Tracing TET1 expression in prostate cancer: discovery of malignant cells with a distinct oncogenic signature.

BACKGROUND: Ten-eleven translocation methylcytosine dioxygenase 1 (TET1) is involved in DNA demethylation and transcriptional regulation, plays a key role in the maintenance of stem cell pluripotency, and is dysregulated in malignant cells. The identification of cancer stem cells (CSCs) driving tumor growth and metastasis is the primary objective of biomarker discovery in aggressive prostate cancer (PCa). In this context, we analyzed TET1 expression in PCa. METHODS: A large-scale immunohistochemical analysis of TET1 was performed in normal prostate (NOR) and PCa using conventional slides (50 PCa specimens) and tissue microarrays (669 NOR and 1371 PCa tissue cores from 371 PCa specimens). Western blotting, RT-qPCR, and 450 K methylation array analyses were performed on PCa cell lines. Genome-wide correlation, gene regulatory network, and functional genomics studies were performed using publicly available data sources and bioinformatics tools. RESULTS: In NOR, TET1 was exclusively expressed in normal cytokeratin 903 (CK903)-positive basal cells. In PCa, TET1 was frequently detected in alpha-methylacyl-CoA racemase (AMACR)-positive tumor cell clusters and was detectable at all tumor stages and Gleason scores. Pearson's correlation analyses of PCa revealed 626 TET1-coactivated genes (r > 0.5) primarily encoding chromatin remodeling and mitotic factors. Moreover, signaling pathways regulating antiviral processes (62 zinc finger, ZNF, antiviral proteins) and the pluripotency of stem cells were activated. A significant proportion of detected genes exhibited TET1-correlated promoter hypomethylation. There were 161 genes encoding transcription factors (TFs), of which 133 were ZNF-TFs with promoter binding sites in TET1 and in the vast majority of TET1-coactivated genes. CONCLUSIONS: TET1-expressing cells are an integral part of PCa and may represent CSCs with oncogenic potential.

[Current biomarkers for gastric cancer]. / Aktuelle Biomarker beim Magenkarzinom.

Gastric cancer is still a relevant malignant disease with high morbidity and mortality. Current molecular genetic data show that gastric cancer, as other solid tumors as well, is not a single entity but consists of several molecular subtypes of gastric cancer with diverse biology. The increasing understanding of molecular pathways is the basis for innovative therapies. These either directly target altered signaling pathways or genes in tumor cells or as in immune checkpoint inhibitors, indirectly target tumor cells by blocking tumor-induced immune inhibition leading to improvement in the prognosis. The selection of eligible patients is a prerequisite for the successful clinical application of these targeted drugs. Pathologists play an important role in integrating tissue-based biomarkers and established pathological parameters (typing, grading and staging) into one comprehensive morphomolecular report.

[HER2 testing in gastric cancer - results of a German expert meeting]. / HER2-Testung beim Magenkarzinom - Ergebnisse eines deutschen Expertentreffens.

Valid HER2 testing is essential for optimal therapy of patients with HER2 positive gastric cancer and the correct use of first-line treatment. While each breast cancer is routinely being tested for the HER2 status, HER2 testing in gastric cancer has still not become part of the routine and is often only done upon request by the therapist. An interdisciplinary German expert group took the challenges of HER2 testing in gastric cancer as an opportunity to address essential aspects and questions for the practical use of HER2 testing in this indication from the perspective of pathologists and therapists. The recommendations made in this manuscript reflect the consensus of all participants and correspond to their opinions and long-term experience.

[The metastatic niche. Mechanisms and prognostic implications]. / Die metastatische Nische. Mechanismen und prognostische Implikationen.

Disseminated tumor cells require a special microenvironment to form metastases. This metastatic niche is organ specific and forms prior to the establishment of visible metastases. The niche is characterized by vascular remodeling and bone marrow-derived cells which have migrated into it. Studies by other groups and our own results have already shown that intranodal lymphangiogenesis is an important prerequisite for regional lymph node metastases in rectal cancer patients, and can be used as a prognostic marker for progression-free survival. Niche cells such as endothelia secrete factors that attract tumor and bone marrow-derived cells. CXCL12 is one of these factors. CXCL12 activates the CXCR4 chemokine axis and induces migration along its gradient. Several factors, such as hypoxia, have been described to regulate CXCR4 function and surface expression on tumor cells. Low molecular weight agents have been used to block CXCR4 activation. This review focuses on the function and regulation of CXCR4 and its ligand CXCL12 in metastases formation. It also discusses potential options for therapeutic blockage.

Neuropilin-2 and its ligand VEGF-C predict treatment response after transurethral resection and radiochemotherapy in bladder cancer patients.

The standard treatment for invasive bladder cancer is radical cystectomy. In selected patients, bladder-sparing therapy can be performed by transurethral resection (TURBT) and radio-chemotherapy (RCT) or radiotherapy (RT). Our published in vitro data suggest that the Neuropilin-2 (NRP2)/VEGF-C axis plays a role in therapy resistance. Therefore, we studied the prognostic impact of NRP2 and VEGF-C in 247 bladder cancer patients (cN0M0) treated with TURBT and RCT (n = 198) or RT (n = 49) and a follow-up time up to 15 years. A tissue microarray was analyzed by immunohistochemistry. NRP2 expression emerged as a prognostic factor in overall survival (OS; HR: 3.42; 95% CI: 1.48 - 7.86; p = 0.004) and was associated with a 3.85-fold increased risk of an early cancer specific death (95% CI: 0.91 - 16.24; p = 0.066) in multivariate analyses. Cancer specific survival (CSS) dropped from 166 months to 85 months when NRP2 was highly expressed (p = 0.037). Patients with high VEGF-C expression have a 2.29-fold increased risk of shorter CSS (95% CI: 1.03-5.35; p = 0.043) in univariate analysis. CSS dropped from 170 months to 88 months in the case of high VEGF-C expression (p = 0.041). Additionally, NRP2 and VEGF-C coexpression is a prognostic marker for OS in multivariate models (HR: 7.54; 95% CI: 1.57-36.23; p = 0.012). Stratification for muscle invasiveness (T1 vs. T2-T4) confirmed the prognostic role of NRP2 and NRP2/VEGF-C co-expression in patients with T2-T4 but also with high risk T1 disease. In conclusion, immunohistochemistry for NRP2 and VEGF-C has been determined to predict therapy outcome in bladder cancer patients prior to TURBT and RCT.

Impact of the adaptor protein GIPC1/Synectin on radioresistance and survival after irradiation of prostate cancer.

PURPOSE: Studies have shown that GIPC1/Synectin is an essential adaptor protein of receptors that play an important role in cancer progression and therapy resistance. This is the first study to explore the role of GIPC1/Synectin in radioresistance of prostate cancer and as a possible predictive marker for outcome of primary radiation therapy. MATERIALS AND METHODS: The effect of RNA interference-mediated GIPC1/Synectin depletion on clonogenic cell survival after irradiation with 0, 2, 4, or 6 Gy was assayed in two different GIPC1/Synectin-expressing human prostate cancer cell lines. The clinical outcome data of 358 men who underwent radiotherapy of prostate cancer with a curative intention were analyzed retrospectively. Uni- and multivariate analysis was performed of prostate-specific antigen recurrence-free survival and overall survival in correlation with protein expression in pretreatment biopsy specimens. Protein expression was evaluated by standard immunohistochemistry methods. RESULTS: In cell culture experiments, no change was detected in radiosensitivity after depletion of GIPC1/Synectin in GIPC1/Synectin-expressing prostate cancer cell lines. Furthermore, there was no correlation between GIPC1/Synectin expression in human pretreatment biopsy samples and overall or biochemical recurrence-free survival after radiotherapy in a retrospective analysis of the study cohort. CONCLUSION: Our results do not show a predictive or prognostic function of GIPC1/Synectin expression for the outcome of radiotherapy in prostate cancer. Furthermore, our in vitro results do not support a role of GIPC1 in the cellular radiation response. However, the role of GIPC1 in the progression of prostate cancer and its precursors should be subject to further research.

What can molecular pathology offer for optimal decision making?

In recent years, colorectal cancer (CRC), once a uniform disease with well-understood carcinogenesis, has been divided into at least five different subgroups with distinct precursor lesions, pathways of carcinogenesis, morphological, and molecular characteristics. Moreover, new therapeutic concepts with 'targeted' substances have added to the complexity of the management of CRC patients. The clinical value of biomarkers in advanced CRC is indisputable ever since activating mutations of the KRAS oncogene have been shown to predict resistance to anti-epidermal growth factor receptor antibodies. Prognostic biomarkers predicting patient outcomes and predictive biomarkers forecasting response to a certain therapy may help us to improve therapeutic agent selection and patient management with the ultimate goal of maximizing the benefit of treatment and minimizing toxicity. Biomarkers with known implications in advanced CRC will be discussed in this paper.

[Barrett's esophagus. An update]. / Barrett-Ösophagus. Aktueller Stand.

Barrett's esophagus (BE), a well-known complication of gastroesophageal reflux disease (GERD), constitutes a precancerous condition for adenocarcinoma of the distal esophagus. The so-called Barrett's carcinoma shows increasing incidences in countries of the western hemisphere; new data, however, indicate that the rise in incidence is not quite as dramatic as previously assumed. The definition of BE is currently changing: despite good reasons for a purely endoscopic definition of BE, goblet cells are still mandatory for this diagnosis in Germany and the USA. Dysplastic changes in the epithelium are the most important risk factor for the development of Barrett's adenocarcinoma and recently dysplasia was subclassified into a more frequent adenomatous (intestinal) and a non-adenomatous (gastric-foveolar) types. The gold standard for diagnosing dysplasia is still H&E staining. The histological diagnosis of dysplasia is still encumbered by a significant interobserver variability, especially regarding the differentiation between low grade dysplasia and inflammatory/reactive changes and the discrimination between high grade dysplasia and adenocarcinoma. Current data, however, show much higher interobserver agreement in endoscopic resection specimens than in biopsies. Nevertheless, the histological diagnosis of dysplasia should be corroborated by an external second opinion because of its clinical consequences. In endoscopic resections of early Barrett's adenocarcinoma, the pathological report has to include a risk stratification for the likelihood of lymphogenic metastases.

[Bacterial colitis]. / Bakterielle Kolitiden.

The question of whether there are inflammatory changes in colorectal biopsy specimens is frequently asked, especially when the patient reports diarrhea or when the mucosa is reddened on endoscopy. The pathologist first has to find out whether there is, in fact, an increase in the inflammatory infiltrate of the colorectal mucosa which warrants the diagnosis of inflammation. If so, the second challenge is to ascertain the etiology of these inflammatory changes, in particular to differentiate between infectious and non-infectious causes. In principle, we can distinguish forms of colitis with distinct morphological hallmarks confirming the diagnosis (e.g. microscopic detection of the causative organism, as well as lymphocytic or collagenous colitis) from other forms of colitis which have a characteristic pattern of findings not necessarily allowing to deduce the etiology (e.g. infectious colitis without microscopic evidence of the germ vs. inflammatory bowel disease). The present article discusses the pathomorphology and differential diagnosis of the most important forms of bacterial colitis.

[Serrated precursor lesions]. / Serratierte Vorläuferläsionen.

The so-called serrated pathway has in recent years been well established as a second route of colorectal carcinogenesis. Sessile serrated polyps, especially sessile serrated adenomas (SSA) and traditional serrated adenomas (TSA) were identified as precursor lesions of this pathway. Activating mutations in either the BRAF (in SSAs) or the KRAS oncogene (in TSAs) have been determined as the initiating molecular alterations, followed by epigenetic methylation of CpG islands in promoter regions of genes which are implicated in cell cycle control or DNA repair. These findings have led to a paradigm shift in gastrointestinal pathology as lesions without cytological dysplasia, such as SSAs and certain forms of hyperplastic polyps, are now accepted to be precancerous lesions. In addition, carcinomas that have developed through the serrated pathway of colorectal carcinogenesis show varying biological behavior relevant for the clinical management of these tumors depending on the molecular aberrations.

[Standardized and structured histopathological evaluation of colorectal polyps: a practical checklist against the background of the new WHO classification]. / Standardisierte und strukturierte histopathologische Befundung kolorektaler Polypen: Eine Checkliste für die Praxis vor dem Hintergrund der neuen WHO-Klassifikation.

Gastroenterologists removing colorectal polyps expect standardized and well-structured pathological reports, providing them with all relevant data for the further clinical management of the patient. Over the last year, a task force of clinicians and pathologists has developed a checklist to improve and harmonize endoscopic and pathological reporting of colorectal polyps. This checklist concentrates more on concrete recommendations from evidence-based guidelines and established international classifications for daily practice rather than detailed molecular pathological pathways of carcinogenesis. These recommendations are based on the current S3 guidelines for colorectal cancer (the chapter entitled "Management of colorectal polyps"), the histomorphological consensus manuscript of the GI working group of the German Society for Pathology, as well as the current WHO classification for tumors of the digestive system.

[Histopathological diagnosis and differential diagnosis of colorectal serrated polys: findings of a consensus conference of the working group "gastroenterological pathology of the German Society of Pathology"]. / Histopathologische Diagnostik und Differenzialdiagnostik serratierter Polypen im Kolorektum : Ergebnisse einer Konsensuskonferenz der AG "Gastroenterologische Pathologie der DGP"

Until recently, two major types of colorectal epithelial polyps were distinguished: the adenoma and the hyperplastic polyp. While adenomas - because of their cytological atypia - were recognized as precursor lesions for colorectal carcinoma, hyperplastic polyps were perceived as harmless lesions without any potential for malignant progression, mainly because hyperplastic polyps lack cytological atypia. Meanwhile, it is evident that the lesions formerly classified as hyperplastic represent a heterogeneous group of polyps, some of which exhibit a significant risk of neoplastic progression. These lesions show characteristic epigenetic alterations not commonly seen in colorectal adenomas and progress to colorectal carcinoma via the so-called serrated pathway (CIMP pathway). This group of polyps is comprised not only of hyperplastic polyps, but also of sessile serrated adenomas (SSA), traditional serrated adenomas (TSA) and mixed polyps, showing serrated and "classical" adenomatous features. In a consensus conference of the working group of gastroenterological pathology of the German Society of Pathology, standardization of nomenclature and diagnostic criteria as well as recommendations for clinical management of these serrated polyps were formulated and are presented herein.

[Serrated pathway of colorectal carcinogenesis]. / Der serratierte Karzinogeneseweg im Kolorektum.

The serrated pathway of colorectal carcinogenesis is heterogeneous with respect to its precursor lesions, molecular alterations and its prognosis. The low-risk-subtype of serrated adenocarcinomas is less frequent (<20% of all serrated adenocarcinomas) and characterized by proximal location, BRAF-mutation, high CpG-island methylation with loss of MLH1-expression and MSI-H phenotype. The assumed precursor lesion of this subtype is the sessile serrated adenoma and the 5-year overall survival is >70%. The high-risk-subtype is more frequent (>80% of all serrated adenocarcinomas) and characterized by distal location, KRAS mutation, MSI-L/MSS phenotype, lower CpG-island methylation, and possible p53 accumulation. The assumed precursor lesion is the traditional serrated adenoma and the prognosis is unfavorable (<30% 5-year overall survival). The analysis of MSI status, KRAS and BRAF mutational status and immunohistochemical analyses of hMLH1 and p53 expression enables the distinction between these two subtypes and is therefore clinically relevant, especially since treatment options for the two subtypes may differ in the future.

[Intraepithelial neoplasia/dysplasia - histopathology in ulcerative colitis]. / Intraepitheliale Neoplasie/Dysplasie - Diagnose bei Colitis ulcerosa.

About 1-2% of all colorectal carcinomas (CRCs) arise from a background of chronic inflammatory bowel diseases (IBDs) such as ulcerative colitis (UC) or Crohn's disease, and around 15% of patients with IBD die from colorectal cancer. Intraepithelial neoplasia/dysplasia in the setting of IBD is considered a precancerous lesion by definition. Intraepithelial neoplasia may present itself either as a flat or polypoid mucosal lesion and is referred to as a dysplasia-associated lesion or mass (DALM) in the latter case. The therapeutic consequence of high-grade intraepithelial neoplasia/dysplasia and/or DALM is proctocolectomy in most centers because of the high risk of synchronous or metachronous CRC. The diagnosis of ulcerative colitis-related intraepithelial neoplasia and its distinction from regenerative changes and sporadic adenomas (ALMs) occurring in UC is still one of the greatest challenges in gastrointestinal pathology. Intra- and interobserver variability for the distinction between low-grade intraepithelial neoplasia and regenerative lesions tends to be quite high. This is partly due to the difficult histomorphology and partly due to the relative rareness of this diagnosis (approximately 10% of patients with IBD).

[Molecular genetic markers for prostate cancer. Evidence in fine needle biopsies for improved confirmation of the diagnosis]. / Molekulargenetische Marker des Prostatakarzinoms. Nachweis in Feinnadelbiopsien zur besseren Diagnoseabsicherung.

Selected transcript markers as well as their combinations were analyzed on minimal prostate tissue specimens with regard to their diagnostic potential. Artificial prostate biopsies from RPE explants were used for evaluation and optimization of the techniques used followed by application to diagnostic prostate needle core biopsies. Minimal prostate specimens were cryopreserved and processed with standardized methods. The RNA amount of a half of each biopsy was sufficient for the analysis of 11 marker genes and one reference gene (TBP) using quantitative PCR assays.The relative transcript amounts obtained were included in several analyses including calculations for each single marker gene like median overexpression rate as well as marker combinations. Two optimized mathematical models based on relative expression levels of EZH2, hepsin, PCA3, prostein, and TRPM8 were evaluated with regard to their diagnostic potential. Compared to single marker analyses these models show higher sensitivity and specificity for prostate cancer detection.Thus biomolecular prostate cancer identification may represent a suitable diagnostic tool to supplement conventional techniques on prostate biopsies. Furthermore, an extension of this approach to PCa prognosis and the transfer to urine samples appear very promising.

[Intraepithelial neoplasia in ulcerative colitis: on the way to more diagnostic confidence]. / Intraepitheliale Neoplasie bei Colitis ulcerosa: Wege zu mehr Diagnosesicherheit.

Ulcerative colitis (UC)-related intraepithelial neoplasia and its distinction from regenerative changes and sporadic adenomas occurring in UC is one of the greatest challenges in gastrointestinal pathology. Recently, the molecular changes in UC-related neoplastic progression have been determined and compared with the molecular changes in sporadic carcinogenesis. Diagnostically promising differences between sporadic and UC-related carcinogenesis are the advent of genetic changes in non-neoplastic UC-related mucosa and the early loss of 18q (harbouring SMAD2, SMAD4, and DCC) and 17p (site of p53) in UC-related tumorigenesis. These studies have given rise to a number of adjunct methods in the determination of UC-related neoplasia. Never the less, conventional histopathology still remains the gold standard in the diagnosis of UC-related neoplasia. Training of histopathologists therefore is one of the most important issues in conquering the diagnostic challenges of UC-related neoplasia. The working group "Gastrointestinal Pathology" of the German Society for Pathology set up a diagnostic multicenter trial which was open to everyone interested. The interobserver variability regarding ulcerative colitis-related neoplasia was quite promising (kappa = 0.63). A consensus diagnosis was reached for all the specimens and diagnostic criteria for UC-related neoplasia were discussed, reevaluated, and agreed on. Adjunct methods and emerging markers for the diagnosis of ulcerative colitis-related neoplasia (p53, Ki67, AMACR) and its distinction from regenerative changes and sporadic adenomas occurring in UC (ALM) will be presented and discussed.

[GIPC: a new target for therapy in pancreatic adenocarcinoma?]. / Ist GIPC ein neuer Kandidat für die Therapie des Pankreaskarzinoms?

GIPC is highly expressed in human pancreatic adenocarcinoma and is a central protein for the stability of IGF-1R in pancreatic adenocarcinoma cell lines (15). The goal of this study was to prove the importance of GIPC in vivo and to evaluate possible therapeutic strategies that target this protein and its PDZ domain. In vivo effects of GIPC knockout were studied after lentiviral transduction of luciferase-expressing MiaPaCa2 pancreatic cancer cells with shRNA against GIPC; growth characteristics were monitored with bioluminiscence. Knockdown of GIPC led to a significant inhibition of pancreatic tumor cell growth in vivo in different mouse models. To test a possible therapeutic approach, the PDZ domain of GIPC was targeted by a short peptide composed of the amino acid sequence PSQSSSEA. This octapeptide was designed based on the C-terminal binding motif of GAIP. Targeting GIPC with this peptide inhibited the association between IGF-1R and GIPC. The subsequent downregulation of IGF-1R decreased proliferation in vitro and in vivo. In conclusion, our findings suggest that targeting GIPC and its PDZ domain-mediated interaction with the tyrosine kinase receptor IGF-1R could be a promising new treatment option for pancreatic cancer.