Evaluation of the Integrated Pulmonary Index® during non-anesthesiologist sedation for percutaneous endoscopic gastrostomy.

Standard monitoring of heart rate, blood pressure and arterial oxygen saturation during endoscopy is recommended by current guidelines on procedural sedation. A number of studies indicated a reduction of hypoxic (art. oxygenation < 90% for > 15 s) and severe hypoxic events (art. oxygenation < 85%) by additional use of capnography. Therefore, U.S. and the European guidelines comment that additional capnography monitoring can be considered in long or deep sedation. Integrated Pulmonary Index® (IPI) is an algorithm-based monitoring parameter that combines oxygenation measured by pulse oximetry (art. oxygenation, heart rate) and ventilation measured by capnography (respiratory rate, apnea > 10 s, partial pressure of end-tidal carbon dioxide [PetCO2]). The aim of this paper was to analyze the value of IPI as parameter to monitor the respiratory status in patients receiving propofol sedation during PEG-procedure. Patients reporting for PEG-placement under sedation were randomized 1:1 in either standard monitoring group (SM) or capnography monitoring group including IPI (IM). Heart rate, blood pressure and arterial oxygen saturation were monitored in SM. In IM additional monitoring was performed measuring PetCO2, respiratory rate and IPI. Capnography and IPI values were recorded for all patients but were only visible to the endoscopic team for the IM-group. IPI values range between 1 and 10 (10 = normal; 8-9 = within normal range; 7 = close to normal range, requires attention; 5-6 = requires attention and may require intervention; 3-4 = requires intervention; 1-2 requires immediate intervention). Results on capnography versus standard monitoring of the same study population was published previously. A total of 147 patients (74 in SM and 73 in IM) were included in the present study. Hypoxic events occurred in 62 patients (42%) and severe hypoxic events in 44 patients (29%), respectively. Baseline characteristics were equally distributed in both groups. IPI = 1, IPI < 7 as well as the parameters PetCO2 = 0 mmHg and apnea > 10 s had a high sensitivity for hypoxic and severe hypoxic events, respectively (IPI = 1: 81%/81% [hypoxic/severe hypoxic event], IPI < 7: 82%/88%, PetCO2: 69%/68%, apnea > 10 s: 84%/84%). All four parameters had a low specificity for both hypoxic and severe hypoxic events (IPI = 1: 13%/12%, IPI < 7: 7%/7%, PetCO2: 29%/27%, apnea > 10 s: 7%/7%). In multivariate analysis, only SM and PetCO2 = 0 mmHg were independent risk factors for hypoxia. IPI (IPI = 1 and IPI < 7) as well as the individual parameters PetCO2 = 0 mmHg and apnea > 10 s allow a fast and convenient conclusion on patients' respiratory status in a morbid patient population. Sensitivity is good for most parameters, but specificity is poor. In conclusion, IPI can be a useful metric to assess respiratory status during propofol-sedation in PEG-placement. However, IPI was not superior to PetCO2 and apnea > 10 s.

Capnography monitoring of non-anesthesiologist provided sedation during percutaneous endoscopic gastrostomy placement: A prospective, controlled, randomized trial.

BACKGROUND AND AIM: A number of studies were able to show a reduction of hypoxemia episodes during procedural sedation through the use of capnography (CA). The present study investigates the number of episodes of hypoxemia during percutaneous endoscopic gastrostomy (PEG) placement with propofol sedation comparing standard monitoring (SM) versus SM with additional CA surveillance. METHODS: In this single center randomized controlled trial, 150 patients were prospectively randomized 1:1 in either the SM group or the CA group after stratification for ASA class, PEG method (push or pull method), presence of head and neck tumor, and tracheostomy. CA analysis was performed for all patients but was blinded for the endoscopic team in the SM group. RESULTS: In the SM group, 57% episodes of hypoxemia (SpO2  < 90% for > 15 s) and 41% episodes of severe hypoxemia (SpO2  < 85% for > 15 s) were observed in comparison with 28% and 20% in the CA group, respectively. Odds ratios for hypoxemia and severe hypoxemia were 0.29 (confidence interval 0.15-0.57; P = 0.0005) and 0.35 (confidence interval 0.17-0.73; P = 0.008) in favor of the CA group. On average, CA was able to detect imminent mild and severe hypoxemia 83 and 99 s before standard monitoring. Standard monitoring represented an independent risk factor for hypoxemia and severe hypoxemia. CONCLUSIONS: Respiratory complications of sedation during PEG placement are frequent events. CA is able to detect imminent hypoxemia at an early time point. This allows an early intervention and consecutively the avoidance of mild and severe hypoxemia. Therefore, CA monitoring can be recommended particularly during PEG insertion procedures.

Interferon-free antiviral treatment in B-cell lymphoproliferative disorders associated with hepatitis C virus infection.

Regression of hepatitis C virus (HCV)-associated lymphoma with interferon (IFN)-based antiviral treatment supports an etiological link between lymphoma and HCV infection. In addition, a favorable impact of antiviral treatment on overall survival of patients with HCV-related lymphoma has been reported. Data on IFN-free regimens combining direct-acting antivirals (DAAs) in HCV-associated lymphoproliferative disorders are scanty. We analyzed the virological and lymphoproliferative disease response (LDR) of 46 patients with indolent B-cell non-Hodgkin lymphomas (NHLs) or chronic lymphocytic leukemia (CLL) and chronic HCV infection treated with DAAs. The histological distribution was 37 marginal zone lymphomas (MZLs), 2 lymphoplasmacytic lymphomas, 2 follicular lymphomas, 4 CLL/small lymphocytic lymphomas (CLL/SLLs), and 1 low-grade NHL not otherwise specified. Thirty-nine patients received a sofosbuvir-based regimen and 7 patients received other DAAs. The median duration of DAA therapy was 12 weeks (range, 6-24 weeks). A sustained virological response at week 12 after finishing DAAs was obtained in 45 patients (98%); the overall LDR rate was 67%, including 12 patients (26%) who achieved a complete response. The LDR rate was 73% among patients with MZL, whereas no response was observed in CLL/SLL patients. Seven patients cleared cryoglobulins out of 15 who were initially positive. After a median follow-up of 8 months, 1-year progression-free and overall survival rates were 75% (95% confidence interval [CI], 51-88] and 98% [95% CI, 86-100], respectively. DAA therapy induces a high LDR rate in HCV-associated indolent lymphomas. These data provide a strong rationale for prospective trials with DAAs in this setting.

Microsatellite Instability Occurs Rarely in Patients with Cholangiocarcinoma: A Retrospective Study from a German Tertiary Care Hospital.

Immune-modulating therapy is a promising therapy for patients with cholangiocarcinoma (CCA). Microsatellite instability (MSI) might be a favorable predictor for treatment response, but comprehensive data on the prevalence of MSI in CCA are missing. The aim of the current study was to determine the prevalence of MSI in a German tertiary care hospital. Formalin-fixed paraffin-embedded tissue samples, obtained in the study period from 2007 to 2015 from patients with CCA undergoing surgical resection with curative intention at Johann Wolfgang Goethe University hospital, were examined. All samples were investigated immunohistochemically for the presence of MSI (expression of MLH1, PMS2, MSH2, and MSH6) as well as by pentaplex polymerase chain reaction for five quasimonomorphic mononucleotide repeats (BAT-25, BAT-26, NR-21, NR-22, and NR-24). In total, 102 patients were included, presenting intrahepatic (n = 35, 34.3%), perihilar (n = 42, 41.2%), and distal CCA (n = 25, 24.5%). In the immunohistochemical analysis, no loss of expression of DNA repair enzymes was observed. In the PCR-based analysis, one out of 102 patients was found to be MSI-high and one out of 102 was found to be MSI-low. Thus, MSI seems to appear rarely in CCA in Germany. This should be considered when planning immune-modulating therapy trials for patients with CCA.

PD-L1 expression in extrahepatic cholangiocarcinoma.

AIMS: To investigate the expression of the programmed cell death 1 (PD-1) receptor-programmed cell death ligand 1 (PD-L1) pathway and the clinicopathological characteristics in extrahepatic cholangiocarcinoma (eCCA). METHODS AND RESULTS: Tissue samples from patients with eCCA [n = 69; perihilar cholangiocarcinoma (CCA), 40; and distal CCA, 29] who underwent surgical resection in the period from 2007 to 2015 were evaluated for PD-1, PD-L1, CD3 and CD163 expression, and correlations with clinicopathological characteristics, including survival data, were determined. PD-L1 was found on both tumour cells of eCCA (8/69, 11.6%) and tumour-associated macrophages (21/69, 30.4%). Significant correlations of PD-L1 expression on cancer cells with venous invasion (P = 0.031) and poor differentiation of the tumour (P = 0.048) were observed. In 19 of 69 (27.5%) samples, tumour-infiltrating lymphocytes (TILs) expressed PD-1, whereas infiltration with CD3-positive and CD163-positive cells was found in 63 of 69 (91.3%) and 68 of 69 (98.6%) cases, respectively. The presence of fewer than five CD3-positive cells per high-power field was significantly correlated with poorer differentiation (P = 0.015) and venous invasion (P < 0.001) of CCA. PD-L1 expression was not correlated with patient survival, but PD-L1 expression on tumour cells combined with low infiltration of CD3-positive TILs was associated with an unfavourable outcome (P = 0.027). CONCLUSION: Only a small number of eCCA patients are PD-L1-positive, which might be important for future application of PD-1/PD-L1-targeted immune-modulating therapy in these patients. A small subgroup of eCCAs with PD-L1 expression and low lymphocytic infiltration showed more invasive growth and worse overall survival.

To treat or not to treat - Successful hepatitis C virus eradication in a patient with advanced hepatocellular carcinoma and complete response to sorafenib.

Background and aims Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide. Infection with the hepatitis C virus (HCV) is one of the most frequent underlying diseases leading to HCC development. Sorafenib is the standard of care for HCC patients not amenable to local treatment, resection, or liver transplantation. Although overall survival can be increased, objective response rates in patients treated with sorafenib are low. In HCC patients who underwent resection or ablation, HCV eradication with interferon-based regimens reduces the risk of recurrence. However, it is not known and under strong debate if patients with HCC should be treated with interferon-free regimens. Furthermore, it is not known if patients with advanced HCC at the time of diagnosis should be treated with antiviral therapy. Methods A patient with histologically confirmed advanced-stage HCC due to HCV-related cirrhosis was treated with sorafenib according to current guideline recommendations. Furthermore, he received subsequent treatment with direct antiviral agents (DAAs). Results The patient achieved a complete response after sorafenib treatment was initiated. Sorafenib treatment was terminated 1 year after complete response. As no recurrence of HCC was evident after treatment cessation, antiviral treatment was initiated with paritaprevir/ritonavir, ombitasvir, dasabuvir, and dose-reduced ribavirin because of chronic kidney disease. The patient achieved a sustained viral response. Conclusions Complete response to sorafenib treatment is scarce. Antiviral treatment should be considered in such patients as well as in patients with HCC who underwent resection or ablation.

Role of CD15 expression in dysplastic and neoplastic tissue of the bile duct - a potential novel tool for differential diagnosis of indeterminate biliary stricture.

BACKGROUND AND AIMS: CD15 is expressed by various cancer types; among these are intrahepatic and perihilar cholangiocarcinoma (CCA). The aim of this study was to elucidate CD15 expression in distal CCA as well as in dysplastic biliary tissue and to determine its prognostic significance. METHODS AND RESULTS: Tissue samples from patients with intrahepatic (iCCA, n = 22), perihilar (pCCA, n = 7) and distal CCA (dCCA, n = 15), who underwent surgical resection in the period from 2010 to 2015 were evaluated for CD15 expression. Tissue of synchronous lymph node metastasis (n = 13), CCA-associated dysplasia (n = 20), dysplasia in intraductal biopsies (n = 10) and benign proliferations (n = 12), as well as inflammatory biliary lesions (n = 28) and non-inflammatory bile ducts (n = 23), were evaluated equally for CD15 expression. CD15 was found to be expressed highly in iCCA (81.8%), pCCA (85.7%), dCCA (73.3%), CCA-associated dysplasia (70.0%), dysplasia in intraductal biopsies (100%) and metastatic tissue (84.6%). CD15 expression was negative in 58 of 64 benign bile duct alterations resulting in an overall sensitivity and specificity of CD15 in CCA of 80.7 and 90.6% patients, respectively. CD15 expression was correlated significantly with a decreased overall survival in patients with CD15-positive CCA associated dysplasia (P = 0.003). However, CD15 expression in the invasive tumour component was not correlated with clinical outcome. CONCLUSION: CD15 is a sensitive and specific marker for intraepithelial and invasive neoplasias of the bile duct. Therefore, it can be helpful in the delineation of dysplastic and neoplastic biliary cells from non-neoplastic tissue, which frequently causes a diagnostic problem in indeterminate biliary stricture.

A possible role of microRNAs as predictive markers for the recurrence of hepatocellular carcinoma after liver transplantation.

With favourable 5-year survival rates up to 75%, liver transplantation (LT) is the treatment of choice for hepatocellular carcinoma (HCC). Nonetheless, tumour recurrence after LT remains a challenge. The aim of this retrospective study was to develop a predictive score for tumour recurrence after LT by combining clinical parameters with HCC biomarkers (microRNA). A microRNA (miRNA) microarray analysis was used to compare miRNA expression patterns in tissue samples of 40 patients with and without HCC recurrence after LT. In a screening cohort (n = 18), the miRNA analysis identified significant differences in the expression of 13 miRNAs in patients with tumour recurrence. Using the most significant miRNAs in this screening cohort, we could develop a predictive score, which combined the expression levels of miR-214, miR-3187 and the Milan criteria, and we could define low- and high-risk groups for tumour recurrence and death. The above score was evaluated in a second and independent cohort (n = 22). In contrast to the Milan criteria alone, this score was significantly associated with tumour recurrence. Our analysis indicated that the use of a specific miRNA expression pattern in combination with a limited tumour burden as defined by the Milan criteria may lead to a more accurate prediction of tumour recurrence.

Whole exome sequencing of microdissected splenic marginal zone lymphoma: a study to discover novel tumor-specific mutations.

BACKGROUND: Splenic marginal zone lymphoma (SMZL) is an indolent B-cell non-Hodgkin lymphoma and represents the most common primary malignancy of the spleen. Its precise molecular pathogenesis is still unknown and specific molecular markers for diagnosis or possible targets for causal therapies are lacking. METHODS: We performed whole exome sequencing (WES) and copy number analysis from laser-microdissected tumor cells of two primary SMZL discovery cases. Selected somatic single nucleotide variants (SNVs) were analyzed using pyrosequencing and Sanger sequencing in an independent validation cohort. RESULTS: Overall, 25 nonsynonymous somatic SNVs were identified, including known mutations in the NOTCH2 and MYD88 genes. Twenty-three of the mutations have not been associated with SMZL before. Many of these seem to be subclonal. Screening of 24 additional SMZL for mutations at the same positions found mutated in the WES approach revealed no recurrence of mutations for ZNF608 and PDE10A, whereas the MYD88 L265P missense mutation was identified in 15% of cases. An analysis of the NOTCH2 PEST domain and the whole coding region of the transcription factor SMYD1 in eight cases identified no additional case with a NOTCH2 mutation, but two additional cases with SMYD1 alterations. CONCLUSIONS: In this first WES approach from microdissected SMZL tissue we confirmed known mutations and discovered new somatic variants. Recurrence of MYD88 mutations in SMZL was validated, but NOTCH2 PEST domain mutations were relatively rare (10 % of cases). Recurrent mutations in the transcription factor SMYD1 have not been described in SMZL before and warrant further investigation.

Feasibility of global miRNA analysis from fine-needle biopsy FFPE material in patients with hepatocellular carcinoma treated with sorafenib.

Sorafenib is the standard treatment for patients with advanced hepatocellular carcinoma (HCC). However, the median overall survival (OS) benefit is only ~3 months, and sufficient biomarkers predicting treatment response are not available. The aim of the present study was to evaluate miRNA expression patterns from HCC tissue biopsies as potential biomarkers in patients under sorafenib treatment. Nineteen patients with advanced HCC treated with sorafenib were included. RNA was extracted from formalin-fixed paraffin-embedded (FFPE) liver biopsies. miRNA expression profiling of 818 mature miRNAs was performed using GeneChip® miRNA Array 2.0 (Affymetrix). Global miRNA patterns were assessed using unsupervised hierarchical clustering analysis (UCA), and specific miRNAs with correlation with disease control rate (DCR) or good OS were evaluated by pairwise supervised analyses. UCA divided the patients into three distinct groups by their miRNA expression patterns. However, DCR or OS did not correlate with these sub-groups. We have identified several miRNAs that correlated with either DCR or OS (P<0.05). However, with correction for multiple testing, these results did not reach statistical significance in this small cohort. Global miRNA analysis from very low input RNA deriving from liver biopsies showed distinctive clustering of molecular sub-groups in patients with intermediate and advanced HCC. Clinical response including OS under sorafenib did not correlate with global miRNA expression patterns, but we have identified candidate miRNAs for the prediction of DCR and OS to be evaluated in prospective studies and larger patient cohorts.

Hepatitis C-associated B-cell non-Hodgkin lymphomas. Epidemiology, molecular signature and clinical management.

There is ample epidemiologic evidence for an association of chronic hepatitis C virus (HCV) infection with B-cell non-Hodgkin lymphoma (B-NHL). B-NHL subtypes most frequently associated with HCV are marginal zone lymphoma and diffuse large B-cell lymphoma. The most convincing evidence for a causal relationship between HCV infection and lymphoma development is the observation of B-NHL regression after HCV eradication by antiviral therapy (AVT). In fact, for indolent HCV-associated B-NHL, first-line AVT instead of standard immune-chemotherapy might be considered. Molecular mechanisms of HCV-NHL development are still poorly understood. Three general theories have emerged to understand the HCV-induced lymphomagenesis: (1) continuous external stimulation of lymphocyte receptors by viral antigens and consecutive proliferation; (2) HCV replication in B cells with oncogenic effect mediated by intracellular viral proteins; (3) permanent B-cell damage, e.g., mutation of tumor suppressor genes, caused by a transiently intracellular virus ("hit and run" theory). This review systematically summarizes the data on epidemiology, interventional studies, and molecular mechanisms of HCV-associated B-NHL.

Bile duct tissue acquisition by cholangioscopy-guided cryobiopsy technique: first-in-human application.

Video 1Case description and video in cholangioscopic view demonstrating bile duct tissue acquisition by cholangioscopy-guided cryobiopsy technique.

A new tool for bile duct tissue sampling: ex vivo clinical evaluation of intraductal cryobiopsy for cholangioscopy.

Background and study aims Indeterminate biliary strictures represent a major challenge in clinical diagnostics. Diagnostic yield of radiological, endoscopic imaging and histopathological diagnosis is insufficient. The cryobiopsy technique is a new method for tissue extraction already used in different clinical settings. The aim of this ex vivo clinical study was to investigate feasibility and tissue quality of cryobiopsy in the bile duct. Patients and methods We included 14 patients who underwent pancreaticoduodenectomy. Bile duct samples were taken with either a new prototype cryoprobe or one of two forceps types. Results were analyzed for general feasibility, specimen size, histological assessability as well as representativity of retrieved tissue. Results Feasibility of cholangioscopic forceps was poor compared to gastric biopsy forceps or cryobiopsy. Significantly larger tissue samples were obtained with cryobiopsy (5.6 ±â€Š4.5 mm 2 ) compared to gastric biopsy forceps (3.3 ±â€Š5.1 mm 2 , P  = 0.006). Furthermore, cryobiopsy was superior in histological assessment quality ( P  = 0.02) and concerning representativity ( P  = 0.03). Conclusions Cryobiopsy in the bile duct is feasible and the quality of the obtained tissue is high. Further investigation of bile duct cryobiopsy in vivo is warranted.

Serum miR-122 as a biomarker of necroinflammation in patients with chronic hepatitis C virus infection.

OBJECTIVES: The liver contains large amounts of microRNA-122 (miR-122), whereas other tissues contain only marginal amounts of this miRNA. MicroRNAs have also been found to circulate in the blood in a cell-free form; their potential as readily accessible disease markers is currently evaluated. Here, we investigated if the serum levels of miR-122 might be useful as disease parameter in patients with chronic hepatitis C virus (HCV) infection. METHODS: RNA was extracted from sera of patients with chronic HCV infection (CHC) and healthy controls and was analyzed for miR-22 content by quantitative real-time reverse-transcription polymerase chain reaction. miR-122 serum levels were correlated with standard parameters of liver function. Liver biopsies from the same patients were examined for the histologic activity index (HAI) and the degree of fibrosis. RESULTS: Sera from patients with CHC contained higher levels of miR-122 than sera from healthy controls. Serum miR-122 levels correlated well with markers of liver inflammatory activity, that is, the serum levels of alanine leucine transaminase (ALT) and aspartate transaminase, and the HAI score. In patients with persistently normal ALT levels, serum miR-122 levels did not differ from healthy controls. There was no correlation of serum miR-122 levels with serum albumin, international normalized ratio, liver fibrosis, or serum HCV RNA. CONCLUSIONS: The serum level of miR-122 strongly correlates with serum ALT activity and with necroinflammatory activity in patients with CHC and elevated ALT levels, but not with fibrosis stage and functional capacity of the liver.

Antiviral therapy of chronic hepatitis C in patients with advanced liver disease and after liver transplantation.

Chronic infection with the hepatitis C virus (HCV) represents one of the major causes for end-stage liver disease worldwide. Although liver transplantation offers an effective treatment, HCV reinfection of the transplanted graft is a critical and almost inevitable complication with major influence on graft- and patient survival. Pre-transplant antiviral therapy in advanced liver disease is limited by poor tolerance and only applicable to mildly decompensated patients but was able to show promising results in patients reaching negative viral load when undergoing transplantation. Prophylactic therapy with HCV antibodies during the anhepatic phase has not been shown to be effective in studies to date. Antiviral therapy after transplantation but before evidence of reinfection, so called pre-emptive treatment, is limited by frequent complications and a high rate of side effects. The mainstay of management represents directed antiviral therapy after evidence of recurrence of chronic Hepatitis C. With a combination therapy of pegylated interferon and ribavirin, sustained virologic response rates of 25-45% are achieved. However, tolerability is often poor, and the need of dose reduction is frequent. To date, there is no general consensus on modality, timing and dosing of antiviral treatment of HCV in patients with advanced liver disease and after liver transplantation. More randomised, controlled trials are needed. Moreover, upcoming new treatment approaches, e.g. specifically targeted antiviral therapy for hepatitis C (STAT-C) with HCV-specific polymerase and protease inhibitors, may represent a therapeutic alternative.

miRNA-26b downregulation in peripheral blood mononuclear cells of patients with hepatitis C associated lymphomas is restored by successful interferon-free antiviral therapy.

BACKGROUND: Patients with chronic HCV infection are at increased risk of developing B-cell non-Hodgkin lymphoma (B-NHL). Regression of HCV-associated B-NHL (HCV-NHL) can be achieved through HCV eradication using interferon (IFN). However, only about two-thirds of patients with sustained virological response (SVR) also had a consecutive lymphoma response. miRNA-26b is associated with HCV-NHL response to antiviral therapy. Recent data suggest that IFN-free direct-acting antiviral (DAA) regimens also have anti-lymphoma activity in this patient population. METHODS: We report four patients with HCV-NHL who were treated with different IFN-free DAA regimens as oncological monotherapy in our centre between 2015 and 2016. We analysed the virological and lymphoproliferative disease response. Moreover, we analysed miRNA-26b expression in peripheral blood mononuclear cells at different time points during antiviral therapy for all included patients as well as for a total of 10 controls with (n=5) and without (n=5) chronic HCV infection. RESULTS: All patients had marginal zone lymphoma subtype and received different DAA regimens for 12-24 weeks. All four patients achieved SVR, but only three patients also had lymphoma response (one complete response, two partial responses). One patient showed progression to a high-grade lymphoma subtype after SVR. miRNA-26b expression was generally decreased in patients with HCV-NHL. Moreover, miRNA-26b expression was restored in those HCV-NHL patients with lymphoma response after 6 months (P=0.009). CONCLUSIONS: We have demonstrated that IFN-free DAA treatment of HCV can improve or even cure NHL. miRNA-26b-levels could be a potentially useful biomarker to predict lymphoma response in HCV-NHL patients.

Genetic heterogeneity of primary lesion and metastasis in small intestine neuroendocrine tumors.

Data on intratumoral heterogeneity of small intestine neuroendocrine tumors (SI-NETs) and related liver metastasis are limited. The aim of this study was to characterize genetic heterogeneity of 5 patients with SI-NETs. Therefore, formalin-fixed, paraffin-embedded tissue samples of primary and metastatic lesions as well as benign liver of five patients with synchronously metastasized, well differentiated SI-NETs were analyzed with whole exome sequencing. For one patient, chip based 850k whole DNA methylome analysis was performed of primary and metastatic tumor tissue as well as control tissue. Thereby, 156 single nucleotide variants (SNVs) in 150 genes were identified and amount of mutations per sample ranged from 9-34 (mean 22). The degree of common (0-94%) and private mutations per sample was strongly varying (6-100%). In all patients, copy number variations (CNV) were found and the degree of intratumoral heterogeneity of CNVs corresponded to SNV analysis. DNA methylation analysis of a patient without common SNVs revealed a large overlap of common methylated CpG sites. In conclusion, SI-NET primary and metastatic lesions show a highly varying degree of intratumoral heterogeneity. Driver events might not be detectable with exome analysis only, and further comprehensive studies including whole genome and epigenetic analyses are warranted.

Sequencing of intraductal biopsies is feasible and potentially impacts clinical management of patients with indeterminate biliary stricture and cholangiocarcinoma.

BACKGROUND: Definite diagnosis and therapeutic management of cholangiocarcinoma (CCA) remains a challenge. The aim of the current study was to investigate feasibility and potential impact on clinical management of targeted sequencing of intraductal biopsies. METHODS: Intraductal biopsies with suspicious findings from 16 patients with CCA in later clinical course were analyzed with targeted sequencing including tumor and control benign tissue (n = 55 samples). A CCA-specific sequencing panel containing 41 genes was designed and a dual strand targeted enrichment was applied. RESULTS: Sequencing was successfully performed for all samples. In total, 79 mutations were identified and a mean of 1.7 mutations per tumor sample (range 0-4) as well as 2.3 per biopsy (0-6) were detected and potentially therapeutically relevant genes were identified in 6/16 cases. In 14/18 (78%) biopsies with dysplasia or inconclusive findings at least one mutation was detected. The majority of mutations were found in both surgical specimen and biopsy (68%), while 28% were only present in biopsies in contrast to 4% being only present in the surgical tumor specimen. CONCLUSION: Targeted sequencing from intraductal biopsies is feasible and potentially improves the diagnostic yield. A profound genetic heterogeneity in biliary dysplasia needs to be considered in clinical management and warrants further investigation. TRANSLATIONAL IMPACT: The current study is the first to demonstrate the feasibility of sequencing of intraductal biopsies which holds the potential to impact diagnostic and therapeutical management of patients with biliary dysplasia and neoplasia.

Eosinophilic cholangitis is a potentially underdiagnosed etiology in indeterminate biliary stricture.

AIM: To investigate presence and extent of eosinophilic cholangitis (EC) as well as IgG4-related disease in patients with indeterminate biliary stricture (IBS). METHODS: All patients with diagnosis of sclerosing cholangitis (SC) and histopathological samples such as biopsies or surgical specimens at University Hospital Frankfurt from 2005-2015 were included. Histopathological diagnoses as well as further clinical course were reviewed. Tissue samples of patients without definite diagnosis after complete diagnostic work-up were reviewed regarding presence of eosinophilic infiltration and IgG4 positive plasma cells. Eosinophilic infiltration was as well assessed in a control group of liver transplant donors and patients with primary sclerosing cholangitis. RESULTS: one hundred and thirty-five patients with SC were included. In 10/135 (13.5%) patients, no potential cause of IBS could be identified after complete diagnostic work-up and further clinical course. After histopathological review, a post-hoc diagnosis of EC was established in three patients resulting in a prevalence of 2.2% (3/135) of all patients with SC as well as 30% (3/10) of patients, where no cause of IBS was identified. 2/3 patients with post-hoc diagnosis of EC underwent surgical resection with suspicion for malignancy. Diagnosis of IgG4-related cholangitis was observed in 7/135 patients (5.1%), whereas 3 cases were discovered in post-hoc analysis. 6/7 cases with IgG4-related cholangitis (85.7%) presented with eosinophilic infiltration in addition to IgG4 positive plasma cells. There was no patient with eosinophilic infiltration in the control group of liver transplant donors (n = 27) and patients with primary sclerosing cholangitis (n = 14). CONCLUSION: EC is an underdiagnosed benign etiology of SC and IBS, which has to be considered in differential diagnosis of IBS.