Periostin is up-regulated in high grade and high stage prostate cancer

BACKGROUND: Expression of periostin is an indicator of epithelial-mesenchymal transition in cancer but a detailed analysis of periostin expression in prostate cancer has not been conducted so far. METHODS: Here, we evaluated periostin expression in prostate cancer cells and peritumoural stroma immunohistochemically in two independent prostate cancer cohorts, including a training cohort (n = 93) and a test cohort (n = 325). Metastatic prostate cancers (n = 20), hormone refractory prostate cancers (n = 19) and benign prostatic tissues (n = 38) were also analyzed. RESULTS: In total, strong epithelial periostin expression was detectable in 142 of 418 (34.0%) of prostate carcinomas and in 11 of 38 benign prostate glands (28.9%). Increased periostin expression in carcinoma cells was significantly associated with high Gleason score (p < 0.01) and advanced tumour stage (p < 0.05) in the test cohort. Whereas periostin expression was weak or absent in the stroma around normal prostate glands, strong periostin expression in tumour stroma was found in most primary and metastatic prostate cancers. High stromal periostin expression was associated with higher Gleason scores (p < 0.001). There was a relationship between stromal periostin expression and shortened PSA relapse free survival times in the training cohort (p < 0.05). CONCLUSIONS: Our data indicate that periostin up-regulation is related to increased tumour aggressiveness in prostate cancer and might be a promising target for therapeutical interventions in primary and metastatic prostate cancer

Skin Electroporation: Effects on Transgene Expression, DNA Persistence and Local Tissue Environment

BACKGROUND: Electrical pulses have been used to enhance uptake of molecules into living cells for decades. This technique, often referred to as electroporation, has become an increasingly popular method to enhance in vivo DNA delivery for both gene therapy applications as well as for delivery of vaccines against both infectious diseases and cancer. In vivo electrovaccination (gene delivery followed by electroporation) is currently being investigated in several clinical trials, including DNA delivery to healthy volunteers. However, the mode of action at molecular level is not yet fully understood. METHODOLOGY/PRINCIPAL FINDINGS: This study investigates intradermal DNA electrovaccination in detail and describes the effects on expression of the vaccine antigen, plasmid persistence and the local tissue environment. Gene profiling of the vaccination site showed that the combination of DNA and electroporation induced a significant up-regulation of pro-inflammatory genes. In vivo imaging of luciferase activity after electrovaccination demonstrated a rapid onset (minutes) and a long duration (months) of transgene expression. However, when the more immunogenic prostate specific antigen (PSA) was co-administered, PSA-specific T cells were induced and concurrently the luciferase expression became undetectable. Electroporation did not affect the long-term persistence of the PSA-expressing plasmid. CONCLUSIONS/SIGNIFICANCE: This study provides important insights to how DNA delivery by intradermal electrovaccination affects the local immunological responses of the skin, transgene expression and clearance of the plasmid. As the described vaccination approach is currently being evaluated in clinical trials, the data provided will be of high significance

Functional characterization of prostate cancer biomarkers

With the onset of prostate-specific antigen (PSA) screening two decades ago the number of prostate cancer suspicious cases has markedly increased. Due to the lack of specificity men with elevated PSA levels need to be biopsied to confirm the diagnosis. In order to reduce the number of the unnecessary taking of biopsies, new specific and non-invasive biomarkers are required to facilitate prostate cancer diagnosis. Here, we suggest calcium-activated nucleotidase (CANT) 1 as a novel serum marker candidate for further validation. As a result of serum PSA measurement, also the number of confirmed prostate cancer cases has risen. Especially more insignificant carcinomas are detected, which would never have affected the patients’ life. Thus, overtreatment of these cases and the therapy-induced morbidity is a big issue in clinical prostate cancer management. Within this study, we identify two potential prognostic biomarkers to predict the risk of disease recurrence. This risk is reduced in patients with very high CANT1 protein levels in their prostatectomy specimens. Applying the histochemical score to quantify CANT1 protein levels, CANT1 is shown to be an independent prognostic biomarker. In contrast, strong forkhead box (FOX) A1 expression is prognostically unfavourable. Since an ideal prognostic marker should be applied to pre- treatment samples, analysis of both immunohistochemical markers in large biopsy cohorts is strongly recommended. Once the tumor has recurred, progression to castration-resistance is likely. Further elucidation of the underlying mechanisms is necessary to progress in the development of life- prolonging therapies for these patients. Here, we show that despite ubiquitous overexpression of Golgi membrane protein (GOLM) 1 in prostate carcinomas, this protein has no functional relevance for the analyzed cancer cells. In contrast, knockdown of FOXA1 reduces in vitro tumorigenicity. Moreover, FOXA1 expression increases with prostate cancer progression and is associated with poor prognosis, especially in cases with low androgen receptor levels. Thus, we conclude that FOXA1 upregulation is a novel mechanism of castration-resistance for prostate cancer cells. Additionally, we reveal that CANT1 expression is already elevated in the precursor lesion prostatic intraepithelial neoplasia. In vitro experiments establish the secretion of CANT1 as well as the reduction of in vitro tumorigenicity accompanied by G1 cell cycle arrest upon CANT1 knockdown. Furthermore, induction of selected unfolded protein response target genes and enhancement of CANT1 mRNA expression upon stress induction in the endoplasmic reticulum (ER) are indicated. We thus propose that CANT1 is upregulated in prostate carcinomas to counteract tumor- associated ER stress. According to our model, this might be achieved by the interplay between soluble truncated CANT1, which prevents calcium release from the ER, and full- length CANT1, which regulates protein folding. Since knockdown of CANT1 heavily impaired proliferation of the hormone-independent cell line PC-3, which is a model for castration- resistant prostate carcinomas, CANT1 might be an interesting therapeutic target for this group of prostate carcinomas. Zusammenfassung Seit Beginn des prostata-spezifischen Antigen (PSA)-Screenings vor zwei Jahrzehnten ist die Zahl der Prostatakrebs-Verdachtsfälle deutlich angestiegen. Aufgrund der fehlenden Spezifität müssen Männer mit einem erhöhten PSA-Level biopsiert werden, um die Diagnose zu bestätigen. Zur Reduktion der unnötigen Entnahmen von Biopsien werden neue spezifische und nicht-invasive Biomarker benötigt, die die Prostatakrebsdiagnose vereinfachen. In der vorliegenden Arbeit schlagen wir Calcium-aktivierte Nukleotidase (CANT) 1 als neuen Serummarkerkandidaten für die weitere Evaluierung vor. Als Folge der Serum-PSA-Messung ist auch die Zahl bestätigter Prostatakarzinomfälle gestiegen. Dabei werden hauptsächlich mehr insignifikante Karzinome detektiert, die niemals das Leben der Patienten beeinträchtigt hätten. Daher ist die Überbehandlung dieser Fälle und die therapieinduzierte Morbidität ein großes Thema im klinischen Management von Prostatakrebs. In dieser Studie identifizieren wir zwei potentielle prognostische Biomarker zur Vorhersage des Rezidivrisikos. Dieses Risiko ist in Patienten, deren Prostatektomie- präparate sehr hohe CANT1-Proteinmengen aufweisen, reduziert. Es wird gezeigt, dass CANT1 unter Anwendung des histochemischen Scores für die Quantifizierung von CANT1- Proteinmengen ein unabhängiger prognostischer Biomarker ist. Im Gegensatz dazu ist eine starke Expression von Forkhead Box (FOX) A1 prognostisch ungünstig. Da ein idealer prognostischer Marker an Proben, die vor der ersten Behandlung entnommen wurden, angewendet werden sollte, wird die Analyse von beiden immunhistochemischen Markern in großen Biopsiekohorten empfohlen. Sobald ein Rezidiv des Tumors auftritt, ist die Progression zur Kastrationsresistenz sehr wahrscheinlich. Die weitere Aufklärung der darunterliegenden Mechanismen ist notwendig, um die Entwicklung lebensverlängernder Therapien für diese Patienten voranzutreiben. Hier zeigen wir, dass Golgimembranprotein (GOLM) 1 trotz ubiquitärer Überexpression in Prostatakarzinomen keine funktionelle Relevanz für die analysierten Krebszellen hat. Im Gegensatz dazu reduziert die Herunterregulierung von FOXA1 die in vitro-Tumorigenität. Darüber hinaus steigt die Expression von FOXA1 mit der Progression des Prostatakarzinoms und ist mit einer schlechten Prognose assoziiert, besonders in Fällen mit niedrigen Androgenrezeptormengen. Daher schlussfolgern wir, dass FOXA1-Aufregulation einen neuartigen Mechanismus der Kastrationsresistenz von Prostatakrebszellen darstellt. Zusätzlich weisen wir eine erhöhte CANT1-Expression schon in der Vorläuferläsion der prostatischen intraepithelialen Neoplasie nach. In vitro-Experimente zeigen die Sekretion von CANT1 sowie eine Reduktion der in vitro-Tumorigenität nach CANT1-Herunterregulierung, die von einem G1-Zellzyklusarrest begleitet wird. Weiterhin deutet sich eine Induktion von ausgewählten Zielgenen der sogenannten „unfolded protein response“ sowie eine Verstärkung der CANT1-mRNA-Expression nach Stressinduktion im endoplasmatischen Retikulum (ER) an. Daher postulieren wir, dass CANT1 in Prostatakarzinomen aufreguliert ist, um dem tumorassoziierten ER-Stress entgegenzuwirken. Unserem Modell zufolge könnte dies erreicht werden durch ein Zusammenspiel zwischen löslichem abgespaltenem CANT1, welches die Calcium-Freisetzung im ER verhindert, und membranverankertem CANT1, welches die Proteinfaltung reguliert. Da die Herunterregulierung von CANT1 die Proliferation der hormonunabhängigen Zelllinie PC-3, welche als Modell für kastrationsresistente Prostatakarzinome gilt, sehr stark beeinträchtigt, könnte CANT1 ein interessantes therapeutisches Zielprotein für diese Gruppe von Prostatakarzinomen sein

Prognostic significance of AGR2 in pancreatic ductal adenocarcinoma

Background/Aims: The human Anterior Gradient-2 (AGR2) is strongly upregulated in various human cancers, including pancreatic ductal adenocarcinomas (PDAC), but its prognostic value in PDAC has not yet been studied. Methods: We analysed 19 microdissected PDAC cases at the mRNA level, and also 148 cases at the protein level by immunohistochemistry based on tissue microarray, using a monoclonal AGR2 antibody, and statistical analyses were applied to test for prognostic associations. Results: Overexpression of AGR2 mRNA was found to be elevated in most pancreatic cell lines and in microdissected pancreatic cancer compared to microdissected normal ductal cells. AGR2 protein was expressed in 109/148 (73.7%) of PDAC, with a higher expression in female patients (p=0.040), whereas no significant associations with other clinical-pathological parameters were found. A prognostic value of AGR2 could not be demonstrated in univariate analyses. Conclusion: Although a prognostic value of AGR2 seems unlikely, further studies are warranted to investigate the biological role of AGR2 in pancreatic adenocarcinomas

Diagnostic and prognostic value of T-cell receptor gamma alternative reading frame protein (TARP) expression in prostate cancer

T-cell receptor gamma chain alternative reading frame protein (TARP) has recently been proposed as being up-regulated in prostate cancer (PCA). Additionally, TARP has been proposed as a potential therapeutic target for cancer therapy. We analysed the protein expression of TARP in a large well characterised prostate cancer cohort to assess its diagnostic and prognostic value. Methodologically, we constructed a tissue microarray comprising more than 600 PCA cases including matching benign prostate tissue. TARP protein expression was carefully analysed and associated with clinico-pathological parameters, PSA-relapse free survival and expression data of established and proposed diagnostic markers (AMACR, p63, GOLPH2). Our results show that TARP is significantly over-expressed in the vast majority (~85%) of PCA in comparison to non neoplastic prostate tissue. Its expression was associated with conventional markers of unfavourable and more aggressive tumour behaviour. However, a prognostic value of TARP could not be found. The diagnostic value of TARP is limited in comparison to AMACR, p63 or GOLPH2. Since TARP specific immunologic therapy regimen are currently being tested, the high frequency of TARP overexpression in PCA conveys a high potential for a predictive and potentially therapeutic use of this biomarker

Insulin-like growth factor II mRNA binding protein 3 (IMP3) is overexpressed in prostate cancer and correlates with higher Gleason scores

BACKGROUND: The oncofetal protein insulin-like growth factor II mRNA binding protein 3 (IMP3) is an important factor for cell-migration and adhesion in malignancies. Recent studies have shown a remarkable overexpression of IMP3 in different human malignant neoplasms and also revealed it as an important prognostic marker in some tumor entities. To our knowledge, IMP3 expression has not been investigated in prostate carcinomas so far. METHODS: Immunohistochemical stainings for IMP3 were performed on tissue microarray (TMA) organized samples from 507 patients: 31 normal prostate tissues, 425 primary carcinomas and 51 prostate cancer metastases or castration-resistant prostate cancers (CRPC). IMP3 immunoreactivity was semiquantitatively scored and correlated with clinical-pathologic parameters including survival. RESULTS: IMP3 is significantly stronger expressed in prostate carcinomas compared to normal prostate tissues (p < 0.0001), but did not show significant correlation with the pT-stage, the proliferation index (MIB1), preoperative serum PSA level and the margin status. Only a weak and slightly significant correlation was found with the Gleason score and IMP3 expression failed to show prognostic significance in clinico-pathological correlation-analyses. CONCLUSIONS: Although IMP3 is overexpressed in a significant proportion of prostate cancer cases, which might be of importance for novel therapeutic approaches, it does not appear to possess any immediate diagnostic or prognostic value, limiting its potential as a tissue biomarker for prostate cancer. These results might be corroborated by the fact, that two independent tumor cohorts were separately reviewed

The Androgen-Regulated Calcium-Activated Nucleotidase 1 (CANT1) Is Commonly Overexpressed in Prostate Cancer and Is Tumor-Biologically Relevant in Vitro

Previously, we identified the calcium-activated nucleotidase 1 (CANT1) transcript as up-regulated in prostate cancer. Now, we studied CANT1 protein expression in a large cohort of nearly 1000 prostatic tissue samples including normal tissue, prostatic intraepithelial neoplasia (PIN), primary carcinomas, metastases, and castrate-resistant carcinomas, and further investigated its functional relevance. CANT1 displayed predominantly a Golgi-type immunoreactivity with additional and variable cytoplasmic staining. In comparison to normal tissues, the staining intensity was significantly increased in PIN lesions and cancer. In cancer, high CANT1 levels were associated with a better prognosis, and castrate-resistant carcinomas commonly showed lower CANT1 levels than primary carcinomas. The functional role of CANT1 was investigated using RNA interference in two prostate cancer cell lines with abundant endogenous CANT1 protein. On CANT1 knockdown, a significantly diminished cell number and DNA synthesis rate, a cell cycle arrest in G1 phase, and a strong decrease of cell transmigration rate and wound healing capacity of CANT1 knockdown cells was found. However, on forced CANT1 overexpression, cell proliferation and migration remained unchanged. In summary, CANT1 is commonly overexpressed in the vast majority of primary prostate carcinomas and in the precursor lesion PIN and may represent a novel prognostic biomarker. Moreover, this is the first study to demonstrate a functional involvement of CANT1 in tumor biology