Search

Background The clinical relevance of hyperamylasemia after distal pancreatectomy (DP) remains unclear and no internationally accepted definition of postoperative acute pancreatitis (POAP) exists. The aim of this study was to characterize POAP after DP and to assess the role of serum amylase (SA) in POAP. Methods Outcomes of 641 patients who had undergone DP between 2015 and 2019 were analyzed. Postoperative SA was determined in all patients. POAP was defined based on contrast-enhanced computed tomography (CT) or intraoperative findings during relaparotomy. Results An elevation of SA on postoperative day 1 (hyperamylasemiaPOD1) was found in 398 patients (62.1%). Twelve patients (1.87%) were identified with POAP. Ten patients demonstrated radiologic criteria for POAP and in two patients POAP was diagnosed during relaparotomy. Outcome of POAP patients was worse than that of patients with hyperamylasemiaPOD1 alone and that with normal SAPOD1 without POAP evidence (postoperative pancreatic fistula 50% vs 30.6% vs 18.5%; length of hospital stay 26 days vs 12 vs 11, respectively). The overall 90-day mortality of all 641 patients was 0.6%. Conclusion POAP is a serious but rare complication after DP. HyperamylasemiaPOD1 is of prognostic relevance after DP, but it seems not sufficient as a single parameter to diagnose POAP.

Radiological reports of pancreatic lesions are currently widely formulated as free texts. However, for optimal characterization, staging and operation planning, a wide range of information is required but is sometimes not captured comprehensively. Structured reporting offers the potential for improvement in terms of completeness, reproducibility and clarity of interdisciplinary communication. Interdisciplinary consensus finding of structured report templates for solid and cystic pancreatic tumors in computed tomography (CT) and magnetic resonance imaging (MRI) with representatives of the German Society of Radiology (DRG), German Society for General and Visceral Surgery (DGAV), working group Oncological Imaging (ABO) of the German Cancer Society (DKG) and other radiologists, oncologists and surgeons. Among experts in the field of pancreatic imaging, oncology and pancreatic surgery, as well as in a public online survey, structured report templates were developed by consensus. These templates are available on the DRG homepage under www.befundung.drg.de and will be regularly revised to the current state of scientific knowledge by the participating specialist societies and responsible working groups. This article presents structured report templates for solid and cystic pancreatic tumors to improve clinical staging (cTNM, ycTNM) in everyday radiology. · Structured report templates offer the potential of optimized radiological reporting with regard to completeness, reproducibility and differential diagnosis.. · This article presents consensus-based, structured reports for solid and cystic pancreatic lesions in CT and MRI.. · These structured reports are available open source on the homepage of the German Society of Radiology (DRG) under www.befundung.drg.de..· Persigehl T, Baumhauer M, Baeßler B et al. Structured Reporting of Solid and Cystic Pancreatic Lesions in CT and MRI: Consensus-Based Structured Report Templates of the German Society of Radiology (DRG). Fortschr Röntgenstr 2020; 192: 641 - 655. In der klinischen Radiologie werden aktuell radiologische Befundberichte bei Pankreasläsionen weitverbreitet als Freitexte formuliert. Für die optimale klinische Versorgung, das Staging und die Operationsplanung sind mannigfaltige Informationen erforderlich, die mitunter nicht umfänglich erfasst werden. Die strukturierte Befundung bietet hier das Potenzial einer Verbesserung in Bezug auf Vollständigkeit, Reproduzierbarkeit sowie Klarheit der interdisziplinären Kommunikation. Interdisziplinäre Erstellung von strukturierten Befundvorlagen für solide und zystische Pankreastumoren in der Computertomografie (CT) und in der Magnetresonanztomografie (MRT) mit Vertretern der Deutschen Röntgengesellschaft (DRG), der Deutschen Gesellschaft für Allgemein- und Viszeralchirurgie (DGAV), der Arbeitsgemeinschaft Onkologische Bildgebung (ABO) der Deutschen Krebsgesellschaft (DKG) sowie weiteren Radiologen, Onkologen und Chirurgen. Unter Experten auf dem Gebiet der Pankreasbildgebung, onkologischen Versorgung und Pankreaschirurgie sowie in einer öffentlichen Online-Umfrage wurden strukturierte Befundvorlagen im Konsensus-Verfahren entwickelt, die im weiteren Verlauf regelmäßig durch die beteiligten Fachgesellschaften und verantwortlichen Arbeitsgemeinschaften auf den aktuellen Stand der wissenschaftlichen Erkenntnis angepasst werden sollen. Dieser Artikel stellt strukturierte Befundvorlagen für solide und zystische Pankreastumoren vor, die im radiologischen Alltag helfen sollen, das klinische Staging (cTNM, ycTNM) zu verbessern. · Strukturierte Befundvorlagen bieten das Potenzial einer optimierten radiologischen Befundung in Bezug auf Vollständigkeit, Reproduzierbarkeit und Differenzialdiagnose.. · Dieser Artikel stellt Konsensus-basierte, strukturierte Befundvorlagen für solide und zystische Pankreasläsionen im CT und MRT vor.. · Die von der Deutsche Röntgengesellschaft entwickelten Befundvorlagen werden unter www.befundung.drg.de zur Verfügung gestellt..· Persigehl T, Baumhauer M, Baeßler B et al. Structured Reporting of Solid and Cystic Pancreatic Lesions in CT and MRI: Consensus-Based Structured Report Templates of the German Society of Radiology (DRG). Fortschr Röntgenstr 2020; 192: 641 – 656.

Esophageal and Gastric Adenocarcinomas (AGE/S) are characterized by early metastasis and poor survival. MACC1 (Metastasis Associated in Colon Cancer 1) acts in colon cancer as a metastasis inducer and is linked to reduced survival. This project illuminates the role and potential for the inhibition of MACC1 in AGE/S. Using 266 of 360 TMAs and survival data of AGE/S patients, we confirm the value of MACC1 as an independent negative prognostic marker in AGE/S patients. MACC1 gene expression is correlated with survival and morphological characteristics. In vitro analysis of lentivirally MACC1-manipulated subclones of FLO-1 and OE33 showed enhanced migration induced by MACC1 in both cell line models, which could be inhibited by the MEK1 inhibitor selumetinib. In vivo, the efficacy of selumetinib on tumor growths and metastases of MACC1-overexpressing FLO-1 cells xenografted intrasplenically in NOG mice was tested. Mice with high-MACC1-expressing cells developed faster and larger distant metastases. Treatment with selumetinib led to a significant reduction in metastasis exclusively in the MACC1-positive xenografts. MACC1 is an enhancer of tumor aggressiveness and a predictor of poor survival in AGE/S. This effect can be inhibited by selumetinib.

Background Tumor mutational burden (TMB) is an emerging biomarker used to identify patients who are more likely to benefit from immuno-oncology therapy. Aside from various unsettled technical aspects, biological variables such as tumor cell content and intratumor heterogeneity may play an important role in determining TMB. Methods TMB estimates were determined applying the TruSight Oncology 500 targeted sequencing panel. Spatial and temporal heterogeneity was analyzed by multiregion sequencing (two to six samples) of 24 pulmonary adenocarcinomas and by sequencing a set of matched primary tumors, locoregional lymph node metastases, and distant metastases in five patients. Results On average, a coding region of 1.28 Mbp was covered with a mean read depth of 609x. Manual validation of the mutation-calls confirmed a good performance, but revealed noticeable misclassification during germline filtering. Different regions within a tumor showed considerable spatial TMB variance in 30% (7 of 24) of the cases (maximum difference, 14.13 mut/Mbp). Lymph node–derived TMB was significantly lower (p = 0.016). In 13 cases, distinct mutational profiles were exclusive to different regions of a tumor, leading to higher values for simulated aggregated TMB. Combined, intratumor heterogeneity and the aggregated TMB could result in divergent TMB designation in 17% of the analyzed patients. TMB variation between primary tumor and distant metastases existed but was not profound. Conclusions Our data show that, in addition to technical aspects such as germline filtering, the tumor content and spatially divergent mutational profiles within a tumor are relevant factors influencing TMB estimation, revealing limitations of single-sample–based TMB estimations in a clinical context.

Tyrosine kinase inhibitors currently confer the greatest survival gain for nonsmall cell lung cancer (NSCLC) patients with actionable genetic alterations. Simultaneously, the increasing number of targets and compounds poses the challenge of reliable, broad and timely molecular assays for the identification of patients likely to benefit from novel treatments. Here, we demonstrate the feasibility and clinical utility of comprehensive, NGS-based genetic profiling for routine workup of advanced NSCLC based on the first 3,000 patients analyzed in our department. Following automated extraction of DNA and RNA from formalin-fixed, paraffin-embedded tissue samples, parallel sequencing of DNA and RNA for detection of mutations and gene fusions, respectively, was performed using PCR-based enrichment with an ion semiconductor sequencing platform. Overall, 807 patients (27%) were eligible for currently approved, EGFR-/BRAF-/ALK- and ROS1-directed therapies, while 218 additional cases (7%) with MET, ERBB2 (HER2) and RET alterations could potentially benefit from experimental targeted compounds. In addition, routine capturing of comutations, e.g. TP53 (55%), KEAP1 (11%) and STK11 (11%), as well as the precise typing of fusion partners and involved exons in case of actionable translocations including ALK and ROS1, are prognostic and predictive tools currently gaining importance for further refinement of therapeutic and surveillance strategies. The reliability, low dropout rates (

Biphenotypic sinonasal sarcoma (BSNS) is a rare, low grade spindle cell sarcoma, recently recognized in the WHO classification of head and neck tumors, which is characterized by a dual myogenic and neural differentiation and recurrent gene fusions, often involving PAX3-MAML3, and less commonly PAX3 fusions with other partners such as NCOA1, NCOA2, or WWTR1. Yet, in about 4% of tumors no gene rearrangements are identified. Herein, we describe a RREB1-MKL2 fusion in a BSNS lesion occurring in a 73-year-old female patient with a right maxillo-ethmoidal angle lesion. The polypoid, moderately cellular tumor with infiltrative submucosal growth was composed of fascicles of relatively bland spindle cells embedded in a loose collagenous matrix. The tumor cells showed moderate amounts of eosinophilic cytoplasm with indistinct borders and uniform, pale, ovoid to slender nuclei. The slowly proliferating neoplastic cells co-expressed smooth muscle actin and S100, and showed focal nuclear positivity for s-catenin, while lacking staining for cytokeratins, desmin, myogenin, caldesmon, glial fibrillary acid protein, and SOX-10. Molecular analysis by targeted RNA-based next-generation sequencing identified an in-frame fusion between exon 8 of RREB1 and exon 11 of MKL2, a genetic event that was reported to be a molecular hallmark of ectomesenchymal chondromyxoid tumor. Gene rearrangements in both genes were independently verified by fluorescence in situ hybridization (FISH). To evaluate its recurrent potential an additional group of 15 fusion negative BSNS were tested for abnormalities in RREB1 and MKL2 genes by FISH, but no additional positive cases were identified. This article is protected by copyright. All rights reserved.

Background Synaptophysin, chromogranin and CD56 are recommended markers to identify pulmonary tumors with neuroendocrine differentiation. Whether the expression of these markers in pulmonary adenocarcinoma and pulmonary squamous cell carcinoma is a prognostic factor has been a matter of debate. Therefore, we investigated retrospectively a large cohort to expand the data on the role of synaptophysin, chromogranin and CD56 in non-small cell lung cancer lacking morphological features of neuroendocrine differentiation. Methods A cohort of 627 pulmonary adenocarcinomas (ADC) and 543 squamous cell carcinomas (SqCC) lacking morphological features of neuroendocrine differentiation was assembled and a tissue microarray was constructed. All cases were stained with synaptophysin, chromogranin and CD56. Positivity was defined as > 1% positive tumor cells. Data was correlated with clinico-pathological features including overall and disease free survival. Results 110 (18%) ADC and 80 (15%) SqCC were positive for either synaptophysin, chromogranin, CD56 or a combination. The most commonly positive single marker was synaptophysin. The least common positive marker was chromogranin. A combination of ≤2 neuroendocrine markers was positive in 2–3% of ADC and 0–1% of SqCC. There was no significant difference in overall survival in tumors with positivity for neuroendocrine markers neither in ADC (univariate: P = 0.4; hazard ratio [HR] = 0.867; multivariate: P = 0.5; HR = 0.876) nor in SqCC (univariate: P = 0.1; HR = 0.694; multivariate: P = 0.1, HR = 0.697). Likewise, there was no significant difference in disease free survival. Conclusions We report on a cohort of 1170 cases that synaptophysin, chromogranin and CD56 are commonly expressed in ADC and SqCC and that their expression has no impact on survival, supporting the current best practice guidelines.

BRAF V600E mutations have been reported as a marker of sporadic microsatellite instability (MSI) colorectal cancer (CRC). Current international diagnostic guidelines recommend BRAF mutation testing in MSI CRC patients to predict low risk of Lynch syndrome (LS). We evaluated the age-specific performance of BRAF testing in LS diagnostics. We systematically compared the prevalence of BRAF mutations in LS-associated CRCs and unselected MSI CRCs in different age groups as available from published studies, databases and population-based patient cohorts. Sensitivity/specificity analysis of BRAF testing for exclusion of LS and cost calculations were performed. Among 969 MSI CRCs from LS carriers in the literature and German HNPCC Consortium, 15 (1.6%) harbored BRAF mutations. Six of seven LS patients with BRAF-mutant CRC and reported age were

AIMS Non-small-cell lung cancer (NSCLC) and breast cancer are common entities. Staining for oestrogen receptor (ER), progesterone receptor (PgR), mammaglobin (MAMG) and GATA-binding protein 3 (GATA3) is frequently performed to confirm a mammary origin in the appropriate diagnostic setting. However, comprehensive data on the immunohistological expression of these markers in NSCLC are limited. Therefore, the aim of this study was to analyse a large cohort of NSCLCs and correlate the staining results with clinicopathological variables. METHODS AND RESULTS A tissue microarray was stained for ER, PgR, MAMG, human epidermal growth factor receptor 2 (HER2), and GATA3, and included 636 adenocarcinomas (ADCs), 536 squamous cell carcinomas (SqCCs), 65 large-cell-carcinomas, 34 pleomorphic carcinomas, and 20 large-cell neuroendocrine carcinomas. HER2 status was determined for immunohistochemically positive cases with chromogenic in-situ hybridisation. Markers with a proportion of ≥5% positive cases in ADC and SqCC were considered for survival analysis. Among ADCs, 62 (10%), 17 (3%), one (

Tumor mutational burden (TMB) represents a new determinant of clinical benefit from immune checkpoint blockade that identifies responders independent of PD-L1 expression levels and is currently being explored in clinical trials. Although TMB can be measured directly by comprehensive genomic approaches such as whole-genome and exome sequencing, broad availability, short turnaround times, costs and amenability to formalin-fixed and paraffin-embedded tissue support the use of gene panel sequencing for approximating TMB in routine diagnostics. However, data on the parameters influencing panel-based TMB estimation are limited. Here, we report an extensive in silico analysis of the TCGA data set that simulates various panel sizes and compositions. We demonstrate that panel size is a critical parameter that influences confidence intervals (CIs) and cutoff values as well as important test parameters including sensitivity, specificity, and positive predictive value. Moreover, we evaluate the Illumina TSO500 panel, which will be made available for TMB estimation, and propose dynamic, entity-specific cutoff values based on current clinical trial data. Optimizing the cost-benefit ratio, our data suggest that panels between 1.5 and 3 Mbp are ideally suited to estimate TMB with small CIs, whereas smaller panels tend to deliver imprecise TMB estimates for low to moderate TMB (0-30 muts/Mbp), connected with insufficient separation of hypermutated tumors from non-hypermutated tumors.

// Heike Loeser 1, *, ** , Simon Schallenberg 1, *, ** , Moritz von Winterfeld 1 , Lars Tharun 1 , Hakan Alakus 3, * , Arnulf Holscher 4 , Elfriede Bollschweiler 3 , Reinhard Buettner 1 , Thomas Zander 2, *, ** and Alexander Quaas 1, *, ** 1 Institute of Pathology, University of Cologne, Cologne, Germany 2 Department I of Internal Medicine, Center for Integrated Oncology (CIO), University of Cologne, Cologne, Germany 3 Department of General, Visceral and Cancer Surgery, University of Cologne, Cologne, Germany 4 Department of Thorax and Oesophageal Surgery, Agaplesion Markus Krankenhaus, Frankfurt/Main, Germany * Gastrointestinal Cancer Group Cologne (GCGC) ** These authors have contributed equally to this work Correspondence to: Alexander Quaas, email: alexander.quaas@uk-koeln.de Keywords: esophageal adenocarcinoma; TUBB3; RNA-in-situ-hybridization; immunohistochemistry Received: May 10, 2017 Accepted: November 20, 2017 Published: December 11, 2017 ABSTRACT Background: Esophageal adenocarcinomas show an increasing incidence in the Western world and their overall survival remains low. Microtubules are multifunctional cytoskeletal proteins involved in crucial cellular roles, including maintenance of cell shape, intracellular transport, meiosis, and mitosis. Microtubulus-TUBB3 was found overexpressed in several carcinomas suggesting a significant role in cancer development. High levels of TUBB3 expression were also described to be associated with poor clinical outcome in various cancers. It was shown that overexpression of TUBB3 could be related to reduced efficiency of taxane-based targeting anticancer drugs in several cancer types. Results: There is a statistically significant association between high TUBB3 protein and TUBB3 mRNA expression and shortened survival (p

This study investigated the correlation between the peritoneal carcinomatosis index (PCI) and patient outcome depending on the tumour type.Peritoneal surface malignancy (PSM) treatment depends on tumour type. Mucinous PSM (m-PSM) is associated with a better prognosis than non-mucinous PSM (nm-PSM). The PCI's predictive ability has not yet been evaluated.We analysed 123 patients with PSM treated with cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) between 2008 and 2015. The m-PSM group (n = 75) included patients with appendiceal cancer (n = 15), colorectal cancer (n = 21), or low-grade appendiceal mucinous neoplasm (n = 39); the nm-PSM group (n = 48) included patients with gastric (n = 18) or colorectal (n = 30) cancer. The PCI's predictive ability was evaluated by multiple Cox-proportional hazard regression analysis and Kaplan-Meier curves.The 5-year survival and PCI were higher in m-PSM patients (67.0%; 20.5 ± 12.1) than in nm-PSM patients (32.6%; p = 0.013; 8.9 ± 6.0; p 0.001). Colorectal nm-PSM patients with PCI ≥16 had a worse 2-year survival (25.0%) vs. patients with PCI16 (79.1%; log rank = 0.009), but no significant effect was observed in patients with m-PSM (66.7% vs. 68.1%; p = 0.935). Underlying disease (HR 5.666-16.240), BMI (HR 1.109), and PCI (HR 1.068) significantly influenced overall survival in all patients.PCI is prognostic in nm-PSM, but not in m-PSM. CRS and HIPEC may benefit not only patients with low PCI, but also those with high PCI and m-PSM.

Oncogenic gene fusions are important drivers in many cancer types, including carcinomas, with diagnostic and therapeutic implications. Hence, sensitive and rapid methods for parallel profiling in formalin-fixed and paraffin-embedded (FFPE) specimens are needed. In this study we analyzed gene fusions in a cohort of 517 cases where standard treatment options were exhausted. To this end the Archer&reg, DX Solid tumor panel (AMP, 285 cases) and the Oncomine Comprehensive Assay v3 (OCA, 232 cases) were employed. Findings were validated by Sanger sequencing, fluorescence in situ hybridization (FISH) or immunohistochemistry. Both assays demonstrated minimal dropout rates (AMP: 2.4%, n = 7/292, OCA: 2.1%, n = 5/237) with turnaround times of 6&ndash, 9 working days (median, OCA and AMP, respectively). Hands-on-time for library preparation was 6 h (AMP) and 2 h (OCA). We detected n = 40 fusion-positive cases (7.7%) with TMPRSS2::ERG in prostate cancer being most prevalent (n = 9/40, 22.5%), followed by other gene fusions identified in cancers of unknown primary (n = 6/40, 15.0%), adenoid cystic carcinoma (n = 7/40, 17.5%), and pancreatic cancer (n = 7/40, 17.5%). Our results demonstrate that targeted RNA-sequencing of FFPE samples is feasible, and a well-suited approach for the detection of gene fusions in a routine clinical setting.

Assessment of Tumor Mutational Burden (TMB) for response stratification of cancer patients treated with immune checkpoint inhibitors is emerging as a new biomarker. Commonly defined as the total number of exonic somatic mutations, TMB approximates the amount of neoantigens that potentially are recognized by the immune system. While whole exome sequencing (WES) is an unbiased approach to quantify TMB, implementation in diagnostics is hampered by tissue availability as well as time and cost constrains. Conversely, panel-based targeted sequencing is nowadays widely used in routine molecular diagnostics, but only very limited data are available on its performance for TMB estimation. Here, we evaluated three commercially available larger gene panels with covered genomic regions of 0.39 Megabase pairs (Mbp), 0.53 Mbp and 1.7 Mbp using i) in silico analysis of TCGA (The Cancer Genome Atlas) data and ii) wet-lab sequencing of a total of 92 formalin-fixed and paraffin-embedded (FFPE) cancer samples grouped in three independent cohorts (non-small cell lung cancer, NSCLC; colorectal cancer, CRC; and mixed cancer types) for which matching WES data were available. We observed a strong correlation of the panel data with WES mutation counts especially for the gene panel >1Mbp. Sensitivity and specificity related to TMB cutpoints for checkpoint inhibitor response in NSCLC determined by wet-lab experiments well reflected the in silico data. Additionally, we highlight potential pitfalls in bioinformatics pipelines and provide recommendations for variant filtering. In summary, our study is a valuable data source for researchers working in the field of immuno-oncology as well as for diagnostic laboratories planning TMB testing.

BACKGROUND: Recent findings indicate that the currently approved predictive biomarker for immune checkpoint blockade (ICB), i.e. PD-L1 expression, is imperfect. Hence, additional biomarkers are warranted, and ICB-related gene expression profiling (GEP) was advocated as a promising approach enabling comprehensive interrogation of the ICB-effector compartment. Here, we assessed inter- and intra-tumor heterogeneity (ITH) of the immunological microenvironment using a comprehensive gene expression profiling approach (770 genes) to further elucidate the biological basis for new diagnostic applications. METHODS: All formalin-fixed paraffin-embedded tissue specimens used for this study were derived from surgically resected lung adenocarinomas at the Thoraxklinik at Heidelberg University Hospital and diagnosed (according to 2015 WHO Classification of lung tumors) at the Institute of Pathology at Heidelberg University Hospital. To evaluate the significance of ITH on gene expression profiling, multi-regional gene expression analysis (2-4 samples per tumor) was performed for 24 ADC using the NanoString IO 360 Panel. ANOVA was used to compare inter-tumor variance to intra-tumor variance of mRNA expression. The F statistics was calculated as ratio of these quantities. A list of genes with significantly higher inter-tumor variance was isolated using the F-test. Multiple testing was addressed using the Benjamini-Hochberg method to control the false discovery rate (FDR). RESULTS: In an unsupervised hierarchical clustering analysis of the entire set of 770 genes, the two to four segments of each of the tumors clustered together. For 752 of the 770 genes (97.7%) the inter-tumor variance was significantly higher than the intra-tumor variance (FDR CONCLUSION: In our preliminary analysis of ICB-GEP, variance of expression levels between tumors was substantial and exceeded ITH for the majority of analyzed genes. In-depth analyses are ongoing to further delineate the influence on gene signatures and ICB related pathways. Citation Format: Daniel Kazdal, Jan Budczies, Martina Kirchner, Klaus Kluck, Michael Allgäuer, Olaf Neumann, Regine Brandt, Eugen Rempel, Carolin Ploeger, Moritz von Winterfeld, Marc Schneider, Steffen Dietz, Hauke Winter, Thomas Muley, Holger Sülmann, Michael Meister, Felix Herth, Claus Heussel, Roland Penzel, Volker Endris, Peter Schirmacher, Michael Thomas, Petros Christopoulos, Albrecht Stenzinger. Inter- and intra- tumorheterogeneity of the microenvironment in pulmonary adenocarcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1571.

Reliable entity subtyping is paramount for therapy stratification in lung cancer. Morphological evaluation remains the basis for entity subtyping and directs the application of additional methods such as immunohistochemistry (IHC). The decision of whether to perform IHC for subtyping is subjective, and access to IHC is not available worldwide. Thus, the application of additional methods to support morphological entity subtyping is desirable. Therefore, the ability of convolutional neuronal networks (CNNs) to classify the most common lung cancer subtypes, pulmonary adenocarcinoma (ADC), pulmonary squamous cell carcinoma (SqCC), and small-cell lung cancer (SCLC), was evaluated. A cohort of 80 ADC, 80 SqCC, 80 SCLC, and 30 skeletal muscle specimens was assembled, slides were scanned, tumor areas were annotated, image patches were extracted, and cases were randomly assigned to a training, validation or test set. Multiple CNN architectures (VGG16, InceptionV3, and InceptionResNetV2) were trained and optimized to classify the four entities. A quality control (QC) metric was established. An optimized InceptionV3 CNN architecture yielded the highest classification accuracy and was used for the classification of the test set. Image patch and patient-based CNN classification results were 95% and 100% in the test set after the application of strict QC. Misclassified cases mainly included ADC and SqCC. The QC metric identified cases that needed further IHC for definite entity subtyping. The study highlights the potential and limitations of CNN image classification models for tumor differentiation.

Purpose Differential diagnosis of ulcerative colitis (UC) and Crohn's disease (CD) is of utmost importance for the decision making of respective therapeutic treatment strategies but in about 10-15% of cases, a clinical and histopathological assessment does not lead to a definite diagnosis. The aim of the study is to characterize proteomic differences between UC and CD. Experimental design Microproteomics is performed on formalin-fixed paraffin-embedded colonic tissue specimens from 9 UC and 9 CD patients. Protein validation is performed using immunohistochemistry (IHC) (nUC =51, nCD =62, nCTRL =10) followed by digital analysis. Results Microproteomic analyses reveal eight proteins with higher abundance in CD compared to UC including proteins related to neutrophil activity and damage-associated molecular patterns. Moreover, one protein, Aldo-keto reductase family 1 member C3 (AKR1C3), is present in eight out of nine CD and absent in all UC samples. Digital IHC analysis reveal a higher percentage and an increased expression intensity of AKR1C3-positive epithelial cells in CD compared to UC and in controls compared to inflammatory bowel disease (IBD). Conclusion and clinical relevance Overall, the results suggest that microproteomics is an adequate tool to highlight protein patterns in IBD. IHC and digital pathology might support future differential diagnosis of UC and CD.

Tumor mutational burden (TMB) is a new biomarker for prediction of response to PD-(L)1 treatment. Comprehensive sequencing approaches (i.e., whole exome and whole genome sequencing) are ideally suited to measure TMB directly. However, as their applicability in routine diagnostics is currently limited by high costs, long turnaround times and poor availability of fresh tissue, targeted next-generation sequencing (NGS) of formalin-fixed and paraffin-embedded (FFPE) samples appears to be a more feasible and straight-forward approach for TMB approximation, which can be seamlessly integrated in already existing diagnostic workflows and pipelines. In this work, we provide an overview of the clinical implications of TMB testing and highlight key parameters including pre-analysis, analysis and post-analytical steps that influence and shape TMB approximation by panel sequencing. Collectively, the data will not only serve as a field guide and state of the art knowledge source for molecular pathologists who consider implementation of TMB measurement in their lab, but also enable clinicians in understanding the specific parameters influencing TMB test results and reporting.

Genetic heterogeneity between and within tumours is a major factor determining cancer progression and therapy response. Here we examined DNA sequence and DNA copy-number heterogeneity in colorectal cancer (CRC) by targeted high-depth sequencing of 100 most frequently altered genes. In 97 samples, with primary tumours and matched metastases from 27 patients, we observe inter-tumour concordance for coding mutations; in contrast, gene copy numbers are highly discordant between primary tumours and metastases as validated by fluorescent in situ hybridization. To further investigate intra-tumour heterogeneity, we dissected a single tumour into 68 spatially defined samples and sequenced them separately. We identify evenly distributed coding mutations in APC and TP53 in all tumour areas, yet highly variable gene copy numbers in numerous genes. 3D morpho-molecular reconstruction reveals two clusters with divergent copy number aberrations along the proximal–distal axis indicating that DNA copy number variations are a major source of tumour heterogeneity in CRC., The contribution of intra-tumour heterogeneity is increasingly associated with resistance to therapy. Here, the authors use genomic analyses to study heterogeneity in colorectal cancer and perform in-depth reconstruction of heterogeneity in one sample.

// Jan Budczies 1,2,6,* , Moritz von Winterfeld 1,* , Frederick Klauschen 1 , Michael Bockmayr 1 , Jochen K. Lennerz 3 , Carsten Denkert 1,6 , Thomas Wolf 4,6 , Arne Warth 4 , Manfred Dietel 1 , Ioannis Anagnostopoulos 1 , Wilko Weichert 4,6,7 , Daniel Wittschieber 5 and Albrecht Stenzinger 4 1 Institute of Pathology, Charite University Hospital, Berlin, Germany 2 German Cancer Research Center (DKFZ), Heidelberg, Germany 3 Massachusetts General Hospital/Harvard Medical School, Department of Pathology, Boston, MA, USA 4 Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany 5 Institute of Legal Medicine, University Hospital Munster, Munster, Germany 6 German Consortium for Translational Cancer Research (DKTK), Heidelberg, Germany 7 National Center for Tumor Diseases (NCT),Heidelberg, Germany * These authors contributed equally to this work Correspondence: Albrecht Stenzinger, email: // Jan Budczies, email: // Keywords : autopsy, metastasis, systemic disease, carcinoma, solid tumor, cancer, survival Received : August 22, 2014 Accepted : November 04, 2014 Published : November 04, 2014 Abstract The majority of patients with solid malignancies die from metastatic burden. However, our current understanding of the mechanisms and resulting patterns of dissemination is limited. Here, we analyzed patterns of metastatic progression across 16 major cancer types in a cohort of 1008 patients with metastatic cancer autopsied between 2000 and 2013 to assess cancer specific progression patterns of disease and related risk predictions. The frequency and location of metastases were evaluated in and across 1) 16 major cancers, 2) smoking- and non-smoking-related cancers and 3) adeno- and squamous cell carcinoma. Associations between primary and secondary sites were analyzed by the fractional and the relative risk methods. We detected significantly different cancer specific patterns of metastatic progression with specific relative risk profiles for secondary site involvement. Histology and smoking etiology influenced these patterns. Backward analysis showed that metastatic patterns help to predict unknown primary sites. Solid malignancies maintain a unique and recurrent organ tropism to specific secondary sites which does not appear to be strongly influenced by advances in cancer medicine as shown by comparison with previous data sets. The delineated landscape of metastatic progression patterns is a comprehensive data resource to both clinical and basic scientists which aids fostering new hypotheses for cancer research and cancer therapies.

Colorectal cancer guidelines recommend adjuvant chemotherapy in stage II disease when less than 12 lymph nodes are assessed. The recommendation bases on previous studies showing an association of a low lymph node count and adverse outcome. Compared to current standards, however, the quality of lymph node examination in the studies was low. We, therefore, investigated the prognostic role of

Introduction: Tumor mutational burden (TMB) is an emerging biomarker to identify patients more likely to benefit from immuno-oncologic therapy. Therefore, academia- and industry-driven endeavors are underway to establish mostly panel-based sequencing solutions. Aside from various unsettled technical aspects, tumor biology itself might play an important role in determining a tumor’s TMB. Intratumor heterogeneity (ITH) is frequent in pulmonary adenocarcinoma (ADC) and seems to be a key factor regarding response failure and resistance development during therapy. In addition, ITH might contribute to inconsistent distribution of a biomarker and therefore an assay’s readout might depend on the region of tumor which is submitted for testing. Therefore, the assessment of ITH is an essential step in the evaluation of new diagnostic applications. Experimental Set-up: In order to investigate potential spatial differences in the TMB of a tumor, we analyzed a comprehensively morpho-molecularly characterized cohort of pulmonary ADC by sequencing 3-4 sections per tumor including lymph node metastases. So far 10 tumors were included in the ongoing analysis. None of the selected tumors had an oncogenic EGFR mutation, six were KRAS positive and four had no known driver mutation. TMB was determined applying the TruSightTM Oncology 500 targeted sequencing panel (TSO 500), covering 500 genes, with a NextSeq 500 sequencing system (both Illumina Inc., San Diego, CA, USA). Results Summary: The current sequencing analysis using the TSO 500 panel covered an average genomic coding region of 1.26 Mbp with a mean read depth of x658. The average TMB of all samples was 8.55 mut/Mbp (range 0.79 - 18.95). Comparing different regions within a given tumor, we detected a considerable variance of TMB (± 5 mut/Mbp) in about half of the analyzed cases, with an overall mean absolute deviation of 2.2 and a maximum difference of 12.63 mut/Mbp. However, in some samples an in depth evaluation of morpho-molecular characteristics revealed several factors impairing TMB read out. Aside from technical issues like insufficient depth of coverage in combination with unadjusted automated variant calling, sample characteristics like tumor cell content had a prominent influence on TMB values. Often, tumor regions with lower TMB turned out to have lower tumor cell content, which led to decreased allele frequencies of tumor specific mutations and subsequently compromised mutation detection. Conclusions: Our data demonstrate spatial heterogeneity of TMB in pulmonary ADC, which can impact clinical decision making and therapy. However, spatial genetic heterogeneity reflecting tumor biology needs to be carefully dissected from technical conditions resulting in pseudo-heterogeneity. Citation Format: Volker Endris, Michael Allgäuer, Jan Budczies, Jonas Leichsenring, Anna-Lena Volckmar, Martina Kirchner, Olaf Neumann, Regine Brandt, Eugen Rempel, Carolin Plöger, Moritz von Winterfeld, Alexander Harms, Mark Kriegsmann, Roland Penzel, Peter Schirmacher, Daniel Kazdal, Albrecht Stenzinger. Spatial heterogeneity of tumor mutational burden (TMB) counts in pulmonary adenocarcinoma: Separating biological effects from technical artifacts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4101.

Hintergund: In dieser Übersichtsarbeit soil der Stellenwert der zytoreduktiven chirurgischen Therapie (CRS) in Kombination mit einer hyperthermen intraperitonealen Chemotherapie (HIPEC) der Peritonealkarzinose bei nichtkolorektalen Karzinomen dargestellt werden. Hierzu zählen als primäre Tumoren des Peritoneums das Mesotheliom und das Pseudomyxoma peritonei. Zu den wichtigsten sekundären malignen Erkrankungen des Peritoneums zählen die Peritonealkarzinose des Magenkarzinoms, des Pankreaskarzinoms und des Ovarialkarzinoms sowie die Sarkomatose. Methode: Pathologische Besonderheiten der einzelnen Entitäten und Therapieempfehlungen anhand der aktuellen Literatur werden im Einzelnen beschrieben. Ergebnisse: Prinzipiell ist die chirurgische Behandlung der Peritonealkarzinose im Sinne einer CRS in Kombination mit einer HIPEC ein sinnvolles Therapiekonzept - vorausgesetzt, dass eine nahezu komplette Tumorentfernung erzielt werden kann. Schlussfolgerung: Aufgrund der erhöhten Morbidität sollte ein chirurgischer Eingriff gut indiziert sein. Bei ausgewählten Patienten ist eine CRS der Peritonealkarzinose in Kombination mit einer HIPEC ein sinnvolles und lebensverlängerndes Verfahren.

Background— Whether adult cardiomyocytes have the capacity to regenerate in response to injury and, if so, to what extent are still issues of intense debate. In human heart failure, cardiomyocytes harbor a polyploid genome. A unique opportunity to study the mechanism of polyploidization is provided through the setting of hemodynamic support by left ventricular assist devices. Hence, the cardiomyocyte DNA content, nuclear morphology, and number of nuclei per cell were assessed before and after left ventricular assist device support. Methods and Results— In 23 paired myocardial samples, cardiomyocyte ploidy was investigated by DNA image cytometry, flow cytometry, and in situ hybridization. Nuclear cross-sectional area and perimeters were measured morphometrically, and the binucleated cardiomyocytes were counted. The median of the cardiomyocyte DNA content and the number of polyploid cardiomyocytes both declined significantly from 6.79 c to 4.7 c and 40.2% to 23%, whereas a significant increase in diploid cardiomyocytes from 33.4% to 50.3% and in binucleated cardiomyocytes from 4.5% to 10% after unloading was observed. Conclusions— The decrease in polyploidy and increase in diploidy after left ventricular assist device suggest a numeric increase in diploid cardiomyocytes (eg, through cell cycle progression with completion of mitosis or by increased stem cells). The cardiac regeneration that follows may serve as a morphological correlate of the recovery observed in some patients after unloading.

Supplemental Digital Content is available in the text., The recently revised ISSVA classification approved in Melbourne in April 2014 recognizes generalized lymphatic anomaly and lymphatic malformation in Gorham-Stout disease. The 2 entities can overlap in presentation, as both are characterized by destructive lymphatic vessel invasion of the axial skeleton and surrounding soft tissues. At least at present, no standard therapeutic options exist, and due to the rarity of the disease, no clinical trials are available. We present 2 patients, 1 with generalized lymphatic anomaly and 1 with lymphatic malformation in Gorham-Stout disease, with severe exacerbation during puberty. The first child presented in florid pulmonary failure and pleural effusion, the other with severe pain due to bone destruction of the pelvis and inability to walk. Both were treated using individualized protocols. The manuscript describes the rationale for choosing sunitinib in combination with low-dose (metronomic) taxol. Both patients experienced clinical and radiologic response without major toxicities, suggesting that patients with rare conditions may benefit from individualized, molecularly based therapies.

Background Inflammatory bowel disease (IBD)-like conditions in glycogen storage disease (GSD) type Ib have been predominantly described in children. Signs and symptoms of GSD type Ib are hypoglycemia, pancytopenia and hepatosplenomegaly. Based on few published cases, there is evidence that granulocyte-colony stimulating factor (G-CSF) in patients with glycogenosis–related pancytopenia might ameliorate the IBD-like disease through leukocyte increase. Case presentation Here we firstly describe a case of an adult 33-year-old Caucasian male patient with GSD type Ib accompanied with IBD-like disease with persistent pancytopenia despite moderate-dose G-CSF treatment. Recent vomiting and abdominal discomfort were due to a high-grade stenosis in the transverse colon. A dose increase of the G-CSF successfully normalized his leukocyte count. However, the stenosis worsened and surgical therapy was needed. Conclusion We suggest that symptomatic patients with GSD type Ib should undergo endoscopic examination in order to detect IBD-like disease and to initiate early treatment.

BackgroundAutopsy rates in Western countries consistently decline to an average of Methods and findingsOf all adult autopsy cases of the Charité Institute of Pathology from the years 1988, 1993, 1998, 2003 and 2008, the pre- and postmortal diagnoses and all demographic data were analyzed retrospectively. Based on power analysis, 1,800 cases were randomly selected to perform discrepancy classification (class I-VI) according to modified Goldman criteria. The rate of discrepancies in major diagnoses (class I) was 10.7% (95% CI: 7.7%-14.7%) in 2008 representing a reduction by 15.1%. Subgroup analysis revealed several influencing factors to significantly correlate with the discrepancy rate. Cardiovascular diseases had the highest frequency among class-I-discrepancies. Comparing the 1988-data of East- and West-Berlin, no significant differences were found in diagnostic discrepancies despite an autopsy rate differing by nearly 50%. A risk profile analysis visualized by intuitive heatmaps revealed a significantly high discrepancy rate in patients treated in low or intermediate care units at community hospitals. In this collective, patients with genitourinary/renal or infectious diseases were at particularly high risk.ConclusionsThis is the current largest and most comprehensive study on diagnostic discrepancies worldwide. Our well-powered analysis revealed a significant rate of class-I-discrepancies indicating that autopsies are still of value. The identified risk profiles may aid both pathologists and clinicians to identify patients at increased risk for a discrepant diagnosis and possibly suboptimal treatment intra vitam.

Human epidermal growth factor receptor 2 (HER2, alias ERBB2)-targeted therapy in breast and gastric cancers depends on the reliable assessment of HER2 protein expression and (in equivocal cases) the quantitative evaluation of HER2 gene amplification. Typically, HER2 and centromere 17 gene copy numbers are evaluated using in situ hybridization (ISH) to calculate ratios for which cutoff values dividing nonamplified and amplified cases have been proposed. Although several studies have investigated how laboratory procedures affect diagnostics, a rigorous quantitative assessment of the diagnostic guidelines for data analysis is still missing. Here, we analyze the dependence of the diagnosed HER2/chromosome 17 ratios on i) sample size (evaluated cells), ii) gene/chromosome signal distributions, and iii) the approach used for quotient calculation using Monte Carlo simulations. Our data show that the current recommendation may lead to statistical HER2/CHR17 ratio variations of up to 0.94 and may therefore lead to incorrect HER2 status diagnoses, given the ratio threshold of 2.0 defined by the Food and Drug Administration. Moreover, borderline cases may receive different amplification diagnoses, depending on the ratio calculation approach: Brightfield-silver ISH with aggregated signal counts may underestimate the HER2/CHR17 ratio compared with two-color fluorescence ISH. Our results provide a basis for quantitative rationales behind HER2 diagnostic guidelines that call for increased numbers of evaluated cells and emphasize the importance of well-designed data analysis methods in diagnostic pathology, especially for predictive clinical application.

In chronic heart failure, atrial and brain natriuretic peptide expression is increased and serves as a clinical marker of cardiac hypertrophy. Chromogranin A is also up-regulated during chronic heart failure and associated with disease severity and prognosis. Significant decrease of both natriuretic peptide and hypertrophy after left ventricular assist device (LVAD) treatment was reported. This study investigated whether chromogranin A and neural cell adhesion molecule (NCAM)/CD56 are associated with cardiac hypertrophy and regulated by LVAD.Expression of atrial and brain natriuretic peptide, chromogranin A, and NCAM/CD56 were investigated by immunohistochemistry and morphometrically quantified in 33 paired myocardial samples before and after LVAD. In a different set of patients, chromogranin A was evaluated in the plasma. Cardiomyocyte colocalization of brain natriuretic peptide and chromogranin A was visualized by immunofluorescence doublestaining.Natriuretic peptide and chromogranin A protein expression is significantly decreased after LVAD (p0.05). NCAM/CD56 expression remains unaltered by unloading. In contrast with natriuretic peptide, chromogranin A and NCAM/CD56 expression is not correlated with cardiomyocyte diameters. Although increased compared with controls, no significant differences for chromogranin A plasma levels were found before and after LVAD. Sarcoplasmic colocalization of chromogranin A and brain natriuretic peptide is considerably decreased after LVAD.Neither chromogranin A nor CD56 is associated with cardiac hypertrophy. Chromogranin A is significantly decreased by ventricular support. Sarcoplasmic colocalization of brain natriuretic peptide and chromogranin A is diminished after unloading. However, owing to its low expression, the negative regulation of chromogranin A is not reflected by plasma levels and thus does not appear to be an appropriate biomarker of reverse cardiac remodeling after unloading.

Complex sets of driver mutations critically control clinical course and treatment response in colorectal cancer (CRC). It is expected that multiple mutations in different signaling pathways interact to establish an individual cancer cell phenotype. Furthermore, mutational patterns can evolve under selective pressure, for instance during tumor metastasis or targeted therapy. Yet only few mutations, such as these in KRAS, NRAS and BRAF, are routinely assessed in the clinic today. With the introduction of next generation sequencing (NGS) many more genes can be simultaneously sequenced, allowing detailed analyses of mutational patterns. We have assembled the CRC5.2 NGS panel for application with Ion Torrent PGM covering 100 frequently mutated genes with 784 amplicons (covering 21,000 COSMIC mutations), going well beyond the scope of commercially available cancer panels. In particular, exon coverage was optimized to embrace all genomic information related to drug sensitivity, using several drug and signaling network databases. The German Consortium for Translational Cancer Research (DKTK) comprises the top cancer centers in Germany. As a part of this group, we employ our custom CRC5.2 panel for the analysis of mutational patterns in CRC. We address several key questions relevant for current diagnostics and treatment in colorectal cancer. In retrospective and prospective clinical studies, we assess mutational heterogeneity in primary tumors, synchronous and metachronous metastases of individual patients. In-depth NGS mutational analyses, transcriptome analysis and methylation studies will identify (epi-)genetic alterations occurring as a consequence of selective pressure during tumor evolution e.g. during metastasis, chemotherapy or targeted tumor therapy. Thus, we will uncover new links between cancer-specific mutations, clinical course and treatment response as well as novel mechanisms of secondary resistance. In addition we aim to identify novel predictive and prognostic mutational markers for the benefit of future CRC patients Citation Format: Soulafa Mamlouk, Liam Childs, Thorben Redmer, Pawel Durek, Dirk Schumacher, Hendrik Bläker, Daniela Aust, Moritz von Winterfeld, Volker Heinemann, Dominik Modest, Kerstin Möhr, Markus Morkel, Reinhold Schäfer, Christine Sers. Linking tumor evolution and therapy response using diagnostic targeted next generation sequencing in colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5170. doi:10.1158/1538-7445.AM2014-5170