Nucleotide resolution analysis of TMPRSS2 and ERG rearrangements in prostate cancer

TMPRSS2–ERG rearrangements occur in approximately 50% of prostate cancers and therefore represent one of the most frequently observed structural rearrangements in all cancers. However, little is known about the genomic architecture of such rearrangements. We therefore designed and optimized a pipeline involving target capture of TMPRSS2 and ERG genomic sequences coupled with paired‐end next‐generation sequencing to resolve genomic rearrangement breakpoints in TMPRSS2 and ERG at nucleotide resolution in a large series of primary prostate cancer specimens (n = 83). This strategy showed > 90% sensitivity and specificity in identifying TMPRSS2–ERG rearrangements, and allowed identification of intra‐ and inter‐chromosomal rearrangements involving TMPRSS2 and ERG with known and novel fusion partners. Our results indicate that rearrangement breakpoints show strong clustering in specific intronic regions of TMPRSS2 and ERG. The observed TMPRSS2–ERG rearrangements often exhibited complex chromosomal architecture associated with several intra‐ and inter‐chromosomal rearrangements. Nucleotide resolution analysis of breakpoint junctions revealed that the majority of TMPRSS2 and ERG rearrangements (～88%) occurred at or near regions of microhomology or involved insertions of one or more base pairs. This architecture implicates non‐homologous end joining (NHEJ) and microhomology‐mediated end joining (MMEJ) pathways in the generation of such rearrangements. These analyses have provided important insights into the molecular mechanisms involved in generating prostate cancer‐specific recurrent rearrangements. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Distinct genomic aberrations associated with ERG rearranged prostate cancer

Emerging molecular and clinical data suggest that ETS fusion prostate cancer represents a distinct molecular subclass, driven most commonly by a hormonally regulated promoter and characterized by an aggressive natural history. The study of the genomic landscape of prostate cancer in the light of ETS fusion events is required to understand the foundation of this molecularly and clinically distinct subtype. We performed genome‐wide profiling of 49 primary prostate cancers and identified 20 recurrent chromosomal copy number aberrations, mainly occurring as genomic losses. Co‐occurring events included losses at 19q13.32 and 1p22.1. We discovered three genomic events associated with ERG rearranged prostate cancer, affecting 6q, 7q, and 16q. 6q loss in nonrearranged prostate cancer is accompanied by gene expression deregulation in an independent dataset and by protein deregulation of MYO6. To analyze copy number alterations within the ETS genes, we performed a comprehensive analysis of all 27 ETS genes and of the 3 Mbp genomic area between ERG and TMPRSS2 (21q) with an unprecedented resolution (30 bp). We demonstrate that high‐resolution tiling arrays can be used to pin‐point breakpoints leading to fusion events. This study provides further support to define a distinct molecular subtype of prostate cancer based on the presence of ETS gene rearrangements. © 2009 Wiley‐Liss,Inc.     

Somatic Alterations Contributing to Metastasis of a Castration‐Resistant Prostate Cancer

Metastatic castration‐resistant prostate cancer (mCRPC) is a lethal disease, and molecular markers that differentiate indolent from aggressive subtypes are needed. We sequenced the exomes of five metastatic tumors and healthy kidney tissue from an index case with mCRPC to identify lesions associated with disease progression and metastasis. An Ashkenazi Jewish (AJ) germline founder mutation, del185AG in BRCA1, was observed and AJ ancestry was confirmed. Sixty‐two somatic variants altered proteins in tumors, including cancer‐associated genes, TMPRSS2‐ERG, PBRM1, and TET2. The majority (n = 53) of somatic variants were present in all metastases and only a subset (n = 31) was observed in the primary tumor. Integrating tumor next‐generation sequencing and DNA copy number showed somatic loss of BRCA1 and TMPRSS2‐ERG. We sequenced 19 genes with deleterious mutations in the index case in additional mCRPC samples and detected a frameshift, two somatic missense alterations, tumor loss of heterozygosity, and combinations of germline missense SNPs in TET2. In summary, genetic analysis of metastases from an index case permitted us to infer a chronology for the clonal spread of disease based on sequential accrual of somatic lesions. The role of TET2 in mCRPC deserves additional analysis and may define a subset of metastatic disease.     

Relevance of cohort design for studying the frequency of the ERG rearrangement in prostate cancer

Braun M, Scheble V J, Menon R, Scharf G, Wilbertz T, Petersen K, Beschorner C, Reischl M, Kuefer R, Schilling D, Stenzl A, Kristiansen G, Rubin M A, Fend F & Perner S
(2011) Histopathology 58 , 1028–1036
Relevance of cohort design for studying the frequency of the ERG rearrangement in prostate cancer Aims: ERG rearrangements, mostly resulting in TMPRSS2–ERG fusions, are frequentalterations in prostate cancer (PCa), with a frequency ranging from 15% to 78%. As the reason for this variability is unknown, our aim was to investigate the ERG rearrangement frequency with a cohort design. Methods and results: We assessed three well‐defined cohorts for ERG rearrangements, using fluorescence in situ hybridization (FISH). The first cohort comprised 119 prostatectomy specimens. The second and third cohorts included incidentally diagnosed PCa [71 cystoprostatectomy specimens, and 105 transurethral resection of the prostate (TURP) specimens]. Seventy of 119 (59%) cases of the prostatectomy cohort harboured ERG rearrangements. Regarding zonal origin, 2/11 (18%) transition zone (TZ) foci and 75/145 (52%) peripheral zone (PZ) foci harboured ERG rearrangements. Within the cystoprostatectomies, 24/71 (34%) cases harboured ERG rearrangements. Regarding zonal origin, 2/9 (22%) TZ foci and 26/86 (30%) PZ foci harboured ERG rearrangements. PCa incidentally identified by TURP harboured ERG rearrangements in 31/105 (29%) cases. Conclusions: ERG rearrangements occur in TZ PCa, although at a lower frequency than in PZ PCa. We confirmed that approximately half of all prostatectomies harbour ERG rearrangements. However, the frequency in incidentally diagnosed PCa cohorts was significantly lower, even if multifocality was considered. Consequently, zonal origin and cohort design are key for studying the clinical implications of ERG rearrangements.     

Upregulation of PTTG1 is associated with poor prognosis in prostate cancer

Pituitary tumor‐transforming gene 1 (PTTG1) is a regulator of chromosome stability. PTTG1 overexpression had been associated with tumor aggressiveness in several cancer types. To examine its prognostic utility in prostate cancer, a tissue microarray including 12 427 tumors with clinical and molecular data was analyzed by immunohistochemistry. PTTG1 immunostaining was largely absent in normal prostate epithelial cells. In cancers, staining was considered weak in 5.4%, moderate in 5.6% and strong in 0.8%. Strong staining was linked to advanced pT stage, high classical and quantitative Gleason grade, high Ki67‐labeling index (all P < 0.0001) and lymph node metastasis (P = 0.0083). The prognostic impact of PTTG1 expression was independent of established preoperative and postoperative prognostic features. Comparison with molecular features revealed that PTTG1 upregulation was associated with nine of 12 common genomic deletions (P < 0.05), p53 alterations and high androgen receptor levels (P < 0.001 each), but was unrelated to the TMPRSS2:ERG fusion status. In conclusion, these data identify PTTG1 as a strong and independent prognostic feature in prostate cancer. PTTG1 measurement, either alone or in combination with other biomarkers might be instrumental for determining prostate cancer aggressiveness.     

Anaerobic bacteria in urine before and after prostatic massage of infertile men

In the present study 25 infertile men delivered urine before and after prostatic massage. Expressed prostatic secretion (EPS) was obtained from 11 men. Aerobic and anaerobic bacterial analyses showed that a total of 33 isolates was found in samples of urine voided before massage as compared to 59 isolates after massage of the prostate. There was an increase in the number of anaerobes whereas there was no change in the number of aerobes. The occurrence of EPS did not influence the number of aerobic and anaerobic isolates in urine voided after massage of the prostate. The most often isolated anaerobes in urine voided after prostatic massage wereEubacterium lentum, Peptococcus asaccharolyticus andBacteroides species and the most common anaerobe in EPS wasPeptostreptococcus micros.     

TMPRSS2-ERG融合基因在转移性前列腺癌中的表达

目的 检测转移性前列腺癌中TMPRSS2-ERG基因融合的发生率,探讨ERG基因重排在前列腺癌进展中的作用.方法 收集32例由细针穿刺诊断的转移性前列腺癌,穿刺部位包括盆腔及远处淋巴结、肝、骨、甲状腺等,回顾相关临床病理学资料.免疫组织化学采用EnVision法标记前列腺特异性抗原、突触素和嗜铬粒素A.运用ERG分离断...     

Identification of frequent BRAF copy number gain and alterations of RAF genes in chinese prostate cancer

We recently found that TMPRSS2:ERG fusion genes and PTEN loss, which are common in Western prostate cancers are infrequent in Chinese cases. As previous studies indicated a higher frequency of RAS and BRAF mutation rates in Eastern Asian than in Western prostate cancers and fusion genes involving the RAF family genes BRAF and RAF1 were recently identified in prostate cancer in the American population, we investigated BRAF and RAF1 alterations in Chinese prostate cancer. Using fluorescence in situ hybridization, we found that BRAF was truncated in five of 200 informative Chinese cases (2.5%) and that RAF1 was truncated in three of 204 informative cases (1.5%) and genomic rearrangements of these genes were significantly correlated with high Gleason scores (>7; P < 0.01) and have a trend to appear in high clinical stage disease. A high frequency of BRAF and RAF1 copy number gain was found (29 and 15%, respectively). BRAF copy number gain in Chinese cancers was significantly higher than in UK cases (9.2%)(P < 0.001) and correlated with a number of clinical parameters. High‐level expression of BRAF was found by immunohistochemistry in Chinese cancer samples compared with adjacent nonmalignant epithelial cells, which was correlated with high BRAF copy number. We also identified KRAS codon 12 mutations in three of 96 Chinese cases, no BRAF V600E mutations were observed. Our finding suggests that the activation of the RAS/RAF/MEK/ERK pathway may be frequent in Chinese prostate cancer, with RAF gene copy number gain potentially being the main contributor. © 2012 Wiley Periodicals, Inc.     

High level of Ets-related gene expression has high specificity for prostate cancer: a tissue microarray study of 11 483 cancers

Minner S, Luebke A M, Kluth M, Bokemeyer C, Jänicke F, Izbicki J, Schlomm T, Sauter G & Wilczak W
(2012) Histopathology  61, 445–453 High level of Ets‐related gene expression has high specificity for prostate cancer: a tissue microarray study of 11 483 cancers Aims: TMPRSS2–ERG fusion resulting in strong Ets‐related gene (ERG) overexpression occurs in about 50% of prostate cancers. This study was undertaken to determine the prevalence of ERG overexpression in other tumour types as well as in normal tissues. Methods and results: A total of 11 483 tumours and 72 different normal tissue types were analysed in a tissue microarray format. Strong nuclear ERG overexpression was found in 36.7% of prostate carcinomas as well as in various vascular tumours, including Kaposi sarcomas (91.7%), angiosarcomas (100%) and haemangiomas (90.9%). Moderate to strong nuclear ERG immunostaining was also observed in thymoma (6.1%). Weak to moderate ERG staining was found in a small number of squamous cell carcinomas of the skin, squamous carcinomas of the lung, malignant mesotheliomas, carcinosarcomas of the uterus, gastrointestinal stromal tumours, hepatocellular carcinomas, teratomas of the testis, anaplastic carcinomas of the thyroid, giant cell tumours of the tendon sheath and benign fibrous histiocytomas of the skin. ERG overexpression was not seen in 8886 samples from 132 other tumour types and subtypes. Within normal tissues, immunohistochemically detectable ERG overexpression was restricted to endothelial cells and subsets of lymphocytes. Conclusions: The high specificity of ERG expression in both normal and neoplastic tissues suggests a very narrow biological role for ERG in highly selected tissues.     

Molecular evidence that invasive adenocarcinoma can mimic prostatic intraepithelial neoplasia (PIN) and intraductal carcinoma through retrograde glandular colonization

Prostate cancer often manifests as morphologically distinct tumour foci and is frequently found adjacent to presumed precursor lesions such as high‐grade prostatic intraepithelial neoplasia (HGPIN). While there is some evidence to suggest that these lesions can be related and exist on a pathological and morphological continuum, the precise clonal and temporal relationships between precursor lesions and invasive cancers within individual tumours remain undefined. Here, we used molecular genetic, cytogenetic, and histological analyses to delineate clonal, temporal, and spatial relationships between HGPIN and cancer lesions with distinct morphological and molecular features. First, while confirming the previous finding that a substantial fraction of HGPIN lesions associated with ERG‐positive cancers share rearrangements and overexpression of ERG, we found that a significant subset of such HGPIN glands exhibit only partial positivity for ERG. This suggests that such ERG‐positive HGPIN cells either rapidly invade to form adenocarcinoma or represent cancer cells that have partially invaded the ductal and acinar space in a retrograde manner. To clarify these possibilities, we used ERG expression status and TMPRSS2–ERG genomic breakpoints as markers of clonality, and PTEN deletion status to track temporal evolution of clonally related lesions. We confirmed that morphologically distinct HGPIN and nearby invasive cancer lesions are clonally related. Further, we found that a significant fraction of ERG‐positive, PTEN‐negative HGPIN and intraductal carcinoma (IDC‐P) lesions are most likely clonally derived from adjacent PTEN‐negative adenocarcinomas, indicating that such PTEN‐negative HGPIN and IDC‐P lesions arise from, rather than give rise to, the nearby invasive adenocarcinoma. These data suggest that invasive adenocarcinoma can morphologically mimic HGPIN through retrograde colonization of benign glands with cancer cells. Similar clonal relationships were also seen for intraductal carcinoma adjacent to invasive adenocarcinoma. These findings represent a potentially undervalued indicator of pre‐existing invasive prostate cancer and have significant implications for prostate cancer diagnosis and risk stratification. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Morphological features of TMPRSS2-ERG gene fusion prostate cancer

The TMPRSS2-ETS fusion prostate cancers comprise 50-70% of the prostate-specific antigen (PSA)-screened hospital-based prostate cancers examined to date, making it perhaps the most common genetic rearrangement in human cancer. The most common variant involves androgen-regulated TMPRSS2 and ERG, both located on chromosome 21. Emerging data from our group and others suggests that TMPRSS2-ERG fusion prostate cancer is associated with higher tumour stage and prostate cancer-specific death. The goal of this study was to determine if this common somatic alteration is associated with a morphological phenotype. We assessed 253 prostate cancer cases for TMPRSS2-ERG fusion status using an ERG break-apart FISH assay. Blinded to gene fusion status, two reviewers assessed each tumour for presence or absence of eight morphological features. Statistical analysis was performed to look for significant associations between morphological features and TMPRSS2-ERG fusion status. Five morphological features were associated with TMPRSS2-ERG fusion prostate cancer: blue-tinged mucin, cribriform growth pattern, macronucleoli, intraductal tumour spread, and signet-ring cell features, all with p-values < 0.05. Only 24% (n=30/125) of tumours without any of these features displayed the TMPRSS2-ERG fusion. By comparison, 55% (n=38/69) of cases with one feature (RR=3.88), 86% (n=38/44) of cases with two features (RR=20.06), and 93% (n=14/15) of cases with three or more features (RR=44.33) were fusion positive (p<0.001). To our knowledge, this is the first study that demonstrates a significant link between a molecular alteration in prostate cancer and distinct phenotypic features. The strength of these findings is similar to microsatellite unstable colon cancer and breast cancer involving BRCA1 and BRCA2 mutations. The biological effect of TMPRSS2-ERG overexpression may drive pathways that favour these common morphological features that pathologists observe daily. These features may also be helpful in diagnosing TMPRSS2-ERG fusion prostate cancer, which may have both prognostic and therapeutic implications.