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Brenner tumors (BT) are rare ovarian tumors encompassing benign, borderline, and malignant variants. While the histopathology of BTs and their clinical course is well described, little is known about the underlying genetic defects. We employed targeted next generation sequencing to analyze the mutational landscape in a cohort of 23 BT cases (17 benign, 2 borderline, and 4 malignant) and 3 ovarian carcinomas with transitional cell histology (TCC). Copy number variations (CNV) were validated by fluorescence in-situ hybridization (FISH) and quantitative PCR-based copy number assays. Additionally, we analyzed the TERT promotor region by conventional Sanger sequencing. We identified 25 different point mutations in 23 of the analyzed genes in BTs and 10 mutations in 8 genes in TCCs. About 57% percent of mutations occurred in genes involved in cell cycle control, DNA repair, and epigenetic regulation processes. All TCC cases harbored TP53 mutations whereas all BTs were negative and none of the mutations observed in BTs were present in TCCs. CNV analysis revealed recurrent MDM2 amplifications in 3 out of 4 of the malignant BT cases with one case harboring a concomitant amplification of CCND1. No mutations were observed in the TERT promoter region in BTs and TCCs, which is mutated in about 50%-75% of urothelial carcinoma and in 16% of ovarian clear-cell carcinomas. In conclusion, our study highlights distinct genetic features of BTs, and detection of the triplet phenotype MDM2 amplification/TP53 wt/TERT wt may aid diagnosis of malignant BT in difficult cases. Moreover, selected genetic lesions may be clinically exploitable in a metastatic setting.

Inhibition of the PD-L1 (CD274) - PD-1 axis has emerged as a powerful cancer therapy that prevents evasion of tumor cells from the immune system. While immunohistochemical detection of PD-L1 was introduced as a predictive biomarker with variable power, much less is known about copy number alterations (CNA) affecting PD-L1 and their associations with expression levels, mutational load, and survival. To gain insight, we employed The Cancer Genome Atlas (TCGA) datasets to comprehensively analyze 22 major cancer types for PD-L1 CNAs. We observed a diverse landscape of PD-L1 CNAs, which affected focal regions, chromosome 9p or the entire chromosome 9. Deletions of PD-L1 were more frequent than gains (31% vs. 12%) with deletions being most prevalent in melanoma and non-small cell lung cancer. Copy number gains most frequently occurred in ovarian cancer, head and neck cancer, bladder cancer, cervical and endocervical cancer, sarcomas, and colorectal cancers. Fine-mapping of the genetic architecture revealed specific recurrently amplified and deleted core regions across cancers with putative biological and clinical consequences. PD-L1 CNAs correlated significantly with PD-L1 mRNA expression changes in many cancer types, and tumors with PD-L1 gains harbored significantly higher mutational load compared to non-amplified cases (median: 78 non-synonymous mutations vs. 40, P = 7.1e-69). Moreover, we observed that, in general, both PD-L1 amplifications and deletions were associated with dismal prognosis. In conclusion, PD-L1 CNAs, in particular PD-L1 copy number gains, represent frequent genetic alterations across many cancers, which influence PD-L1 expression levels, are associated with higher mutational loads, and may be exploitable as predictive biomarker for immunotherapy regimens. © 2016 Wiley Periodicals, Inc.

Sinn B V, Darb-Esfahani S, Wirtz R M, Budczies J, Sehouli J, Chekerov R, Dietel M & Denkert C (2011) Histopathology 59, 918–927 Evaluation of a hormone receptor-positive ovarian carcinoma subtype with a favourable prognosis by determination of progesterone receptor and oestrogen receptor 1 mRNA expression in formalin-fixed paraffin-embedded tissue Aims: In vitro and epidemiological studies indicate an essential role for progesterone in the aetiology and progression of ovarian carcinoma. The aim of this study was to examine the prognostic role of progesterone receptor (PR) protein and mRNA expression. Methods and results: PR expression was examined by immunohistochemistry (n = 143) and kinetic reverse transcription–polymerase chain reaction (RT–PCR) from formalin-fixed and paraffin-embedded tissue (n = 55). PR mRNA and protein expression correlated (P

Darb-Esfahani S, Sinn B V, Weichert W, Budczies J, Lehmann A, Noske A, Buckendahl A-C, Muller B M, Sehouli J, Koensgen D, Gyorffy B, Dietel M & Denkert C (2010) Histopathology 56. 727–739 Expression of classical NF-κB pathway effectors in human ovarian carcinoma Aims: Functional studies have demonstrated that nuclear factor (NF)-κB promotes tumour progression in ovarian cancer cells. However, surprisingly little is known of the expression of effectors of the NF-κB pathway in human ovarian cancer in vivo. Methods and results: Immunohistochemistry and in situ hybridization revealed that in a cohort of 85 primary ovarian carcinomas, total p65 expression was inversely correlated to nuclear and cytoplasmic phospho-IκBα (P = 0.002 and P = 0.05, respectively), and IκBα mRNA expression (P = 0.032). In contrast, phospho-p65 expression was paralleled by the expression of nuclear (P = 0.027) and cytoplasmic phospho-IκBα (P = 0.01). Total p65 expression was an adverse prognostic factor for overall survival (P = 0.018). In contrast, total IκBα and phosphorylated nuclear and cytoplasmic IκBα expression were favourable prognostic markers (P = 0.001, P = 0.031, P = 0.001, respectively). Cytoplasmic phospho-IκBα expression remained a significant prognostic factor on multivariate analysis (P = 0.010). In cultured, stimulated OVCAR-3 ovarian cancer cells the cytoplasmic retranslocation of p65 was delayed by inhibition of the nuclear membrane transporter chromosomal region maintenance/exportin1 protein (CRM1). A positive association of p65 and CRM1 expression was demonstrated in ovarian cancer tissue (P

MicroRNA (miRNA) deregulation is a hallmark of human cancer. However, the mechanisms underlying miRNA alteration and the specific role of proteins involved in miRNA processing remains to be elucidated. Dicer is a key enzyme in the miRNA processing pathway that is essential for the production of mature miRNAs from their precursors. We tested the hypothesis that Dicer has biological and clinical relevance in ovarian cancer, using a range of methods including in vitro manipulation of Dicer expression. We observed down-regulation of Dicer in a subgroup of ovarian carcinomas, and found that decreased Dicer expression correlates significantly with reduced patient survival in serous cancers and advanced disease stages. Moreover, microarray and functional analysis suggest that reduced Dicer expression is connected with a global down-regulation of the microRNAome and with gene expression changes, particularly reduced expression of oestrogen receptor (ER) mRNA and protein in tumour tissue and in cell culture. Our data suggest a common mechanism for miRNAs changes by alterations in the basic machinery controlling miRNA biogenesis, of which Dicer is a central enzyme. These alterations of miRNA processing are of prognostic value and may play a role in the molecular pathogenesis of ovarian carcinoma and, possibly, other tumours. Knowledge of these molecular pathways may help toward new targeted therapeutic approaches for ovarian cancer.

Ovarian carcinoma has the highest mortality rate among gynaecological malignancies. In this project, we investigated the hypothesis that molecular markers are able to predict outcome of ovarian cancer independently of classical clinical predictors, and that these molecular markers can be validated using independent data sets. We applied a semi-supervised method for prediction of patient survival. Microarrays from a cohort of 80 ovarian carcinomas (TOC cohort) were used for the development of a predictive model, which was then evaluated in an entirely independent cohort of 118 carcinomas (Duke cohort). A 300-gene ovarian prognostic index (OPI) was generated and validated in a leave-one-out approach in the TOC cohort (Kaplan-Meier analysis, p = 0.0087). In a second validation step, the prognostic power of the OPI was confirmed in an independent data set (Duke cohort, p = 0.0063). In multivariate analysis, the OPI was independent of the post-operative residual tumour, the main clinico-pathological prognostic parameter with an adjusted hazard ratio of 6.4 (TOC cohort, CI 1.8-23.5, p = 0.0049) and 1.9 (Duke cohort, CI 1.2-3.0, p = 0.0068). We constructed a combined score of molecular data (OPI) and clinical parameters (residual tumour), which was able to define patient groups with highly significant differences in survival. The integrated analysis of gene expression data as well as residual tumour can be used for optimized assessment of the prognosis of platinum-taxol-treated ovarian cancer. As traditional treatment options are limited, this analysis may be able to optimize clinical management and to identify those patients who would be candidates for new therapeutic strategies.

Aims: Multidrug resistance is a major impediment in chemotherapeutic treatment of ovarian carcinoma patients. The aim of this study was to investigate the expression of multidrug resistance-associated protein 1 (MRP1) and to assess the possible associations with clinicopathological variables and patient outcome in primary ovarian carcinoma. Methods and results: Tumour specimens from 129 patients were obtained before chemotherapy and analysed by immunohistochemistry on tissue microarrays, and by real-time reverse transcriptase-polymerase chain reaction on RNA extracted from formalin-fixed paraffin-embedded tissue specimens using a new technique. Significantly increased MRP1 protein expression was observed in high-grade tumours (P = 0.005) and advanced International Federation of Gynaecology and Obstetrics stages (P = 0.036). On univariate Kaplan–Meier analysis, patients with higher expression of MRP1 protein had significantly decreased overall survival (P = 0.006). On multivariate Cox regression analysis, MRP1 protein expression retained its significance as an independent negative prognostic marker for overall survival (hazard ratio = 6.52, P = 0.003). Furthermore, MRP1 expression correlated with topoisomerase IIα expression both at mRNA and protein level (P