Expression Patterns and Corepressor Function of Retinoic Acid-induced 2 in Prostate Cancer.

BACKGROUND: Revealing molecular mechanisms linked to androgen receptor activity can help to improve diagnosis and treatment of prostate cancer. Retinoic acid-induced 2 (RAI2) protein is thought to act as a transcriptional coregulator involved in hormonal responses and epithelial differentiation. We evaluated the clinical relevance and biological function of the RAI2 protein in prostate cancer. METHODS: We assessed RAI2 gene expression in the Cancer Genome Atlas prostate adenocarcinoma PanCancer cohort and protein expression in primary tumors (n = 199) by immunohistochemistry. We studied RAI2 gene expression as part of a multimarker panel in an enriched circulating tumor cell population isolated from blood samples (n = 38) of patients with metastatic prostate cancer. In prostate cancer cell lines, we analyzed the consequences of androgen receptor inhibition on RAI2 protein expression and the consequences of RAI2 depletion on the expression of the androgen receptor and selected target genes. RESULTS: Abundance of the RAI2 protein in adenocarcinomas correlated with the androgen receptor; keratins 8, 18, and 19; and E-cadherin as well as with an early biochemical recurrence. In circulating tumor cells, detection of RAI2 mRNA significantly correlated with gene expression of FOLH1, KLK3, RAI2, AR, and AR-V7. In VCaP and LNCaP cell lines, sustained inhibition of hormone receptor activity induced the RAI2 protein, whereas RAI2 depletion augmented the expression of MME, STEAP4, and WIPI1. CONCLUSIONS: The RAI2 protein functions as a transcriptional coregulator of the androgen response in prostate cancer cells. Detection of RAI2 gene expression in blood samples from patients with metastatic prostate cancer indicated the presence of circulating tumor cells.

EGFR as a stable marker of prostate cancer dissemination to bones.

BACKGROUND: Prostate cancer (PCa) is among the most commonly diagnosed malignancies in men. Although 5-year survival in patients with localised disease reaches nearly 100%, metastatic disease still remains incurable. Therefore, there is a need for markers indicating metastatic dissemination. METHODS: EGFR overexpression (EGFRover) was tracked in 1039 primary tumours, circulating tumour cells from 39 d'Amico high-risk patients and metastatic samples from 21 castration-resistant PCa cases. EGFR status was compared to clinical parameters and multiple molecular factors were assessed using immunohistochemistry and gene ontology analysis. The functional aspect of EGFR was evaluated by plating PC-3 cells on soft and rigid matrices. RESULTS: EGFRover was found in 14% of primary tumours, where it was associated with shorter metastasis-free survival and was an independent indicator of worse overall survival. EGFRover correlated with a pro-migratory and pro-metastatic phenotype of tumour cells as well as rich collagen fibre content. All circulating tumour cells (detected in 13% of cases) were positive for EGFR, independent of their EMT-related phenotype. EGFRover was more prevalent in castration-resistant bone metastases (29% of patients) and supported growth of human PCa cells on rigid matrices mimicking bone stiffness. CONCLUSIONS: EGFRover is a stable, EMT-independent marker of PCa disseminating to rigid organs, preferentially bones.

Somatic aberrations of BRCA1 gene are associated with ALDH1, EGFR, and tumor progression in prostate cancer.

BRCA1 is a pivotal tumor suppressor. Its dysfunction is known to play a role in different tumors. Among others, BRCA1 germline mutations account for higher risk and more aggressive course of prostate cancer (PCa). In addition, somatic BRCA1 gene loss was demonstrated to be a signature of PCa dissemination to lymph nodes and peripheral blood, and indicate worse clinical outcome. In order to substantiate the data for BRCA1 gene loss in PCa and reveal its phenotypical background, BRCA1 gene status was assessed in a large cohort of PCa patients and compared to different molecular factors. BRCA1 gene dosage was assessed in 2398 tumor samples from 1,199 PCa patients using fluorescent in situ hybridization. It was compared to clinico-pathological parameters, patients' outcome as well as selected proteins (Ki-67, apoptosis marker, cytokeratins, vimentin, E- and N-cadherin, ALDH1 and EGFR) examined immunohistochemically. BRCA1 losses were found in 10%, whereas gains appeared in 7% of 603 informative PCa patients. BRCA1 losses correlated to higher T stage (p = 0.027), Gleason score (p = 0.039), shorter time to biochemical recurrence in patients with Gleason score > 7 independently of other factors (multivariate analysis, p = 0.005) as well as expression of proteins regulating stemness and epithelial-mesenchymal transition, that is, ALDH1 (p = 0.021) and EGFR (p = 0.011), respectively. BRCA1 gains correlated to shorter time to metastasis (p = 0.012) and expression of ALDH1 (p = 0.014). These results support the assumption that BRCA1 gene losses contribute to a progressive and stem cell-like phenotype of PCa. Furthermore, they reveal that also BRCA1 gain conceivably representing loss-of-function might mark more invasive tumors.

Blind Biobanking of the Prostatectomy Specimen: Critical Evaluation of the Existing Techniques and Development of the New 4-Level Tissue Extraction Model With High Sampling Efficacy.

BACKGROUND: Fresh tissue is mandatory to perform high-quality translation studies. Several models for tissue extraction from prostatectomy specimens without guidance by frozen sections are already introduced. However, little is known about the sampling efficacy of these models, which should provide representative tissue in adequate volumes, account for multifocality and heterogeneity of tumor, not violate the routine final pathological examination, and perform quickly without frozen section-based histological control. The aim of the study was to evaluate the sampling efficacy of the existing tissue extraction models without guidance by frozen sections ("blind") and to develop an optimized model for tissue extraction. METHODS: Five hundred thirty-three electronic maps of the tumor distribution in prostates from a single-center cohort of the patients subjected to radical prostatectomy were used for analysis. Six available models were evaluated in silico for their sampling efficacy. Additionally, a novel model achieving the best sampling efficacy was developed. RESULTS: The available models showed high efficacies for sampling "any part" from the tumor (up to 100%), but were uniformly low in efficacy to sample all tumor foci from the specimens (with the best technique sampling only 51.6% of the all tumor foci). The novel 4-level extraction model achieved a sampling efficacy of 93.1% for all tumor foci. CONCLUSIONS: The existing "blind" tissue extraction models from prostatectomy specimens without frozen sections control are suitable to target tumor tissues but these tissues do not represent the whole tumor. The novel 4-level model provides the highest sampling efficacy and a promising potential for integration into routine. Prostate 77: 396-405, 2017. © 2016 Wiley Periodicals, Inc.

Analysis of topographical distribution of prostate cancer and related pathological findings in prostatectomy specimens using cMDX document architecture.

INTRODUCTION: Understanding the topographical distribution of prostate cancer (PCa) foci is necessary to optimize the biopsy strategy. This study was done to develop a technical approach that facilitates the analysis of the topographical distribution of PCa foci and related pathological findings (i.e., Gleason score and foci dimensions) in prostatectomy specimens. MATERIAL & METHODS: The topographical distribution of PCa foci and related pathologic evaluations were documented using the cMDX documentation system. The project was performed in three steps. First, we analyzed the document architecture of cMDX, including textual and graphical information. Second, we developed a data model supporting the topographic analysis of PCa foci and related pathologic parameters. Finally, we retrospectively evaluated the analysis model in 168 consecutive prostatectomy specimens of men diagnosed with PCa who underwent total prostate removal. The distribution of PCa foci were analyzed and visualized in a heat map. The color depth of the heat map was reduced to 6 colors representing the PCa foci frequencies, using an image posterization effect. We randomly defined 9 regions in which the frequency of PCa foci and related pathologic findings were estimated. RESULTS: Evaluation of the spatial distribution of tumor foci according to Gleason score was enabled by using a filter function for the score, as defined by the user. PCa foci with Gleason score (Gls) 6 were identified in 67.3% of the patients, of which 55 (48.2%) also had PCa foci with Gls between 7 and 10. Of 1173 PCa foci, 557 had Gls 6, whereas 616 PCa foci had Gls>6. PCa foci with Gls 6 were mostly concentrated in the posterior part of the peripheral zone of the prostate, whereas PCa foci with Gls>6 extended toward the basal and anterior parts of the prostate. The mean size of PCa foci with Gls 6 was significantly lower than that of PCa with Gls>6 (P<0.0001). CONCLUSION: The cMDX-based technical approach facilitates analysis of the topographical distribution of PCa foci and related pathologic findings in prostatectomy specimens.

Prostate cancers detected on repeat prostate biopsies show spatial distributions that differ from those detected on the initial biopsies.

OBJECTIVE: To evaluate the spatial distribution of prostate cancer detected at a single positive biopsy (PBx) and a repeat PBx (rPBx). PATIENTS AND METHODS: We evaluated 533 consecutive men diagnosed with prostate cancer who underwent radical prostatectomy using a clinical map document based on XML (cMDX©)-based map model of the prostate. We determined the number of cancer foci, relative tumour volume, Gleason score, zone of origin, localisation, and pathological stage after stratification according to the number of PBx sessions (PBx vs rPBx). The distribution of 3966 prostate cancer foci was analysed and visualised on heat maps. The colour gradient of the heat map was reduced to six colours representing the frequency classification of prostate cancer using an image posterisation effect. Additionally, the spatial distribution of organ-confined prostate cancer between PBx and rPBx was evaluated. RESULTS: Prostate cancer diagnosed on PBx was mostly localised to the apical portion and the peripheral zone of the prostate. Prostate cancer diagnosed on rPBx was more frequently found in the anterior portion and the base of the prostate. Organ-confined prostate cancer foci were mostly localised in the dorsolateral zone of the prostate in men at PBx, whereas men at rPBx had more prostate cancer foci in the anterior portion. CONCLUSIONS: The spatial distribution of prostate cancer with rPBx differs significantly from the spatial distribution of prostate cancer with PBx. The whole anterior portion of the prostate should be considered by rPBx.

Exploring prostate cancer genome reveals simultaneous losses of PTEN, FAS and PAPSS2 in patients with PSA recurrence after radical prostatectomy.

The multifocal nature of prostate cancer (PCa) creates a challenge to patients' outcome prediction and their clinical management. An approach that scrutinizes every cancer focus is needed in order to generate a comprehensive evaluation of the disease, and by correlating to patients' clinico-pathological information, specific prognostic biomarker can be identified. Our study utilized the Affymetrix SNP 6.0 Genome-wide assay to investigate forty-three fresh frozen PCa tissue foci from twenty-three patients. With a long clinical follow-up period that ranged from 2.0-9.7 (mean 5.4) years, copy number variation (CNV) data was evaluated for association with patients' PSA status during follow-up. From our results, the loss of unique genes on 10q23.31 and 10q23.2-10q23.31 were identified to be significantly associated to PSA recurrence (p < 0.05). The implication of PTEN and FAS loss (10q23.31) support previous reports due to their critical roles in prostate carcinogenesis. Furthermore, we hypothesize that the PAPSS2 gene (10q23.2-10q23.31) may be functionally relevant in post-operative PSA recurrence because of its reported role in androgen biosynthesis. It is suggestive that the loss of the susceptible region on chromosome 10q, which implicates PTEN, FAS and PAPSS2 may serve as genetic predictors of PSA recurrence after radical prostatectomy.

Multiple recognition assay reveals prostasomes as promising plasma biomarkers for prostate cancer.

Prostasomes are microvesicles (mean diameter, 150 nm) that are produced and secreted by normal and malignant prostate acinar cells. It has been hypothesized that invasive growth of malignant prostate cells may cause these microvesicles, normally released into seminal fluid, to appear in interstitial space and therewith into peripheral circulation. The suitability of prostasomes as blood biomarkers in patients with prostate cancer was tested by using an expanded variant of the proximity ligation assay (PLA). We developed an extremely sensitive and specific assay (4PLA) for detection of complex target structures such as microvesicles in which the target is first captured via an immobilized antibody and subsequently detected by using four other antibodies with attached DNA strands. The requirement for coincident binding by five antibodies to generate an amplifiable reporter results in both increased specificity and sensitivity. The assay successfully detected significantly elevated levels of prostasomes in blood samples from patients with prostate cancer before radical prostatectomy, compared with controls and men with benign biopsy results. The medians for prostasome levels in blood plasma of patients with prostate cancer were 2.5 to sevenfold higher compared with control samples in two independent studies, and the assay also distinguished patients with high and medium prostatectomy Gleason scores (8/9 and 7, respectively) from those with low score (≤ 6), thus reflecting disease aggressiveness. This approach that enables detection of prostasomes in peripheral blood may be useful for early diagnosis and assessment of prognosis in organ-confined prostate cancer.

From Satirical Poems and Invisible Poisons to Radical Surgery and Organized Cervical Cancer Screening-A Historical Outline of Cervical Carcinoma and Its Relation to HPV Infection.

Over the last century, the narrative of cervical cancer history has become intricately tied to virus research, particularly the human papillomavirus (HPV) since the 1970s. The unequivocal proof of HPV's causal role in cervical cancer has placed its detection at the heart of early screening programs across numerous countries. From a historical perspective, sexually transmitted genital warts have been already documented in ancient Latin literature; the remarkable symptoms and clinical descriptions of progressed cervical cancer can be traced back to Hippocrates and classical Greece. However, in the new era of medicine, it was not until the diagnostic-pathological accomplishments of Aurel Babes and George Nicolas Papanicolaou, as well as the surgical accomplishments of Ernst Wertheim and Joe Vincent Meigs, that the prognosis and prevention of cervical carcinoma were significantly improved. Future developments will likely include extended primary prevention efforts consisting of better global access to vaccination programs as well as adapted methods for screening for precursor lesions, like the use of self-sampling HPV-tests. Furthermore, they may also advantageously involve additional novel diagnostic methods that could allow for both an unbiased approach to tissue diagnostics and the use of artificial-intelligence-based tools to support decision making.

High-grade prostatic intraepithelial neoplasia (HGPIN) and topographical distribution in 1,374 prostatectomy specimens: existence of HGPIN near prostate cancer.

PURPOSE: High-grade prostatic intraepithelial neoplasia (HGPIN) is believed to be a precursor of prostate cancer (PCa). This study evaluated whether HGPIN was located close to PCa in whole radical prostatectomy specimens (RPSs). MATERIALS AND METHODS: We evaluated 1,374 prostate specimens from 1999 to 2010 using a cMDX-based map model of the prostate. The distribution of 10,439 PCa foci was analyzed and visualized on a heat map. The color gradient of the heat map was reduced to six colors representing the frequency classification of the relative frequency of PCa using an image posterization effect. We defined 22 regions in the prostate according to the frequency of PCa occurrence. Seven hundred ninety RPSs containing 6,374 PCa foci and 4,502 HGPIN foci were evaluated. The topographical association between PCa and HGPIN in the RPSs was analyzed by estimating the frequencies of PCa and HGPIN in 22 regions. A logistic regression analysis was performed to assess the odds ratios of HGPIN for the presence of PCa in 22 regions. RESULTS: Fifty-eight percent of PCa specimens included HGPIN and had significantly more favorable Gleason scores, lower PSA levels and smaller relative tumor volumes than isolated PCa specimens. HGPIN (68%) and PCa (69%) were predominantly localized to the apical half of the prostate. HGPIN was mainly concentrated in the peripheral zone medial to regions with high PCa frequencies. Upon logistic regression analysis, HGPIN was a significant predictor of PCa co-existence in 11 regions. CONCLUSIONS: HGPIN was located adjacent to PCa in whole RPSs. PCa concomitant with HGPIN had more favorable pathologic features than isolated PCa.

Genome-wide investigation of multifocal and unifocal prostate cancer-are they genetically different?

Prostate cancer is widely observed to be biologically heterogeneous. Its heterogeneity is manifested histologically as multifocal prostate cancer, which is observed more frequently than unifocal prostate cancer. The clinical and prognostic significance of either focal cancer type is not fully established. To investigate prostate cancer heterogeneity, the genetic profiles of multifocal and unifocal prostate cancers were compared. Here, we report observations deduced from tumor-tumor comparison of copy number alteration data of both focal categories. Forty-one fresh frozen prostate cancer foci from 14 multifocal prostate cancers and eight unifocal prostate cancers were subjected to copy number variation analysis with the Affymetrix SNP 6.0 microarray tool. With the investigated cases, tumors obtained from a single prostate exhibited different genetic profiles of variable degrees. Further comparison identified no distinct genetic pattern or signatures specific to multifocal or unifocal prostate cancer. Our findings suggest that samples obtained from multiple sites of a single unifocal prostate cancer show as much genetic heterogeneity and variability as separate tumors obtained from a single multifocal prostate cancer.

MicroRNA profiles of prostate carcinoma detected by multiplatform microRNA screening.

MicroRNAs (miRNAs) are small RNA molecules that regulate gene expression via posttranscriptional inhibition of protein synthesis. They play a vital role in tumorigenesis. To characterize the diagnostic potential of miRNAs in prostate cancer, a leading cause of cancer mortality, we performed screening of miRNA expression profiles. We used commercially available microarrays to establish miRNA expression profiles from a cohort of 20 cancer samples. The expression of selected miRNAs was analyzed by quantitative real-time PCR and the identity of miRNA expressing cells was determined by miRNA in situ hybridization. We identified 25 miRNAs that showed a significant differential expression in cancer samples. The comparison with previously published data generated by deep sequencing of cDNA libraries of small RNA molecules revealed a concordance rate of 47% among miRNAs identified with both techniques. The differential expression of miRNAs miR-375, miR-143 and miR-145 was validated by quantitative PCR. MiRNA in situ hybridization revealed that the differential expression is cancer-cell associated. A combination of three miRNAs correctly classified tissue samples with an accuracy of 77.6% with an area under the receiver-operator characteristic curve of 0.810. Our data extend the knowledge about the deregulation of miRNAs in prostate cancer. The differential expression of several miRNAs is highly consistent using independent cohorts of tumor samples, different tissue preservation methods and different experimental methods. Our results indicate that combinations of miRNAs are promising biomarkers for the diagnosis of prostate cancer.

CMDX©-based single source information system for simplified quality management and clinical research in prostate cancer.

BACKGROUND: Histopathological evaluation of prostatectomy specimens is crucial to decision-making and prediction of patient outcomes in prostate cancer (PCa). Topographical information regarding PCa extension and positive surgical margins (PSM) is essential for clinical routines, quality assessment, and research. However, local hospital information systems (HIS) often do not support the documentation of such information. Therefore, we investigated the feasibility of integrating a cMDX-based pathology report including topographical information into the clinical routine with the aims of obtaining data, performing analysis and generating heat maps in a timely manner, while avoiding data redundancy. METHODS: We analyzed the workflow of the histopathological evaluation documentation process. We then developed a concept for a pathology report based on a cMDX data model facilitating the topographical documentation of PCa and PSM; the cMDX SSIS is implemented within the HIS of University Hospital Muenster. We then generated a heat map of PCa extension and PSM using the data. Data quality was assessed by measuring the data completeness of reports for all cases, as well as the source-to-database error. We also conducted a prospective study to compare our proposed method with recent retrospective and paper-based studies according to the time required for data analysis. RESULTS: We identified 30 input fields that were applied to the cMDX-based data model and the electronic report was integrated into the clinical workflow. Between 2010 and 2011, a total of 259 reports were generated with 100% data completeness and a source-to-database error of 10.3 per 10,000 fields. These reports were directly reused for data analysis, and a heat map based on the data was generated. PCa was mostly localized in the peripheral zone of the prostate. The mean relative tumor volume was 16.6%. The most PSM were localized in the apical region of the prostate. In the retrospective study, 1623 paper-based reports were transferred to cMDX reports; this process took 15 ± 2 minutes per report. In a paper-based study, the analysis data preparation required 45 ± 5 minutes per report. CONCLUSIONS: cMDX SSIS can be integrated into the local HIS and provides clinical routine data and timely heat maps for quality assessment and research purposes.