miR-124 exhibits antiproliferative and antiaggressive effects on prostate cancer cells through PACE4 pathway.

INTRODUCTION: PACE4 plays an important role in prostate cancer (PCa) proliferation and aggression, which might provide a useful target against prostate cancer. In this study, we had strived to find some key miRNAs to decrease malignancy and invasiveness of PCa through regulating PACE4 expression. METHODS: Clinically pathological analysis of immunohistochemistry/in situ hybridization was carried out to detect the relationship between PACE4 expression/miRNAs and the malignancy of prostate mass. Prostate cell lines (DU145, C4-2, and BPH-1) were cultured for growth curve, immunocytochemistry analysis, colony formation, Matrigel invasion, and transcriptional/translational expression assay of PACE4-related signaling molecules for confirming the relationship. MiRNAs targeting PACE4 were predicted, validated and further-corroborated using bio-software, real-time PCR, luciferase reporter assay and transfection of miRNA mimics and inhibitor. RESULTS: It was suggested that PACE4 might reflect the pathological malignancy of prostate lesion from pathology analysis. Moreover, DU145 cells, the highest PACE4-level and related TF expression indicated of the strongest malignancy and invasiveness. It was significantly found that miR-124 was presented with the biggest odd to target PACE4-3'UTR, the capability of decreasing PACE expression and slowing down cell growth and cell invasion. CONCLUSIONS: It was clear that PACE4 level was closely associated with malignancy and invasiveness of PCa in vivo or in vitro MiR-124, played a crucial role inhibiting PACE4 transcription thus exhibiting obvious effects of antiproliferation and antiaggression of PCa.

Sprouty genes function in suppression of prostate tumorigenesis.

Expression of Sprouty genes is frequently decreased or absent in human prostate cancer, implicating them as suppressors of tumorigenesis. Here we show they function in prostate tumor suppression in the mouse. Concomitant inactivation of Spry1 and Spry2 in prostate epithelium causes ductal hyperplasia and low-grade prostatic intraepithelial neoplasia (PIN). However, when Spry1 and Spry2 loss-of-function occurs in the context of heterozygosity for a null allele of the tumor suppressor gene Pten, there is a striking increase in PIN and evidence of neoplastic invasion. Conversely, expression of a Spry2 gain-of-function transgene in Pten null prostatic epithelium suppresses the tumorigenic effects of loss of Pten function. We show that Sprouty gene loss-of-function results in hyperactive RAS/ERK1/2 signaling throughout the prostate epithelium and cooperates with heterozygosity for a Pten null allele to promote hyperactive PI3K/AKT signaling. Furthermore, Spry2 gain-of-function can suppress hyperactivation of AKT caused by the absence of PTEN. Together, these results point to a key genetic interaction between Sprouty genes and Pten in prostate tumorigenesis and provide strong evidence that Sprouty genes can function to modulate signaling via the RAS/ERK1/2 and PI3K/AKT pathways. The finding that Sprouty genes suppress tumorigenesis caused by Pten loss-of-function suggests that therapeutic approaches aimed at restoring normal feedback mechanisms triggered by receptor tyrosine kinase signaling, including Sprouty gene expression, may provide an effective strategy to delay or prevent high-grade PIN and invasive prostate cancer.

Relationship between IL-6/ERK and NF-κB: a study in normal and pathological human prostate gland.

BACKGROUND: There is growing evidence that inflammation is a causal factor in cancer, where pro-inflammatory cytokines such as IL-6, IL-1 or TNF-α could induce cellular proliferation by activation of NF-κB. This study focuses on the IL-6/ERK transduction pathway, its relationship with NF-κB, and the consequences of dysregulation in the development of prostate pathologies such as benign prostate hyperplasia (BPH), prostate intraepithelial neoplasia (PIN) and prostate cancer (PC). METHODS: Immunohistochemical and Western blot analyses for IL-6, gp-130, Raf-1, MEK-1, ERK-1, p-MEK, ERK-2, p-ERK, NF-κB/p-50 and NF-κB/p-65 were carried out in 20 samples of normal prostate glands, 35 samples of BPH, 27 samples with a diagnosis of PIN (low-grade PIN or high-grade PIN), and 95 samples of PC (23 with low, 51 with medium and 21 with high Gleason scores). RESULTS: Immunoreaction to IL-6, gp-130, ERK-1, ERK-2, p-ERK and NF-κB/p50 was found in the cytoplasm of epithelial cells in normal prostate samples; p-MEK was found in the nucleus of epithelial cells; but not expression to Raf-1, MEK-1 and NF-κB/p65. In BPH, all of these proteins were immunoexpressed, while there was increased immunoexpression of IL-6, gp-130, p-MEK, ERK-1, ERK-2 and NF-κB/p50 (cytoplasm). In PC, immunoexpression of IL-6 and gp-130 were similar to that found in BPH; while immunoexpression of Raf-1, MEK-1, p-MEK, ERK-1, ERK-2, p-ERK, NF-κB/p50 (nucleus and cytoplasm), and NF-κB/p65 (nucleus and cytoplasm) was higher than in BPH. CONCLUSION: Translocation of NF-κB to the nucleus in PC and high-grade PIN could be stimulated by the IL-6/ERK transduction pathway, but might also be stimulated by other transduction pathways, such as TNF-α/NIK, TNF/p38, IL-1/NIK or IL-1/p38. Activation of NF-κB in PC could regulate IL-6 expression. These transduction pathways are also related to activation of other transcription factors such as Elk-1, ATF-2 or c-myc (also involved in cell proliferation and survival). PC is a heterogeneous disease, where multiple transduction pathways might alter the apoptosis/proliferation balance. Significant attention should be give to the combination of novel agents directed towards inactivation of pro-inflammatory cytokines than can disrupt tumour cell growth.

VEGF elicits epithelial-mesenchymal transition (EMT) in prostate intraepithelial neoplasia (PIN)-like cells via an autocrine loop.

Vascular endothelial growth factor (VEGF) is overexpressed during the transition from prostate intraepithelial neoplasia (PIN) to invasive carcinoma. We have mimicked such a process in vitro using the PIN-like C3(1)/Tag-derived Pr-111 cell line, which expresses low levels of VEGF and exhibits very low tumorigenicity in vivo. Elevated expression of VEGF164 in Pr-111 cells led to a significant increase in tumorigenicity, invasiveness, proliferation rates and angiogenesis. Moreover, VEGF164 induced strong changes in cell morphology and cell transcriptome through an autocrine mechanism, with changes in TGF-beta1- and cytoskeleton-related pathways, among others. Further analysis of VEGF-overexpressing Pr-111 cells or following exogenous addition of recombinant VEGF shows acquisition of epithelial-mesenchymal transition (EMT) features, with an increased expression of mesenchymal markers, such as N-cadherin, Snail1, Snail2 (Slug) and vimentin, and a decrease in E-cadherin. Administration of VEGF led to changes in TGF-beta1 signaling, including reduction of Smad7 (TGF-beta inhibitory Smad), increase in TGF-betaR-II, and translocation of phospho-Smad3 to the nucleus. Our results suggest that increased expression of VEGF in malignant cells during the transition from PIN to invasive carcinoma leads to EMT through an autocrine loop, which would promote tumor cell invasion and motility. Therapeutic blockade of VEGF/TGF-beta1 in PIN lesions might impair not only tumor angiogenesis, but also the early dissemination of malignant cells outside the epithelial layer.

[Apoptosis in pathologic prostatic processes]. / Apoptoza u patoloskim procesima prostate.

Numerous studies have demonstrated the apoptotic index to be significantly higher in benign prostatic lesions than in PIN and adenocarcinoma. Furthermore, a lower apoptotic index in epithelial cell populations has been shown to correlate with decreased immunoreactivity for prostate specific antigen that may be seen in higher Gleason grade tumors. It is suggested that the loss of function of some apoptotic regulators contributes to the development of PIN and prostatic adenocarcinoma. Of these, three signaling molecules involved in apoptotic functions ofTGF-beta have now been intensively investigated, including transmembrane receptor II (T betaRII), cell cycle inhibitor p27(Kp1) and Smad4, effectors of TGF-beta signaling pathway. Increased expression of p53 and decreased expression of maspin, a serine protease inhibitor, also play a major role in the apoptotic process. Changes in the expression of these markers may serve as a reliable criterion on making the diagnosis in biopsy material and may help choose an appropriate therapeutic approach.

Activation of beta-Catenin in mouse prostate causes HGPIN and continuous prostate growth after castration.

BACKGROUND: The role of Wnt/beta-Catenin signaling in embryogenesis and carcinogenesis has been extensively studied in organs such as colon, lung and pancreas, but little is known about Wnt/beta-Catenin signaling in the prostate. Although stabilizing mutations in APC and beta-Catenin are rare in primary prostate tumors, recent studies suggest that cytoplasmic/nuclear beta-Catenin is associated with advanced, metastatic, hormone-refractory prostate carcinoma. METHODS: To better understand the role of beta-Catenin in prostatic development and carcinogenesis, we studied Wnt expression during prostate development and activated Wnt/beta-Catenin signaling in the developing and adult prostate. RESULTS: Our results demonstrated that during prostate development Wnt ligands display a dynamic expression pattern. Activation of beta-Catenin during prostate development caused epithelial hyperplasia followed by prostatic intraepithelial neoplasia (PIN) in prostate. In the adult prostate, activation of beta-Catenin resulted in high grade PIN (HGPIN) and continuous prostatic growth after castration. As a result of activation of beta-Catenin, AR was first up-regulated with the emergence of epithelial hyperplasia, but was later down-regulated when HGPIN developed. Furthermore, activation of beta-Catenin induced Foxa2 re-expression in adult prostate which normally is only expressed in the embryonic budding stage during prostate development. CONCLUSIONS: The results from this study strongly suggest that Wnt/beta-Catenin signaling is involved in the regulation of prostate development and confirm that constitutive activation of this pathway enables the mouse prostate to grow after castration.

Advances in mouse models of prostate cancer.

Advances in science and technology have allowed us to manipulate the mouse genome and analyse the effect of specific genetic alterations on the development of prostate cancer in vivo. We can now analyse the molecular basis of initiation, invasion and progression to metastatic disease. The current mouse models utilise knockout, knock-in or conditional regulation of expression using Cre-loxP technology. Genes that have been targeted include homeobox genes, tumour suppressors and oncogenes, growth factors (and their receptors), steroid hormones and cell-cycle regulators, as well as pro- and anti-apoptotic proteins. Bigenic models indicate that that two 'hits' are required for progression from intra-epithelial neoplasia (PIN) to invasion carcinoma, and two to five hits are needed for metastasis. Here, we discuss the numerous models that mimic various aspects of the disease process, such as PIN, locally invasive adenocarcinoma and metastatic disease. Currently the PB-Cre4 x PTEN(loxP/loxP) mouse is the only model that spans the entire continuum from initiation to local invasion and metastasis. Such mouse models increase our understanding of the disease process and provide targets for novel therapeutic approaches. Hopefully, the transgenic models will become inducible and ultimately allow both temporal and spatial gene inactivation. Compound mutational models will also develop further, with double and triple knock-in or knockout systems adding to our knowledge of the interaction between different signalling cascades.

Overexpression of cyclooxygenase-2 in human prostate carcinoma and prostatic intraepithelial neoplasia-association with increased expression of Polo-like kinase-1.

BACKGROUND: Cyclooxygenases (COX) as well as Polo-like kinases (PLK) are involved in proliferation and cell cycle regulation and have been suggested for preventive and therapeutic approaches in prostate carcinoma. METHODS: In this study, we studied expression and prognostic impact of COX-2 in invasive prostate carcinoma, prostatic intraepithelial neoplasia (PIN), atrophic glands, and normal prostatic glands, and investigated the association between COX-2 and PLK-1. RESULTS: We observed a positivity for COX-2 in 72.1% of PIN and in 44.7% of prostate carcinomas with an overexpression of COX-2 in prostate cancer and PIN compared to benign prostatic tissue (P < 0.0005). Furthermore, we observed a strong correlation between expression of PLK-1 and COX-2 (P < 0.0005). CONCLUSIONS: To our knowledge, this is the first report of a correlation between COX-2 and PLK-1 in a malignant tumor. COX-2 and PLK-1 may be interesting targets for new molecular therapies in prostate cancer.

DNA methylation paradigm shift: 15-lipoxygenase-1 upregulation in prostatic intraepithelial neoplasia and prostate cancer by atypical promoter hypermethylation.

Fifteen (15)-lipoxygenase type 1 (15-LO-1, ALOX15), a highly regulated, tissue- and cell-type-specific lipid-peroxidating enzyme has several functions ranging from physiological membrane remodeling, pathogenesis of atherosclerosis, inflammation and carcinogenesis. Several of our findings support a possible role for 15-LO-1 in prostate cancer (PCa) tumorigenesis. In the present study, we identified a CpG island in the 15-LO-1 promoter and demonstrate that the methylation status of a specific CpG within this island region is associated with transcriptional activation or repression of the 15-LO-1 gene. High levels of 15-LO-1 expression was exclusively correlated with one of the CpG dinucleotides within the 15-LO-1 promoter in all examined PCa cell-lines expressing 15-LO-1 mRNA. We examined the methylation status of this specific CpG in microdissected high grade prostatic intraepithelial neoplasia (HGPIN), PCa, metastatic human prostate tissues, normal prostate cell lines and human donor (normal) prostates. Methylation of this CpG correlated with HGPIN, PCa and metastatic human prostate tissues, while this CpG was unmethylated in all of the normal prostate cell lines and human donor (normal) prostates that either did not display or had minimal basal 15-LO-1 expression. Immunohistochemistry for 15-LO-1 was performed in prostates from PCa patients with Gleason scores 6, 7 [(4+3) and (3+4)], >7 with metastasis, (8-10) and 5 normal (donor) individual males. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to detect 15-LO-1 in PrEC, RWPE-1, BPH-1, DU-145, LAPC-4, LNCaP, MDAPCa2b and PC-3 cell lines. The specific methylated CpG dinucleotide within the CpG island of the 15-LO-1 promoter was identified by bisulfite sequencing from these cell lines. The methylation status was determined by COBRA analyses of one specific CpG dinucleotide within the 15-LO-1 promoter in these cell lines and in prostates from patients and normal individuals. Fifteen-LO-1, GSTPi and beta-actin mRNA expression in BPH-1, LNCaP and MDAPCa2b cell lines with or without 5-aza-2'-deoxycytidine (5-aza-dC) and trichostatin-A (TSA) treatment were investigated by qRT-PCR. Complete or partial methylation of 15-LO-1 promoter was observed in all PCa patients but the normal donor prostates showed significantly less or no methylation. Exposure of LNCAP and MDAPCa2b cell lines to 5-aza-dC and TSA resulted in the downregulation of 15-LO-1 gene expression. Our results demonstrate that 15-LO-1 promoter methylation is frequently present in PCa patients and identify a new role for epigenetic phenomenon in PCa wherein hypermethylation of the 15-LO-1 promoter leads to the upregulation of 15-LO-1 expression and enzyme activity contributes to PCa initiation and progression.

Expression and role of Foxa proteins in prostate cancer.

The molecular mechanism(s) for prostate cancer progression to androgen independence are poorly understood. We have recently shown that Foxa1 and Foxa2 proteins are differentially expressed in epithelial cells during murine prostate development, growth, and adult function. Currently, the role of Foxa proteins in prostate cancer development and progression is unknown. Foxa protein expression was investigated in the LPB-Tag LADY mouse prostate cancer models, in human prostate cancer specimens, and various prostate cancer cell lines using Western blot and immunostaining analysis. In vitro transient transfection, studies were performed to investigate Foxa/prostate-specific gene regulation. Foxa1 was strongly expressed in areas of prostatic intraepithelial neoplasia (PIN) in both the androgen dependent 12T-7f and in the metastatic, androgen independent 12T-10 LADY models. Prominent Foxa1 and Foxa2 expression was observed in 12T-10 invasive undifferentiated neuroendocrine carcinomas, in the hormone independent and metastasizing 12T-10 derived, NE-10 allograft tumors, and in all metastatic lesions isolated from 12T-10 mice. Foxa1 protein expression was always observed in human prostate carcinomas, regardless of Gleason grade score, while Foxa2 was only detected in neuroendocrine small cell carcinomas and in some high Gleason score adenocarcinomas. Foxa proteins were also differentially expressed in three prostate cancer cell lines. Importantly, in vitro functional assays demonstrated that Foxa2 could activate androgen-dependent prostate-specific genes in an androgen receptor and ligand-independent manner. These results suggest that Foxa proteins are important in prostate carcinogenesis. In particular, Foxa2 may be involved in progression of prostate cancer to androgen independence. As such, Foxa proteins may represent novel targets for therapeutic intervention.

Serum insulin-like growth factor-I levels and prostatic intraepithelial neoplasia: a clue to the relationship between IGF-I physiology and prostate cancer risk.

Serum insulin-like growth factor-I (IGF-I) levels at the higher end of the reference range have been associated with increased risk for the future development of prostate cancer. We determined whether high serum IGF-I levels are associated with precancerous lesions of the prostate. We conducted a case-control study to determine whether high serum IGF-I levels were associated with the presence of high-grade prostatic intraepithelial neoplasia (HGPIN) among patients who presented for prostate biopsy because of an abnormal serum prostate-specific antigen level or digital rectal exam. We measured serum IGF-I and insulin-like growth factor binding protein-3 (IGFBP-3) prior to prostate biopsy and compared them between 103 men with HGPIN (cases) and 205 men with normal prostate histology (controls). The mean IGF-I level in patients with HGPIN (130.2 ng/mL) was significantly higher compared with controls (118.8 ng/mL, P = 0.01). The mean IGFBP-3 level in patients with HGPIN (2,393.9 ng/mL) was also higher compared with controls (2,276.0 ng/mL, P = 0.06). After adjusting for age, prostate-specific antigen, digital rectal examination, and ethnic background, the odds ratio for a HGPIN diagnosis among men in the highest relative to the lowest quartile of serum IGF-I level was 1.94 (95% confidence interval, 1.0-3.7; P = 0.04). The potential association between a high serum IGF-I level and the presence of HGPIN may represent an important clue to understanding the basis for the relationship between IGF-I physiology and prostate cancer risk. Larger studies will be required to confirm this relationship.

Decreased retinoid X receptor-alpha protein expression in basal cells occurs in the early stage of human prostate cancer development.

The development of prostatic intraepithelial neoplasia (PIN)-like lesions in the prostate-specific retinoid X receptor-alpha (RXRalpha) null mouse suggests that RXRalpha may protect against neoplasia. The purpose of this study was to characterize RXRalpha protein expression in human prostate to determine if RXRalpha is altered in early stages of tumor progression. Immunohistochemistry with anti-RXRalpha antibody was performed on 138 fresh frozen prostate specimens collected from 27 noncarcinomatous prostates and 111 radical prostatectomy samples of prostate adenocarcinoma (CA). The RXRalpha signal intensity was scored using a scale of 0-3. In normal glands, RXRalpha was expressed strongly in basal cells and only weakly in secretory epithelial cells. This finding was confirmed by double immunofluorescence labeling of RXRalpha and Keratin-903, a basal cell marker, followed by confocal microscopic examination. In basal cells, a gradual decrease of RXRalpha expression was noted from normal glands of noncarcinomatous prostate (3.0 +/- 0) to "normal" glands distant to CA (2.13 +/- 0.44) to "normal" glands adjacent to CA (1.25 +/- 0.53) and high-grade PIN (0.56 +/- 0.58). While nearly all "normal" glands from 138 specimens were positive for RXRalpha in basal cells, only 48% (13 of 27) of the high-grade PIN glands appeared positive. Moreover, basal cell expression of RXRalpha in "normal" tissue was less in specimens with poorly differentiated tumor (Gleason score >/= 8; 1.83 +/- 0.36) compared with well-differentiated tumor (Gleason score < 6; 2.35 +/- 0.34; P = 0.04). Thus, a decrease of RXRalpha in the basal cells may serve as a marker for prostate CA-associated field change, which may represent an early event in the prostate carcinogenic process. These findings suggest that chemoprevention strategies with retinoids may be most effective if applied during the early stages of transformation.

High-grade prostatic intraepithelial neoplasia.

High-grade prostatic intraepithelial neoplasia (PIN) is now accepted as the most likely preinvasive stage of adenocarcinoma, almost two decades after its first formal description. PIN has a high predictive value as a marker for adenocarcinoma, and its identification warrants repeat biopsy for concurrent or subsequent invasive carcinoma. The only method of detection is biopsy; PIN does not significantly elevate serum prostate-specific antigen (PSA) concentration or its derivatives and cannot be detected by current imaging techniques, including ultrasound. Most patients with PIN will develop carcinoma within 10 years. PIN is associated with progressive abnormalities of phenotype and genotype, which are similar to cancer rather than normal prostatic epithelium, indicating impairment of cell differentiation with advancing stages of prostatic carcinogenesis. Androgen deprivation therapy decreases the prevalence and extent of PIN, suggesting that this form of treatment may play a role in chemoprevention.

Myc-driven murine prostate cancer shares molecular features with human prostate tumors.

Increased Myc gene copy number is observed in human prostate cancer. To define Myc's functional role, we generated transgenic mice expressing human c-Myc in the mouse prostate. All mice developed murine prostatic intraepithelial neoplasia followed by invasive adenocarcinoma. Microarray-based expression profiling identified a Myc prostate cancer expression signature, which included the putative human tumor suppressor NXK3.1. Human prostate tumor databases revealed modules of human genes that varied in concert with the Myc prostate cancer signature. This module includes the Pim-1 kinase, a gene known to cooperate with Myc in tumorigenesis, and defines a subset of human, "Myc-like" human cancers. This approach illustrates how genomic technologies can be applied to mouse cancer models to guide evaluation of human tumor databases.

Expression study of three secretory proteins (prostatic secretory protein of 94 amino acids, probasin, and seminal vesicle secretion II) in dysplastic and neoplastic rat prostates.

BACKGROUND: Prostatic secretory protein of 94 amino acids (PSP94), probasin, and seminal vesicle secretion II (SVSII) are the three major proteins secreted by the lateral lobe of the rat prostate gland. Among these proteins, rodent PSP94 but not probasin and SVSII has a human homologue and it is also a major secretory protein of the human prostate, in addition to prostatic acid phosphatase and prostate-specific antigen. METHODS: In this study, we examined and compared the mRNA expression of these three secretory markers in three rat models of prostate cancer including the sex steroid-induced dysplasia (prostatic intraepithelial neoplasia or PIN) in Noble (Nb) rat model, an androgen-independent Nb rat prostatic tumor (AIT) and Dunning rat prostatic adenocarcinomas (both androgen-dependent and -independent) by in situ hybridization (ISH), reverse transcriptase-polymerase chain reaction (RT-PCR), and immunohistochemistry. RESULTS: The transcripts for the three markers were highly expressed in the secretory epithelium of normal lateral prostate (LP). Their hybridization signals became reduced in the epithelial cells in the low-grade PINs and significantly weakened or lost in the high-grade PINs induced in the LP. Interestingly, we observed that some dysplastic cells located at the basal compartment of the PIN lesions, and nests of outpouching epithelial cells in the vicinity of PINs, expressed positive hybridization signals of three markers. In the adenocarcinoma, signals of probasin but not PSP94 and SVSII were detected. No hybridization signals were detected in both Dunning and AIT tumors. By RT-PCR, transcripts for these proteins were still detected but significantly reduced in the Dunning tumors, whereas in the AIT tumor, only SVSII transcripts were detected. Immunohistochemistry of PSP94 also showed a reduced staining in the PIN lesions, but no immunoreactivity was seen in the rat prostatic tumors. CONCLUSIONS: The mRNA expression of the three prostatic secretory markers were decreased in the hormone-induced PINs and in two rat prostatic tumors, indicating that the androgen-regulated secretory differentiation was impaired during the development of the premalignant lesion and further reduced in advanced tumors. The abnormal expression pattern of these secretory markers and androgen receptor (AR) in the basal compartment of the PIN lesions suggests that there is a population of cell types with secretory phenotype appearing in the basal cell layer during the early malignant transformation of the prostatic epithelium.

[Prostate gland-transition zone lesions. Etiology, growth regulation, growth factors, genetic changes]. / Prostata--promjene prelazne zone. Etiologija, regulacija rasta, cimbenici rasta, geneticke promjene.

Human prostate gland is a composite organ made up of several glandular and nonglandular components. There are three distinct glandular regions. These are the peripheral, the central, and the transition zone. They differ histologically and biologically. The central zone is relatively resistant to carcinoma and other diseases. The peripheral zone is the site of origin of most carcinomas and prostatic intraepithelial neoplasia (PIN). Most authors now agree that PIN is a likely precursor of at least a proportion of carcinomas arising in this zone. The transition zone is the exclusive site of origin of benign prostatic hyperplasia (BPH). A subset of prostate cancers arises in this zone as well. Most are found incidentally at transuretral resection (TUR). A morphological lesion, atypical adenomatous hyperplasia (AH), arising also in transition zone, provides a possible link between BPH and transition zone cancers. However, further studies of biological markers of neoplastic transformation are needed to address this issue.

The role of cyclooxygenase-2 in prostate cancer.

Cyclooxygenase-2 (COX-2) is the inducible isozyme of COX, a key enzyme in the conversion of arachidonic acid to prostaglandins and other eicosanoids. COX-2 is highly expressed in a number of human cancers and cancer cell lines, including prostate cancer. We studied the immunohistochemical expression of COX-2 in the human prostate gland. The enzyme is strongly expressed in smooth muscle cells of both the normal and cancerous prostate. Its expression in noncancerous epithelial cells is limited to the basal cell layer. In prostatic inflammation, luminal epithelial cells surrounded by lymphocytes are induced to express the enzyme. COX-2 is expressed in the epithelial cells of high-grade prostatic intraepithelial neoplasia and cancer. We have demonstrated that treatment of human prostate-cancer cell lines with a selective COX-2 inhibitor induces apoptosis both in vitro and in vivo. The in vivo results also indicate that the COX-2 inhibitor decreases tumor microvessel density and angiogenesis. COX-2 inhibitors can prevent the hypoxic upregulation of a potent angiogenic factor, vascular endothelial growth factor. These results indicate that COX-2 inhibitors may, therefore, serve as effective chemopreventive and therapeutic agents in cancer of the prostate.

Antioxidant enzyme expression and reactive oxygen species damage in prostatic intraepithelial neoplasia and cancer.

BACKGROUND: Oxidative stress results in damage to cellular structures and has been linked to many diseases, including cancer. The authors sought to determine whether the expression of three major antioxidant enzymes, copper-zinc superoxide dismutase (SOD1), manganese superoxide dismutase (SOD2), and catalase, was altered in human prostate carcinoma and its likely precursor, high grade prostatic intraepithelial neoplasia (PIN). The level of reactive oxygen species damage was evaluated by measuring the expression of the DNA adduct 8-hydroxydeoxyguanosine. METHODS: The authors evaluated the tissue expression of the antioxidant enzymes in prostate carcinoma by immunohistochemistry, immunogold electron microscopy, and enzymatic assay. The polymerase chain reaction was used to amplify and screen tissue specimens for the genes of SOD1, SOD2, and extracellular SOD (SOD3). Matched paraffin embedded tissue sections were evaluated by RNA in situ hybridization for expression of SOD1 and immunohistochemically for the DNA adduct 8-hydroxydeoxyguanosine. RESULTS: All prostatic tissues, including cancer, displayed immunoreactivity for the three antioxidant enzymes in epithelial cells, with no staining of the stroma, inflammatory cells, or endothelial cells. The number of immunoreactive cells was greater in benign epithelium than in PIN and cancer for each enzyme. The mean percentage and intensity of immunoreactive cells was greatest for SOD2, intermediate for SOD1, and lower for catalase. Staining in cancer was heterogeneous. Immunogold ultrasound studies revealed strong mitochondrial labeling for SOD2, which was greater in benign epithelium than in cancer; SOD1 labeling was invariably weaker, with nuclear labeling in benign epithelium and cytoplasmic labeling in cancer cells. There was no difference in enzyme activity for the three antioxidant enzymes between benign epithelium and cancer. No mutations were found in the 5 exons of SOD1, 5 exons of SOD2, and 3 exons of SOD3, except for 3 of 20 cases with polymorphisms for exon 3 of SOD1. Intense nuclear immunoreactivity for 8-hydroxydeoxyguanosine was present in fewer than 3% of epithelial cells, with no apparent differences among benign epithelium, PIN, and cancer. CONCLUSIONS: SOD1, SOD2, and catalase had lower expression in PIN and prostate carcinoma than in benign epithelium. The number of immunoreactive cells in PIN was similar to cancer, indicating that these are closely related. Enzyme activities were variable, with no difference between benign epithelial cells and cancer, although this lack of change in enzyme activity could have been due to the presence of contaminating benign cells within the cancer specimens. The results of reactive oxygen species damage were found only in the epithelium and not in the stroma. Expression of the DNA adduct 8-hydroxydeoxyguanosine was present in fewer than 3% of cells, with no apparent differences among benign epithelium, PIN, and cancer. These findings suggest that oxidative stress is an early event in carcinogenesis.

Expression of Bcl-2, Bax, and p53 in high-grade prostatic intraepithelial neoplasia and localized prostate cancer: relationship with apoptosis and proliferation.

BACKGROUND: Apoptosis-regulating genes have been shown to be important in the biology of prostate cancer. The aim of this study was to examine and correlate the expression of the apoptosis-regulating genes bcl-2, bax, and p53 with the frequency of apoptosis and rate of proliferation in benign prostatic epithelium (BP), prostate cancer, and high-grade prostatic intraepithelial neoplasia (HGPIN), which is currently considered the most likely precursor of prostate cancer. METHODS: Forty-four patients with histologically proven prostate cancer were investigated. All the men underwent radical prostatectomy. Immunohistochemistry was performed to assess expression of bcl-2, bax, and p53, and proliferation rate, as measured by the Ki-67 index. The frequency of apoptotic bodies was assessed by morphological criteria. RESULTS: The apoptotic index (AI) was highest in prostate cancer, and was significantly greater in HGPIN compared to benign prostate. The Ki-67 index was greatest in cancer, intermediate in HGPIN, and lowest in BP. The AI was increased in areas of BP in patients treated with neoadjuvant androgen ablation. No change in AI was seen in treated cases of HGPIN or cancer. Accumulation of p53 protein was infrequent in prostate cancer (2/43: 4.6%), and was absent in HGPIN. Bcl-2 overexpression was present in 2.3% of cancers (1/43) and in 34.9% of cases of HGPIN (15/43). Bax expression was seen in all cases of cancer and HGPIN. There was no correlation between bcl-2 expression and the apoptotic and Ki-67 indices in HGPIN. CONCLUSIONS: p53 and bcl-2 expression is infrequent in clinically organ confined prostate cancer. Bcl-2 expression is significantly higher in HGPIN than in both the associated prostate cancer and BP. The AI and Ki-67 index appeared intermediate in the putative precursor lesion HGPIN compared to prostate cancer and BP.