TMPRSS2:ERG fusion identifies a subgroup of prostate cancers with a favorable prognosis.

PURPOSE: Our aim was to assess the frequency of ERG overexpression and TMPRSS2:ERG rearrangement in prostate cancer and their association with clinicopathologic variables and outcome. EXPERIMENTAL DESIGN: The presence of the TMPRSS2:ERG rearrangement was studied by reverse transcription-PCR and fluorescence in situ hybridization in 19 prostate cancer xenografts and 7 prostate cancer cell lines. The expression of ERG was studied in the xenografts and cell lines and in 49 freshly frozen clinical prostate samples by quantitative reverse transcription-PCR. The frequency of the TMPRSS2:ERG fusion in clinical prostate cancer (n = 253) on tissue microarrays was assessed by three-color fluorescence in situ hybridization. RESULTS: Seven of 19 (37%) of the xenografts overexpressed ERG and had TMPRSS2:ERG rearrangement. Two xenografts, representing small cell carcinomas, also contained the fusion but did not express ERG. In clinical tumor specimens, the overexpression of ERG was associated with the rearrangement (P = 0.0019). Fifty of 150 (33%) of the prostatectomy specimens and 28 of 76 (37%) of the hormone-refractory prostate cancers on the tissue microarrays carried the TMPRSS2:ERG rearrangement. It was associated with longer progression-free survival in patients treated by prostatectomy (P = 0.019), and according to multivariate analysis, it was an independent predictor of favorable outcome (relative risk, 0.54; 95% confidence interval, 0.30-0.98). The fusion was not associated with Gleason score, pT stage, diagnostic prostate-specific antigen, or cell proliferation activity in prostatectomy specimens nor with the AR gene amplification in hormone-refractory tumors. CONCLUSIONS: The TMPRSS2:ERG rearrangement can be found in about one third of prostate cancers. A subgroup of prostate cancer patients with a good prognosis may be identified by the rearrangement.

EZH2, Ki-67 and MCM7 are prognostic markers in prostatectomy treated patients.

The aim of the study was to evaluate the prognostic value of Ki-67, EZH2, MCM7 and EIF3S3 in prostatectomy treated patients. A retrospective population-based material of 249 radical prostatectomy specimens on tissue microarrays was utilized. The median follow-up of the patients was approximately 5.5 years and the main end-point was biochemical progression. The expression of Ki-67, EZH2 and MCM7 was determined by immunohistochemistry and the gene copy number of EIF3S3 was analyzed by fluorescence in situ hybridization (FISH). In the whole material, increased immunostainings of EZH2, MCM7 and Ki-67 were significantly associated with a high Gleason score and a short progression-free survival. In multivariate analysis, MCM7 and Ki-67 showed independent prognostic value with relative risks (RR) of 2.65 (95%-confidence interval of 1.22-5.70), and 1.85 (1.14-3.01), respectively. In subgroup analysis of patients, whose treatment was evaluated to be truly radical (n = 226), EZH2 (3.14, 1.38-7.16), MCM7 (2.70, 1.16-6.30) and PSA (1.5, 1.03-2.20) showed independent prognostic value. In subgroup analysis of cases with a Gleason score <7, low Ki-67 staining was associated with favorable prognosis with RR of 0.09 (0.01-0.69). In conclusion, Ki-67, EZH2 and MCM7 are potential prognostic biomarkers in prostatectomy treated patients.

A new biodegradable braided self-expandable PLGA prostatic stent: an experimental study in the rabbit.

PURPOSE: The biodegradable PLGA (a copolymer of L-lactide and glycolide) urethral stent with a spiral configuration has been used clinically for the prevention of postoperative urinary retention after different types of thermal therapy for benign prostatic hyperplasia. A new braiding pattern for this stent has recently been developed by our group. The aim here was to investigate the in situ degradation and biocompatibility of the new braided stent in the rabbit urethra. MATERIALS AND METHODS: PLGA stents with a one-over-one braiding pattern and steel stents served as controls that were inserted into the posterior urethras of 24 male rabbits using a special delivery instrument. The animals were sacrificed after 1 week, 1 month, 2 months, or 4 months, and light microscopy and histologic analyses were performed. RESULTS: The delivery instrument worked well and cystoscopy was not needed in the insertion process. The braided PLGA stents degraded smoothly in 1 to 2 months. The metallic stents induced more epithelial hyperplasia and epithelial changes than the biodegradable stents at all time points analyzed. These differences increased during follow-up. CONCLUSION: The degradation process was well controlled and the biodegradable stents were more biocompatible than the metallic stents. The new stent can be inserted into the posterior urethra without cystoscopic aid.

Activation of signal transducer and activator of transcription 5 in human prostate cancer is associated with high histological grade.

We have recently identified signal transducer and activator of transcription 5 (Stat5) as a critical survival factor for prostate cancer cells. We now report that activation of Stat5 is associated with high histological grade of human prostate cancer. Specifically, immunohistochemical analysis demonstrated a strong positive correlation with activation of Stat5 and high Gleason score in 114 human prostate cancers. To investigate the mechanisms underlying constitutive activation of Stat5 in prostate cancer, a dominant-negative mutant of Janus kinase 2 (Jak2) was delivered by adenovirus to CWR22Rv cells. Dominant-negative-Jak2 effectively blocked the activation of Stat5 whereas wild-type Jak2 enhanced activation, indicating that Jak2 is the main kinase that phosphorylates Stat5 in human prostate cancer cells. A ligand-induced mechanism for activation of Stat5 in prostate cancer was suggested by the ability of prolactin (Prl) to stimulate activation of both Jak2 and Stat5 in CWR22Rv human prostate cancer cells and in CWR22Rv xenograft tumors. In addition, Prl restored constitutive activation of Stat5 in five of six human prostate cancer specimens in ex vivo long-term organ cultures. Finally, Prl protein was locally expressed in the epithelium of 54% of 80 human prostate cancer specimens with positive correlation with high Gleason scores and activation of Stat5. In conclusion, our data indicate that increased activation of Stat5 was associated with more biologically aggressive behavior of prostate cancer. The results further suggest that Jak2 is the principal Stat5 tyrosine kinase in human prostate cancer, possibly activated by autocrine/paracrine Prl.

Expression of nuclear receptors and cofactors in human endometrium and myometrium.

OBJECTIVE: To study the expression of nuclear receptors and cofactors in human endometrium and myometrium in proliferative and secretory phases of the menstrual cycle. METHODS: Multiprobe ribonuclease protection assay and real-time reverse transcriptase polymerase chain reaction were used to quantitate mRNA levels of steroid receptors, vitamin D receptor (VDR), retinoic acid receptors (RAR), and cofactors AIB1 (amplified in breast cancer-1), CBP (cyclic adenosine monophosphate response element binding protein), pCAF (p300/CBP-associated factor), TIF2 (transcription intermediary factor-2), N-CoR (nuclear receptor corepressor), and SMRT (silencing mediator of repressed transcription). Cyclin A expression was analyzed to determine the proliferation status of the tissues. RESULTS: The expression of androgen receptor, estrogen receptors alpha and beta, progesterone receptor, and RARalpha followed cyclin A expression. There was more abundant expression in the proliferative phase endometrium than in the secretory phase endometrium. Glucocorticoid receptor, VDR, RARbeta, and RARgamma were stably expressed during the menstrual cycle in both endometrium and myometrium. Cofactors N-CoR, SMRT, pCAF, CBP, TIF2, AIB1, and p300 mRNAs were expressed in all samples in both endometrium and myometrium. N-CoR, pCAF, AIB1, and p300 appeared not to be regulated when comparing proliferative and secretory phases of the cycle. Individual differences were found in the expression levels of both nuclear receptors and cofactors. CONCLUSION: The menstrual cycle-dependent regulation of nuclear receptor expression was more apparent in the endometrium than in the myometrium, whereas cofactor expression was not cycle dependent. There were individual differences in the expression levels of different receptors and cofactors. In hormonal therapy these differences might result in different responses, depending on the patient as well as the ligand used.

Cellular changes in prostate cancer cells induced by intermittent androgen suppression.

OBJECTIVES: This study used tumour specimens from a clinical trial on intermittent androgen suppression (IAS) to study the effects of castration on tissue morphology, cell proliferation, apoptosis, and the expression of androgen receptor (AR). METHODS: A total of 113 representative needle biopsy specimens were available from 29 patients for evaluation of the Gleason score as well as Ki-67, cleaved caspase-3, and AR immunostaining. RESULTS: At 6 mo from the beginning of the first androgen withdrawal, cell proliferation activity was significantly (p = 0.002) reduced, whereas no effect on apoptosis was found. A nonsignificant trend to an increase in Gleason score after castration was found. Subsequent cycles of withdrawals had no significant effect on any of the measured parameters. The Gleason score, proliferation activity, and apoptosis rate showed only heterogeneous, nonsignificant changes after the first progression. Strong nuclear AR staining was evident in all cancer specimens. CONCLUSIONS: The findings suggest that the long-term (months) effect of androgen withdrawal on tumour growth is due to the inhibition of proliferation. Because the tumours seem to become resistant to castration at the first cycle of treatment withheld, it is possible that IAS does not postpone the emergence of ablation-resistant tumours. There was no consistent sign of increased proliferation or Gleason score during the treatment, suggesting that the biologic aggressiveness of a particular tumour is defined already at an early stage of disease. The constant nuclear expression of AR in cancer cells indicates that the AR signalling remains active despite of the androgen withdrawal.

Bax and Bcl-2 are focally overexpressed in the normal epithelium of cancerous prostates.

OBJECTIVE: Development of prostate cancer is connected with a disturbance of apoptosis. Prostate cancer is multifocal, suggesting that the control of apoptosis is impaired at multiple foci. We wanted to know whether apoptosis is generally disturbed in cancerous prostates and if changes in apoptotic control could be detected even in the absence of any morphologically visible changes. Therefore, we compared expression of two common apoptotic markers, Bax and Bcl-2, in normal epithelium of cancerous prostates and controls. We also evaluated the expression of these proteins in hyperplasia, prostatic intraepithelial neoplasia (PIN), carcinomas of different Gleason grades and capsular perineural invasion. MATERIAL AND METHODS: The tissue material was obtained from radical prostatectomies, transurethral resection chips and autopsies. Individual tissue arrays were done for each patient. The intensity of Bax and Bcl-2 immunostaining was estimated semiquantitatively. The data were analyzed using a linear mixed-models analysis as well as dichotomized staining indices. RESULTS: Normal epithelium of cancerous prostates contained foci with high expression of Bax and Bcl-2. The expression of Bax in Gleason grades 3-5 carcinoma was significantly higher than that in Gleason grade 2, and was highest in foci with perineural invasion. The expression of Bcl-2 was strongest in PIN foci. CONCLUSIONS: Expression of Bax and Bcl-2 in normal epithelium of cancerous prostates suggests that increases in these indirect markers may reflect altered apoptotic control in these foci. Further studies are needed to show whether these changes represent the earliest step of the multifocal carcinogenetic process. Control of apoptosis seems to be involved and modulated during local progression of prostate cancer.

Tissue biocompatibility of new biodegradable drug-eluting stent materials.

Drug-eluting stents are a recent innovation for endovascular and endourethral purposes. The aim of this study was to assess the biocompatibility of new biodegradable drug-eluting stent materials in vivo. Rods made of SR-PLDLA (self-reinforced poly-96L,4D: -lactic acid) covered with P(50L/50D)LA and rods made of 96L/4D SR-PLA and covered with P(50L/50D)LA including indomethacin 3.3 microg/mm(2)or dexamethasone 1.5 microg/mm(2), were inserted into the dorsal muscles of 20 rabbits serving as test animals. Rods made of silicone and organotin-positive polyvinylchloride were used as negative and positive controls. The animals were sacrificed after 1 week, 1 month, 2 months or 4 months. Histological changes attributable to the operative trauma were seen in all specimens at 1 week and 1 month. At 2 months both dexamethasone and indomethacin induced less fibrosis than the plain SR-PLDLA covered with P(50L/50D)LA without drug. At 4 months dexamethasone induced both chronic inflammatory changes and foreign body reaction, whereas the reactions in the indomethacin and drug-free plain SR-PLDLA groups were insignificant. The new biodegradable drug-eluting stent materials are highly biocompatible. Drug-eluting biodegradable stents may offer a promising new treatment modality for vascular and urethral diseases. However, further studies are needed to demonstrate their feasibility and efficacy.

The gene for polycomb group protein enhancer of zeste homolog 2 (EZH2) is amplified in late-stage prostate cancer.

Overexpression of the polycomb group protein enhancer of zeste homologue 2 (EZH2) has been found in several malignancies, including prostate cancer, with an aggressive phenotype. Amplification of the gene has previously been demonstrated in several malignancies, but not in prostate cancer. Our goal was to evaluate the gene copy number and expression alterations of EZH2 in prostate cancer. The copy number of EZH2 in cell lines (LNCaP, DU145, PC-3, 22Rv1), xenografts (n = 10), and clinical tumors (n = 191) was studied with fluorescence in situ hybridization. All cell lines had a gain of EZH2. Eight of the ten xenografts showed an increased copy number of the gene, including one case of high-level amplification (>or=5 copies of the gene and EZH2/centromere ratio >or=2). 34/125 (27%) of untreated prostate carcinomas showed increased copy number, but only one case of low-level amplification (>or=5 copies of the gene and EZH2/centromere ratio <2), whereas half (25/46) of the hormone-refractory carcinomas showed increased copy number, including seven cases of low-level amplification and three cases of high-level amplification (P < 0.0001). Expression of EZH2 was significantly (P = 0.0009) higher in hormone-refractory prostate cancer compared with that in benign prostatic hyperplasia or untreated cancer, according to quantitative real-time RT-PCR assay. Also, the expression of EZH2 protein was found to be higher in hormone-refractory tumors than in hormone-naïve tumors by immunohistochemistry. The EZH2 gene amplification was significantly (P < 0.05) associated with increased EZH2 protein expression. The data show that amplification of the EZH2 gene is rare in early prostate cancer, whereas a fraction of late-stage tumors contains the gene amplification leading to the overexpression of the gene, thus indicating the importance of EZH2 in the progression of prostate cancer.

Biocompatibility properties of a new braided biodegradable urethral stent: a comparison with a biodegradable spiral and a braided metallic stent in the rabbit urethra.

OBJECTIVE: To compare the biocompatibility properties of a new braided biodegradable self-reinforced poly-L-lactic acid (SR-PLLA) urethral stent to the former spiral biodegradable SR-PLLA stent and the stainless steel stent in a rabbit model. MATERIALS AND METHODS: In all, 54 male New Zealand White rabbits were anaesthetized and stents inserted into the prostatic urethra, three of each kind for each sample time. The rabbits were killed after 1, 3, 6, 9, 12 or 15 months and light microscopy and scanning electron microscopy used to analyse the effects. RESULTS: The disintegration of the braided SR-PLLA stent was more closely controlled than that of the spiral SR-PLLA stent. The metallic stent induced epithelial hyperplasia and polyposis earlier than the biodegradable stents, and in these rabbits the polyposis disappeared after the disintegration process. There were no differences in the histological analyses between the biodegradable stents, whereas the metallic stents caused the strongest inflammatory reactions. CONCLUSIONS: The braided SR-PLLA urethral stent functioned well in the rabbit urethra and clinical studies are already planned.

Screening of genetic and expression alterations of SRC1 gene in prostate cancer.

BACKGROUND: Genetic alterations of the SRC1 gene have not been thoroughly studied in prostate cancer. MATERIALS AND METHODS: Five prostate cancer cell lines and 32 xenografts were screened for mutations and gene copy number alterations. Subsequently, frequencies of detected sequence variations were further analyzed in 44 clinical prostate cancers, 6 benign prostate hyperplasias, and 48 normal controls. Finally, the protein expression of SRC1 in 254 clinical prostate tumors was investigated. RESULTS: Three non-recurrent sequence variations, and one single nucleotide polymorphism in the coding region of SRC1, as well as one case of SRC1 gene amplification were found. The protein expression of SRC1 was higher in androgen ablation resistant than untreated prostate carcinomas, but the difference was not statistically significant (P = 0.0796). CONCLUSIONS: Genetic alterations of SRC1 are rare in prostate cancer. The nuclear protein accumulation of SRC1 seems to be mildly increased in androgen ablation resistant prostate cancers. .

Detection rates of high-grade prostate cancer during subsequent screening visits. Results of the European Randomized Screening Study for Prostate Cancer.

Screening for prostate cancer using prostate-specific antigen (PSA) tests has led to a stage and grade shift as compared to the pre-PSA era. Effectiveness of screening for prostate cancer should be manifested by a reduction in detection rate of aggressive cancers during subsequent screening. In 6 centers of the European Randomized Screening study for Prostate Cancer, a total of 58,710 men were tested for prostate cancer. Screening centers differed with regard to age-range, screening interval and biopsy indications. During the 2nd visit, the proportion of Gleason score 6 cancers increased from 62.5 to 75%, mainly at the expense of Gleason score 7 cancers. High-grade (Gleason score 8-10) cancer detection rates varied per screening center during the 1st visit from 5.1 to 41.1, and during the 2nd visit from 6.4 to 29.3/10,000 men. The overall detection rate of high-grade cancers showed a reduction during the 2nd visit from 26 to 12/10,000 men, an effect mainly attributable to the screening center with the highest cancer detection rate (i.e. 507/10,000 men). Variations in detection rates among screening centers related among others to biopsy compliance and age range.

Dual-specificity phosphatase 1 and serum/glucocorticoid-regulated kinase are downregulated in prostate cancer.

Inactivation of tumor suppressor genes through deletion, mutation and epigenetic silencing has been shown to occur in cancer. In our study, we combined DNA demethylation and histone deacetylation inhibition treatments with suppression subtraction hybridization (SSH) and cDNA microarrays to identify potentially epigenetically downregulated genes in PC-3 prostate cancer cell line. We found 11 genes whose expression was upregulated after relieving epigenetic regulation. Expression of 3 genes [dual-specificity phosphatase 1 (DUSP1), serum/glucocorticoid regulated kinase (SGK) and spermidine/spermine N1-acetyltransferase (SAT)] was subsequently studied in clinical sample material using real-time quantitative RT-PCR and immunohistochemistry. The DUSP1 and SGK mRNA expression was lower in hormone-refractory prostate carcinomas compared to benign prostate hyperplasia (BPH) or untreated prostate carcinomas. BPH, normal prostate and high-grade prostate intraepithelial neoplasia (PIN) expressed high levels of DUSP1 and SGK proteins. Ninety-two percent and 48% of the prostate carcinomas showed almost complete lack of DUSP1 and SGK proteins, respectively, indicating common downregulation of these genes. The genomic bisulphite sequencing did not reveal dense hypermethylation in the promoter regions of either DUSP1 or SGK. In conclusion, the data suggest that downregulation of DUSP1 and SGK is an early event and could be important in the tumorigenesis of prostate cancer.

Biocompatibility and implantation properties of 2 differently braided, biodegradable, self-reinforced polylactic acid urethral stents: an experimental study in the rabbit.

PURPOSE: Biodegradable urethral stents have been in clinical use for more than 10 years. To solve the problems connected with the helical spiral configuration of the stents used to date we developed a new tubular mesh configuration and evaluated the biocompatibility properties and degradation time of 2 differently braided stents in the rabbit urethra. MATERIALS AND METHODS: The biodegradable, self-expanding stents were made of self-reinforced polylactic acid polymer blended with BaSO4 (Alfa Chem, Kings Point, New York). Two braiding patterns, namely a diamond 1/1 and a regular 2/2 + 1 (Prodesco, Perkasie, Pennsylvania), were used to produce a tubular mesh configuration. Stainless steel stents with 1/1 braiding served as controls. The stents were inserted into the posterior urethra of 36 male rabbits. The animals were sacrificed after 1 week, 1 month, 6 months or 12 months. Light microscopy and scanning electron microscopy analyses were done. RESULTS: Tissue reactions to operative trauma were seen in all specimens at week 1. The changes gradually abated in the biodegradable stent groups, whereas chronic inflammatory changes and fibrosis were increasingly seen with metallic stents after 6 months. Epithelial hyperplasia increased with time for all stent types and materials. As expected, stent fragmentation started at 6 months. CONCLUSIONS: Biodegradable polymers are suitable materials for braided urethral stents. However, the braided configuration of the stent with a decreased mass of material does not prevent the development of epithelial hyperplasia. The biodegradable, self-expanding, braided stents functioned well in the rabbit urethra and are suitable for clinical studies.

Renal cell carcinoma MIB-1, Bax and Bcl-2 expression and prognosis.

PURPOSE: Proliferation and programmed cell death (apoptosis) are key factors in oncogenesis and tumor progression. In carcinogenesis important regulators of apoptosis are members of the Bcl-2 family. In this family the Bcl-2 gene has an inhibitory effect on apoptosis, while Bax promotes cell death. In renal cell carcinoma (RCC) the associations between Bcl-2 proteins and RCC prognosis have been controversial. We evaluated Bax and Bcl-2 levels in RCC, and their associations with prognosis, proliferation and traditional prognostic factors. MATERIALS AND METHODS: Our prospective study population comprised 138 consecutive patients who underwent radical nephrectomy for RCC. Immunostaining and semiquantitative indices for Ki-67 (MIB-1), Bax and Bcl-2 were estimated. Their associations with prognosis were explored. RESULTS: On univariate analysis according to survival statistically significant differences were achieved by Bax (positive vs negative HR 3.04, 95% CI 1.27 to 7.23), Bcl-2 (positive vs negative HR 0.43, 95% CI 0.23 to 0.81), MIB-1 (continuous HR 1.03, 95% CI 1.001 to 1.064), Fuhrman nuclear class (4 vs 1 plus 2 HR 8.15, 95% CI 3.13 to 21.20) and stage (4 vs 1 HR 60.04, 95% CI 13.99 to 257.68). Only stage (HR 47.96, 95% CI 10.85 to 212.03) and Fuhrman classification (HR 4.32, 95% CI 1.60 to 11.65) attained statistical significance on Cox regression multivariate analysis. CONCLUSIONS: In our prospective study Bax and Bcl-2 showed a statistically significant association with prognosis in RCC but did not achieve the status of independent prognostic factors. Further studies are needed to clarify the role of the apoptotic process in tumor progression and prognosis.