelial cells did not affect tumor growth. Furthermore, tamoxifen-induced deletion of EGFR from epithelial cells of established intestinal tumors in mice given AOM and DSS did not reduce tumor size. EGFR signaling in myeloid cells promoted activation of STAT3 and expression of survivin in intestinal tumor cells. Mice with deletion of EGFR from myeloid cells developed more severe colitis after DSS administration, characterized by increased intestinal inflammation and intestinal barrier disruption, than control mice or mice with deletion of EGFR from intestinal epithelial cells. EGFR-deficient myeloid cells in the colon of DSS-treated LysM-Cre; Egfrf/f mice had reduced expression of interleukin 6 (IL6), and epithelial STAT3 activation was reduced compared with controls. Administration of recombinant IL6 to LysM-Cre; Egfrf/f mice given DSS protected them from weight loss and restored epithelial proliferation and STAT3 activation, compared with administration of DSS alone to these mice. CONCLUSIONS: Increased expression of EGFR in myeloid cells from the colorectal tumor stroma associates with tumor progression and reduced survival time of patients with metastatic colorectal cancer. Deletion of EGFR from myeloid cells, but not intestinal epithelial cells, protects mice from colitis-induced intestinal cancer and ApcMin-dependent intestinal tumorigenesis. Myeloid cell expression of EGFR increases activation of STAT3 and expression of survivin in intestinal epithelial cells and expression of IL6 in colon tissues. These findings indicate that expression of EGFR by myeloid cells of the colorectal tumor stroma, rather than the cancer cells themselves, contributes to tumor development.

Feasibility of personalized treatment concepts in gastrointestinal malignancies: Sub-group results of prospective clinical phase II trial EXACT.

OBJECTIVE: Advances in high-throughput genomic profiling and the development of new targeted therapies improve patient's survival. In gastrointestinal (GI) malignancies, the concept of personalized medicine (PM) was not investigated so far. The aim of this prospective study was to evaluate the efficacy of a personalized treatment in GI patients who failed standard treatment. METHODS: Out of the original prospective clinical phase II EXACT trial, 21 (38%) GI cancer patients who had no further treatment options were identified. A molecular profile (MP) via a 50 gene next generation sequencing (NGS) panel in combination with immunohistochemistry (IHC) was conducted using real-time biopsy tumor material. Results were discussed by a multidisciplinary team (MDT) to translate the individual MP in an experimental treatment. RESULTS: Of the 55 patients originally included in the EXACT trial, 21 (38%) suffered from GI malignancies. The final analysis showed that 15 (71%) patients had experienced a longer progression-free survival (PFS) upon experimental targeted treatment (124 d, quartiles 70/193 d), when compared with the PFS achieved by the previous conventional therapy (62 d, quartiles 55/83 d) (P=0.014). Thirteen (62%) patients receiving targeted treatment experienced a disease control according to Response Evaluation Criteria in Solid Tumors (RECIST). Median overall survival (OS) from the start of experimental therapy to time of censoring or death was 193 d (quartiles 115/374 d). CONCLUSIONS: PM was not investigated in GI malignancies so far in a prospective trial. This study shows that treatment based on real-time molecular tumor profiling led to a superior clinical benefit, and survival as well as response was significantly improved when compared with previous standard medications.

Ultra-high-field magnetic resonance spectroscopy in non-alcoholic fatty liver disease: Novel mechanistic and diagnostic insights of energy metabolism in non-alcoholic steatohepatitis and advanced fibrosis.

BACKGROUND & AIMS: With the rising prevalence of non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH) non-invasive tools obtaining pathomechanistic insights to improve risk stratification are urgently needed. We therefore explored high- and ultra-high-field magnetic resonance spectroscopy (MRS) to obtain novel mechanistic and diagnostic insights into alterations of hepatic lipid, cell membrane and energy metabolism across the spectrum of NAFLD. METHODS: MRS and liver biopsy were performed in 30 NAFLD patients with NAFL (n=8) or NASH (n=22). Hepatic lipid content and composition were measured using 3-Tesla proton (1 H)-MRS. 7-Tesla phosphorus (31 P)-MRS was applied to determine phosphomonoester (PME) including phosphoethanolamine (PE), phosphodiester (PDE) including glycerophosphocholine (GPC), phosphocreatine (PCr), nicotinamide adenine dinucleotide phosphate (NADPH), inorganic phosphate (Pi), γ-ATP and total phosphorus (TP). Saturation transfer technique was used to quantify hepatic ATP flux. RESULTS: Hepatic steatosis in 1 H-MRS highly correlated with histology (P<.001) showing higher values in NASH than NAFL (P<.001) without differences in saturated or unsaturated fatty acid indices. PE/TP ratio increased with advanced fibrosis (F3/4) (P=.002) whereas GPC/PME+PDE decreased (P=.05) compared to no/mild fibrosis (F0-2). γ-ATP/TP was lower in advanced fibrosis (P=.049), while PCr/TP increased (P=.01). NADPH/TP increased with higher grades of ballooning (P=.02). Pi-to-ATP exchange rate constant (P=.003) and ATP flux (P=.001) were lower in NASH than NAFL. CONCLUSIONS: Ultra-high-field MRS, especially saturation transfer technique uncovers changes in energy metabolism including dynamic ATP flux in inflammation and fibrosis in NASH. Non-invasive profiling by MRS appears feasible and may assist further mechanistic and therapeutic studies in NAFLD/NASH.

A retrospective study on the safety, diagnostic yield, and therapeutic effects of endoscopic unroofing for small gastric subepithelial tumors.

BACKGROUND AND AIMS: Accurate diagnosis of small gastric subepithelial tumors (SETs) is essential to assess their malignant potential. Endoscopic unroofing has been reported to yield sufficient tissue samples for histologic evaluation. This study aimed to evaluate the safety, diagnostic yield, and potential therapeutic effects of this technique over time. METHODS: This retrospective analysis of prospectively collected clinical data identified patients who underwent endoscopic unroofing at the Medical University of Vienna from January 2003 to December 2012. Demographic data, indications for endoscopic unroofing, intraprocedural adverse events, hospital stay, histologic results, and follow-up procedures were reviewed. RESULTS: A total of 14 patients (7 men; 7 women; median age, 70 years; range, 51-95 years) underwent endoscopic unroofing of 14 gastric SETs with a mean diameter of 26 ± 13 mm at EUS. In 9 of 14 cases, endoscopic unroofing was done exclusively for diagnostic purposes; in the remaining cases, it was performed with therapeutic intent because of bleeding from the gastric SETs. Unroofing was technically successful in 13 of 14 cases and revealed 8 cases of GI stromal tumor (GIST) and 1 case each of leiomyoma, fibroid polyp, glomus tumor, pancreatic rest, and nondiagnostic material at histology. Intraprocedural bleeding was the only adverse event (4 cases) and could be managed endoscopically. A follow-up EUS was available (median, 8 months) for 10 of the 14 patients. Notably, most patients showed complete regression of their gastric SETs after unroofing (on white light and EUS), including the glomus tumor, the leiomyoma, and 6 of the 8 cases of GIST. CONCLUSIONS: Endoscopic unroofing was safe and had a very favorable diagnostic yield in this study. Unexpectedly, it led to complete regression in most gastric SETs. Although it is not an oncologically curative treatment, endoscopic unroofing can be a valuable option to treat local adverse events in patients unfit for surgical therapy. (Clinical trial registration number: NCT02587923.).

Potential of DNA methylation in rectal cancer as diagnostic and prognostic biomarkers.

BACKGROUND: Aberrant DNA methylation is more prominent in proximal compared with distal colorectal cancers. Although a number of methylation markers were identified for colon cancer, yet few are available for rectal cancer. METHODS: DNA methylation differences were assessed by a targeted DNA microarray for 360 marker candidates between 22 fresh frozen rectal tumour samples and 8 controls and validated by microfluidic high-throughput and methylation-sensitive qPCR in fresh frozen and formalin-fixed paraffin-embedded (FFPE) samples, respectively. The CpG island methylator phenotype (CIMP) was assessed by MethyLight in FFPE material from 78 patients with pT2 and pT3 rectal adenocarcinoma. RESULTS: We identified and confirmed two novel three-gene signatures in fresh frozen samples that can distinguish tumours from adjacent tissue as well as from blood with a high sensitivity and specificity of up to 1 and an AUC of 1. In addition, methylation of individual CIMP markers was associated with specific clinical parameters such as tumour stage, therapy or patients' age. Methylation of CDKN2A was a negative prognostic factor for overall survival of patients. CONCLUSIONS: The newly defined methylation markers will be suitable for early disease detection and monitoring of rectal cancer.

Improved survival in HPV/p16-positive oropharyngeal cancer patients treated with postoperative radiotherapy.

INTRODUCTION: In the literature, HPV infection and/or p16 positivity have been consistently demonstrated to correlate with improved response rates in oropharyngeal squamous cell carcinoma (OPSCC) patients treated with primary radiotherapy (RT) alone and in combination with chemotherapy. However, the exact role of HPV/p16 positivity in patients treated with postoperative RT is still unclear. METHODS: We analyzed tumor samples for HPV-DNA and p16 expression and correlated these variables with treatment outcome in a series of 63 consecutively treated oropharyngeal cancer patients (95% stage III/IV). HPV and p16 analysis were performed using validated test systems. Survival was estimated by the Kaplan-Meier method. Cox proportional hazard regression models were applied to compare the risk of death among patients stratified according to risk factors. RESULTS: Expression of p16 or high-risk HPV-DNA was detected in 60.3% and 39.6% of the tumors, respectively. p16 expression [overall survival (OS) at 2 years: 91%] as well as HPV infection (OS at 2 years: 95%) was associated with improved OS. Mean survival in p16-positive patients was 112 months compared to 64.6 months in case of p16 negativity. All HPV-positive tumors stained positive for p16. In a multivariable analysis, p16 positivity was associated with improved OS and with disease-free survival. CONCLUSION: p16 expression and HPV infection are strongly associated with the outcome of postoperatively irradiated OPSCC patients. HPV and p16 double-negative OPSCC patients should be regarded as a distinct "very high-risk patient group" that may benefit from intensified or novel treatment combinations.

Differential survival trends of stage II colorectal cancer patients relate to promoter methylation status of PCDH10, SPARC, and UCHL1.

Surgical excision of colorectal cancer at early clinical stages is highly effective, but 20-30% of patients relapse. Therefore, it is of clinical relevance to identify patients at high risk for recurrence, who would benefit from adjuvant chemotherapy. The objective of this study was to identify prognostic and/or predictive methylation markers in stage II colorectal cancer patients. Therefore, we selected six gene promoters (FZD9, PCDH10 (protocadherin 10), SFRP2, SPARC (secreted protein acidic and rich in cysteine), UCHL1 (ubiquitin carboxyl-terminal hydrolase 1), and WIF1) for methylation analysis in formalin-fixed, paraffin-embedded primary tumor samples of colorectal cancer patients (n=143) who were enrolled in a prospective randomized phase III trial of the Austrian Breast and Colorectal cancer Study Group. Patients were randomized to adjuvant chemotherapy with 5-fluorouracil and leucovorin or surveillance only. Survival analyses revealed that combined evaluation of three promoters (PCDH10, SPARC, and UCHL1) showed differential effects with regard to disease-free survival and overall survival in the two treatment groups (significance level 0.007). In the chemotherapy arm, a statistically insignificant trend for patients without methylation toward longer survival was observed (P=0.069 for disease-free survival and P=0.139 for overall survival). Contrary, patients in the surveillance arm without methylation in their gene promoters had shorter disease-free survival and overall survival (P=0.031 for disease-free survival and P=0.003 for overall survival), indicating a prognostic effect of methylation in this group (test for interaction, P=0.006 for disease-free survival and P=0.018 for overall survival). These results indicate that promoter methylation status of PCDH10, SPARC, and UCHL1 may be used both as prognostic and predictive molecular marker for colorectal cancer patients and, therefore, may facilitate treatment decisions for stage II colorectal cancer.

Significance of p16 expression in head and neck cancer patients treated with radiotherapy and cetuximab.

BACKGROUND: HPV-infection, p16 positivity, and EGFR expression have been correlated with favorable responses of head and neck cancer patients treated with radiotherapy (RT) with or without chemotherapy. However, a possible correlation of HPV/p16 and EGFR status on the effect of RT in combination with cetuximab has not been sufficiently investigated. MATERIALS AND METHODS: We analyzed tumor samples for p16 and EGFR expression and correlated these variables with treatment outcome. Cox-proportional-hazard regression models were applied to compare the risk of death among patients stratified according to risk factors. Survival was estimated by the Kaplan-Meier method. Results were compared with an institutional historical control group treated without cetuximab and with published data. RESULTS: Expression of p16 was predominantly found in oropharyngeal squamous cell cancer patients (OPSCC; 36.6% positivity; 92% of all cases), while EGFR was expressed at high levels in all tumor subsites (82%). p16 expression was associated with improved overall survival in irradiated OPSCC patients (2-year overall survival of 80% in p16-positive vs. 33% overall survival in p16-negative patients). In a multivariable analysis covering all tumor sites, nodal stage (> N2a vs. ≤ N2a) and tumor site (OPSSC vs. non-OPSCC) had an impact on overall survival. CONCLUSION: Our results show that p16 positivity is associated with a favorable outcome in OPSCC patients treated with RT and cetuximab.

Role of FDFT1 polymorphism for fibrosis progression in patients with chronic hepatitis C.

BACKGROUND: In chronic hepatitis C (CHC), steatosis is associated with fibrosis and impaired response to antiviral therapy. Recently, a polymorphism of single nucleotide polymorphism SNP rs2645424 of farnesyl diphosphate farnesyl transferase 1 (FDFT1) was identified in NAFLD/NASH as a possible causal link to steatosis and fibrosis progression. SNP rs738409 in the adiponutrin gene (PNPLA3) is a well described factor for steatosis. This study evaluated the relation of these SNPs on steatosis, fibrosis and treatment response in CHC. METHODS: The SNPs rs738409478 and rs2645424 were determined by real-time PCR in 478 patients with CHC (m/f: 314/164; mean age: 44.9 ± 10.7; GT1: 387, GT4: 91) who completed treatment with peg-IFN-α-2a/ribavirin. All had a pretreatment liver biopsy. Steatosis and fibrosis were graded by board-certified pathologists according to Brunt and METAVIR respectively. RESULTS: The distribution of FDFT1 rs2645424 was GG: 186 (38.9%), AG: 222 (46.4%) and AA: 70 (14.6%) and of the rs738409 PNPLA3 allele: CC: 269 (56.3%), CG: 177 (37.0%) and GG: 32 (6.7%). Overall, FDTF1 polymorphism was not linked to the extent of steatosis or fibrosis. However, in patients without steatosis the AA genotype was associated with advanced fibrosis [AA: 8/20 (40.0%), AG: 6/70 (8.5%), GG: 9/57 (16.1%), P = 0.003]. In contrast, the minor PNPLA3 allele was associated with both steatosis and advanced fibrosis (P < 0.001). Both SNPs did not influence treatment response. CONCLUSION: The minor allele in FDFT1 was associated with advanced fibrosis in the non-steatotic but not in the steatotic subgroup. This may reflect different metabolic pathways in fibrosis progression for steatotic and non-steatotic patients with CHC.

Anti-LFA-1 or rapamycin overcome costimulation blockade-resistant rejection in sensitized bone marrow recipients.

While costimulation blockade-based mixed chimerism protocols work well for inducing tolerance in rodents, translation to preclinical large animal/nonhuman primate models has been less successful. One recognized cause for these difficulties is the high frequency of alloreactive memory T cells (Tmem) found in the (pre)clinical setting as opposed to laboratory mice. In the present study, we therefore developed a murine bone marrow transplantation (BMT) model employing recipients harboring polyclonal donor-reactive Tmem without concomitant humoral sensitization. This model was then used to identify strategies to overcome this additional immune barrier. We found that B6 recipients that were enriched with 3 × 10(7) T cells isolated from B6 mice that had been previously grafted with Balb/c skin, rejected Balb/c BM despite costimulation blockade with anti-CD40L and CTLA4Ig (while recipients not enriched developed chimerism). Adjunctive short-term treatment of sensitized BMT recipients with rapamycin or anti-LFA-1 mAb was demonstrated to be effective in controlling Tmem in this model, leading to long-term mixed chimerism and donor-specific tolerance. Thus, rapamycin and anti-LFA-1 mAb are effective in overcoming the potent barrier that donor-reactive Tmem pose to the induction of mixed chimerism and tolerance despite costimulation blockade.

Gastrointestinal stromal tumors: diagnosis, therapy and follow-up care in Austria.

Optimal treatment for patients suffering from gastrointestinal stromal tumors (GIST) is based on an interdisciplinary treatment approach. Austrian representatives of Medical and Surgical Oncology, Pathology, Radiology, Nuclear Medicine, Gastroenterology, and Laboratory Medicine issued this manuscript on a consensual base within the context of currently available and published literature. This paper contains guidelines and recommendations for diagnosis, therapy, and follow-up of GIST patients in Austria.

Association of the IL28B genotype with insulin resistance in patients with chronic hepatitis C.

BACKGROUND & AIMS: Insulin resistance, fibrosis and steatosis are established predictors of response to peg-interferon/ribavirin therapy in chronic hepatitis C (CHC). Several host genetic polymorphisms (IL28B, PNPLA3) modify treatment-outcome, the degree of steatosis or fibrosis. The aim of our study was to evaluate the role of these polymorphisms on insulin resistance (IR) in treatment-naïve patients with chronic hepatitis C. METHODS: Two hundred and two non-diabetic CHC patients (GT1: 181, GT4: 21; m = 126, f = 76) undergoing liver biopsy in two tertiary academic centers were studied. The SNPs rs12979860 (IL28B) and rs738409 (PNPLA3) were investigated by RT-PCR. HOMA-IR, BMI, stage of fibrosis, extent of steatosis, and genetic data were analyzed. RESULTS: Insulin resistance (HOMA-IR ≥ 3.0) was associated with rs12979860 genotype, presence of advanced fibrosis, and higher BMI. HOMA-IR in CC and in TC/TT was 2.08 ± 1.61 (mean ± SD) and 2.94 ± 2.89 (p=0.041), respectively. HOMA-IR was higher in advanced than in mild fibrosis (F3-4: 3.92 ± 3.15; F0-2: 2.38 ± 2.38; p=0.004). The percentage of steatotic hepatocytes was higher in patients with advanced fibrosis (21.3 ± 21.5 vs. 9.1 ± 14.2; p<0.001), HOMA-IR ≥ 3.0 (17.7 ± 17.8 vs. 8.8 ± 15.4%; p<0.001), and BMI > 25.0 kg/m(2) (14.7 ± 17.0 vs. 9.1 ± 16.1; p<0.001). The rs738409 GG genotype was associated with advanced fibrosis and steatosis, but not with HOMA-IR. Multivariable logistic regression identified advanced fibrosis (OR: 2.820, 95% CI: 1.344-5.917; p = 0.006) and the IL28B genotype non-CC (OR: 3.000, 1.348-6.676; p = 0.007) as independent risk factors for insulin resistance. CONCLUSIONS: Insulin resistance is more common in carriers of the T allele of SNP rs12979860 than in CC homozygotes and may partly explain the poor outcome of peginterferon/ribavirin therapy in these patients.

Up-regulation of the fibroblast growth factor 8 subfamily in human hepatocellular carcinoma for cell survival and neoangiogenesis.

UNLABELLED: Fibroblast growth factors (FGFs) and their high-affinity receptors [fibroblast growth factor receptors (FGFRs)] contribute to autocrine and paracrine growth stimulation in several non-liver cancer entities. Here we report that at least one member of the FGF8 subfamily (FGF8, FGF17, and FGF18) was up-regulated in 59% of 34 human hepatocellular carcinoma (HCC) samples that we investigated. The levels of the corresponding receptors (FGFR2, FGFR3, and FGFR4) were also elevated in the great majority of the HCC cases. Overall, 82% of the HCC cases showed overexpression of at least one FGF and/or FGFR. The functional implications of the deregulated FGF/FGFR system were investigated by the simulation of an insufficient blood supply. When HCC-1.2, HepG2, or Hep3B cells were subjected to serum withdrawal or the hypoxia-mimetic drug deferoxamine mesylate, the expression of FGF8 subfamily members increased dramatically. In the serum-starved cells, the incidence of apoptosis was elevated, whereas the addition of FGF8, FGF17, or FGF18 impaired apoptosis, which was associated with phosphorylation of extracellular signal-regulated kinase 1/2 and ribosomal protein S6. In contrast, down-modulation of FGF18 by small interfering RNA (siRNA) significantly reduced the viability of the hepatocarcinoma cells. siRNA targeting FGF18 also impaired the cells' potential to form clones at a low cell density or in soft agar. With respect to the tumor microenvironment, FGF17 and FGF18 stimulated the growth of HCC-derived myofibroblasts, and FGF8, FGF17, and FGF18 induced the proliferation and tube formation of hepatic endothelial cells. CONCLUSION: FGF8, FGF17, and FGF18 are involved in autocrine and paracrine signaling in HCC and enhance the survival of tumor cells under stress conditions, malignant behavior, and neoangiogenesis. Thus, the FGF8 subfamily supports the development and progression of hepatocellular malignancy.

Evaluation of immunohistochemical expression of p53, p21, p27, cyclin D1, and Ki67 in oral and oropharyngeal squamous cell carcinoma.

BACKGROUND: The purpose of this study was to evaluate whether the immunohistochemical expression of p53, p21, p27, cyclin D1, and Ki67 can predict therapy response and survival in patients with oral and oropharyngeal squamous cell carcinoma treated with preoperative chemoradiation. METHODS: Biomarker expression was evaluated by immunohistochemistry in formalin-fixed, paraffin-embedded pretreatment biopsies of 111 homogenously treated patients. We assessed the association between clinicopathological variables including response to neoadjuvant chemoradiotherapy as well as the survival of the patients and the expression of the biomarkers as both dichotomized (positive vs. negative) and continuous variables. RESULTS: Biomarker overexpression on the basis of pre-selected cutoff points was seen in 66 of 111 (59%) cases for p53, in 77 (69%) for p21, in 48 (43%) for p27, in 81 (73%) for cyclin D1, and in 54 (49%) cases for Ki67, respectively. None of the examined biomarkers was able to predict response to neoadjuvant chemoradiotherapy or was associated with survival outcome. Post-treatment pathologic TNM stage (P < 0.001), pathologic response (P < 0.001), and perineural invasion (P < 0.001) were the only factors having a significant effect on recurrence-free survival. Post-treatment pathologic N stage (P = 0.005), post-treatment pathologic TNM stage (P < 0.001), pathologic response (P < 0.001), and perineural invasion (P = 0.001) had a significant impact on overall survival. CONCLUSIONS: Our results suggest that the biomarkers p53, p21, p27, cyclin D1, and Ki67 have no impact on treatment response and survival in patients with oral and oropharyngeal cancer treated with preoperative chemoradiation.

Hiatal hernia, Barrett's esophagus, and long-term symptom control after laparoscopic fundoplication for gastroesophageal reflux.

BACKGROUND: The aim of this study was to determine the long-term symptom control after laparoscopic fundoplication for gastroesophageal reflux disease (GERD), and possible prognostic factors. METHODS: A cohort of 271 patients, operated on at a university hospital from 1996 through 2002, was eligible for evaluation after a median interval of 102 months (range = 12-158). The time between surgery and recurrence of reflux symptoms (i.e., time to treatment failure) served as the end point for statistical analysis. Putative risk factors for symptom recurrence were analyzed by univariate analysis and by using Cox's multiple-hazards regression. RESULTS: According to Kaplan-Meier estimates, the rate of reflux symptom recurrence was 15 % after 108 months, 11 % in cases without intestinal metaplasia, but 43 % in patients with long-segment (≥ 3 cm) Barrett's esophagus (BE; p < 0.0001). Reflux symptoms recurred in 22 % of cases with a hiatal hernia (HH) ≥ 3 cm before operation, but only in 7 % with smaller or absent HH (p = 0.005). Multivariate analysis revealed a relative risk of 6.6 (CI = 3.0-13.0) for long-segment BE and 3.0 (CI = 1.7-10.1) for HH ≥ 3 cm. A strong statistical interaction was found between HH ≥ 3 cm and long-segment BE: the small group (n = 18) of cases exhibiting both risk factors had an exaggerated recurrence rate of 72 % at 108 months. CONCLUSIONS: Laparoscopic fundoplication for symptomatic GERD provided a long-lasting abolition of reflux symptoms in 231 of 271 (85 %) patients. HH ≥ 3 cm and long-segment BE were shown as independent prognostic factors favoring recurrence.

HPV infection and p16 expression in carcinomas of the minor salivary glands.

The correlation between p16 upregulation and high-risk HPV infection is well known in head and neck squamous cell cancer. However, no information is available on rare carcinomas of the minor salivary glands. We analyzed 38 samples of minor salivary gland malignancies. p16 expression was determined by immunohistochemistry, HPV DNA by using in situ hybridization to detect low (type 6 and 11) and high (type 16 and 18) risk HPV types. In 71% of samples p16 expression was found. Low-risk HPV DNA could not be detected in any of the samples, whereas high-risk HPV DNA was found in two samples of mucoepidermoid carcinoma, which also showed the highest immunoreactivity with p16. HPV infection does not seem to be a common event in minor salivary gland carcinoma. Upregulation of p16 is present in the majority of cases but only diffuse staining correlates with HPV 16 and 18 infections.