Archival Prostate Cancer Tissue Research Articles

The Brazilian national prospective active surveillance (AS) cohort of patients with low-risk prostate cancer in the public health system: vigiaSUS study protocol

BackgroundProstate cancer exhibits a very diverse behaviour, with some patients dying from the disease and others never needing treatment. Active surveillance (AS) consists of periodic PSA assessment (prostate-specific antigen), DRE (digital rectal examination) and periodic prostate biopsies. According to the main guidelines, AS is the preferred strategy for low-risk patients, to avoid or delay definitive treatment. However, concerns remain regarding its applicability in certain patient subgroups, such as African American men, who were underrepresented in the main cohorts. Brazil has a very racially diverse population, with 56.1% self-reporting as brown or black. The aim of this study is to evaluate and validate the AS strategy in low-risk prostate cancer patients following an AS protocol in the Brazilian public health system.MethodsThis is a multicentre AS prospective cohort study that will include 200 patients from all regions of Brazil in the public health system. Patients with prostate adenocarcinoma and low-risk criteria, defined as clinical staging T1–T2a, Gleason score ≤ 6, and PSA < 10 ng/ml, will be enrolled. Archival prostate cancer tissue will be centrally reviewed. Patients enrolled in the study will follow the AS strategy, which involves PSA and physical examination every 6 months as well as multiparametric MRI (mpMRI) every two years and prostate biopsy at month 12 and then every two years. The primary objective is to evaluate the reclassification rate at 12 months, and secondary objectives include determining the treatment-free survival rate, metastasis-free survival, and specific and overall survival. Exploratory objectives include the evaluation of quality of life and anxiety, the impact of PTEN loss and the economic impact of AS on the Brazilian public health system.DiscussionThis is the first Brazilian prospective study of patients with low-risk prostate cancer under AS. To our knowledge, this is one of the largest AS study cohort with a majority of nonwhite patients. We believe that this study is an opportunity to better understand the outcomes of AS in populations underrepresented in studies. Based on these data, an AS national clinical guideline will be developed, which may have a beneficial impact on the quality of life of patients and on public health.Trial registrationClinicaltrials registration is NCT05343936.

Ductal variant of prostate adenocarcinoma harbor Xenotropic murine leukemia virus related virus (XMRV) infection: a novel finding in subtype of prostate cancer.

Xenotropic murine leukemia virus related virus (XMRV), is the first gammaretrovirus identified a decade ago, in human tissue bearing adenocarcinoma of prostate, followed by several researches documenting little or no prevalence of XMRV in prostate cancer samples. However, the status of XMRV within subtype of prostate adenocarcinoma has not been investigated yet. In this study, we investigated the relationship between XMRV and broad spectrum morphological entities of prostate adenocarcinoma, including acinar, ductal and other rare subtypes. The prevalence of XMRV DNA in different histological subtypes of prostate adenocarcinoma was examined after characterizing the tumors into groups, using formalin-fixed, paraffin-embedded tissue samples from newly diagnosed prostate adenocarcinomas and archival prostate cancer tissue from our XMRV case control analysis. Broad-spectrum XMRV DNA amplification was performed by end-point polymerase chain reaction, using commercially available primer set. The study included 100 patients with prostate cancer. XMRV DNA was detected in 4 of 8 (50%) ductal adenocarcinomas, exhibiting papillary and cribriform histological features. XMRV DNA was not detected in any other variant of adenocarcinoma including acinar (0/91) and mucinous carcinomas (0/1). Majority of XMRV positive cases were biologically aggressive and present cancer at an early age upon diagnosis. Ductal adenocarcinomas demonstrate a significant association of XMRV DNA while other histological variants of prostate adenocarcinoma seem unrelated to XMRV infection.

Abstract 4947: Branched chain RNA in situ hybridization for the detection of androgen receptor splice variants within archival prostate cancer tissue

Abstract Introduction: Androgen receptor (AR) mRNA splice variants have emerged as predictive biomarkers for response to AR targeted therapies. There are no commercially available assays to detect AR splice variants. The branched chain RNA in situ hybridization (ISH) platform enables the highly sensitive detection of RNA transcripts in formalin-fixed, paraffin-embedded (FFPE) tissues. In this pilot study, we developed an RNA ISH assay to detect the constitutively active AR-V7 splice variant, and we correlated AR-V7 expression with clinical outcomes after first-line androgen deprivation therapy (ADT) for metastatic prostate cancer. Methods: We designed a branched chain RNA ISH probe to target the unique cryptic exon CE3 of AR-V7 using multiple tiling probes (Affymetrix). Automated ISH assays were performed using ViewRNA eZl Detection Kit on the BOND RX IHC and ISH Staining System. A semiquantitative scoring method was used to score the RNA ISH signal in FFPE tissue. We analyzed AR-V7 and full length AR within two prostate cancer cohorts that we hypothesized to represent “low likelihood” and “high likelihood” to have detectable AR-V7. “Low likelihood” men had undergone prostatectomy for Gleason 6 adenocarcinoma, and “high likelihood” men had metastatic castration resistant prostate cancer with tumor tissue obtained after disease progression despite at least one subsequent line of hormonal therapy (abiraterone, enzalutamide, or bicalutamide). We then analyzed two additional cohorts who experienced a “sustained response” (&amp;gt;2.5 years; n = 13) or “brief response” (&amp;lt;9 months; n = 9) to first-line ADT for metastatic prostate cancer and for whom baseline tumor tissue was available. Results: “High likelihood” cohort samples had detectable AR-V7 with a score of 1+ to 3+ in all samples (n = 12). “Low likelihood” cohort samples had no detectable AR-V7 (n = 10). Given the apparent binary distinction of AR-V7 signal among the above groups, we analyzed pre-treatment AR-V7 status as a predictive and prognostic biomarker in men with treatment-naive metastatic disease. Detectable AR-V7 was more common among “brief response” samples (6 of 9) than among “sustained response” samples (4 of 13) (not significant; P = 0.19). Patients without detectable AR-V7 RNA had significantly longer overall survival (OS, logrank P = 0.044), with a non-significant trend toward superior progression-free survival (PFS, logrank P = 0.055). Conclusions: Within an institutional cohort, the RNA ISH assay identified AR-V7 in all tested samples of high clinical likelihood for the splice variant RNA and no tested samples of low clinical likelihood. AR-V7 RNA was detectable in some pretreatment samples, and its presence was associated with significantly shorter overall survival. The potential prognostic and predictive utility of AR-V7 status as determined by RNA ISH from conventionally prepared FFPE tissue warrants further study in larger cohorts. Citation Format: Philip J. Saylor, Richard J. Lee, Kshitij S. Arora, David T. Ting, Vikram Deshpande, Miguel N. Rivera, Rong Hu, Chin-Lee Wu, David T. Miyamoto. Branched chain RNA in situ hybridization for the detection of androgen receptor splice variants within archival prostate cancer tissue. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4947.

Detection of infectious organisms in archival prostate cancer tissues.

BackgroundSeroepidemiological studies have reported associations between exposure to sexually transmitted organisms and prostate cancer risk. This study sought DNA evidence of candidate organisms in archival prostate cancer tissues with the aim of assessing if a subset of these cancers show any association with common genital infections.Methods221 archival paraffin-embedded tissue blocks representing 128 histopathologically confirmed prostate cancers comprising 52 “aggressive” (Gleason score ≥ 7) and 76 “non-aggressive” (Gleason score ≤ 6) TURP or radical prostatectomy specimens were examined, as well as unaffected adjacent tissue when available. Representative tissue sections were subjected to DNA extraction, quality tested and screened by PCR for HSV-1, HSV-2, XMRV, BKV, HPV, Chlamydia trachomatis, Ureaplasma parvum, Ureaplasma urealyticum, Mycoplasma genitalium, and Trichomonas vaginalis.Results195 of 221 DNA samples representing 49 “aggressive” and 66 “non-aggressive” prostate cancer cases were suitable for analysis after DNA quality assessment. Overall, 12.2% (6/49) aggressive and 7.6% (5/66) non-aggressive cases were positive for any of the candidate organisms. Mycoplasma genitalium DNA was detected in 4/66 non-aggressive, 5/49 aggressive cancers and in one cancer-unaffected adjacent tissue block of an aggressive case. Ureaplasma urealyticum DNA was detected in 0/66 non-aggressive and 1/49 aggressive cancers and HSV DNA in 1/66 non-aggressive and 0/49 aggressive cancers. This study did not detect BKV, XMRV, T. vaginalis, U. parvum, C. trachomatis or HPV DNA.ConclusionsThe low prevalence of detectable microbial DNA makes it unlikely that persistent infection by the selected candidate microorganisms contribute to prostate cancer risk, regardless of tumour phenotype.

Rapid and reliable detection of LINE-1 hypomethylation using high-resolution melting analysis

Objectives The aim of the present study was to evaluate the precision and reproducibility of the LINE-1 high-resolution melting (HRM) assay to detect LINE-1 hypomethylation. Design and methods We first evaluated a methylated DNA dilution matrix and a panel of human cancer cell lines. We then applied this LINE-1 HRM assay to a set of 37 archival prostate cancer tissue samples. Results Our LINE-1 HRM assay revealed small and reproducible run-to-run and bisulfite-to-bisulfite variations. As expected, we found a large variation in methylation levels between different cancer cell lines. All results were confirmed with MethyLight and pyrosequencing as indicated by the high correlation coefficient. Finally, we successfully applied the LINE-1 HRM assay to archival prostate cancer tissues. Conclusions The present LINE-1 HRM assay represents a novel, accurate, and cost-effective method to measure global hypomethylation, which makes it suitable for high- and low-throughput laboratories.

Detection of TMPRSS2-ETS Fusions by a Multiprobe Fluorescence in Situ Hybridization Assay for the Early Diagnosis of Prostate Cancer : A Pilot Study

Fusion of the prostate-specific and androgen-regulated transmembrane-serine protease gene (TMPRSS2) with the erythroblast transformation-specific (ETS) family members is the most common genetic alteration in prostate cancer. However, the biological and clinical role of TMPRSS2-ETS fusions in prostate cancer, especially in problematic prostate needle core biopsies, has not been rigorously evaluated. We randomly collected 85 specimens including 50 archival prostate cancer tissue blocks, 15 normal prostate specimens, and 20 benign prostatic hyperplasia specimens for TMPRSS2-ETS fusion analyses. Moreover, the fusion status in an additional 20 patients with initial negative biopsies who progressed to biopsy-positive prostate cancer at subsequent follow-ups was also characterized. Fluorescently labeled probes specific for ERG-related rearrangements involving the TMPRSS2-ERG fusion as well as TMPRSS2-ETV1 and TMPRSS2-ETV4 were used to assess samples for gene rearrangements indicative of malignancy under a design of sequential trial. Rearrangements involving TMPRSS2-ETS fusions were detected in 90.0% of the 50 postoperative prostate cancer samples. The positive rate for the rearrangements in the initial prostate cancer-negative biopsies of 20 patients who eventually progressed to prostate cancer was 60.0% (12/20). Our preliminary study demonstrates that the clinical utility of TMPRSS2-ETS fusion detection as a biomarker and ancillary diagnostic tool for the early diagnosis of prostate cancer is promising, given this approach shows significant high sensitivity and specificity in detection.

Somatic Mosaicism and Cancer: A Micro-Genetic Examination into the Role of the Androgen Receptor Gene in Prostate Cancer

Recent evidence has shown that the androgen receptor (AR) plays a major role in all prostate cancer stages, including both androgen-dependent and -independent tumors. A large number of studies have examined the possible effects of a functional polymorphism in the AR gene, a variable-length CAG repeat, on the development of prostate cancer, but the results to date have been inconclusive. We have considered the fact that the tissue heterogeneity present in almost all prostate cancer tumors has rarely been regarded as an indicator of AR genetic heterogeneity. To determine if genetic heterogeneity exists and is a significant event in prostate cancer development, we have examined prostate cancer tumors for somatic shortening of the AR gene CAG repeat. All 72 laser capture microdissected samples from archival prostate cancer tissues, as well as samples from freshly prepared prostate cancer tissues, showed some genetic heterogeneity (somatic mosaicism) for AR CAG repeat length. Cancerous tissues showed a much greater degree of genetic heterogeneity than adjacent benign tissues, as well as a very significant shortening of their CAG repeat lengths. However, CAG repeat length heterogeneity was not observed in normal prostate tissues. It is hypothesized that somatic mosaicism of the AR CAG repeat in prostate cancer tumors may be found to be an important genetic event in precancerous tissue, which may subsequently lead to the development of prostate cancer.

Application of a yeast assay to detect functional p53 mutations in archival prostate cancer tissue.

Detection and functional evaluation of mutant p53 alleles using a yeast assay could yield significant information for predicting the prognosis of patients with prostate cancer (CaP). Since the current version of this yeast assay is not applicable to archival tissues, we developed a modified assay for use on formalin-fixed, paraffin-embedded tissue and have applied it to the study of patient samples. Using this modified assay, we examined archival CaP samples from 10 patients for mutations in exons 5-8 of p53 gene. Mutations were detected in four samples: three resulted in the formation of red yeast colonies indicating complete loss of function, while one gave pink yeast colonies, indicating that this mutant retained partial function. In parallel, we analyzed these samples for p53 abnormalities using a single-strand conformational polymorphism (SSCP) approach. Only three of the four yeast-positive samples gave abnormal SSCP bands. In each case where abnormal p53 was found by both methods, DNA sequencing revealed the identical base change. These results suggest that the modified yeast assay may be more sensitive than SSCP for detection of p53 mutations, and demonstrate that the modified method can be used to detect and evaluate the function of p53 mutants present in archival tissue.

HER-2/neu oncogene amplification in prostate cancer

Oncogene amplification is a manifestation of genetic instability, which has been implicated in the pathogenesis of many cancers. Work performed in this laboratory has focused on HER-2/neu amplification in breast cancer, ovarian cancer, cervical cancer and rhabdomyosarcoma, all of which revealed HER-2/neu amplification in at least a proportion of tumors. In the present project, we studied HER-2/neu oncogene amplification in prostate carcinoma, 244,000 new cases of which are diagnosed yearly, and which affects a significant number of men. Fluorescent in situ hybridization (FISH) using the Vysis HER-2/neu probe with a chromosome 17 centromere control probe was performed on formalin-fixed paraffin-embedded archival prostate cancer tissues. Out of a total of 85 cases studied, 7 cases (8.2%) were found to be amplified. Of those 7 amplified cases, 2 were also trisomic for chromosome 17. Amplification is defined as a signal ratio of HER-2/neu to chromosome 17 of >1.5, after Mark et al. (1998). Clinicopathologic characteristics of the amplified cases versus the nonamplified cases will be compared. Although the results of the present study still need to be confirmed and extended by additional cases in other centers, the data thus far do indicate that a small subset of prostate cancer is characterized by HER-2/neu gene amplification, as in other cancers. (This study was partially funded by Vysis, Inc., Downers Grove, IL).

Immunohistochemical determination of tumor angiogenesis measured by the maximal microvessel density in human prostate cancer.

Tumor growth beyond a certain size requires angiogenesis. Experimental evidence shows that once tumors leave the pre-angiogenic phenotype to become angiogenic, metastases often start to evolve. The aim of this study was to develop a reproducible immunohistochemical technique and method to characterize the neovascularization in archival prostate cancer tissue by quantifying the microvessel density (MVD). Archival tumor specimens from 64 consecutively diagnosed prostate cancer patients were immunostained for von Willebrand Factor (vWF), endothelial antigen and for CD31 combined with the use of different digestive enzymes (trypsin and pronase) and heating in a microwave oven. Both the mean and the maximal MVD, and the reproducibility of the method were estimated. Finally, the mean MVD, the maximal MVD, and clinical characteristics were correlated with the crude survival of the patient population. The immunohistochemical staining for vWF to measure the maximal MVD was found to be a reproducible method of characterizing the individual tumor. Both a univariate and a multivariate analysis demonstrated that the maximal MVD, in contrast to the mean MVD, was significantly associated with survival in prostate cancer patients. We conclude that evaluation of angiogenesis by immunostaining the endothelial cells for vWF measured by the MVD in the most vascularized areas of the tumor is a reproducible method of characterizing the individual prostate tumor. Maximal MVD proved to be an independent prognostic parameter useful in conjunction with other known prognostic markers in human prostate cancer.