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Supplementary Figure S1 and Legend

Purpose: We had previously identified a new transmembrane collagen, type XXIII, in metastatic rat prostate carcinoma cells. The purpose of this study was to determine the expression of collagen XXIII in human prostate cancer and investigate its relationship with disease progression.Experimental Design: We investigated collagen XXIII expression in prostate cancer tissue and did a retrospective analysis of association with prostate-specific antigen (PSA)–defined disease recurrence. The presence of collagen XXIII in prostate cancer patient urine was also assessed before and after prostatectomy.Results: Collagen XXIII protein was detected at very low levels in benign prostate tissue and was significantly increased in prostate cancer. Distant metastases exhibited significantly higher collagen XXIII levels compared with either localized prostate cancer or regional (lymph node) metastases. Patients with high collagen XXIII levels had a 2.8-fold higher risk of PSA failure with median time to failure of 8.1 months, compared with low collagen XXIII patients with a median time to failure of 5 years. Multivariate Cox regression showed that the presence of collagen XXIII was significantly associated with time to PSA recurrence, independent of other clinical variables. Collagen XXIII was also detected in prostate cancer patient urine, with reduced levels after prostatectomy, indicating potential as a noninvasive fluid biomarker.Conclusions: We present the first report demonstrating increased collagen XXIII expression in prostate cancer tissue. We show that collagen XXIII level is a significant independent predictor of PSA-defined disease recurrence, suggesting a potential role as a molecular biomarker of prostate cancer progression and metastasis.

Supplementary Figure Legends 1-2, Methods from SLC45A3-ELK4 Is a Novel and Frequent Erythroblast Transformation–Specific Fusion Transcript in Prostate Cancer

Supplementary Figure 2 from SLC45A3-ELK4 Is a Novel and Frequent Erythroblast Transformation–Specific Fusion Transcript in Prostate Cancer

Supplementary Figure 1 from TMPRSS2:ERG Fusion-Associated Deletions Provide Insight into the Heterogeneity of Prostate Cancer

Supplementary Figure 2 from Treatment-Dependent Androgen Receptor Mutations in Prostate Cancer Exploit Multiple Mechanisms to Evade Therapy

Supplementary Figure 3 from TMPRSS2:ERG Fusion-Associated Deletions Provide Insight into the Heterogeneity of Prostate Cancer

Prostate cancer is a common and clinically heterogeneous disease with marked variability in progression. The recent identification of gene fusions of the 5′-untranslated region of TMPRSS2 (21q22.3) with the ETS transcription factor family members, either ERG (21q22.2), ETV1 (7p21.2), or ETV4 (17q21), suggests a mechanism for overexpression of the ETS genes in the majority of prostate cancers. In the current study using fluorescence in situ hybridization (FISH), we identified the TMPRSS2:ERG rearrangements in 49.2% of 118 primary prostate cancers and 41.2% of 18 hormone-naive lymph node metastases. The FISH assay detected intronic deletions between ERG and TMPRSS2 resulting in TMPRSS2:ERG fusion in 60.3% (35 of 58) of the primary TMPRSS2:ERG prostate cancers and 42.9% (3 of 7) of the TMPRSS2:ERG hormone-naive lymph node metastases. A significant association was observed between TMPRSS2:ERG rearranged tumors through deletions and higher tumor stage and the presence of metastatic disease involving pelvic lymph nodes. Using 100K oligonucleotide single nucleotide polymorphism arrays, a homogeneous deletion site between ERG and TMPRSS2 on chromosome 21q22.2-3 was identified with two distinct subclasses distinguished by the start point of the deletion at either 38.765 or 38.911 Mb. This study confirms that TMPRSS2:ERG is fused in approximately half of the prostate cancers through deletion of genomic DNA between ERG and TMPRSS2. The deletion as cause of TMPRSS2:ERG fusion is associated with clinical features for prostate cancer progression compared with tumors that lack the TMPRSS2:ERG rearrangement. (Cancer Res 2006; 66(17): 8337-41)

Supplementary Figure 1 from Treatment-Dependent Androgen Receptor Mutations in Prostate Cancer Exploit Multiple Mechanisms to Evade Therapy

Supplementary Figure 2 from TMPRSS2:ERG Fusion-Associated Deletions Provide Insight into the Heterogeneity of Prostate Cancer

Fusion of the 5′-untranslated region of androgen-regulated TMPRSS2 promoter with ETS transcription factor family members is found frequently in prostate cancers, and recent work suggests that the most common TMPRSS2-ERG fusion is associated with an aggressive clinical phenotype compared with fusion-negative prostate cancer. Thus far, analysis of the fusion has been limited to sporadic cases of prostate cancer. In the current study, we explore for an enrichment of TMPRSS2-ERG fusion in familial prostate cancer. TMPRSS2-ERG fusion was identified using a break-apart fluorescence in situ hybridization assay on tissue microarrays. Presence of TMPRSS2-ERG fusion was associated with higher Gleason scores (P = 0.027). Of 75 patients with established history of prostate cancer, we detected the TMPRSS2-ERG fusion in 44 (59%) patients. Almost three quarters (73%) of fusion-positive patients accumulated within 16 specific families whereas only 27% were single fusion-positive cases within one family. Based on reported prevalence rates, we calculated a sibling recurrence risk ratio of up to 18.9. A subset (63%) of families with uniformly TMPRSS2-ERG–positive prostate cancer underwent a genome-wide linkage scan at 500 markers. This revealed several loci located on chromosomes #9, #18, and X that were suggestive of linkage to the TMPRSS2-ERG fusion-positive prostate cancer phenotype with linkage-of-disease scores up to 2.16 and nonparametric linkage scores up to 2.77. This suggests the presence of an inherited susceptibility to developing the TMPRSS2-ERG fusion. Given the association of TMPRSS2-ERG fusion and aggressive prostate cancer, close surveillance of relatives of patients with established fusion-positive prostate cancer or a family history of prostate cancer in general would be warranted. [Cancer Res 2009;69(4):640–6]

Chromosomal rearrangements account for all erythroblast transformation–specific (ETS) family member gene fusions that have been reported in prostate cancer and have clinical, diagnostic, and prognostic implications. Androgen-regulated genes account for the majority of the 5′ genomic regulatory promoter elements fused with ETS genes. TMPRSS2-ERG, TMPRSS2-ETV1, and SLC45A3-ERG rearrangements account for roughly 90% of ETS fusion prostate cancer. ELK4, another ETS family member, is androgen regulated, involved in promoting cell growth, and highly expressed in a subset of prostate cancer, yet the mechanism of ELK4 overexpression is unknown. In this study, we identified a novel ETS family fusion transcript, SLC45A3-ELK4, and found it to be expressed in both benign prostate tissue and prostate cancer. We found high levels of SLC45A3-ELK4 mRNA restricted to a subset of prostate cancer samples. SLC45A3-ELK4 transcript can be detected at high levels in urine samples from men at risk for prostate cancer. Characterization of the fusion mRNA revealed a major variant in which SLC45A3 exon 1 is fused to ELK4 exon 2. Based on quantitative PCR analyses of DNA, unlike other ETS fusions described in prostate cancer, the expression of SLC45A3-ELK4 mRNA is not exclusive to cases harboring a chromosomal rearrangement. Treatment of LNCaP cancer cells with a synthetic androgen (R1881) revealed that SLC45A3-ELK4, and not endogenous ELK4, mRNA expression is androgen regulated. Altogether, our findings show that SLC45A3-ELK4 mRNA expression is heterogeneous, highly induced in a subset of prostate cancers, androgen regulated, and most commonly occurs through a mechanism other than chromosomal rearrangement (e.g., trans-splicing). [Cancer Res 2009;69(7):2734–8]

Supplementary Figure 1 from SLC45A3-ELK4 Is a Novel and Frequent Erythroblast Transformation–Specific Fusion Transcript in Prostate Cancer

Supplementary Table 1 from Genome-Wide Linkage Analysis of TMPRSS2-ERG Fusion in Familial Prostate Cancer

Nucleoporins as components of the nuclear pore complex (NCP) are known for regulating nuclear-cytoplasmatic transport. Recently, the nucleoporin POM121 was found to have an important impact on intranuclear translocation of prostate cancer (PCa)-specific tumor drivers including the androgen receptor (AR). The aim of our study was to assess the potential of POM121 as a prognostic biomarker.Therefore, we performed immunohistochemistry (IHC) for POM121 on a large clinically, well characterized PCa tissue cohort comprising benign prostatic samples, radical prostatectomy (RPE) samples, lymph node metastases, local recurrent tumors and distant metastases of 289 patients. Using a semi automated tissue image analysis software we evaluated POM121 protein expression level based on IHC.We could show that POM121 expression increases during tumor progression. Expression levels were significantly higher in primary tumors compared to benign samples (P = 0.001), and substantially higher in advanced tumors (P0.001) and in distant metastases (P = 0.006) compared to primary tumors. Furthermore, POM121 expression predicts biochemical recurrence free survival (BFS) after surgery independent of the WHO group and other clinicopathological markers. 5-years BFS with primary tumors lacking POM121 and expressing POM121 was 88.8% and 68.9%, respectively.Our study reveals the potential of POM121 as a potential biomarker for PCa, predicting BFS independent of other common clinicopathological parameters. Furthermore, POM121 might be a new targetable structure for patients suffering from advanced PCa.

The Mediator complex is a transcriptional regulator interacting with transcription factors and RNA-polymerase-II. Recently, we identified its subunit CDK19 to be specifically expressed in prostate cancer (PCa) and to be functionally implicated in PCa aggressiveness. Aim of our study was to comprehensively characterize the protein expression of CDK19 and its paralog CDK8 in PCa. We performed immunohistochemistry (IHC) for CDK19/CDK8 on a large cohort including needle biopsies from 202 patients, 799 primary tumor foci of radical prostatectomy specimens from 415 patients, 120 locally advanced tumor foci obtained by palliative transurethral resection, 140 lymph node metastases, 67 distant metastases and 82 benigns. Primary tumors were stained for the proliferation marker Ki67, androgen receptor (AR) and ERG. For 376 patients, clinic-pathologic data were available. Primary endpoint was disease-recurrence-free survival (DFS). Nuclear CDK19 and CDK8 expression increases during progression showing the highest intensity in metastatic and castration-resistant tumors. High CDK19 expression on primary tumors correlates with DFS independently from Gleason grade and PSA. Five-year-DFS rates of patients with primary tumors expressing no, moderate and high CDK19 are 73.7, 56.9 and 30.4%, respectively. CDK19 correlates with Gleason grade, T-stage, Ki67 proliferation-index, nuclear AR expression and ERG-status. Therapeutic options for metastatic and castration-resistant PCa remain limited. In the current study, we confirmed an important role of the Mediator subunit CDK19 in advanced PCa supporting current developments to target CDK19 and its paralog CDK8. Furthermore, CDK19 protein expression has the potential to predict disease recurrence independently from established biomarkers thus contributing to individual management for PCa patients.

Background: Stratifying prostate cancer (PCa) patients into risk groups at time of initial diagnosis enabling a risk-adapted disease management is still a major clinical challenge. Existing studies evaluating the prognostic potential of PSMA (prostate-specific membrane antigen) for PCa were performed on radical prostatectomy specimens (RPE), i.e., decision making for disease management was already completed at time of sample analysis. Aim of our study was to assess the prognostic value of PSMA expression for PCa patients on biopsies at time of initial diagnosis. Methods: PSMA expression was assessed by immunohistochemistry on 294 prostate biopsies with corresponding RPE, 621 primary tumor foci from 242 RPE, 43 locally advanced or recurrent tumors, 34 lymph node metastases, 78 distant metastases and 52 benign prostatic samples. PSMA expression was correlated with clinico-pathologic features. Primary endpoint was recurrence free survival. Other clinicopathologic features included WHO/ISUP grade groups, PSA serum level, TNM-stage, and R-status. Chi-square test, ANOVA-analyses, Cox-regression, and log-rank tests were performed for statistical analyses. Results: High PSMA expression on both biopsy and RPE significantly associates with a higher risk of disease recurrence following curative surgery. The 5-year-recurrence free survival rates were 88.2, 74.2, 67.7 and 26.8% for patients exhibiting no, low, medium, or high PSMA expression on biopsy, respectively. High PSMA expression on biopsy was significant in multivariate analysis predicting a 4-fold increased risk of disease recurrence independently from established prognostic markers. PSMA significantly increases during PCa progression. Conclusion: PSMA is an independent prognostic marker on biopsies at time of initial diagnosis and can predict disease recurrence following curative therapy for PCa. Our study proposes the application of the routinely used IHC marker PSMA for outcome prediction and decision making in risk-adapted PCa management on biopsies at time of initial diagnosis.

Background: Simply applicable biomarkers for prostate cancer (PCa) predicting the clinical course are urgently needed. Recently, TRIM24 has been identified to promote androgen-receptor signaling an ...

Gleason grading is the best independent predictor for prostate cancer (PCa) progression. Recently, a new PCa grading system has been introduced by the International Society of Urological Pathology (ISUP) and is recommended by the World Health Organization (WHO). Following studies observed more accurate and simplified grade stratification of the new system. Aim of this study was to compare the prognostic value of the new Grade Groups compared to the former Gleason Grading, and to determine whether re-definition of Gleason Pattern 4 might reduce up-grading from prostate biopsy to radical prostatectomy (RP) specimen. A cohort of men undergoing radical prostatectomy from 2002 to 2015 at the Hospital of Goeppingen (Goeppingen, Germany) was used for this study. In total, 339 pre-operative prostatic biopsies and corresponding RP specimens, as well as additional 203 RP specimens were re-reviewed for Grade Groups according to the ISUP. Biochemical recurrence free survival (BFS) after surgery was used as endpoint to analyze prognostic significance. Other clinicopathological data included TNM-stage and pre-operative PSA level. Kaplan-Meier analysis revealed risk stratification of patients based on both former Gleason Grading and ISUP Grade Groups, and was statistically significant using the log-rank test (p

Introduction Simply applicable biomarkers for prostate cancer patients predicting the clinical course are urgently needed. Recently, TRIM24 has been identified to promote androgen receptor signaling and to correlate with an aggressive prostate cancer phenotype. Based on these data, we proofed TRIM24 as a prognostic biomarker for risk stratification. Materials and Methods We performed TRIM24 immunohistochemistry on 2 independent cohorts including a total of 806 primary tumors, 26 locally advanced/recurrent tumors, 30 lymph node metastases, 30 distant metastases, and 129 benign prostatic samples from 497 patients as well as on 246 prostate needle biopsies. Expression data were correlated with clinic-pathological data including biochemical recurrence-free survival (bRFS) as endpoint. Results Benign samples show no or low TRIM24 expression in 94%, while tumor tissues demonstrate significant higher levels. Strongest expression is observed in advanced and metastatic tumors. In multivariate analyses, TRIM24 up-regulation on radical prostatectomy specimens correlates with shorter bRFS independent of other prognostic parameters. 5-(10-) year bRFS rates for TRIM24 negative, low, medium and high expressing tumors are 93.1(93.1)%, 75.4(68.5)%, 54.9(47.5)% and 43.1(32.3)%, respectively. Of interest, tumors diagnosed as indolent disease, TRIM24 expression stratifies patients into specific risk groups. Increased TRIM24 expression associates with higher grade group, positive nodal status and extraprostatic tumor growth. TRIM24 assessment on prostate needle biopsies taken prior to treatment decision at time of initial diagnosis significantly correlates with recurrence after surgery. Conclusion Using 2 large independent radical prostatectomy specimen cohorts, we found that TRIM24 expression predicts patients’ risk to develop disease recurrence with high accuracy and independent from other established biomarkers. Further, this is the first study exploring TRIM24 expression on prostate needle biopsies which represents the clinically relevant tissue type on which biomarkers guide treatment decisions. Thus, we strongly suggest introducing TRIM24 evaluation in prostate needle biopsies in clinical routine as an inexpensive and simple immunohistochemical test.

6 Background: Stratifying prostate cancer (PCa) patients into risk groups at time of initial diagnosis for individual treatment decision is still a major clinical challenge. PSMA expression has emerged as a promising prognostic biomarker since its overexpression in radical prostatectomy specimens (RP) has been linked to disease recurrence. Aim of our study was to assess the prognostic value of PSMA on prostate biopsies (Bx) thus improving risk stratification at time of initial diagnosis. Methods: Immunohistochemistry for PSMA expression was performed on 294 Bx with corresponding RP, 621 primary tumor foci from 242 RP, 43 locally advanced or recurrent tumors, 34 lymph node metastases, 78 distant metastases, and 52 benign prostatic samples. Grade group, PSA, TNM-, and R-status were assessed as clinico-pathologic features. Primary endpoint was recurrence-free survival (RFS). Chi-square, ANOVA-analyses, Cox regression and log rank tests were performed for statistical analyses. Results: PSMA expression significantly associates with grade group and initial PSA level. Elevated PSMA expression on both RP and Bx significantly correlates with an increased risk of disease recurrence after curative surgery. 5-year RFS rates are 88.2%, 74.2%, 67.7%, and 26.8% for patients with no, low, medium, or high PSMA expression on Bx, respectively. Elevated PSMA level on Bx predict a 4-fold increased risk of disease recurrence independently from initial PSA and grade group on Bx. PSMA expression significantly increases during PCa progression. Conclusions: PSMA qualifies as an independent prognostic biomarker on Bx at time of initial diagnosis in addition to the established markers PSA and grade group. PSMA predicts disease recurrence following curative surgery and potentially improves the discrimination indolent vs. aggressive disease. We propose the routine assessment of PSMA expression on Bx for outcome prediction and risk stratification at time of initial diagnosis prior to treatment decision.

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.

OBJECTIVES To interrogate multifocal prostate cancer (PCa) to determine its predilection for metastasis, using ERG rearrangement as marker of clonality. A hallmark of PCa is that distinct tumor foci may arise independently, which has important biological and clinical implications. Recent studies characterizing ERG-rearranged PCa possessing intrafocal homogeneity but interfocal heterogeneity support this hypothesis. METHODS We studied 26 patients who underwent prostatectomy and lymphadenectomy with at least 2 distinct PCa foci and 1 lymph node (LN) metastasis. Each focus was assessed for size, Gleason score, ERG rearrangement, and TMPRSS2-ERG transcript. RESULTS Fifteen of 26 cases exhibited interfocal homogeneity with regard to ERG rearrangement (ie, presence vs absence of ERG rearrangement). ERG rearrangement was present in all foci for 6 and absent in all foci for 9 cases. Two cases revealed interfocal heterogeneity with regard to rearrangement mechanism (ie, rearrangement through insertion or deletion). Eight of 26 cases revealed interfocal heterogeneity with regard to rearrangement status. In all cases with at least 1 ERG rearranged focus, we found the corresponding LN metastasis harboring an ERG rearrangement. Interestingly, in a subset of cases the rearrangement status in the LN did not correspond to size or Gleason score. All but 2 ERG rearranged foci had detectable TMPRSS2-ERG transcript levels. CONCLUSIONS When multifocal PCa demonstrates both ERG-positive and ERG-negative foci, the positive foci have a greater predilection for metastasis. Larger studies are needed to confirm the potential additional risk an ERG rearranged focus confers on the likelihood of disease progression.

Localized prostate cancer (CaP) can be cured using several strategies. However, the need to identify active substances in advanced tumor stages is tremendous, as the outcome in such cases is still disappointing. One approach is to deliver human tumor antigen-targeted therapy, which is recognized by T cells or antibodies. We used data mining of the Cancer Immunome Database (CID), which comprises potential immunologic targets identified by serological screening of expression libraries. Candidate antigens were screened by DNA microarrays. Genes were then validated at the protein level by tissue microarrays, representing various stages of CaP disease. Of 43 targets identified by CID, 10 showed an overexpression on the complementary DNA array in CaP metastases. The RHAMM (CD168) gene, earlier identified by our group as an immunogenic antigen in acute and chronic leukemia, also showed highly significant overexpression in CaP metastases compared with localized disease and benign prostatic hyperplasia. At the protein level, RHAMM was highest in metastatic tissue samples and significantly higher in neoplastic localized disease compared with benign tissue. High RHAMM expression was associated with clinical parameters known to be linked to better clinical outcome. Patients with high RHAMM expression in the primaries had a significantly lower risk of biochemical failure. The number of viable cells in cell cultures was reduced in blocking experiments using hormone-sensitive and hormone-insensitive metastatic CaP cell lines. Acknowledging the proven immunogenic effects of RHAMM in leukemia, this antigen is intriguing as a therapeutic target in far-advanced CaP.

Mutations in the androgen receptor (AR) that enable activation by antiandrogens occur in hormone-refractory prostate cancer, suggesting that mutant ARs are selected by treatment. To validate this hypothesis, we compared AR variants in metastases obtained by rapid autopsy of patients treated with flutamide or bicalutamide, or by excision of lymph node metastases from hormone-naïve patients. AR mutations occurred at low levels in all specimens, reflecting genetic heterogeneity of prostate cancer. Base changes recurring in multiple samples or multiple times per sample were considered putative selected mutations. Of 26 recurring missense mutations, most in the NH2-terminal domain (NTD) occurred in multiple tumors, whereas those in the ligand binding domain (LBD) were case specific. Hormone-naïve tumors had few recurring mutations and none in the LBD. Several AR variants were assessed for mechanisms that might underlie treatment resistance. Selection was evident for the promiscuous receptor AR-V716M, which dominated three metastases from one flutamide-treated patient. For the inactive cytoplasmically restricted splice variant AR23, coexpression with AR enhanced ligand response, supporting a decoy function. A novel NTD mutation, W435L, in a motif involved in intramolecular interaction influenced promoter-selective, cell-dependent transactivation. AR-E255K, mutated in a domain that interacts with an E3 ubiquitin ligase, led to increased protein stability and nuclear localization in the absence of ligand. Thus, treatment with antiandrogens selects for gain-of-function AR mutations with altered stability, promoter preference, or ligand specificity. These processes reveal multiple targets for effective therapies regardless of AR mutation. [Cancer Res 2009;69(10):4434–42]

Tumor recurrence after radical cystectomy for bladder cancer can be detected in an asymptomatic patient by regular followup or in a symptomatic patient by symptom guided examination. To our knowledge it is still unknown whether detecting tumor recurrence at an asymptomatic stage offers a better survival rate.A total of 1,270 radical cystectomies for bladder cancer were performed at a single institution between January 1, 1986 and December 2006. All patients had regular followup examinations with chest x-ray and abdominal ultrasound every 3 months, computerized tomography of the abdomen every 6 months, and bone scan and excretory urography every 12 months. Additional examinations were required for symptomatic disease. We analyzed the first site and date of tumor recurrence. Survival was compared using the log rank test.The 20-year recurrence rate was 48.6% in the complete series. Tumor recurrence developed in 444 patients, including 154 asymptomatic and 290 symptomatic patients, with a mean time after radical cystectomy of 20 and 17.5 months, respectively. The most frequent symptoms were pain, ileus, acute urinary retention, hydronephrosis with flank pain, hematuria, neurological symptoms and a palpable mass. Of the 444 patients 182 (41%) had local recurrence and 324 (73%) had distant failure at the time of first recurrence. The overall survival rate 1, 2 and 5 years after first recurrence was 22.5%, 10.1% and 5.5% in asymptomatic patients, and 18.9%, 8.2% and 2.9% in symptomatic patients, respectively (log rank not significant).This study fails to demonstrate a survival benefit for detecting tumor recurrence early at an asymptomatic stage by regular followup examinations. These data show that symptom guided followup examinations may provide similar results at lower cost.

Chromosomal rearrangements account for all erythroblast transformation–specific (ETS) family member gene fusions that have been reported in prostate cancer and have clinical, diagnostic, and prognostic implications. Androgen-regulated genes account for the majority of the 5′ genomic regulatory promoter elements fused with ETS genes. TMPRSS2-ERG, TMPRSS2-ETV1, and SLC45A3-ERG rearrangements account for roughly 90% of ETS fusion prostate cancer. ELK4, another ETS family member, is androgen regulated, involved in promoting cell growth, and highly expressed in a subset of prostate cancer, yet the mechanism of ELK4 overexpression is unknown. In this study, we identified a novel ETS family fusion transcript, SLC45A3-ELK4, and found it to be expressed in both benign prostate tissue and prostate cancer. We found high levels of SLC45A3-ELK4 mRNA restricted to a subset of prostate cancer samples. SLC45A3-ELK4 transcript can be detected at high levels in urine samples from men at risk for prostate cancer. Characterization of the fusion mRNA revealed a major variant in which SLC45A3 exon 1 is fused to ELK4 exon 2. Based on quantitative PCR analyses of DNA, unlike other ETS fusions described in prostate cancer, the expression of SLC45A3-ELK4 mRNA is not exclusive to cases harboring a chromosomal rearrangement. Treatment of LNCaP cancer cells with a synthetic androgen (R1881) revealed that SLC45A3-ELK4, and not endogenous ELK4, mRNA expression is androgen regulated. Altogether, our findings show that SLC45A3-ELK4 mRNA expression is heterogeneous, highly induced in a subset of prostate cancers, androgen regulated, and most commonly occurs through a mechanism other than chromosomal rearrangement (e.g., trans-splicing). [Cancer Res 2009;69(7):2734–8]

A recent genome-wide association study found that genetic variants on chromosomes 3, 6, 7, 10, 11, 19 and X were associated with prostate cancer risk. We evaluated the most significant single-nucleotide polymorphisms (SNP) in these loci using a worldwide consortium of 13 groups (PRACTICAL). Blood DNA from 7,370 prostate cancer cases and 5,742 male controls was analyzed by genotyping assays. Odds ratios (OR) associated with each genotype were estimated using unconditional logistic regression. Six of the seven SNPs showed clear evidence of association with prostate cancer (P = 0.0007-P = 10−17). For each of these six SNPs, the estimated per-allele OR was similar to those previously reported and ranged from 1.12 to 1.29. One SNP on 3p12 (rs2660753) showed a weaker association than previously reported [per-allele OR, 1.08 (95% confidence interval, 1.00-1.16; P = 0.06) versus 1.18 (95% confidence interval, 1.06-1.31)]. The combined risks associated with each pair of SNPs were consistent with a multiplicative risk model. Under this model, and in combination with previously reported SNPs on 8q and 17q, these loci explain 16% of the familial risk of the disease, and men in the top 10% of the risk distribution have a 2.1-fold increased risk relative to general population rates. This study provides strong confirmation of these susceptibility loci in multiple populations and shows that they make an important contribution to prostate cancer risk prediction. (Cancer Epidemiol Biomarkers Prev 2008;17(8):2052–61)

The chromosomal region 7q was repeatedly found to be rearranged in prostate carcinoma. It harbors several well described candidate tumor suppressor and oncogenes. We addressed two genes with opposite roles in cancer; CAV1, a putative tumor suppressor gene at 7q31, and EZH2 at 7q36, which is believed to promote tumor progression. Our primary aim was to assess their expression changes in primary tumors, and then to elucidate the underlying mechanism, assuming that genomic alterations of either locus could affect the other gene as well. In 35 prostate tumor samples, compared with adjacent tissues, CAV1 was overall downregulated (P < 10(-06)), whereas EZH2 was significantly overexpressed (P < 10(-06)). The observed dysregulations were coincident in nearly 70% of the cases. Copy number changes occurred in few tumors. Loss of CAV1 DNA was only marginally associated with reduced expression (P = 0.07), however, and genomic amplification of EZH2 could not explain its upregulation. Through bisulfite sequencing of four tumor samples, CpG-hypermethylation was verified as an alternative mechanism for CAV1 silencing, as reported previously. Moreover, it could also be involved in the reactivation of EZH2.

Introduction: In this prospective study we set out to investigate the diagnostic value of [11C]choline-PET/CT in patients with suspected lymph node metastases before salvage lymph node dissection. Patients and Methods: 15 consecutive patients with rising PSA underwent [11C]choline-PET/CT and consecutive open salvage pelvic/retroperitoneal extended lymph node dissection due to uptake of [11C]choline in at least 1 lymph node. Mean age was 62.1 (range 53–73). Results: [11C]choline-PET/CT results were compared with the histopathology reports and clinical follow-up (mean 13.7 months, range 6–24). Mean time to progression was 23.6 months (range 4–81). [11C]choline uptake was observed in nodes along the external and internal and common iliac arteries and in the paraaortic region. A positive histology was reported in 8/15 patients. Only one patient had a PSA nadir of Conclusions: This interim analysis indicates that [11C]choline-PET/CT may be a useful technique in detection of lymph node metastases when rising PSA occurs after definite prostate cancer therapy. The presented cohort is limited in size, but there is still strong evidence that the patients benefit from [11C]choline-PET/CT and consecutive salvage lymph node dissection is rather small.

Die extrakorporale Stoswellenlithotripsie (ESWL) gilt als sehr sichere, nicht-invasive Behandlungsform der Urolithiasis. Durch technische Weiterentwicklungen sind Komplikationen im Verhaltnis zur Anwendungshaufigkeit immer seltener geworden. Aus unserem Patientenkollektiv entwickelte eine Patientin mit Zystinurie in Folge mehrfacher ESWL-Sitzungen eine temporare Ureterstriktur. Auf der Basis dieser Beobachtung wurde eine Medline-Recherche durchgefuhrt, um das Spektrum seltener aber relevanter Komplikationen bei der modernen Lithotripsie abzubilden.

Hintergrund Bisher galten rekurrente Translokationen und Genfusionen bei den sehr haufigen epithelialen Tumoren als ein seltenes Ereignis. Uberraschenderweise hat unsere Arbeitsgruppe vor kurzem mittels eines neuen bioinformatischen Ansatzes eine neuartige Genfusion beim Prostatakarzinom identifiziert. Dabei fusioniert das androgenregulierte Gen TMPRSS2 mit einem der 3 Gene der ETS-Gengruppe. Bei den ETS-Genen handelt es sich um eine Gruppe von Transkriptionsfaktoren, die bereits in Ewing-Sarkomen als transloziert beschrieben wurden. Ahnlich wie bei Ewing-Tumoren scheint die Genfusion auch beim Prostatakarzinom nachgeschaltete genetische Veranderungen und das klinischen Verhalten zu beeinflussen.

Purpose: We have previously shown that 5-aza-2′-deoxycytidine (5-aza) is an effective chemopreventive agent capable of preventing early disease progression in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. The purpose of this study was to determine the effect of 5-aza on preexisting TRAMP prostate cancers and prevention of androgen-independent prostate cancer. Experimental Design: TRAMP mice with established prostate cancers were treated with 5-aza, castration, castration + 5-aza, or vehicle control (PBS). One cohort of 22 mice per treatment was euthanized after 10 weeks of treatment, whereas a second cohort of 14 mice per group was followed until death to determine survival. Histologic sections of prostate, pelvic lymph nodes, lung, and liver were blinded and analyzed by a certified genitourinary pathologist (K.J.W.). Results: Combined treatment (castration + 5-aza) provided significant survival benefits over either single treatment (combined versus castration P = 0.029, combined versus 5-aza P = 0.036). At 24 weeks of age, 86% of mice within the PBS cohort exhibited histologic evidence of prostate cancer, whereas only 47% of the combined cohort exhibited malignant disease (P < 0.0001). Additionally, whereas 43% of the PBS treatment group exhibited lymph node metastases, these were only observed in 21% of the combined treatment mice. Conclusions: This is the first study to examine the effect of 5-aza and combined castration + 5-aza on preexisting prostate cancer in an animal model. Based on these preclinical findings, we suggest that 5-aza treatment may prolong the time to an androgen-independent status and thus survival in a hormone-deprived setting in prostate cancer.

Butyrates and retinoids are promising antineoplastic agents. Here we analyzed effects of sodium butyrate and N -(4-hydroxyphenyl)-retinamide (4-HPR) on prostate cancer cells as monotherapy or in combination in vitro and in vivo . Sodium butyrate and 4-HPR induced concentration-dependent growth inhibition in prostate cancer cells in vitro . The isobologram analysis revealed that sodium butyrate and 4-HPR administered together antagonize effects of each other. For the in vivo studies, a water-soluble complex (4-HPR with a cyclodextrin) was created. A single dose of sodium butyrate and 4-HPR showed a peak level in chicken plasma within 30 minutes. Both compounds induced inhibition of proliferation and apoptosis in xenografts of the chicken chorioallantoic membrane. Analysis of the cytotoxic effects of the drugs used in combination demonstrated an antagonistic effect on inhibition of proliferation and on induction of apoptosis. Prolonged jun N-terminal kinase phosphorylation induced by sodium butyrate and 4-HPR was strongly attenuated when both compounds were used in combination. Both compounds induced inhibition of NF-κ,B. This effect was strongly antagonized in LNCaP cells when the compounds were used in combination. These results indicate that combinational therapies have to be carefully investigated due to potential antagonistic effects in the clinical setting despite promising results of a monotherapy.

Hydronephrosis in patients with bladder cancer is caused by tumour at the ureteral orifice, secondary ureteral tumours, intramural or extravesical tumour infiltration, or compression of the ureter. This study investigated the prognostic impact of hydronephrosis in bladder cancer.A series of 788 patients were treated with radical cystectomy with curative intent for transitional cell carcinoma of the bladder without neoadjuvant/adjuvant radiotherapy/chemotherapy between January 1986 and September 2003. All patients had a complete follow-up until death or until the study's end date. Survival rates were calculated using the Kaplan-Meier method. A multivariate analysis with a Cox regression model was performed with respect to potential influencing factors.A total of 108 patients (13.7%) had unilateral and 25 patients (3.2%) had bilateral hydronephrosis. The rate of organ-confined tumours was significantly higher in patients without hydronephrosis (67.9% vs. 37.6%; p0.001). Forty-three (32.3%) of the 133 hydronephrotic patients had a tumour involving the ureteral orifice. In this group the rate of organ-confined tumours was significantly higher than in the other patients with hydronephrosis (53.5% vs. 30.0%; p=0.009). In the multivariate analysis, preoperative hydronephrosis was determined as an independent prognostic marker for recurrence-free survival besides the pT classification and lymph node status (p=0.0015). The etiology of hydronephrosis did not affect the tumour-specific survival.Hydronephrosis at the time of diagnosis of bladder cancer is associated with a high probability of advanced tumours. It is an independent prognostic factor for recurrence-free survival.

Activation of E2F transcription factors, through disruption of the retinoblastoma (Rb) tumor-suppressor gene, is a key event in the development of many human cancers. Previously, we showed that homozygous deletion of Rb in a prostate tissue recombination model exhibits increased E2F activity, activation of E2F-target genes, and increased susceptibility to hormonal carcinogenesis. In this study, we examined the expression of E2F1 in 667 prostate tissue cores and compared it with the expression of the androgen receptor (AR), a marker of prostate epithelial differentiation, using tissue microarray analysis. We show that E2F1 expression is low in benign and localized prostate cancer, modestly elevated in metastatic lymph nodes from hormone-naïve patients, and significantly elevated in metastatic tissues from hormone-resistant prostate cancer patients (P = 0.0006). In contrast, strong AR expression was detected in benign prostate (83%), localized prostate cancer (100%), and lymph node metastasis (80%), but decreased to 40% in metastatic hormone-resistant prostate cancer (P = 0.004). Semiquantitative reverse transcription-PCR analysis showed elevated E2F1 mRNA levels and increased levels of the E2F-target genes dihyrofolate reductase and proliferating cell nuclear antigen in metastatic hormone–independent prostate cancer cases compared with benign tissues. To identify a role of E2F1 in hormone-independent prostate cancer, we examined whether E2F1 can regulate AR expression. We show that exogenous expression of E2F1 significantly inhibited AR mRNA and AR protein levels in prostate epithelial cells. E2F1 also inhibited an AR promoter-luciferase construct that was dependent on the transactivation domain of E2F1. Furthermore, using chromatin immunoprecipitation assays, we show that E2F1 and the pocket protein family members p107 and p130 bind to the AR promoter in vivo. Taken together, these results show that elevated E2F1, through its ability to repress AR transcription, may contribute to the progression of hormone-independent prostate cancer. (Cancer Res 2006; 66(24): 11897-906)

Objectives To characterize lymph node metastasis of prostate cancer (PCa) and identify the parameters associated with patient outcome. The incidence of clinically localized PCa with concurrent lymph node metastasis has decreased to less than 1% in the United States but is between 10% and 15% in other countries. Methods Our study cohort of 1148 patients underwent radical prostatectomy in Ulm, Germany, between 1986 and 2002, and 201 (18%) had lymph node-positive PCa. Results The metastases showed growth architecture resembling primary PCa. We assigned a Gleason pattern and evaluated for size, extranodal extension, and lymphovascular invasion (LVI). Of 201 patients, 155 had original pathology slides available; 36 of the 155 were excluded because of preoperative hormonal ablation therapy. Of the remaining 119 patients, 22 (19%) were assigned Gleason pattern 3, 93 (78%) Gleason pattern 4, and 4 (3%) Gleason pattern 5. Extranodal extension was present in 66 (55%) of 119 patients and LVI in 29 (25%). An increased risk of prostate-specific antigen (PSA) recurrence was found for Gleason pattern 4/5 (hazard ratio [HR] 2.5, P = 0.038), LVI in the lymph nodes (HR 1.9, P = 0.038), and nuclear grade of the primary tumor (HR 2, P = 0.025). Independent predictors of PSA recurrence included LVI and nuclear grade (HR 1.9, P = 0.03 and HR 2, P = 0.03, respectively). Conclusions Lymph node metastases of PCa are heterogeneous and have a close relation to the corresponding primary tumor. Most patients with lymph node-positive PCa remained disease free for up to 13 years after radical prostatectomy. Independent predictors of PSA recurrence among those with lymph node-positive PCa included LVI in the lymph nodes and the nuclear grade of the primary tumor. These parameters may be useful in predicting PSA recurrence in lymph node-positive PCa and could be included in patient follow-up.

For patients with metastatic renal cell cancer (RCC), therapeutic options after cytokine failure are rather limited. There is a considerable need to identify new substances for systemic therapy. Due to upregulation after the loss of a functional von Hippel Lindau gene product, the vascular endothelial growth factor (VEGF) pathway is a promising target for a molecular based therapy. Over the last few years, therapeutic agents have been developed which inhibit this pathway at various levels. Here, we provide an overview of the molecular background and currently used drugs which have entered clinical trials in the setting of metastatic RCC disease. Until now, the results from early clinical trials are very promising, however, the best schedule, dosage, potential combination regimens, as well as long time efficacy, are still to be determined.