Dutasteride, the dual 5alpha-reductase inhibitor, inhibits androgen action and promotes cell death in the LNCaP prostate cancer cell line

BACKGROUND: Reduction of T to DHT by 5alphaR in the prostate enhances androgenic activity for most targets. Inhibition of 5alphaR activity with finasteride attenuates androgen action in men and animal models. The objective of this study was to compare and contrast the effects of a potent new 5alphaR inhibitor, dutasteride, with finasteride in the LNCaP prostate cancer cell line. METHODS: LNCaP cells were incubated for varying times with T or DHT in steroid-free medium in the absence or presence of increasing doses of dutasteride or finasteride and the effects on 5alphaR activity, PSA accumulation in the medium, and on cell proliferation were determined. Drug effects on apoptosis were investigated using Annexin V staining and a cell death ELISA assay. Effects of the drugs on AR ligand-binding activity and on AR protein levels were determined. RESULTS: Dutasteride inhibited (3)H-T conversion to (3)H-DHT and, as anticipated, inhibited T-induced secretion of PSA and proliferation. However the drug also inhibited DHT-induced PSA secretion and cell proliferation (IC(50) approximately 1 microM). Finasteride also inhibited DHT action but was less potent than dutasteride. Dutasteride competed for binding the LNCaP cell AR with an IC(50) approximately 1.5 microM. High concentrations of dutasteride (10-50 microM), but not finasteride, in steroid-free medium, resulted in enhanced cell death, possibly by apoptosis. This was accompanied by loss of AR protein and decreased AR ligand-binding activity. Occupation of AR by R1881 partly protected against cell death and loss of AR protein. PC-3 prostate cancer cells, which do not contain AR, also were killed by high concentrations of dutasteride, as well as by 50 microM finasteride. CONCLUSIONS: Dutasteride exhibited some inhibitory actions in LNCaP cells possibly related to 5alphaR inhibition but also had antiandrogenic effects at relatively low concentrations and cell death-promoting effects at higher concentrations. Finasteride also was antiandrogenic, but less than dutasteride. The antiandrogenic effects may be mediated by the mutant LNCaP cell AR. Promotion of cell death by dutasteride can be blocked, but only in part, by androgens.     

Isolation and characterization of transforming growth factor beta response variants from human prostatic tumor cell lines.

In this study we examined the relation between the response to transforming growth factor beta (TGF beta 1) in vitro and the growth in vivo of 1-LN-PC3-1A (1-LN) human prostatic carcinoma cells. 1-LN cells resistant to the growth-inhibitory effects of TGF beta 1 were isolated after exposure to 2 ng/ml TGF beta 1 in an anchorage-independent growth assay. Cloning of TGF beta 1-resistant and -sensitive populations produced 2 clones (R2-6 and 1-LN clone 4), which maintained relatively stable resistance or sensitivity, respectively, in the absence of TGF beta 1 for up to 12 passages. Colony formation by the R2-6 cells in the presence of TGF beta 1 was 2-10 times greater than that of 1-LN clone 4, depending upon the TGF beta 1 concentration. Injection of 1 x 10(5) R2-6 cells into athymic nude mice produced tumors with a significantly shorter latency interval as compared with 1-LN clone 4 tumors (P < 0.0001). Western immunoblotting showed that higher levels of latent TGF beta 1 protein were secreted into the culture medium by 1-LN clone 4 cells. Acidified conditioned media from both clones inhibited mink lung epithelial cell DNA synthesis. Neutralizing monoclonal antibody to TGF beta 1 but not TGF beta 2 abrogated this inhibitory effect. Comparison of the different sensitive and resistant clones showed that in vitro sensitivity to TGF beta 1 and in vivo tumor latency interval were not invariably correlated. Thus, the TGF beta 1 response phenotype in vitro was not always predictive of growth delay in vivo.     

Differential expression of 17beta-hydroxysteroid dehydrogenase isozyme genes in prostate cancer and noncancer tissues

BACKGROUND: The adrenal steroids dehydroepiandrosterone and androstenediones are converted into active androgen testosterone in prostatic tissues. Different 17beta-hydroxysteroid dehydrogenase (17betaHSD) isozymes are characterized by either oxidation or reduction reactions. These redox reactions represent an important step in both biosynthesis and metabolism of androgens. This study presents the differential expression of 17betaHSD isozyme genes in cancerous and noncancerous prostate tissues of in vivo samples. METHODS: Thirty-four fresh specimens of transrectal prostatic needle biopsy were obtained; 11 were pathologically diagnosed as adenocarcinoma and 23 as without malignancy. The gene expression levels of five isozymes (type 1-5) of 17betaHSD were evaluated. The quantification of gene expression was assessed by means of the real-time polymerase chain reaction. RESULTS: The expression levels of the type 3 17betaHSD gene with malignancy were significantly higher than those in prostatic tissues without malignancy, and those of type 2 17betaHSD with malignancy were significantly lower than those in nonmalignant tissues. There were no significant differences in 17betaHSD type 1, type 4, and type 5 gene expression in cancerous and noncancerous tissues. CONCLUSION: Our results suggest that 17betaHSD type 2 and type 3 play an important role in the conversion of adrenal steroids into potential androgens in prostate cancer tissue.     

Small interference RNA targeting heat-shock protein 27 inhibits the growth of prostatic cell lines and induces apoptosis via caspase-3 activation in vitro

OBJECTIVES: To evaluate synthetic small interference RNA (siRNA) compounds targeting heat-shock protein 27 (Hsp27) as an alternative approach to Hsp27 'knockdown' in prostate cancer cells, as Hsp27 expression is highly up-regulated in prostate cancer cells after androgen withdrawal or chemotherapy, to become uniformly highly expressed in androgen-independent (AI) prostate cancer. MATERIALS AND METHODS: We recently showed that targeting Hsp27 by a 2'-methoxyethyl modified phosphorothioate antisense oligonucleotide, OGX-427, inhibits Hsp27 expression and enhances hormone- and chemotherapy in prostate cancer xenograft models. In the present study, a 'gene walk' screening different siRNAs was initially used in PC-3 and LNCaP cells to determine the most potent sequence to down-regulate Hsp27 mRNA and protein levels. The effects of Hsp27 silencing on in vitro growth rates were studied by tetrazolium-blue and crystal violet assays. Apoptosis was determined by single-stranded DNA nuclear and cleaved caspase-3 immunostaining, as well as flow cytometry. Spotted microarrays with 14,000 human oligonucleotides were used to examine changes in gene expression. RESULTS: Low concentrations of 1 nm siRNA decreased Hsp27 mRNA levels by 19-fold and suppressed protein expression to undetectable levels. Silencing of Hsp27 in prostate cancer cells by siRNA # 2 increased apoptotic rates 2.4-4 fold and caused 40-76% inhibition of cell growth in LNCaP and PC-3 cells. Characteristic cleavage of caspase-3 occurred after treatment with Hsp27 siRNA (1 nm). cDNA microarray analysis from LNCaP and PC-3 cell lines revealed differential gene expression profiles after Hsp27 down-regulation that could be used to identify various survival pathways involved in androgen-dependent and AI growth. CONCLUSIONS: These findings illustrate the potential utility of Hsp27-silencing therapy and highlight Hsp27 siRNA strategies as a novel and highly effective tool, with the potential for future targeted therapy in enhancing the efficacy of chemotherapy in advanced prostate cancer.     

Raloxifene, an oestrogen-receptor-beta-targeted therapy, inhibits androgen-independent prostate cancer growth: results from preclinical studies and a pilot phase II clinical trial

OBJECTIVES: To determine, in preclinical in vivo animal and in clinical studies, whether raloxifene (a selective oestrogen-receptor (ER) modulator that targets ER-beta and induces apoptosis in vitro in androgen-independent prostate cancer, AIPC cells) affects prostate cell differentiation, proliferation and carcinogenesis, and in the pilot phase II clinical trial, the response rate and duration of patients with AIPC treated with a daily oral dose of raloxifene. PATIENTS, MATERIALS AND METHODS: Tumour proliferation rate in response to raloxifene treatment, and molecular markers of cell cycle and apoptosis, were evaluated in established ER-beta-positive androgen-dependent (AD) CWR22 and AI CWRSA9 human xenograft prostate cancer models. Twenty-one patients with AIPC and evidence of disease progression were enrolled into the clinical trial and given daily oral raloxifene. RESULTS: There was significant growth inhibition by raloxifene in the ADPC and AIPC xenograft models (CWR22 68%, P < 0.010; CWRSA9 64%, P < 0.001), with no tumour regression. There was evidence of G1 arrest by increased p27kip1 expression in the raloxifene-treated group. Eighteen patients comprised the efficacy analysis, as three withdrew before the first evaluation. At the first evaluation, five men had stable disease and continued on the study for a median of five cycles. The longest response was 17 cycles. Drug related toxicity was minimal. CONCLUSION: Raloxifene has activity in xenograft models, slowing disease progression. This translated to possible disease stabilization in patients with AIPC. Further studies are warranted.     

The effect of 5 alpha-androstane-3 alpha, 17 beta-diol and 17 beta-estradiol on the adult and immature chacma baboon prostate

We have studied the Chacma baboon prostate in an attempt to develop a primate model of benign prostatic hyperplasia. Anatomically, the baboon prostate can be divided into caudal, cranial, and prostatic urethral segments. A peri-urethral group of glands has been identified in the prostatic urethra. Following treatment with either 5 alpha-androstane-3 alpha, 17 beta-diol or 17 beta-estradiol, or the hormones in combination, the mature baboon prostate showed very little response by gravimetric or morphometric analysis. In contrast, the stimulatory effect of 5 alpha-androstane-3 alpha, 17 beta-diol on the caudal lobe of the immature baboon prostate and the prostatic urethral segment (a structure that is largely fibromuscular) was potentiated by the addition of 17 beta-estradiol. The weight of the immature baboon cranial prostate was increased by 5 alpha-androstane-3 alpha, 17 beta-diol treatment, but 17 beta estradiol did not potentiate androgen-induced growth. By morphometric analysis it could be shown that both the epithelial and stromal component of the immature baboon caudal prostate responded to 5 alpha-androstane-3 alpha, 17 beta-diol (androstanediol) treatment and that the addition of 17 beta-estradiol had a slight, but significant, potentiating effect on the androstanediol-induced increase of epithelial volume.     

Mechanism of estrogens-induced increases in intracellular Ca(2+) in PC3 human prostate cancer cells

BACKGROUND: The effect of estrogens (diethylstilbestrol [DES], 17 beta-estradiol) on intracellular Ca(2+) concentrations ([Ca(2+)](i)) in hormone-insensitive PC3 human prostate cancer cells was examined. METHODS: [Ca(2+)](i) changes in suspended cells were measured by using the Ca(2+)-sensitive fluorescent dye fura-2. RESULTS: Estrogens (1--20 microM) increased [Ca(2+)](i) concentration-dependently with DES being more potent. Ca(2+) removal inhibited 50 +/- 10% of the signal. In Ca(2+)-free medium, pretreatment with 20 microM estrogens abolished the [Ca(2+)](i) increases induced by 2 microM carbonylcyanide m-chlorophenylhydrazone (CCCP, a mitochondrial uncoupler) and 1 microM thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor), but pretreatment with CCCP and thapsigargin did not alter DES-induced Ca(2+) release and partly inhibited 17 beta-estradiol-induced Ca(2+) release. Addition of 3 mM Ca(2+) increased [Ca(2+)](i) in cells pretreated with 1- 20 microM estrogens in Ca(2+)-free medium. Pretreatment with 1 microM U73122 to block phospholipase C-coupled inositol 1,4,5-trisphosphate formation did not alter estrogens-induced Ca(2+) release. The effect of 20 microM estrogen on [Ca(2+)](i) was not affected by pretreatment with 0.1 microM estrogens. CONCLUSIONS: Estrogen induced significant Ca(2+) release and Ca(2+) influx in an inositol 1,4,5-trisphosphate-independent manner in PC3 cells. These effects of estrogens on Ca(2+) signaling appear to be nongenomic. Prostate 47:141-148, 2001.     

Production, clearance, and metabolism of testosterone in men with prostatic cancer

It was previously unknown whether the production and metabolism of testosterone was altered in men with prostatic cancer. We recently observed a familial influence on the plasma concentration of sex steroids in men with the cancer. We have now determined, by isotope dilution techniques, the blood testosterone production and clearance rates and testosterone metabolism to potent androgen metabolites in men with prostatic cancer, their brothers, and unrelated controls. Nineteen men had a diagnosis of prostatic cancer before age 63 (probands), 23 were brothers of these index cases, and nine controls matched for age were selected randomly from the general population. None had received endocrine therapy. The plasma content of testosterone, dihydrotestosterone, sex hormone binding globulin, apparent free testosterone concentration, follicle-stimulating hormone, and luteinizing hormone were not significantly different between the groups. The metabolic clearance rate of testosterone was significantly (P = .04) higher in probands (458 liters/day/body surface area, median) than in controls (306 liters/day/body surface area). The conversion ratios of both testosterone (1.8%) and dihydrotestosterone (16.9%) to 3 alpha-androstanediol were significantly greater (P = .04 and P = .004, respectively) in probands than in controls (0.95%, 7.8%). These results indicate that men with prostatic cancer have elevated clearance rates of testosterone and an increased conversion ratio of testosterone to its potent 5 alpha-reduced metabolites.     

Association of the G289S single nucleotide polymorphism in the HSD17B3 gene with prostate cancer in Italian men

BACKGROUND: Prostate cancer is a significant public health problem in this country. Substantial data support a plausible role for androgens in the etiology of this disease. The human HSD17B3 gene encodes the testicular (or type III) 17 beta-hydroxysteroid dehydrogenase enzyme, which catalyzes testosterone biosynthesis in men. METHODS: We have investigated the G289S (glycine at codon 289 replaced by serine) polymorphism at the HSD17B3 locus as a candidate single nucleotide polymorphism (SNP) for prostate cancer risk in constitutional DNA from 103 Italian prostate cancer patients and 109 Italian disease-free centenarians to assess the role of this SNP in susceptibility to prostate cancer. RESULTS: The G289S polymorphism confers a significant increase in risk for prostate cancer (odds ratio = 2.5; 95% confidence interval, 1.03-6.07) in our study population. CONCLUSION: Our data are consistent with a plausible role of the G289S SNP in prostate cancer susceptibility. Therefore, the HSD17B3 gene may be a plausible candidate gene for prostate cancer risk.     

5alpha-dihydrotestosterone inhibits 1alpha,25-dihydroxyvitamin D3-induced expression of CYP24 in human prostate cancer cells

BACKGROUND: A cross-talk between 1alpha,25-dihydroxyvitamin D(3) [1alpha,25-(OH)(2)D(3)] and 5alpha-dihydrotestosterone (DHT) in the growth inhibition has been demonstrated, but the mechanism is unknown. METHODS: The expression of 25-hydroxyvitamin D(3) 24-hydroxylase (24-hydroxylase) was measured using a real-time quantitative RT-PCR assay and the catabolism of 1alpha,25-(OH)(2)D(3) was measured using a radioreceptor assay. RESULTS: Real-time RT-PCR showed that DHT at 1-100 nM significantly inhibited 1alpha,25-(OH)(2)D(3)-induced expression of 24-hydroxylase in LNCaP cells. Furthermore, the catabolism of 1alpha,25-(OH)(2)D(3) was decreased by 10 nM DHT. An androgen receptor (AR) antagonist, Casodex antagonized the DHT effect, whereas an AR agonist (due to the mutant AR in LNCaP cells) hydroxyflutamide did not. CONCLUSIONS: We demonstrated, for the first time, that DHT reduces the ability of 1alpha,25-(OH)(2)D(3) to induce 24-hydroxylase expression. Our results not only support the earlier finding of a cross-talk between androgen and vitamin D in human prostate cancer cells but also provide a possible mechanism how androgen and vitamin D signaling pathways may interact.     

Effect of permixon on human prostate cell growth: lack of apoptotic action

BACKGROUND: Permixon, a phytotherapeutic agent derived from the saw palmetto or Serenoa repens plant, is a lipid/sterol extract that is believed to interfere with 5alpha-reductase activity, thus inhibiting prostate growth. In this study, we investigated the magnitude and specificity of the effect of Permixon on cell proliferation and apoptosis in human prostate cancer cells. METHODS: The effect of Permixon was examined in androgen-independent PC-3 prostate cancer cells, androgen-sensitive LNCaP prostate cancer cells, and MCF-7 breast cancer cells in vitro. Cell growth, apoptosis induction, and cell proliferation was studied after exposure to Permixon at two concentrations (10 and 100 microg/ml). Cell proliferation and cell cycle progression were determined after 24 hr on the basis of (3)[H]-thymidine incorporation assay and flowcytometric analysis, respectively. Apoptosis induction was evaluated in treated and untreated cultures using the Hoescht staining and caspase-3 activation. RESULTS: Exposure of prostate and breast cancer cells to a high dose of Permixon (100 microg/ml) resulted in a significant decrease in the rate of cell growth; an effect that was not time-dependent and was not associated with cell cycle arrest. Permixon treatment (at either high or low dose) had no effect on apoptosis induction in prostate cancer cell lines (P > 0.6). Furthermore, in vitro Permixon was a weak inhibitor of 5alpha-reductase activity type 2 in prostatic homogenates. CONCLUSIONS: The results indicate the ability of Permixon to affect prostate cancer cell growth without inducing apoptosis or cell cycle arrest. This effect was not prostate-specific and was only manifested at high concentrations of Permixon. Furthermore our findings indicate that Permixon is weak inhibitor of 5alpha-reductase compared to finasteride. This study challenges previous evidence on the anti-growth effect of Permixon in the prostate and its ability to inhibit 5alpha-reductase activity, while questioning apoptosis as a mechanism of action of this phytotherapeutic against prostate growth, a concept that may have therapeutic significance.     

Plasma levels of transforming growth factor-1beta and alpha2-macroglobulin before and after radical prostatectomy: association to clinicopathological parameters

BACKGROUND: To study the levels of transforming growth factor-1beta (TGF-beta1) and of alpha2-macroglobulin (alpha2-M), a high affinity binding protein of TGF-beta1, in comparison to prostate-specific antigen (PSA) in prostate cancer (PCa) patients before and up to 12 months after prostatectomy, and to correlate the results with clinicopathological parameters. METHODS: Eighty-one patients who underwent radical prostatectomy for PCa were included in this study. Pre- and postoperatively, plasma levels of TGF-beta1, alpha2-M and PSA were measured in the same samples by ELISA, and were correlated with pathological parameters and clinical outcomes. RESULTS: The preoperative TGF-beta1 levels were significantly elevated as compared to the controls; they showed a positive correlation with the Gleason score. Patients with initial androgen-deprivation therapy had lower TGF-beta1 levels than untreated patients. Elevated concentrations of TGF-beta1 levelled off 12 months after prostatectomy approaching values of healthy individuals. Decreased plasma levels of total and transformed alpha2-M (proteinase-complexed form) were observed in PCa. Preoperative levels of TGF-beta1 but not of alpha2-M seem to be influenced by the body mass index (BMI). CONCLUSIONS: Elevated TGF-beta1 and decreased alpha2-M were consistently found in patients with PCa, and may be considered as risk factors for tumor development and progression. In comparison to PSA, the TGF-beta1 levels displayed a slow decline after radical prostatectomy; this indicates that TGF-beta1 is mainly produced outside the prostatic tissue. Since TGF-beta1 levels are influenced by the BMI, this indicates that PCa might be sensitive to diet.     

Bcl-2 antagonizes the combined apoptotic effect of transforming growth factor-beta and dihydrotestosterone in prostate cancer cells

BACKGROUND: We previously demonstrated that dihydrotestosterone (DHT) enhances transforming growth factor-beta (TGF-beta) -induced apoptosis in human prostate cancer cells (Endocrinology 2001;142:2419-2426). METHODS: In this study, the ability of the apoptosis suppressor bcl-2 to directly antagonize the combined apoptotic effect of TGF-beta and DHT in the androgen-sensitive LNCaP TbetaRII prostate cancer cells was examined. The previously cloned TGF-RbetaII receptor LNCaP cells, responsive to both TGF-beta and androgens, were engineered to overexpress the bcl-2 oncoprotein and the profile of apoptosis induction was analyzed in response to TGF-beta alone (5.0 ng/ml) or in combination with DHT (1 nM). RESULTS: Biological characterization of cloned LNCaP TbetaRII hygromycin/bcl-2 transfectants demonstrated that bcl-2 overexpression resulted in a significant inhibition of the combined TGF-beta and DHT apoptotic effect in prostate cancer cells (P < 0.01). Furthermore, molecular analysis indicated that this antagonistic effect of bcl-2 on apoptosis was due to partial suppression of TGF-beta and DHT-mediated induction of caspase-1 expression and activation in LNCaP TbetaRII-hygro/bcl-2 transfectants. These results support a potential bcl-2 interference with the TGF-beta and androgen apoptotic signaling in prostate cancer cells by means of an antagonistic effect on caspase-1 activation. CONCLUSION: This evidence may have mechanistic significance in understanding the contribution of bcl-2 overexpression in the development of androgen-independent prostate cancer by means of conferring resistance to TGF-beta-mediated apoptosis.     

Lithium suppresses cell proliferation by interrupting E2F-DNA interaction and subsequently reducing S-phase gene expression in prostate cancer

BACKGROUND: Lithium is an existing drug for bipolar disorder and its uptake was recently linked to reduced tumor incidence compared to the general population. The major target of lithium action is glycogen synthase kinase 3 (GSK-3). Since GSK-3 expression and activation are associated with prostate cancer progression, the anti-cancer potential of lithium on prostate cancer was investigated in this study. METHODS: Multiple prostate cancer cell lines were treated with lithium chloride (LiCl). Cell proliferation and cell cycle distribution were analysed. DNA replication was determined using BrdU labeling assay. Genome-wide screening of gene expression was performed using cDNA microarray assay. GSK-3beta gene-specific silencing was conducted using small interferencing RNA (siRNA) transfection. E2 factor (E2F) transactivation was evaluated using reporter gene assay and E2F-DNA interaction was determined with chromatin-immunoprecipitation assay (ChIP). RESULTS: LiCl significantly inhibited cell proliferation, which was associated with reduced DNA replication and S-phase cell cycle arrest. LiCl significantly decreased the expression of multiple DNA replication-related genes, including cell division cycle 6 (cdc6), cyclin A, cyclin E, and cdc25C, which are regulated by E2F factor during cell cycle. A novel GSK-3-specific inhibitor TDZD-8 and GSK-3beta siRNA also suppressed the expression of these E2F target genes, indicating that LiCl-induced anti-cancer effect was associated with GSK-3beta inhibition. Furthermore, LiCl suppressed E2F transactivation by interrupting the interaction of E2F1 factor with its target gene promoter. CONCLUSIONS: These data indicated that LiCl suppresses cancer cell proliferation by disrupting E2F-DNA interaction and subsequent E2F-mediated gene expression in prostate cancer.     

Effect of a 4-methyl-4-aza steroid on androgen metabolism by rat ventral prostate epithelial and stromal cell cultures: selective inhibition of 5 alpha-reductase activity

The effect of a potent steroid metabolic inhibitor, 17 beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5 alpha-androstan-3-one (DMAA), on androgen metabolism was investigated in primary monolayer cultures of rat ventral prostate epithelial and stromal cells. Using testosterone (T) as substrate, 5 alpha-reductase (5 alpha-R) activity in both cell types was inhibited by greater than 98% at an inhibitor concentration of 1000 nM. The concentrations required to produce a 50% inhibition (IC50) were 7.4 and 9.0 nM for epithelial and stromal cells, respectively. To examine the specificity of this compound, its effect on other steroid-metabolic enzymes was examined. DMAA at a concentration of 1,000 nM had no effect on 3 alpha-hydroxysteroid oxidase (3 alpha-HSORox), 3-ketosteroid reductase (3 alpha-HSORred), and 6/7-hydroxylase (6/7-HSH) activities in both cell types; 17 beta-hydroxysteroid oxidase (17 beta-HSORox) activity, located primarily in epithelial cells, also was not influenced by DMAA. In contrast, epithelial 3 beta-hydroxysteroid oxidase (3 beta-HSORox) and 3-ketosteroid reductase (3 beta-HSORred) activities were inhibited by 65% (P less than .001) and 58% (P greater than .05), respectively, albeit the latter result was not statistically significant. Stromal 3 beta-HSORox and 3 beta-HSORred activities were negligible; hence the effect of the inhibitor of these enzymes could not be assessed. In conclusion, DMAA is a relatively selective and potent inhibitor of 5 alpha-R activity in primary cultures of rat ventral prostate epithelial and stromal cells and should be a useful compound for antagonizing androgen-mediated actions in the prostate and other androgen target tissues.     

Osteopontin enhances the cell proliferation induced by the epidermal growth factor in human prostate cancer cells

BACKGROUND: Susceptibility to extracellular matrix and growth factors has been demonstrated to play a critical role in the development of prostate cancer (PCa) metastases. The aim of this study was to elucidate some mechanisms by which stroma controls tumor progression. METHODS: In our study we tested the growth ability of the LNCaP human prostatic cell line in steroid-free culture conditions in response to osteopontin (OPN), a non-collageneous matrix protein, localized in large amounts in the bone. RESULTS: In the LNCaP cell model, OPN stimulates cell proliferation in serum-free medium and colony growth at high dilution but this effect is visible only in presence of epidermal growth factor (EGF). Proliferation induced by OPN is accompanied by a sustained activation of EGF receptor (EGFR) whose phosphorylation is detectable up to 12 hr after treatment in association with EGF. The colocalization of integrin beta1, a ligand of OPN, and of EGFR on the cellular membrane, suggests that the association of these cell surface receptors may be the principal mechanism involved in the long-term activation of the EGFR. CONCLUSIONS: Our data describe a new possible mechanism involved in the establishment of bone metastases which may also account for the formation of androgen-independent cellular clones, frequently responsible of the clinical progression of PCa.