Publications by authors named "Jutta Kirfel"

64 Publications

Recurrent HNSCC Harbor an Immunosuppressive Tumor Immune Microenvironment Suggesting Successful Tumor Immune Evasion.

Clin Cancer Res 2021 Jan 27;27(2):632-644. Epub 2020 Oct 27.

Institute of Pathology, University Hospital Schleswig-Holstein, Campus Lübeck, Lübeck, Germany.

Purpose: Recurrent tumors (RT) of head and neck squamous cell carcinoma (HNSCC) occur in up to 60%, with poor therapeutic response and detrimental prognosis. We hypothesized that HNSCC RTs successfully evade antitumor immune response and aimed to reveal tumor immune microenvironment (TIME) changes of primary tumors (PT) and corresponding RTs.

Experimental Design: Tumor-infiltrating leukocytes (TIL) of 300 PTs and 108 RTs from two large independent and clinically well-characterized HNSCC cohorts [discovery cohort (DC), validation cohort (VD)] were compared by IHC. mRNA expression analysis of 730 immune-related genes was performed for 18 PTs and RTs after adjuvant chemoradiotherapy (CRT). The effect of chemotherapy and radiation resistance was assessed with an spheroid/immunocyte coculture model.

Results: TIME analysis revealed overall decrease of TILs with significant loss of CD8 T cells (DC = 0.045/VC < 0.0001) and B lymphocytes (DC = 0.036/VC < 0.0001) in RTs compared with PTs in both cohorts. Decrease predominantly occurred in RTs after CRT. Gene expression analysis confirmed loss of TILs ( = 0.0004) and B lymphocytes ( < 0.0001) and showed relative increase of neutrophils ( = 0.018), macrophages ( < 0.0001), dendritic cells ( = 0.0002), and mast cells ( = 0.0057) as well as lower overall expression of immune-related genes ( = 0.018) in RTs after CRT. Genes involved in B-lymphocyte functions and number of tertiary lymphoid structures showed the strongest decrease. and were upregulated and , indicating their potential suitability as therapeutic targets in CRT resistance.

Conclusions: HNSCC RTs have an immunosuppressive TIME, which is particularly apparent after adjuvant CRT and might substantially contribute to poor therapeutic response and prognosis.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0197DOI Listing
January 2021

NR2F6 as a Prognostic Biomarker in HNSCC.

Int J Mol Sci 2020 Aug 1;21(15). Epub 2020 Aug 1.

Institute of Pathology, University of Luebeck and University Hospital Schleswig-Holstein, 23538 Luebeck, Germany.

Head and neck squamous cell carcinoma (HNSCC)is the 6th most common cancer in humans worldwide and is associated with a poor prognosis for patients. NR2F6 has been identified as an immune checkpoint molecule in tumor-infiltrating T lymphocytes and is associated with a poor prognostic outcome in various cancers. The prognostic value of NR2F6 in HNSCC has not been described yet. We used a large, representative and clinically well-characterized cohort of 383 HNSCC patients, of which 22.4% developed a local recurrence. The NR2F6 expression was analyzed by using immunohistochemistry and was afterward correlated with clinical characteristics and clinicopathological features of HNSCC patients. Primary tumors from patients who develop a local recurrence have a higher NR2F6 expression than primary tumors which do not develop a local recurrence. Furthermore, a high NR2F6 expression is associated with poorer recurrence-free survival, although there is no correlation with overall survival. NR2F6 expression is independent of the T stage and UICC stage. NR2F6 might be a new prognostic biomarker for the early detection of local recurrences in HNSCC patients. Therefore, it may help to improve the recognition of patients who would benefit from more frequent follow-up examinations.
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http://dx.doi.org/10.3390/ijms21155527DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7432340PMC
August 2020

In silico analysis reveals EP300 as a panCancer inhibitor of anti-tumor immune response via metabolic modulation.

Sci Rep 2020 06 10;10(1):9389. Epub 2020 Jun 10.

Pathology of the Research Center Borstel, Leibniz Lung Center, Borstel, Germany.

The tumor immune microenvironment (TIME) of head and neck squamous cell carcinomas (HNSCC) and other solid malignancies is a key determinant of therapy response and prognosis. Among other factors, it is shaped by the tumor mutational burden and defects in DNA repair enzymes. Based on the TCGA database we aimed to define specific, altered genes associated with different TIME types, which might represent new predictive markers or targets for immuno-therapeutic approaches. The HNSCC cohort of the TCGA database was used to define 3 TIME types (immune-activated, immune-suppressed, immune-absent) according to expression of immune-related genes. Mutation frequencies were correlated to the 3 TIME types. Overall survival was best in the immune-activated group. 9 genes were significantly differentially mutated in the 3 TIME types with strongest differences for TP53 and the histone-acetyltransferase EP300. Mutations in EP300 correlated with an immune-activated TIME. In panCancer analyses anti-tumor immune activity was increased in EP300 mutated esophageal, stomach and prostate cancers. Downregulation of EP300 gene expression was associated with higher anti-tumor immunity in most solid malignancies. Since EP300 is a promoter of glycolysis, which negatively affects anti-tumor immune response, we analyzed the association of EP300 with tumor metabolism. PanCancer tumor metabolism was strongly shifted towards oxidative phosphorylation in EP300 downregulated tumors. In silico analyses of of publicly available in vitro data showed a decrease of glycolysis-associated genes after treatment with the EP300 inhibitor C646. Our study reveals associations of specific gene alterations with different TIME types. In detail, we defined EP300 as a panCancer inhibitor of the TIME most likely via metabolic modulation. In this context EP300 represents a promising predictive biomarker and an immuno-therapeutic target.
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http://dx.doi.org/10.1038/s41598-020-66329-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7287052PMC
June 2020

High Levels of Chromosomal Copy Number Alterations and TP53 Mutations Correlate with Poor Outcome in Younger Breast Cancer Patients.

Am J Pathol 2020 08 13;190(8):1643-1656. Epub 2020 May 13.

Genetics Branch, National Cancer Institute, NIH, Bethesda, Maryland. Electronic address:

Prognosis in young patients with breast cancer is generally poor, yet considerable differences in clinical outcomes between individual patients exist. To understand the genetic basis of the disparate clinical courses, tumors were collected from 34 younger women, 17 with good and 17 with poor outcomes, as determined by disease-specific survival during a follow-up period of 17 years. The clinicopathologic parameters of the tumors were complemented with DNA image cytometry profiles, enumeration of copy numbers of eight breast cancer genes by multicolor fluorescence in situ hybridization, and targeted sequence analysis of 563 cancer genes. Both groups included diploid and aneuploid tumors. The degree of intratumor heterogeneity was significantly higher in aneuploid versus diploid cases, and so were gains of the oncogenes MYC and ZNF217. Significantly more copy number alterations were observed in the group with poor outcome. Almost all tumors in the group with long survival were classified as luminal A, whereas triple-negative tumors predominantly occurred in the short survival group. Mutations in PIK3CA were more common in the group with good outcome, whereas TP53 mutations were more frequent in patients with poor outcomes. This study shows that TP53 mutations and the extent of genomic imbalances are associated with poor outcome in younger breast cancer patients and thus emphasize the central role of genomic instability vis-a-vis tumor aggressiveness.
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http://dx.doi.org/10.1016/j.ajpath.2020.04.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7397461PMC
August 2020

EVI1 as a Marker for Lymph Node Metastasis in HNSCC.

Int J Mol Sci 2020 Jan 28;21(3). Epub 2020 Jan 28.

Institute of Pathology, University Hospital Schleswig-Holstein, Campus Luebeck, Ratzeburger Allee 160, 23538 Luebeck, Germany.

Background: HNSCC is the sixth most common cancer in humans and has still a very poor prognosis. The treatment methods so far are very often associated with mutilation and impairment in the quality of life. Except for p16 expression, there are no reliable prognostic markers in HNSCC so far. Ecotropic Viral Integration Site 1 (EVI1) is a well-described prognostic marker in leukemia and different types of solid cancers. In these, a high EVI1 expression is associated with a poor prognosis. In HNSCC, it is not known so far if EVI1 has any prognostic relevance.

Materials And Methods: We used our representative tissue cohort of 389 primary HNSCCs, of which 57.2% had one or more lymph node metastases. Here EVI1 expression was analyzed via immunohistochemistry and correlated with the clinical characteristics of these patients.

Results: Although in HNSCC EVI1 expression does not predict poor survival, a high EVI1 expression in the primary tumor correlates with a lymph node metastatic disease.

Conclusion: Consequently, EVI1 may serve as a biomarker to predict an occult lymph node metastasis in a clinical nodal negative (cN0) HNSCC.
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http://dx.doi.org/10.3390/ijms21030854DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7038015PMC
January 2020

Increased mediator complex subunit CDK19 expression associates with aggressive prostate cancer.

Int J Cancer 2020 01 19;146(2):577-588. Epub 2019 Jul 19.

Pathology of the University Hospital Schleswig-Holstein, Campus Luebeck, Luebeck, Germany.

The Mediator complex is a transcriptional regulator interacting with transcription factors and RNA-polymerase-II. Recently, we identified its subunit CDK19 to be specifically expressed in prostate cancer (PCa) and to be functionally implicated in PCa aggressiveness. Aim of our study was to comprehensively characterize the protein expression of CDK19 and its paralog CDK8 in PCa. We performed immunohistochemistry (IHC) for CDK19/CDK8 on a large cohort including needle biopsies from 202 patients, 799 primary tumor foci of radical prostatectomy specimens from 415 patients, 120 locally advanced tumor foci obtained by palliative transurethral resection, 140 lymph node metastases, 67 distant metastases and 82 benigns. Primary tumors were stained for the proliferation marker Ki67, androgen receptor (AR) and ERG. For 376 patients, clinic-pathologic data were available. Primary endpoint was disease-recurrence-free survival (DFS). Nuclear CDK19 and CDK8 expression increases during progression showing the highest intensity in metastatic and castration-resistant tumors. High CDK19 expression on primary tumors correlates with DFS independently from Gleason grade and PSA. Five-year-DFS rates of patients with primary tumors expressing no, moderate and high CDK19 are 73.7, 56.9 and 30.4%, respectively. CDK19 correlates with Gleason grade, T-stage, Ki67 proliferation-index, nuclear AR expression and ERG-status. Therapeutic options for metastatic and castration-resistant PCa remain limited. In the current study, we confirmed an important role of the Mediator subunit CDK19 in advanced PCa supporting current developments to target CDK19 and its paralog CDK8. Furthermore, CDK19 protein expression has the potential to predict disease recurrence independently from established biomarkers thus contributing to individual management for PCa patients.
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http://dx.doi.org/10.1002/ijc.32551DOI Listing
January 2020

TRIM24 as an independent prognostic biomarker for prostate cancer.

Urol Oncol 2019 09 7;37(9):576.e1-576.e10. Epub 2019 Jun 7.

Institute of Pathology, University Hospital Schleswig-Holstein, Luebeck, Germany; Pathology of the Research Center Borstel, Leibniz Lung Center, Borstel, Germany. Electronic address:

Introduction: Simply applicable biomarkers for prostate cancer patients predicting the clinical course are urgently needed. Recently, TRIM24 has been identified to promote androgen receptor signaling and to correlate with an aggressive prostate cancer phenotype. Based on these data, we proofed TRIM24 as a prognostic biomarker for risk stratification.

Materials And Methods: We performed TRIM24 immunohistochemistry on 2 independent cohorts including a total of 806 primary tumors, 26 locally advanced/recurrent tumors, 30 lymph node metastases, 30 distant metastases, and 129 benign prostatic samples from 497 patients as well as on 246 prostate needle biopsies. Expression data were correlated with clinic-pathological data including biochemical recurrence-free survival (bRFS) as endpoint.

Results: Benign samples show no or low TRIM24 expression in 94%, while tumor tissues demonstrate significant higher levels. Strongest expression is observed in advanced and metastatic tumors. In multivariate analyses, TRIM24 up-regulation on radical prostatectomy specimens correlates with shorter bRFS independent of other prognostic parameters. 5-(10-) year bRFS rates for TRIM24 negative, low, medium and high expressing tumors are 93.1(93.1)%, 75.4(68.5)%, 54.9(47.5)% and 43.1(32.3)%, respectively. Of interest, tumors diagnosed as indolent disease, TRIM24 expression stratifies patients into specific risk groups. Increased TRIM24 expression associates with higher grade group, positive nodal status and extraprostatic tumor growth. TRIM24 assessment on prostate needle biopsies taken prior to treatment decision at time of initial diagnosis significantly correlates with recurrence after surgery.

Conclusion: Using 2 large independent radical prostatectomy specimen cohorts, we found that TRIM24 expression predicts patients' risk to develop disease recurrence with high accuracy and independent from other established biomarkers. Further, this is the first study exploring TRIM24 expression on prostate needle biopsies which represents the clinically relevant tissue type on which biomarkers guide treatment decisions. Thus, we strongly suggest introducing TRIM24 evaluation in prostate needle biopsies in clinical routine as an inexpensive and simple immunohistochemical test.
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http://dx.doi.org/10.1016/j.urolonc.2019.05.006DOI Listing
September 2019

Expression of Prostate-Specific Membrane Antigen (PSMA) on Biopsies Is an Independent Risk Stratifier of Prostate Cancer Patients at Time of Initial Diagnosis.

Front Oncol 2018 20;8:623. Epub 2018 Dec 20.

Pathology of the University Hospital Schleswig-Holstein and Research Center Borstel, Leibniz Lung Center, Luebeck, Germany.

Stratifying prostate cancer (PCa) patients into risk groups at time of initial diagnosis enabling a risk-adapted disease management is still a major clinical challenge. Existing studies evaluating the prognostic potential of PSMA (prostate-specific membrane antigen) for PCa were performed on radical prostatectomy specimens (RPE), i.e., decision making for disease management was already completed at time of sample analysis. Aim of our study was to assess the prognostic value of PSMA expression for PCa patients on biopsies at time of initial diagnosis. PSMA expression was assessed by immunohistochemistry on 294 prostate biopsies with corresponding RPE, 621 primary tumor foci from 242 RPE, 43 locally advanced or recurrent tumors, 34 lymph node metastases, 78 distant metastases and 52 benign prostatic samples. PSMA expression was correlated with clinico-pathologic features. Primary endpoint was recurrence free survival. Other clinicopathologic features included WHO/ISUP grade groups, PSA serum level, TNM-stage, and R-status. Chi-square test, ANOVA-analyses, Cox-regression, and log-rank tests were performed for statistical analyses. High PSMA expression on both biopsy and RPE significantly associates with a higher risk of disease recurrence following curative surgery. The 5-year-recurrence free survival rates were 88.2, 74.2, 67.7 and 26.8% for patients exhibiting no, low, medium, or high PSMA expression on biopsy, respectively. High PSMA expression on biopsy was significant in multivariate analysis predicting a 4-fold increased risk of disease recurrence independently from established prognostic markers. PSMA significantly increases during PCa progression. PSMA is an independent prognostic marker on biopsies at time of initial diagnosis and can predict disease recurrence following curative therapy for PCa. Our study proposes the application of the routinely used IHC marker PSMA for outcome prediction and decision making in risk-adapted PCa management on biopsies at time of initial diagnosis.
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http://dx.doi.org/10.3389/fonc.2018.00623DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6307416PMC
December 2018

Exome sequencing characterizes the somatic mutation spectrum of early serrated lesions in a patient with serrated polyposis syndrome (SPS).

Hered Cancer Clin Pract 2017 29;15:22. Epub 2017 Nov 29.

Institute of Human Genetics, University of Bonn, Bonn, Germany.

Background: Serrated or Hyperplastic Polyposis Syndrome (SPS, HPS) is a yet poorly defined colorectal cancer (CRC) predisposition characterised by the occurrence of multiple and/or large serrated polyps throughout the colon. A serrated polyp-CRC sequence (serrated pathway) of CRC formation has been postulated, however, to date only few molecular signatures of serrated neoplasia (, mutations, CpG Island Methylation, MSI) have been described in a subset of SPS patients and neither the etiology of the syndrome nor the distinct genetic alterations during tumorigenesis have been identified.

Methods: To identify somatic point mutations in potential novel candidate genes of SPS-associated lesions and the involved pathways we performed exome sequencing of eleven early serrated polyps obtained from a 41 year-old female patient with clinically confirmed SPS. For data filtering and analysis, standard pipelines were used. Somatic mutations were identified by comparison with leukocyte DNA and were validated by Sanger sequencing.

Results: The p.V600E or p.G12D mutation was identified in six polyps (~50%) and not found in polyps from the distal colon. In addition, we found seven unique rare somatic alterations of seven different genes in four serrated tumours, all of which are missense variants. The variant in and are predicted to be deleterious No established cancer gene or candidate genes related to serrated tumorigenesis were affected.

Conclusions: Somatic mutations seem to be rare events in early hyperplastic and serrated lesions of SPS patients. Neither frequently affected genes nor enrichment of specific pathways were observed. Thus, other alterations such as non-coding variants or epigenetic changes might be the major driving force of tumour progression in SPS.
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http://dx.doi.org/10.1186/s13053-017-0082-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5707812PMC
November 2017

Tumoral PD-L1 expression defines a subgroup of poor-prognosis vulvar carcinomas with non-viral etiology.

Oncotarget 2017 Nov 6;8(54):92890-92903. Epub 2017 Oct 6.

Department of Obstetrics and Gynecology, Center for Integrated Oncology, University of Bonn, Bonn, Germany.

Vulvar cancer is rare but incidence rates are increasing due to an aging population and higher frequencies of young women being affected. In locally advanced, metastatic or recurrent disease prognosis is poor and new treatment modalities are needed. Immune checkpoint blockade of the PD-1/PD-L1 pathway is one of the most important advancements in cancer therapy in the last years. The clinical relevance of PD-L1 expression in vulvar cancer, however, has not been studied so far. We determined PD-L1 expression, numbers of CD3 T cells, CD20 B cells, CD68 monocytes/macrophages, Foxp3 regulatory T cells and CD163 tumor-associated macrophages by immunohistochemistry in 103 patients. Correlation analysis with clinicopathological parameters was undertaken; the cause-specific outcome was modeled with competing risk analysis; multivariate Cox regression was used to determine independent predictors of survival. Membranous PD-L1 was expressed in a minority of tumors, defined by HPV-negativity. Its presence geographically correlated with immunocyte-rich regions of cancer islets and was an independent prognostic factor for poor outcome. Our data support the notion that vulvar cancer is an immunomodulatory tumor that harnesses the PD-1/PD-L1 pathway to induce tolerance. Accordingly, immunotherapeutic approaches might have the potential to improve outcome in patients with vulvar cancer and could complement conventional cancer treatment.
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http://dx.doi.org/10.18632/oncotarget.21641DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5696230PMC
November 2017

Prognostic Value of the New Prostate Cancer International Society of Urological Pathology Grade Groups.

Front Med (Lausanne) 2017 29;4:157. Epub 2017 Sep 29.

Pathology of the University Hospital Schleswig-Holstein, Campus Luebeck and Research Center Borstel, Leibniz Center for Medicine and Biosciences, Luebeck, Germany.

Gleason grading is the best independent predictor for prostate cancer (PCa) progression. Recently, a new PCa grading system has been introduced by the International Society of Urological Pathology (ISUP) and is recommended by the World Health Organization (WHO). Following studies observed more accurate and simplified grade stratification of the new system. Aim of this study was to compare the prognostic value of the new grade groups compared to the former Gleason Grading and to determine whether re-definition of Gleason Pattern 4 might reduce upgrading from prostate biopsy to radical prostatectomy (RP) specimen. A cohort of men undergoing RP from 2002 to 2015 at the Hospital of Goeppingen (Goeppingen, Germany) was used for this study. In total, 339 pre-operative prostatic biopsies and corresponding RP specimens, as well as additional 203 RP specimens were re-reviewed for Grade Groups according to the ISUP. Biochemical recurrence-free survival (BFS) after surgery was used as endpoint to analyze prognostic significance. Other clinicopathological data included TNM-stage and pre-operative PSA level. Kaplan-Meier analysis revealed risk stratification of patients based on both former Gleason Grading and ISUP Grade Groups, and was statistically significant using the log-rank test ( < 0.001). Both grading systems significantly correlated with TNM-stage and pre-operative PSA level ( < 0.001). Higher tumor grade in RP specimen compared to corresponding pre-operative biopsy was observed in 44 and 34.5% of cases considering former Gleason Grading and ISUP Grade Groups, respectively. Both, former Gleason Grading and ISUP Grade Groups predict survival when applied on tumors in prostatic biopsies as well as RP specimens. This is the first validation study on a large representative German community-based cohort to compare the former Gleason Grading with the recently introduced ISUP Grade Groups. Our data indicate that the ISUP Grade Groups do not improve predictive value of PCa grading and might be less sensitive in deciphering tumors with 3 + 4 and 4 + 3 pattern on RP specimen. However, the Grade Group system results less frequently in an upgrading from biopsy to the corresponding RP specimens, indicating a lower risk to miss potentially aggressive tumors not represented on biopsies.
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http://dx.doi.org/10.3389/fmed.2017.00157DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5626925PMC
September 2017

Implication of the Receptor Tyrosine Kinase AXL in Head and Neck Cancer Progression.

Int J Mol Sci 2016 Dec 22;18(1). Epub 2016 Dec 22.

Institute of Pathology, University Hospital of Luebeck, 23538 Luebeck, Germany.

Head and neck squamous cell carcinoma (HNSCC) remains a clinical challenge and identification of novel therapeutic targets is necessary. The receptor tyrosine kinase AXL has been implicated in several tumor entities and a selective AXL small molecule inhibitor (BGB324) is currently being tested in clinical trials for patients suffering from non-small cell lung cancer or acute myeloid leukemia. Our study investigates AXL expression during HNSCC progression and its use as a potential therapeutic target in HNSCC. AXL protein expression was determined in a HNSCC cohort ( = 364) using immunohistochemical staining. For functional validation, AXL was either overexpressed or inhibited with BGB324 in HNSCC cell lines to assess proliferation, migration and invasion. We found AXL protein expression increasing during tumor progression with highest expression levels in recurrent tumors. In HNSCC cell lines in vitro, AXL overexpression increased migration as well as invasion. Both properties could be reduced through treatment with BGB324. In contrast, proliferation was neither affected by AXL overexpression nor by inhibition with BGB324. Our patient-derived data and in vitro results show that, in HNSCC, AXL is important for the progression to more advanced tumor stages. Moreover, they suggest that AXL could be a target for precision medicine approaches in this dismal tumor entity.
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http://dx.doi.org/10.3390/ijms18010007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5297642PMC
December 2016

MED15 overexpression in prostate cancer arises during androgen deprivation therapy via PI3K/mTOR signaling.

Oncotarget 2017 Jan;8(5):7964-7976

Pathology of the University Medical Center Schleswig-Holstein, Campus Luebeck and Research Center Borstel, Leibniz Center for Medicine and Biosciences, Borstel, Germany.

Androgen deprivation therapy (ADT) is the main therapeutic option for advanced prostate cancer (PCa). After initial regression, most tumors develop into castration-resistant PCa (CRPC). Previously, we found the Mediator complex subunit MED15 to be overexpressed in CRPC and to correlate with clinical outcome. Therefore, we investigated whether MED15 is implicated in the signaling changes taking place during progression to CRPC. Immunohistochemistry (IHC) for MED15 on matched samples from the same patients before and after ADT reveals significantly increased MED15 expression after ADT in 72%. A validation cohort comprising samples before and after therapy confirmed our observations. Protein analysis for pAKT and pSMAD3 shows that MED15 correlates with PI3K and TGFß activities, respectively, and that hyper-activation of both pathways simultaneously correlates with highest levels of MED15. We further show that MED15 protein expression increases in LNCaP cells under androgen deprivation, and via EGF mediated PI3K activation. PI3K/mTOR and TGFß-receptor inhibition results in decreased MED15 expression. MED15 knockdown reduces LNCaP cell viability and induces apoptosis during androgen deprivation, while cell cycle is not affected. Collectively, MED15 overexpression arises during ADT via hyper-activation of PI3K/mTOR signaling, thus MED15 may serve as a predictive marker for response to PI3K/mTOR inhibitors. Furthermore, MED15 is potentially a therapeutic target for the treatment of CRPC.
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http://dx.doi.org/10.18632/oncotarget.13860DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5352374PMC
January 2017

Overexpression of histone demethylase Fbxl10 leads to enhanced migration in mouse embryonic fibroblasts.

Exp Cell Res 2016 Nov 17;348(2):123-131. Epub 2016 Sep 17.

Institute of Pathology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn, Germany. Electronic address:

Cell migration is a central process in the development and maintenance of multicellular organisms. Tissue formation during embryonic development, wound healing, immune responses and invasive tumors all require the orchestrated movement of cells to specific locations. Histone demethylase proteins alter transcription by regulating the chromatin state at specific gene loci. FBXL10 is a conserved and ubiquitously expressed member of the JmjC domain-containing histone demethylase family and is implicated in the demethylation of H3K4me3 and H3K36me2 and thereby removing active chromatin marks. However, the physiological role of FBXL10 in vivo remains largely unknown. Therefore, we established an inducible gain of function model to analyze the role of Fbxl10 and compared wild-type with Fbxl10 overexpressing mouse embryonic fibroblasts (MEFs). Our study shows that overexpression of Fbxl10 in MEFs doesn't influence the proliferation capability but leads to an enhanced migration capacity in comparison to wild-type MEFs. Transcriptome and ChIP-seq experiments demonstrated that Fbxl10 binds to genes involved in migration like Areg, Mdk, Lmnb1, Thbs1, Mgp and Cxcl12. Taken together, our results strongly suggest that Fbxl10 plays a critical role in migration by binding to the promoter region of migration-associated genes and thereby might influences cell behaviour to a possibly more aggressive phenotype.
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http://dx.doi.org/10.1016/j.yexcr.2016.08.026DOI Listing
November 2016

Pan-Cancer Analysis of the Mediator Complex Transcriptome Identifies CDK19 and CDK8 as Therapeutic Targets in Advanced Prostate Cancer.

Clin Cancer Res 2017 Apr 27;23(7):1829-1840. Epub 2016 Sep 27.

Pathology of the University Medical Center Schleswig-Holstein, Campus Lübeck and the Research Center Borstel, Leibniz Center for Medicine and Biosciences, Lübeck and Borstel, Germany.

The Mediator complex is a multiprotein assembly, which serves as a hub for diverse signaling pathways to regulate gene expression. Because gene expression is frequently altered in cancer, a systematic understanding of the Mediator complex in malignancies could foster the development of novel targeted therapeutic approaches. We performed a systematic deconvolution of the Mediator subunit expression profiles across 23 cancer entities ( = 8,568) using data from The Cancer Genome Atlas (TCGA). Prostate cancer-specific findings were validated in two publicly available gene expression cohorts and a large cohort of primary and advanced prostate cancer ( = 622) stained by immunohistochemistry. The role of and was evaluated by siRNA-mediated gene knockdown and inhibitor treatment in prostate cancer cell lines with functional assays and gene expression analysis by RNAseq. Cluster analysis of TCGA expression data segregated tumor entities, indicating tumor-type-specific Mediator complex compositions. Only prostate cancer was marked by high expression of In primary prostate cancer, was associated with increased aggressiveness and shorter disease-free survival. During cancer progression, highest levels of CDK19 and of its paralog CDK8 were present in metastases. , inhibition of CDK19 and CDK8 by knockdown or treatment with a selective CDK8/CDK19 inhibitor significantly decreased migration and invasion. Our analysis revealed distinct transcriptional expression profiles of the Mediator complex across cancer entities indicating differential modes of transcriptional regulation. Moreover, it identified and to be specifically overexpressed during prostate cancer progression, highlighting their potential as novel therapeutic targets in advanced prostate cancer. .
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http://dx.doi.org/10.1158/1078-0432.CCR-16-0094DOI Listing
April 2017

A signaling cascade including ARID1A, GADD45B and DUSP1 induces apoptosis and affects the cell cycle of germ cell cancers after romidepsin treatment.

Oncotarget 2016 Nov;7(46):74931-74946

Institute of Pathology, Department of Developmental Pathology, University Medical School, Bonn, Germany.

In Western countries, the incidence of testicular germ cell cancers (GCC) is steadily rising over the last decades. Mostly, men between 20 and 40 years of age are affected. In general, patients suffering from GCCs are treated by orchiectomy and radio- or chemotherapy. Due to resistance mechanisms, intolerance to the therapy or denial of chemo- / radiotherapy by the patients, GCCs are still a lethal threat, highlighting the need for alternative treatment strategies.In this study, we revealed that germ cell cancer cell lines are highly sensitive to the histone deacetylase inhibitor romidepsin in vitro and in vivo, highlighting romidepsin as a potential therapeutic option for GCC patients.Romidepsin-mediated inhibition of histone deacetylases led to disturbances of the chromatin landscape. This resulted in locus-specific histone-hyper- or hypoacetylation. We found that hypoacetylation at the ARID1A promotor caused repression of the SWI/SNF-complex member ARID1A. In consequence, this resulted in upregulation of the stress-sensors and apoptosis-regulators GADD45B, DUSP1 and CDKN1A. RNAi-driven knock down of ARID1A mimicked in parts the effects of romidepsin, while CRISPR/Cas9-mediated deletion of GADD45B attenuated the romidepsin-provoked induction of apoptosis and cell cycle alterations.We propose a signaling cascade involving ARID1A, GADD45B and DUSP1 as mediators of the romidepsin effects in GCC cells.
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http://dx.doi.org/10.18632/oncotarget.11647DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5342713PMC
November 2016

Exome Sequencing Identifies Biallelic MSH3 Germline Mutations as a Recessive Subtype of Colorectal Adenomatous Polyposis.

Am J Hum Genet 2016 Aug 28;99(2):337-51. Epub 2016 Jul 28.

Institute of Human Genetics, University of Bonn, 53127 Bonn, Germany; Center for Hereditary Tumor Syndromes, University of Bonn, 53127 Bonn, Germany. Electronic address:

In ∼30% of families affected by colorectal adenomatous polyposis, no germline mutations have been identified in the previously implicated genes APC, MUTYH, POLE, POLD1, and NTHL1, although a hereditary etiology is likely. To uncover further genes with high-penetrance causative mutations, we performed exome sequencing of leukocyte DNA from 102 unrelated individuals with unexplained adenomatous polyposis. We identified two unrelated individuals with differing compound-heterozygous loss-of-function (LoF) germline mutations in the mismatch-repair gene MSH3. The impact of the MSH3 mutations (c.1148delA, c.2319-1G>A, c.2760delC, and c.3001-2A>C) was indicated at the RNA and protein levels. Analysis of the diseased individuals' tumor tissue demonstrated high microsatellite instability of di- and tetranucleotides (EMAST), and immunohistochemical staining illustrated a complete loss of nuclear MSH3 in normal and tumor tissue, confirming the LoF effect and causal relevance of the mutations. The pedigrees, genotypes, and frequency of MSH3 mutations in the general population are consistent with an autosomal-recessive mode of inheritance. Both index persons have an affected sibling carrying the same mutations. The tumor spectrum in these four persons comprised colorectal and duodenal adenomas, colorectal cancer, gastric cancer, and an early-onset astrocytoma. Additionally, we detected one unrelated individual with biallelic PMS2 germline mutations, representing constitutional mismatch-repair deficiency. Potentially causative variants in 14 more candidate genes identified in 26 other individuals require further workup. In the present study, we identified biallelic germline MSH3 mutations in individuals with a suspected hereditary tumor syndrome. Our data suggest that MSH3 mutations represent an additional recessive subtype of colorectal adenomatous polyposis.
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http://dx.doi.org/10.1016/j.ajhg.2016.06.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4974087PMC
August 2016

Targeting DDR2 in head and neck squamous cell carcinoma with dasatinib.

Int J Cancer 2016 11 30;139(10):2359-69. Epub 2016 Jul 30.

Pathology of the University Medical Center Schleswig-Holstein, Campus Luebeck and the Research Center Borstel, Leibniz Center for Medicine and Biosciences, Luebeck and Borstel, Germany.

Squamous cell carcinoma of the head and neck (HNSCC) is the tenth most common tumor entity in men worldwide. Nevertheless therapeutic options are mostly limited to surgery and radio-chemotherapy resulting in 5-year survival rates of around 50%. Therefore new therapeutic options are urgently needed. During the last years, targeting of receptor tyrosine kinases has emerged as a promising strategy that can complement standard therapeutical approaches. Here, we aimed at investigating if the receptor tyrosine kinase DDR2 is a targetable structure in HNSCC. DDR2 expression was assessed on a large HNSCC cohort (554 patients) including primary tumors, lymph node metastases and recurrences and normal mucosa as control. Subsequently, DDR2 was stably overexpressed in two different cell lines (FaDu and HSC-3) using lentiviral technology. Different tumorigenic properties such as proliferation, migration, invasion, adhesion and anchorage independent growth were assessed with and without dasatinib treatment using in-vitro cell models and in-vivo zebrafish xenografts. DDR2 was overexpressed in all tumor tissues when compared to normal mucosa. DDR2 overexpression led to increased migration, invasion, adhesion and anchorage independent growth whereas proliferation remained unaltered. Upon dasatinib treatment migration, invasion and adhesion could be inhibited in-vitro and in-vivo whereas proliferation was unchanged. Our data suggest treatment with dasatinib as a promising new therapeutic option for patients suffering from DDR2 overexpressing HNSCC. Since dasatinib is already FDA-approved we propose to test this drug in clinical trials so that patients could directly benefit from this new treatment option.
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http://dx.doi.org/10.1002/ijc.30279DOI Listing
November 2016

MERTK as a novel therapeutic target in head and neck cancer.

Oncotarget 2016 May;7(22):32678-94

Pathology of The University Hospital of Luebeck, Luebeck, Germany.

Although head and neck cancer (HNSCC) is the sixth most common tumor entity worldwide therapy options remain limited leading to 5-year survival rates of only 50 %. MERTK is a promising therapeutic target in several tumor entities, however, its role in HNSCC has not been described yet. The aim of our study was to investigate the biological significance of MERTK and to evaluate its potential as a novel therapeutic target in this dismal tumor entity. In two large HNSCC cohorts (n=537 and n=520) we found that MERTK is overexpressed in one third of patients. In-vitro, MERTK overexpression led to increased proliferation, migration and invasion whereas MERTK inhibition with the small molecule inhibitor UNC1062 or MERTK knockdown reduced cell motility via the small GTPase RhoA.Taken together, we are the first to show that MERTK is frequently overexpressed in HNSCC and plays an important role in tumor cell motility. It might therefore be a potential target for selected patients suffering from this dismal tumor entity.
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http://dx.doi.org/10.18632/oncotarget.8724DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5078043PMC
May 2016

Evaluation of FGFR3 as a Therapeutic Target in Head and Neck Squamous Cell Carcinoma.

Target Oncol 2016 10;11(5):631-642

Pathology of the University Hospital of Luebeck, Luebeck, Germany.

Background: Although head and neck squamous cell carcinoma (HNSCC) is the sixth most common tumour entity worldwide, it remains a clinical challenge. Large-scale explorative genomic projects have identified several genes as potential targets for therapy, including fibroblast growth factor receptor 3 (FGFR3).

Aims: The aim of this study was to investigate the biological significance of wild-type and mutated FGFR3 to evaluate its potential as a novel therapeutic target in HNSCC.

Methods: FGFR3 protein expression was analysed in a large HNSCC tissue cohort (n = 536) and FGFR3 mRNA expression from The Cancer Genome Atlas (TCGA; n = 520). Moreover, FGFR3 wild-type and mutant versions were overexpressed in vitro, and both proliferation and migration was assessed with and without BGJ398 (a specific FGFR1-3 inhibitor) treatment.

Results: Although FGFR3 expression for both cohorts decreased during tumour progression, high FGFR3 expression levels were observed in a small subset of patients. In vitro, FGFR3 overexpression led to increased proliferation, whereas migration was not altered. Moreover, FGFR3-overexpressing cells were more sensitive to BGJ398. Cells overexpressing FGFR3 mutant versions showed increased proliferation compared to wild-type FGFR3 under serum-reduced conditions and were largely as sensitive as the wild-type protein to BGJ398.

Conclusions: Taken together, the results of this study demonstrate that although FGFR3 expression decreases during HNSSC progression, it plays an important role in tumour cell proliferation and thus may be a potential target for therapy in selected patients suffering from this dismal tumour entity.
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http://dx.doi.org/10.1007/s11523-016-0431-zDOI Listing
October 2016

IL-6 Overexpression in ERG-Positive Prostate Cancer Is Mediated by Prostaglandin Receptor EP2.

Am J Pathol 2016 Apr;186(4):974-84

Section of Prostate Cancer Research, Institute of Pathology, Center for Integrated Oncology Cologne/Bonn, University Hospital of Bonn, Bonn, Germany. Electronic address:

Prostate cancer is the most diagnosed cancer in men and multiple risk factors and genetic alterations have been described. The TMPRSS2-ERG fusion event and the overexpression of the transcription factor ERG are present in approximately 50% of all prostate cancer patients, however, the clinical outcome is still controversial. Prostate tumors produce various soluble factors, including the pleiotropic cytokine IL-6, regulating cellular processes such as proliferation and metastatic segregation. Here, we used prostatectomy samples in a tissue microarray format and analyzed the co-expression and the clinicopathologic data of ERG and IL-6 using immunohistochemical double staining and correlated the read-out with clinicopathologic data. Expression of ERG and IL-6 correlated strongly in prostate tissue samples. Forced expression of ERG in prostate tumor cell lines resulted in significantly increased secretion of IL-6, whereas the down-regulation of ERG decreased IL-6 secretion. By dissecting the underlying mechanism in prostate tumor cell lines we show the ERG-mediated up-regulation of the prostanoid receptors EP2 and EP3. The prostanoid receptor EP2 was overexpressed in human prostate cancer tissue. Furthermore, the proliferation rate and IL-6 secretion in DU145 cells was reduced after treatment with EP2-receptor antagonist. Collectively, our study shows that the expression of ERG in prostate cancer is linked to the expression of IL-6 mediated by the prostanoid receptor EP2.
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http://dx.doi.org/10.1016/j.ajpath.2015.12.009DOI Listing
April 2016

Loss of the LIM-only protein Fhl2 impairs inflammatory reaction and scar formation after cardiac ischemia leading to better hemodynamic performance.

Life Sci 2016 Apr 24;151:348-358. Epub 2016 Feb 24.

Institute of Pathology, University of Cologne, Kerpener Str. 62, 50937 Cologne, Germany. Electronic address:

Aims: The pathogenesis of myocardial ischemia-reperfusion injury (MI/R) involves an inflammatory response. Since the four-and-a-half LIM domain-containing protein 2 (Fhl2) has been observed to modulate immune cell migration, we aimed to study the consequences of Fhl2(-/-) under MI/R with respect to immune reaction, scar formation, and hemodynamic performance.

Material And Methods: In a closed chest model of 1h MI/R, immune cell invasion of phagocytic monocytes was characterized by flow cytometry and immunohistochemistry. In addition, infarct size was assessed by triphenyltetrazolium chloride/Masson trichrome staining 24h/21days after reperfusion and a set of hemodynamic parameters was recorded by catheterisation in Fhl2(-/-) mice and controls.

Key Findings: While flow cytometry did not reveal differences in myocardial CD45(high) immune cell infiltrate, histological analysis showed that infiltrating immune cells in Fhl2(-/-) animals were preferentially located in the perivascular area, whereas in wild type, immune cells were well dispersed within the area at risk. After 24h and 21days of reperfusion, infarct size was significantly reduced in Fhl2(-/-) compared to WT animals. In addition, hemodynamic performance was better in Fhl2(-/-) mice, compared to WT mice up to day 21 of reperfusion. The loss of Fhl2 leads to an altered immune response to myocardial ischemia, which results in smaller infarcts and better hemodynamic performance up to 21days after myocardial ischemia reperfusion.

Significance: Immune cell invasion plays a pivotal role in the context of MI/R. Fhl2 significantly influences immune cell function and immune cell interaction with injured cardiac tissue leading to altered scar composition.
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http://dx.doi.org/10.1016/j.lfs.2016.02.084DOI Listing
April 2016

Low-level APC mutational mosaicism is the underlying cause in a substantial fraction of unexplained colorectal adenomatous polyposis cases.

J Med Genet 2016 Mar 27;53(3):172-9. Epub 2015 Nov 27.

Institute of Human Genetics, University of Bonn, Bonn, Germany Center for Hereditary Tumor Syndromes, University of Bonn, Bonn, Germany.

Background: In 30-50% of patients with colorectal adenomatous polyposis, no germline mutation in the known genes APC, causing familial adenomatous polyposis, MUTYH, causing MUTYH-associated polyposis, or POLE or POLD1, causing polymerase-proofreading-associated polyposis can be identified, although a hereditary aetiology is likely. This study aimed to explore the impact of APC mutational mosaicism in unexplained polyposis.

Methods: To comprehensively screen for somatic low-level APC mosaicism, high-coverage next-generation sequencing of the APC gene was performed using DNA from leucocytes and a total of 53 colorectal tumours from 20 unrelated patients with unexplained sporadic adenomatous polyposis. APC mosaicism was assumed if the same loss-of-function APC mutation was present in ≥ 2 anatomically separated colorectal adenomas/carcinomas per patient. All mutations were validated using diverse methods.

Results: In 25% (5/20) of patients, somatic mosaicism of a pathogenic APC mutation was identified as underlying cause of the disease. In 2/5 cases, the mosaic level in leucocyte DNA was slightly below the sensitivity threshold of Sanger sequencing; while in 3/5 cases, the allelic fraction was either very low (0.1-1%) or no mutations were detectable. The majority of mosaic mutations were located outside the somatic mutation cluster region of the gene.

Conclusions: The present data indicate a high prevalence of pathogenic mosaic APC mutations below the detection thresholds of routine diagnostics in adenomatous polyposis, even if high-coverage sequencing of leucocyte DNA alone is taken into account. This has important implications for both routine work-up and strategies to identify new causative genes in this patient group.
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http://dx.doi.org/10.1136/jmedgenet-2015-103468DOI Listing
March 2016

SRC inhibition represents a potential therapeutic strategy in liposarcoma.

Int J Cancer 2015 Dec 6;137(11):2578-88. Epub 2015 Jul 6.

Department of Pathology, University Hospital Cologne, Cologne, Germany.

Liposarcomas (LS) are the most common malignant mesenchymal tumors, with an overall long-term mortality rate of 60%. LS comprise three major subtypes, i.e., well-differentiated/dedifferentiated liposarcoma (WDLS/DDLS), myxoid/round cell liposarcoma (MLS) and pleomorphic liposarcoma (PLS). Aiming at the preclinical identification of novel therapeutic options, we here investigate the functional significance of SRC in primary human LS and in LS-derived cell lines. Immunohistochemical and Western blot analyses reveal relevant levels of activated p-(Tyr416)-SRC in LS of the different subtypes with particular activation in MLS and PLS. Dysregulation of the SRC modifiers CSK and PTP1B was excluded as major reason for the activation of the kinase. Consistent siRNA-mediated knockdown of SRC or inhibition by the SRC inhibitor Dasatinib led to decreased proliferation of LS cell lines of the different subtypes, with MLS cells reacting particularly sensitive in MTT assays. Flow cytometric analyses revealed that this effect was due to a significant decrease in mitotic activity and an induction of apoptosis. SRC inhibition by Dasatinib resulted in dephosphorylation of SRC itself, its interacting partners FAK and IGF-IR as well as its downstream target AKT. Consistent with a particular role of SRC in cell motility, Dasatinib reduced the migratory and invasive potential of MLS cells in Boyden chamber and Matrigel chamber assays. In summary, we provide evidence that SRC activation plays an important role in LS biology and therefore represents a potential therapeutic target, particularly in MLS and PLS.
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http://dx.doi.org/10.1002/ijc.29645DOI Listing
December 2015

Α₁-antitrypsin PiMZ heterozygosity has an independent aggravating effect on liver fibrosis in alcoholic liver disease.

Virchows Arch 2014 Nov 29;465(5):539-46. Epub 2014 Jul 29.

Institute of Pathology, University of Bonn, Sigmund-Freud-Str. 25, 53105, Bonn, Germany,

Heterozygous α1-antitrypsin deficiency type PiZ (PiMZ) results in chronic liver injury and predisposes to hepatocellular carcinoma. Gene frequency of the PiZ allele ranges from 0.005 to 0.027 in Western and Central Europe; therefore, there is a substantial risk of coincidence with chronic alcohol abuse. This retrospective case-control study evaluates the impact of PiMZ genotype on the development of chronic liver disease in alcohol consuming patients. Six thousand eight hundred eighty-six consecutive liver specimens were immunohistochemically tested for PiZ-deposits. From 254 PiZ-positive patients, the liver biopsies of 30 PiMZ adults without concomitant liver disease other than alcoholic liver disease (ALD) were selected and matched to PiMM (wild type) patients with respect to age, gender and lifetime daily alcohol ingestion (LDAI). Histomorphological changes were assessed using the SAF score and by digital image analysis. Liver cirrhosis was significantly more frequent in PIMZ patients than in matched PiMM patients (PiMM 9/30 vs. PiMZ 14/30, p = 0.04). Comparison of the extent of fibrosis in PiMZ and PiMM livers by two-way ANOVA indicated that the amount of LDAI has a major effect in PiMZ and PiMM patients (30.04 % of total variation, p < 0.0001), whereas PIMZ genotype has a minor but independent effect on liver fibrosis as assessed by digital planimetric evaluation (9.27 % of total variation, p = 0.005). Semiquantitative assessment was in agreement with this finding. Histomorphological findings support that PiMZ heterozygosity has an independent aggravating effect on liver fibrosis, even though the pathogenic effect of alcohol consumption is much stronger.
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http://dx.doi.org/10.1007/s00428-014-1633-3DOI Listing
November 2014

KDM5C is overexpressed in prostate cancer and is a prognostic marker for prostate-specific antigen-relapse following radical prostatectomy.

Am J Pathol 2014 Sep 9;184(9):2430-7. Epub 2014 Jul 9.

Institute of Pathology, University of Bonn, Bonn, Germany. Electronic address:

Currently, few prognostic factors are available to predict the emergence of castration-resistant prostate cancer and no curative options are available. Epigenetic gene regulation has been shown to trigger prostate cancer metastasis and androgen independence. Histone lysine demethylases (KDMs) are epigenetic enzymes that can remove both repressive and activating histone marks. KDM5 family members are capable of removing the histone H3 lysine 4 dimethylation-activating mark, rendering them potential players in the down-regulation of tumor suppressors and suggesting that their activity could repress oncogenes. Here, we systematically investigated KDM5C expression patterns in two independent radical prostatectomy cohorts (822 prostate tumors in total) by immunohistochemistry. Positive nuclear KDM5C staining was significantly associated with a reduced prostate-specific antigen relapse-free survival. Our study confirmed that nuclear KDM5C expression is an independent prognostic parameter. Most strikingly, the prognostic value of nuclear KDM5C expression for progression-free survival was exclusively pronounced for the Gleason group 7. In addition, KDM5C knockdown resulted in growth retardation of prostate cancer cells in vitro and induced regulation of several proliferation-associated genes. Our data indicate that KDM5C is functionally involved in proliferation control of prostate cancer cells and might represent a novel attractive therapy target. Moreover, overexpression of KDM5C is an independent new predictive marker for therapy failure as determined by biochemical recurrence in patients after prostatectomy.
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http://dx.doi.org/10.1016/j.ajpath.2014.05.022DOI Listing
September 2014

Lysine-specific demethylase 1 (LSD1) in hematopoietic and lymphoid neoplasms.

Blood 2014 Jul;124(1):151-2

Institute of Pathology, University of Bonn, Bonn, Germany.

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http://dx.doi.org/10.1182/blood-2014-04-569525DOI Listing
July 2014

MED12 overexpression is a frequent event in castration-resistant prostate cancer.

Endocr Relat Cancer 2014 Aug 17;21(4):663-75. Epub 2014 Jun 17.

Department of Prostate Cancer ResearchInstitute of PathologyUniversity Hospital of Bonn, Sigmund-Freud Strasse 25, 53127 Bonn, GermanyInstitute for PathologyUniversity Hospital Basel, Basel, SwitzerlandDepartment of UrologySt. Claraspital, Basel, SwitzerlandDepartment of UrologyUniversity Hospital Basel, Basel, SwitzerlandDepartment of UrologyUniversity Hospital of Örebro, Örebro, SwedenSchool of Health and Medical SciencesÖrebro University, Örebro, SwedenCenter for Genomics and TranscriptomicsCeGaT GmbH, Tuebingen, GermanyClinic for Urology and Pediatric UrologyUniversity Hospital of Bonn, Sigmund-Freud Strasse 25, 53127 Bonn, GermanySection of Molecular UrooncologyDepartment of Urology, School of Medicine, University of Heidelberg, Heidelberg, GermanyDepartment of Prostate Cancer ResearchInstitute of PathologyUniversity Hospital of Bonn, Sigmund-Freud Strasse 25, 53127 Bonn, GermanyInstitute for PathologyUniversity Hospital Basel, Basel, SwitzerlandDepartment of UrologySt. Claraspital, Basel, SwitzerlandDepartment of UrologyUniversity Hospital Basel, Basel, SwitzerlandDepartment of UrologyUniversity Hospital of Örebro, Örebro, SwedenSchool of Health and Medical SciencesÖrebro University, Örebro, SwedenCenter for Genomics and TranscriptomicsCeGaT GmbH, Tuebingen, GermanyClinic for Urology and Pediatric UrologyUniversity Hospital of Bonn, Sigmund-Freud Strasse 25, 53127 Bonn, GermanySection of Molecular UrooncologyDepartment of Urology, School of Medicine, University of Heidelberg, Heidelberg, Germany

In a recent effort to unravel the molecular basis of prostate cancer (PCa), Barbieri and colleagues using whole-exome sequencing identified a novel recurrently mutated gene, MED12, in 5.4% of primary PCa. MED12, encoding a subunit of the Mediator complex, is a transducer of Wnt/β-catenin signaling, linked to modulation of hedgehog signaling and to the regulation of transforming growth factor beta (TGFβ)-receptor signaling. Therefore, these studies prompted us to investigate the relevance of MED12 in PCa. Expression of MED12, SMAD3 phosphorylation, and proliferation markers was assessed by immunohistochemistry on tissue microarrays from 633 patients. siRNA-mediated knockdown of MED12 was carried out on PCa cell lines followed by cellular proliferation assays, cell cycle analysis, apoptosis assays, and treatments with recombinant TGFβ3. We found nuclear overexpression of MED12 in 40% (28/70) of distant metastatic castration-resistant prostate cancer (CRPC(MET)) and 21% (19/90) of local-recurrent CRPC (CRPC(LOC)) in comparison with frequencies of less than 11% in androgen-sensitive PCa, and no overexpression in benign prostatic tissues. MED12 expression was significantly correlated with high proliferative activity in PCa tissues, whereas knockdown of MED12 decreased proliferation, reduced G1- to S-phase transition, and increased the expression of the cell cycle inhibitor p27. TGFβ signaling activation associates with MED12 nuclear overexpression in tissues and results in a strong increase in MED12 nuclear expression in cell lines. Furthermore, MED12 knockdown reduced the expression of the TGFβ target gene vimentin. Our findings show that MED12 nuclear overexpression is a frequent event in CRPC in comparison with androgen-sensitive PCa and is directly implicated in TGFβ signaling.
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http://dx.doi.org/10.1530/ERC-14-0171DOI Listing
August 2014

ATZ11 recognizes not only Z-α1-antitrypsin-polymers and complexed forms of non-Z-α1-antitrypsin but also the von Willebrand factor.

PLoS One 2014 19;9(3):e91538. Epub 2014 Mar 19.

Department of Pathology, University Bonn, Bonn, Germany.

Aims: The ATZ11 antibody has been well established for the identification of α1-anti-trypsin (AAT) molecule type PiZ (Z-AAT) in blood samples and liver tissue. In this study, we systematically analyzed the antibody for additional binding sites in human tissue.

Methods And Results: Ultrastructural ATZ11 binding was investigated immunoelectron microscopically in human umbilical vein endothelial cells (HUVECs) and in platelets of a healthy individual. Human embryonic kidney (HEK293) cells were transiently transfected with Von Willebrand factor (VWF) and analyzed immunocytochemically using confocal microscopy and SDS-PAGE electrophoresis followed by western blotting (WB). Platelets and serum samples of VWF-competent and VWF-deficient patients were investigated using native PAGE and SDS-PAGE electrophoresis followed by WB. The specificity of the ATZ11 reaction was tested immunohistochemically by extensive antibody-mediated blocking of AAT- and VWF-antigens. ATZ11-positive epitopes could be detected in Weibel-Palade bodies (WPBs) of HUVECs and α-granules of platelets. ATZ11 stains pseudo-WBP containing recombinant wild-type VWF (rVWF-WT) in HEK293 cells. In SDS-PAGE electrophoresis followed by WB, anti-VWF and ATZ11 both identified rVWF-WT. However, neither rVWF-WT-multimers, human VWF-multimers, nor serum proteins of VWF-deficient patients were detected using ATZ11 by WB, whereas anti-VWF antibody (anti-VWF) detected rVWF-WT-multimers as well as human VWF-multimers. In human tissue specimens, AAT-antigen blockade using anti-AAT antibody abolished ATZ11 staining of Z-AAT in a heterozygous AAT-deficient patient, whereas VWF-antigen blockade using anti-VWF abolished ATZ11 staining of endothelial cells and megakaryocytes.

Conclusions: ATZ11 reacts with cellular bound and denatured rVWF-WT and human VWF as shown using immunocytochemistry and subsequent confocal imaging, immunoelectron microscopy, SDS-PAGE and WB, and immunohistology. These immunoreactions are independent of the binding of Z-AAT-molecules and non-Z-AAT complexes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0091538PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3960128PMC
December 2014

Absence of TERT promoter mutations in primary melanocytic tumours of the central nervous system.

Neuropathol Appl Neurobiol 2014 Oct;40(6):794-7

Institute of Neuropathology, University of Bonn Medical Center, Bonn, Germany.

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http://dx.doi.org/10.1111/nan.12138DOI Listing
October 2014