Long noncoding RNA MALAT1 enhances the docetaxel resistance of prostate cancer cells via miR‐145‐5p‐mediated regulation of AKAP12

Our present work was aimed to study on the regulatory role of MALAT1/miR‐145‐5p/AKAP12 axis on docetaxel (DTX) sensitivity of prostate cancer (PCa) cells. The microarray data (GSE33455) to identify differentially expressed lncRNAs and mRNAs in DTX‐resistant PCa cell lines (DU‐145‐DTX and PC‐3‐DTX) was retrieved from the Gene Expression Omnibus (GEO) database. QRT‐PCR analysis was performed to measure MALAT1 expression in DTX‐sensitive and DTX‐resistant tissues/cells. The human DTX‐resistant cell lines DU145‐PTX and PC3‐DTX were established as in vitro cell models, and the expression of MALAT1, miR‐145‐5p and AKAP12 was manipulated in DTX‐sensitive and DTX‐resistant cells. Cell viability was examined using MTT assay and colony formation methods. Cell apoptosis was assessed by TUNEL staining. Cell migration and invasion was determined by scratch test (wound healing) and Transwell assay, respectively. Dual‐luciferase assay was applied to analyse the target relationship between lncRNA MALAT1 and miR‐145‐5p, as well as between miR‐145‐5p and AKAP12. Tumour xenograft study was undertaken to confirm the correlation of MALAT1/miR‐145‐5p/AKAP12 axis and DTX sensitivity of PCa cells in vivo. In this study, we firstly notified that the MALAT1 expression levels were up‐regulated in clinical DTX‐resistant PCa samples. Overexpressed MALAT1 promoted cell proliferation, migration and invasion but decreased cell apoptosis rate of PCa cells in spite of DTX treatment. We identified miR‐145‐5p as a target of MALAT1. MiR‐145‐5p overexpression in PC3‐DTX led to inhibited cell proliferation, migration and invasion as well as reduced chemoresistance to DTX, which was attenuated by MALAT1. Moreover, we determined that AKAP12 was a target of miR‐145‐5p, which significantly induced chemoresistance of PCa cells to DTX. Besides, it was proved that MALAT1 promoted tumour cell proliferation and enhanced DTX‐chemoresistance in vivo. There was an lncRNA MALAT1/miR‐145‐5p/AKAP12 axis involved in DTX resistance of PCa cells and provided a new thought for PCa therapy.     

Transcriptome analysis of differentially expressed genes and pathways associated with mitoxantrone treatment prostate cancer

The global physiological function of specifically expressed genes of mitoxantrone (MTX)‐resistant prostate cancer (PCa) is unclear. In this study, gene expression pattern from microarray data was investigated for identifying differentially expressed genes (DEGs) in MTX‐resistant PCa xenografts. Human PCa cell lines DU145 and PC3 were cultured in vitro and xenografted into severe combined immunodeficiency (SCID) mice, treated with MTX intragastrically, three times a week until all mice relapsed. Gene expression profiles of the xenografts from castrated mice were performed with Affymetrix human whole genomic oligonucleotide microarray. The Cytoscape software was used to investigate the relationship between proteins and the signalling transduction network. A total of 355 overlapping genes were differentially expressed in MTX‐resistant DU145R and PC3R xenografts. Of these, 16 genes were selected to be validated by quantitative real‐time PCR (qRT‐PCR) in these xenografts, and further tested in a set of formalin‐fixed, paraffin‐embedded and optimal cutting temperature (OCT) clinical tumour samples. Functional and pathway enrichment analyses revealed that these DEGs were closely related to cellular activity, androgen synthesis, DNA damage and repair, also involved in the ERK/MAPK, PI3K/serine‐threonine protein kinase, also known as protein kinase B, PKB (AKT) and apoptosis signalling pathways. This exploratory analysis provides information about potential candidate genes and may bring new insights into the molecular cascade involvement in MTX‐resistant PCa.     

Cabazitaxel exhibits more favorable molecular changes compared to other taxanes in androgen‐independent prostate cancer cells

Taxane‐based chemotherapy drugs (cabazitaxel, docetaxel, and paclitaxel) are microtubule inhibitors, which are effectively and frequently used to treat metastatic prostate cancer (PCa). Among these, cabazitaxel is offered as a new therapeutic option for patients with metastatic castration‐resistant PC as that are resistant to other taxanes. Here, we investigated the cellular and molecular changes in response to cabazitaxel in comparison with docetaxel and paclitaxel in androgen‐independent human PCas. The androgen‐independent human PCa cell lines, PC3 and DU145, were treated with 1 to 5nM cabazitaxel, docetaxel, or paclitaxel, and assessed for cell viability (MTT assay), colony forming ability and migration (scratch assay). The induction of apoptosis was determined through measurement of mitochondrial membrane potential (JC‐1 assay) and caspase‐3 activity assay. The protein expression changes (caspase‐3, caspase‐8, Bax, Bcl‐2, β‐tubulin, nuclear factor‐κB [NF‐κB/p50, NF‐κB/p65], vascular endothelial growth factor, WNT1‐inducible signaling pathway protein‐1 [WISP1], transforming growth factor β [TGF‐β]) in response to drug treatment were screened via western blotting. Under our experimental conditions, all taxanes significantly reduced WISP1 and TGF‐β expressions, suggesting an anti‐metastatic/antiangiogenic effect for these drugs. On the other hand, cabazitaxel induced more cell death and inhibited colony formation compared to docetaxel or paclitaxel. The highest fold change in caspase‐3 activity and Bax/Bcl‐2 ratio was also detected in response to cabazitaxel. Furthermore, the induction of β‐tubulin expression was lower in cabazitaxel‐treated cells relative to the other taxanes. In summary, cabazitaxel shows molecular changes in favor of killing PCa cells compared to other taxanes, at least for the parameters analyzed herein. The differences with other taxanes may be important while designing other studies or in clinical settings.     

Resistance to paclitaxel increases the sensitivity to other microenvironmental stresses in prostate cancer cells

The microenvironment is central to many aspects of cancer pathobiology and has been proposed to play a role in the development of cancer cell resistance to therapy. To examine the response to microenvironmental conditions, two paclitaxel resistant prostate cancer (PCa) cell lines (stable and reversible) and one reversible heat resistant cell line were studied. In comparison to their parental cell lines, both paclitaxel resistant cell lines (stable and reversible) were more sensitive to microenvironmental heat, potentially yielding a synergistic therapeutic opportunity. In the two phenotypic cells repopulated after acute heat or paclitaxel treatments, there was an inverse correlation between paclitaxel and heat resistance: resistance to paclitaxel imparted sensitivity to heat; resistance to heat imparted sensitivity to paclitaxel. These studies indicate that as cancer cells evolve resistance to single microenvironmental stress they may be more sensitive to others, perhaps allowing us to design new approaches for PCa therapy.     

Combination effect of PectaSol and Doxorubicin on viability, cell cycle arrest and apoptosis in DU-145 and LNCaP prostate cancer cell lines

The effect of PectaSol on Dox (Doxorubicin) cytotoxicity in terms of apoptosis and cell cycle changes in PCa (prostate cancer) cell lines (DU‐145 and LNCaP) has been investigated. Combination of PectaSol and Dox resulted in a viability of 29.4 and 32.6% (P<0.001) in DU‐145 and LNCaP cells. The IC₅₀ values decreased 1.5‐fold and 1.3‐fold in the DU‐145 and LNCaP cells respectively. In the DU‐145 cells, combination of PectaSol and Dox resulted in a reduction in p27 gene and protein expression (P<0.001). In LNCaP cells, this combination increased p53, p27 and Bcl‐2 expression. Treatment with both drugs in DU‐145 cells led to an increase in sub‐G₁ arrest (54.6% compared with 12.2% in Dox). In LNCaP cells, combination of the drugs led to an increased in G₂/M arrest (61.7% compared with 53.6% in Dox). Based on these findings, progressive cytotoxicity effect of Dox and PectaSol together rapidly induce cell death in DU‐145 through apoptosis and in LNCaP cells through cell cycle arrest (G₂/M arrest).     

Comparative proteome analysis identified CD44 as a possible serum marker for docetaxel resistance in castration‐resistant prostate cancer

Baseline or acquired resistance to docetaxel (DOC) represents a significant risk for patients with metastatic prostate cancer (PC). In the last years, novel therapy regimens have been approved providing reasonable alternatives for DOC‐resistant patients making prediction of DOC resistance of great clinical importance. We aimed to identify serum biomarkers, which are able to select patients who will not benefit from DOC treatment. DOC‐resistant PC3‐DR and DU145‐DR sublines and their sensitive parental cell lines (DU145, PC3) were comparatively analyzed using liquid chromatography‐coupled tandem mass spectrometry (LC‐MS/MS). Results were filtered using bioinformatics approaches to identify promising serum biomarkers. Serum levels of five proteins were determined in serum samples of 66 DOC‐treated metastatic castration‐resistant PC patients (mCRPC) using ELISA. Results were correlated with clinicopathological and survival data. CD44 was subjected to further functional cell culture analyses. We found at least 177 two‐fold significantly overexpressed proteins in DOC‐resistant cell lines. Our bioinformatics method suggested 11/177 proteins to be secreted into the serum. We determined serum levels of five (CD44, MET, GSN, IL13RA2 and LNPEP) proteins in serum samples of DOC‐treated patients and found high CD44 serum levels to be independently associated with poor overall survival (p = 0.001). In accordance, silencing of CD44 in DU145‐DR cells resulted in re‐sensitization to DOC. In conclusion, high serum CD44 levels may help identify DOC‐resistant patients and may thereby help optimize clinical decision‐making regarding type and timing of therapy for mCRPC patients. In addition, our in vitro results imply the possible functional involvement of CD44 in DOC resistance.     

Apolipoprotein C1 promotes prostate cancer cell proliferation in vitro

Here, we aimed to investigate the carcinogenic effects of apolipoprotein C1 (APOC1) in prostate cancer (PCa). APOC1 expression was evaluated in PCa and normal prostate specimens, and lentivirus‐mediated RNA interference was used to knockdown APOC1 in DU145 cells. The effects of APOC1 silencing on cell proliferation, cell cycle arrest, and apoptosis were assessed. APOC1 expression was much higher in PCa tissues than in normal tissues. Moreover, APOC1 silencing inhibited cell proliferation and colony formation, arrested cell cycle progression, and enhanced apoptosis in DU145 cells. Additionally, APOC1 silencing decreased survivin, phospho‐Rb, and p21 levels and increased cleaved caspase‐3 expression. These data supported the procarcinogenic effects of APOC1 in the pathogenesis of PCa and suggested that targeting APOC1 may have applications in the treatment of PCa.     

Activity of different anthracycline formulations in hormone-refractory prostate cancer cell lines: role of Golgi apparatus

The efficacy of therapy for hormone‐refractory prostate cancer (HRPC) is still unsatisfactory and new agents and therapeutic modalities are needed. The aims of the present work were to examine the in vitro activity and mechanisms of action of doxorubicin (DX), pegylated liposomal DX (PLDX), and non‐pegylated liposomal DX (NPLDX) in DU145 and taxane‐resistant DU145‐R HRPC cell lines. Drug activity and incorporation, apoptosis, and expression of cell death‐related markers were evaluated by SRB test, cytofluorimetric assays, and Western blot, respectively. Among the different DX formulations, NPLDX showed the highest cytotoxic activity in both cell lines, with more than 50% of apoptotic cells at only 1/10 of the plasma peak concentration after 72 h exposure. Anthracyclines, in particular NPLDX, were highly concentrated in the Golgi apparatus. Moreover, a significant increase was observed in the expression of CD95 receptor, GD3 ganglioside and, caspase‐2 and ‐8 active forms in both cell lines followed by caspase‐3 activation and mitochondrial membrane depolarization. The Golgi apparatus, probably acting as a stress sensor, intensified the conventional apoptotic mechanism induced by anthracyclines. Our data support the hypothesis that organelle‐dependent initiation of cell death other than that induced by mitochondria and nucleus is a research area worthy of pursuing and suggest that the Golgi apparatus could be an ideal target for anti‐cancer therapy. Of note, the activity of NLPDX in taxane‐resistant DU145‐R cells warrants further evaluation as second‐line treatment of advanced HRPC after taxane failure. J. Cell. Physiol. 226: 3035–3042, 2011. © 2011 Wiley‐Liss, Inc.     

Analysis of the expression and association of retinoblastoma binding protein 6 with the JNK signaling pathway in prostate cancers

This study aims to investigate the expression of retinoblastoma binding protein 6 (RBBP6) in prostate cancer (PCa) and its association with the c‐Jun N‐terminal kinase (JNK) pathway. Immunohistochemistry was used to detect RBBP6 and JNK1/2 expression in PCa and benign prostatic hyperplasia tissues. RBBP6 expression in PCa cells (LNCap, PC3, and DU145) and noncancerous prostate epithelial cells (RWPE‐1) was determined by quantitative real‐time polymerase chain reaction and western blot analysis. PC3 and DU145 cells were transfected with RBBP6 small interfering RNAs (siRNAs) to examine the biological characteristics. Anisomycin (a JNK activator) with/without RBBP6 siRNA was used to treat PC3 cells for further investigating the ramification of the RBBP6‐mediated JNK pathway in PCa. PCa tissues and cells showed higher RBBP6 and JNK1/2 expression. RBBP6 was positively correlated with JNK1/2 in PCa tissues. Besides, RBBP6 expression was correlated to clinical tumor stage, lymph node metastasis, Gleason grade, preoperative prostate‐specific antigen level, as well as prognosis of PCa. RBBP6 siRNA reduced cell proliferation, arrested cells at G2/M, and promoted cell apoptosis, and suppressed JNK pathway. In addition, migration and invasion decreased after the RBBP6 siRNA transfection with downregulated matrix metallopeptidase‐2 (MMP‐2) and MMP‐9. Anisomycin promoted the proliferation, invasion, and migration of PC3 cells and inhibited PC3 cell apoptosis, which could be reversed by RBBP6 siRNA. RBBP6 expression was upregulated in PCa tissues and positively correlated with expression level of JNK1/2. With inhibition of RBBP6 expression, the proliferation, invasion, and migration of PCa cells decreased dramatically, while PC3 cell apoptosis increased appreciably, accompanied by the suppression of the JNK pathway.     

Genistein Inhibits Prostate Cancer Cell Growth by Targeting miR-34a and Oncogenic HOTAIR

Objective

Genistein is a soy isoflavone that has antitumor activity both in vitro and in vivo. It has been shown that genistein inhibits many type of cancers including prostate cancer (PCa) by regulating several cell signaling pathways and microRNAs (miRNAs). Recent studies suggest that the long non-coding RNAs (lncRNAs) are also involved in many cellular processes. At present there are no reports about the relationship between gensitein, miRNAs and lncRNAs. In this study, we focused on miRNAs, lncRNA that are regulated by genistein and investigated their functional role in PCa.

Method

Microarray (SurePrint G3 Human GE 8×60K) was used for expression profiling of genistein treated and control PCa cells (PC3 and DU145). Functional assay (cell proliferation, migration, invasion, apoptosis and cell cycle assays) were performed with the PCa cell lines, PC3 and DU145. Both in vitro and in vivo (nude mouse) models were used for growth assays. Luciferase reporter assays were used for binding of miR-34a to HOTAIR.

Results

LncRNA profiling showed that HOTAIR was highly regulated by genistein and its expression was higher in castration-resistant PCa cell lines than in normal prostate cells. Knockdown (siRNA) of HOTAIR decreased PCa cell proliferation, migration and invasion and induced apoptosis and cell cycle arrest. miR-34a was also up-regulated by genistein and may directly target HOTAIR in both PC3 and DU145 PCa cells.

Conclusions

Our results indicated that genistein inhibited PCa cell growth through down-regulation of oncogenic HOTAIR that is also targeted by tumor suppressor miR-34a. These findings enhance understanding of how genistein regulates lncRNA HOTAIR and miR-34a in PCa.     

Phenylbutyrate-induced apoptosis and differential expression of Bcl-2, Bax, p53 and Fas in human prostate cancer cell lines

OBJECTIVE: To assess the mechanisms of action of phenylbutyrate (PB), an investigational chemotherapeutic agent for prostate cancer (PCa), in apoptosis induction in PCa cell lines in vitro. STUDY DESIGN: We analyzed the differential expression of different apoptosis modulators, Bcl-2, Bax, p53 and Fas, for their potential role in PB-induced apoptosis using relative quantitative flow cytometry (FCM). Both androgen-dependent (LNCaP) and androgen-independent (C-4-2, PC-3-PF and DU145) human PCa cell lines were examined. RESULTS: PB induced apoptosis in PCa cell lines in a dose-dependent manner. Fifty percent apoptosis could be induced by 5-10 mM PB. Bcl-2 was down-regulated 30-75% and the Bax:Bcl-2 ratio elevated in apoptotic PCa cell lines regardless of their androgen dependency or p53 status. FCM revealed a heterogeneous stimulatory effect on the expression of Bax and Bcl-2 in PC3-PF cells at 0.5-2.5 mM PB. In a p53-positive cell line (DU145), p53 was repressed by 70% and Fas elevated sixfold with 10 mM PB. CONCLUSION: Our data show that PB-induced PCa apoptosis is associated with the relative repression of Bcl-2 and with up-regulation of Bax and Fas proteins and that this PB-induced apoptosis is independent of p53 and androgen-dependency status of PCa cell lines.     

Dual roles of E-cadherin in prostate cancer invasion

The role(s) of E-cadherin in tumor progression, invasion, and metastasis remains somewhat enigmatic. In order to investigate various aspects of E-cadherin biological activity, particularly in prostate cancer progression, our laboratory cloned unique subpopulations of the heterogeneous DU145 human prostatic carcinoma cell line and characterized their distinct biological functions. The data revealed that the highly invasive, fibroblastic-like subpopulation of DU145 cells (designated DU145-F) expressed less than 0.1-fold of E-cadherin protein when compared to the parental DU145 or the poorly invasive DU145 cells (designated DU145-E). Experimental disruption of E-cadherin function stimulated migration and invasion of DU145-E and other E-cadherin-positive prostate cancer cell lines, but did not affect the fibroblastic-like DU145-F subpopulation. Within the medium of parental DU145 cells, the presence of an 80 kDa E-cadherin fragment was detected. Subsequent functional analyses revealed the stimulatory effect of this fragment on the migratory and invasive capability of E-cadherin-positive cells. These results suggest that E-cadherin plays an important role in regulating the invasive potential of prostate cancer cells through an unique paracrine mechanism.     

Protein kinase Cdelta signaling downstream of the EGF receptor mediates migration and invasiveness of prostate cancer cells

Tumor progression to the invasive phenotype occurs secondary to upregulated signaling from growth factor receptors that drive key cellular responses like proliferation, migration, and invasion. We hypothesized that Protein kinase Cdelta (PKCdelta)-mediated transcellular contractility is required for migration and invasion of prostate tumor cells. Two invasive human prostate cancer cell lines, DU145 cells overexpressing wildtype human EGFR (DU145WT) and PC3 cells, were studied. PKCdelta is overexpressed in these cells relative to normal prostate epithelial cells, and is activated downstream of EGFR leading to cell motility via modulation of myosin light chain activity. Abrogation of PKCdelta using Rottlerin and specific siRNA significantly decreased migration and invasion of both cell lines in vitro. Both PKCdelta and phosphorylated PKCdelta protein levels were higher in human prostate cancer tissue relative to normal donor prostate as assessed by Western blotting and immunohistochemistry. Thus, we conclude that PKCdelta inhibition can limit migration and invasion of prostate cancer cells.     

The Mitogen-activated Protein (MAP) Kinases p38 and Extracellular Signal-regulated Kinase (ERK) Are Involved in Hepatocyte-mediated Phenotypic Switching in Prostate Cancer Cells

The greatest challenge for the seeding of cancer in metastatic sites is integration into the ectopic microenvironment despite the lack of an orthotopic supportive environment and presence of pro-death signals concomitant with a localized “foreign-body” inflammatory response. In this metastatic location, many carcinoma cells display a reversion of the epithelial-to-mesenchymal transition that marks dissemination in the primary tumor mass. This mesenchymal to epithelial reverting transition (MErT) is thought to help seeding and colonization by protecting against cell death. We have previously shown that hepatocyte coculture induces the re-expression of E-cadherin via abrogation of autocrine EGFR signaling pathway in prostate cancer (PCa) cells and that this confers a survival advantage. Herein, we show that hepatocytes educate PCa to undergo MErT by modulating the activity of p38 and ERK1/2. Hepatocytes inhibited p38 and ERK1/2 activity in prostate cancer cells, which allowed E-cadherin re-expression. Introduction of constitutively active MEK6 and MEK1 to DU145 cells cocultured with hepatocytes abrogated E-cadherin re-expression. At least a partial phenotypic reversion can be achieved by suppression of p38 and ERK1/2 activation in DU145 cells even in the absence of hepatocytes. Interestingly, these mitogen-activated protein kinase activities were also triggered by re-expressed E-cadherin leading to p38 and ERK1/2 activity in PCa cells; these signals provide protection to PCa cells upon challenge with chemotherapy and cell death-inducing cytokines. We propose that distinct p38/ERK pathways are related to E-cadherin levels and function downstream of E-cadherin allowing, respectively, for hepatocyte-mediated MErT and tumor cell survival in the face of death signals.     

Orphan nuclear receptor RORγ confers doxorubicin resistance in prostate cancer

Prostate cancer (PCa) is a malignant tumor with an extremely high prevalence. Doxorubicin is the first‐line clinical treatment for castration‐resistant PCa. Clinically, relapse is almost inevitable due to the cancer cells' increasing resistance to doxorubicin. Our previous studies have revealed that retinoic acid‐related orphan nuclear receptor γ (RORγ) is a key protein for cancer progression and a promising target for PCa therapy. Though, RORγ's role and mechanism in doxorubicin‐resistant PCa remain unclear. To study the mechanism of doxorubicin resistance, we generated a doxorubicin‐resistant PCa cell line C4‐2B (C4‐2B DoxR) in this study, by culturing cells in an increasing doxorubicin concentration. Here, we show that RORγ expression was upregulated in C4‐2B DoxR cells compared with that in normal C4‐2B cells. The RORγ‐stably‐overexpressing PCa cell line constructed by lentiviral transfection showed an obvious improvement in doxorubicin resistance and a trend toward castration resistance. Furthermore, RORγ‐specific small molecule inhibitors XY018, GSK805, and SR2211 can significantly inhibit the proliferation of C4‐2B DoxR cells and promote their apoptosis. Collectively, these results have demonstrated the correlation between the upregulation of RORγ and the development of PCa's doxorubicin resistance, thus providing new ideas for solving the problem of chemotherapy drug resistance in PCa.     

The E1 copper binding domain of full-length amyloid precursor protein mitigates copper-induced growth inhibition in brain metastatic prostate cancer DU145 cells

Copper plays an important role in the aetiology and growth of tumours and levels of the metal are increased in the serum and tumour tissue of patients affected by a range of cancers including prostate cancer (PCa). The molecular mechanisms that enable cancer cells to proliferate in the presence of elevated copper levels are, therefore, of key importance in our understanding of tumour growth progression. In the current study, we have examined the role played by the amyloid precursor protein (APP) in mitigating copper-induced growth inhibition of the PCa cell line, DU145. A range of APP molecular constructs were stably over-expressed in DU145 cells and their effects on cell proliferation in the presence of copper were monitored. Our results show that endogenous APP expression was induced by sub-toxic copper concentrations in DU145 cells and over-expression of the wild-type protein was able to mitigate copper-induced growth inhibition via a mechanism involving the cytosolic and E1 copper binding domains of the full-length protein. APP likely represents one of a range of copper binding proteins that PCa cells employ in order to ensure efficient proliferation despite elevated concentrations of the metal within the tumour microenvironment. Targeting the expression of such proteins may contribute to therapeutic strategies for the treatment of cancers.     

Expression profile of WNT molecules in prostate cancer and its regulation by aminobisphosphonates

Skeletal metastases represent a frequent complication in patients with advanced prostate cancer (PCa) and often require bisphosphonate treatment to limit skeletal‐related events. Metastasized PCa cells disturb bone remodeling. Since the WNT signaling pathway regulates bone remodeling and has been implicated in tumor progression and osteomimicry, we analyzed the WNT profile of primary PCa tissues and PCa cell lines and assessed its regulation by bisphosphonates. Prostate tissue (n = 18) was obtained from patients with benign prostate hyperplasia (BPH) and PCa patients with different disease stages. Serum samples were collected from 62 patients. Skeletal metastases were present in 17 patients of whom 6 had been treated with zoledronic acid. The WNT profile and its regulation by bisphoshonates were analyzed in tissue RNA extracts and serum samples as well as in osteotropic (PC3) and non‐osteotropic (DU145, LNCaP) PCa cell lines. Several members of the WNT pathway, including WNT5A, FZD5, and DKK1 were highly up‐regulated in PCa tissue from patients with advanced PCa. Interestingly, osteotropic cells showed a distinct WNT profile compared to non‐osteotropic cells. While WNT5A, FZD5, and DKK1 were highly expressed in PC3 cells, WNT1 and SFRP1 mRNA levels were higher in DU145 cells. Moreover, zoledronic acid down‐regulated mRNA levels of WNT5A (?34%), FZD5 (?60%), and DKK1 (?46%) in PC3 cells. Interestingly, patients with skeletal metastases who received zoledronic acid had twofold higher DKK1 serum levels compared to bisphosphonate‐naive patients. The WNT signaling pathway is up‐regulated in advanced PCa, differentially expressed in osteotropic versus non‐osteotropic cells, and is regulated by zoledronic acid. J. Cell. Biochem. 112: 1593–1600, 2011. © 2011 Wiley‐Liss, Inc.