Biomedical subjects

Donna M Peehl

Publications and source records attributed to Donna M Peehl.

At least 19 recordsLinked to original sources

PubMed2007

Novel pathways that contribute to the anti-proliferative and chemopreventive activities of calcitriol in prostate cancer.

Calcitriol, the hormonally active form of Vitamin D, inhibits the growth and development of many cancers through multiple mechanisms. Our recent research supports the contributory role of several new and diverse pathways that add to the mechanisms already established as playing a role in the actions of calcitriol to inhibit the development and progression of prostate cancer (PCa). Calcitriol increases the expression of insulin-like growth factor binding protein-3 (IGFBP-3), which plays a critical role in the inhibition of PCa cell growth by increasing the expression of the cell cycle inhibitor p21. Calcitriol inhibits the prostaglandin (PG) pathway by three actions: (i) the inhibition of the expression of cyclooxygenase-2 (COX-2), the enzyme that synthesizes PGs, (ii) the induction of the expression of 15-prostaglandin dehydrogenase (15-PGDH), the enzyme that inactivates PGs and (iii) decreasing the expression of EP and FP PG receptors that are essential for PG signaling. Since PGs have been shown to promote carcinogenesis and progression of multiple cancers, the inhibition of the PG pathway may add to the ability of calcitriol to prevent and inhibit PCa development and growth. The combination of calcitriol and non-steroidal anti-inflammatory drugs (NSAIDs) result in a synergistic inhibition of PCa cell growth and offers a potential therapeutic strategy. Mitogen activated protein kinase phosphatase 5 (MKP5) is a member of a family of phosphatases that are negative regulators of MAP kinases. Calcitriol induces MKP5 expression in prostate cells leading to the selective dephosphorylation and inactivation of the stress-activated kinase p38. Since p38 activation is pro-carcinogenic and is a mediator of inflammation, this calcitriol action, especially coupled with the inhibition of the PG pathway, contributes to the chemopreventive activity of calcitriol in PCa. Mullerian Inhibiting Substance (MIS) has been evaluated for its inhibitory effects in cancers of the reproductive tissues and is in development as an anti-cancer drug. Calcitriol induces MIS expression in prostate cells revealing yet another mechanism contributing to the anti-cancer activity of calcitriol in PCa. Thus, we conclude that calcitriol regulates myriad pathways that contribute to the potential chemopreventive and therapeutic utility of calcitriol in PCa.

Animals↗

PubMed2007

Distinctive gene expression of prostatic stromal cells cultured from diseased versus normal tissues.

To obtain a comprehensive view of the transcriptional programs in prostatic stromal cells of different histological/pathological origin, we profiled 18 adult human stromal cell cultures from normal transition zone (TZ), normal peripheral zone (PZ), benign prostatic hyperplasia (BPH), and prostate cancer (CA) using cDNA microarrays. A hierarchical clustering analysis of 714 named unique genes whose expression varied at least threefold from the overall mean abundance in at least three samples in all 18 samples demonstrated that cells of different origin displayed distinct gene expression profiles. Many of the differentially expressed genes are involved in biological processes known to be important in the development of prostatic diseases including cell proliferation and apoptosis, cell adhesion, and immune response. Significance Analysis of Microarrays (SAM) analysis identified genes that showed differential expression with statistical significance including 24 genes between cells from TZ versus BPH, 34 between BPH versus CA, and 101 between PZ versus CA. S100A4 and SULF1, the most up- and downregulated genes in BPH versus TZ, respectively, showed expression at the protein level consistent with microarray analysis. In addition, sulfatase assay showed that BPH cells have lower SULF1 activity compared to TZ cells. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis confirmed differential expression of ENPP2/autotoxin and six other genes between PZ versus CA, as well as differential expression of six genes between BPH versus CA. Our results support the hypothesis that prostatic stromal cells of different origin have unique transcriptional programs and point towards genes involved in actions of stromal cells in BPH and CA.

Adult↗

PubMed2007

Calcitriol and genistein actions to inhibit the prostaglandin pathway: potential combination therapy to treat prostate cancer.

We present an overview of the prostaglandin (PG) pathway as a novel target for the treatment of prostate cancer (PCa) using a combination of calcitriol and genistein, both of which have known antiproliferative properties. Calcitriol inhibits the PG pathway in PCa cells in 3 separate ways: by decreasing cyclooxygenase-2 (COX-2) expression, stimulating 15-hydroxyprostaglandin dehydrogenase (15-PGDH) expression, and decreasing EP (PGE2) and FP (PGF(2alpha)) receptors. These actions of calcitriol result in reduced levels of biologically active PGE2, leading ultimately to growth inhibition of the PCa cells. We also demonstrate the advantages of using calcitriol in combination with genistein for the treatment of PCa. Genistein, a major component of soy, is a potent inhibitor of the activity of CYP24, the enzyme that initiates the degradation of calcitriol. This leads to increased half-life of bioactive calcitriol, thereby enhancing all of calcitriol's actions including those on the PG pathway. In addition to inhibiting CYP24 enzyme activity, genistein has its own independent actions on the PG pathway in PCa cells. Like calcitriol it inhibits COX-2 expression and activity, leading to decreased synthesis of PGE2. It also inhibits the EP and FP receptors, thereby reducing the biological function of PGE2. Thus, the combination of calcitriol and genistein acts additively to inhibit the PG pathway. Both calcitriol and genistein are relatively safe and have little toxicity associated with their intake. We postulate that the combination of calcitriol and genistein is an attractive therapeutic option for the treatment of PCa.

Animals↗

PubMed2006

Chemopreventive anti-inflammatory activities of curcumin and other phytochemicals mediated by MAP kinase phosphatase-5 in prostate cells.

As inflammation emerges as a risk factor for prostate cancer (PCa), there is potential for chemoprevention by anti-inflammatory agents. Dietary phytochemicals have been shown to have chemopreventive properties which may include anti-inflammatory activities. In this study, we demonstrate a role for mitogen-activated protein kinase phosphatase-5 (MKP5) in mediating anti-inflammatory activities of the phytochemicals curcumin, resveratrol and [6]-gingerol. We utilized the cytokines tumor necrosis factor-alpha (TNFalpha) and interleukin (IL)-1beta to increase p38-dependent nuclear factor kappa-B (NFkappaB) activation and expression of pro-inflammatory genes cyclooxygenase-2 (COX-2), IL-6 and IL-8 in normal prostatic epithelial cells. MKP5 over-expression decreased cytokine-induced NFkappaB activation, COX-2, IL-6 and IL-8 in normal prostatic epithelial cells, suggesting potent anti-inflammatory activity of MKP5. Pretreatment of cells with a p38 inhibitor mimicked the results observed with MKP5 over-expression, further implicating p38 inhibition as the main activity of MKP5. Curcumin, the phytochemical found in turmeric, up-regulated MKP5, subsequently decreasing cytokine-induced p38-dependent pro-inflammatory changes in normal prostatic epithelial cells. Resveratrol and [6]-gingerol, phytochemicals present in red wine and ginger, respectively, also up-regulated MKP5 in normal prostate epithelial cells. Moreover, we found that PCa cell lines DU 145, PC-3, LNCaP and LAPC-4 retained the ability to up-regulate MKP5 following curcumin, resveratrol and [6]-gingerol exposure, suggesting utility of these phytochemicals in PCa treatment. In summary, our findings show direct anti-inflammatory activity of MKP5 in prostate cells and suggest that up-regulation of MKP5 by phytochemicals may contribute to their chemopreventive actions by decreasing prostatic inflammation.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2006

Transcript profiling of the androgen signal in normal prostate, benign prostatic hyperplasia, and prostate cancer.

Human prostate adenocarcinoma (CaP) and benign prostatic hyperplasia (BPH) have epithelial and stromal cell origins, respectively. To determine whether the androgen signal is processed differently in these cell types the expression of transcripts for enzymes that control ligand access to the androgen receptor (AR) were measured. Transcripts for type 2 5alpha-reductase, ketosteroid reductases [aldo-keto reductase (AKR)1C1-AKR1C4], the major oxidative 3alpha-hydroxysteroid dehydrogenase (HSD) retinol dehydrogenase (RODH)-like 3alpha-HSD (RL-HSD) and nuclear receptors [AR, estrogen receptor (ER)alpha, and ERbeta] were determined in whole human prostate and in cultures of primary epithelial cells (PEC) and primary stromal cells (PSC) from normal prostate, CaP and BPH by real-time RT-PCR. Normal PEC (n=14) had higher levels of AKR1C1 (10-fold, P<0.001), AKR1C2 (115-fold, P<0.001) and AKR1C3 (6-fold, P<0.001) than normal PSC (n=15), suggesting that reductive androgen metabolism occurs. By contrast, normal PSC had higher levels of AR (8-fold, P<0.001) and RL-HSD (21-fold, P<0.001) than normal PEC, suggesting that 3alpha-androstanediol is converted to 5alpha-dihydrotestosterone to activate AR. In CaP PEC (n=14), no significant changes in transcript levels vs. normal PEC were observed. In BPH PSC (n=21) transcripts for AR (2-fold, P<0.001), AKR1C1 (4-fold, P<0.001), AKR1C2 (10-fold P<0.001), AKR1C3 (4-fold, P<0.001) and RL-HSD (3-fold, P<0.003) were elevated to increase androgen response. Differences in the AR:ERbeta transcript ratios (eight in normal PEC vs. 280 in normal PSC) were maintained in PEC and PSC in diseased prostate. These data suggest that CaP may be more responsive to an ERbeta agonist and BPH may be more responsive to androgen ablation.

20-Hydroxysteroid Dehydrogenases↗

PubMed2006

Inhibition of p38 by vitamin D reduces interleukin-6 production in normal prostate cells via mitogen-activated protein kinase phosphatase 5: implications for prostate cancer prevention by vitamin D.

Although numerous studies have implicated vitamin D in preventing prostate cancer, the underlying mechanism(s) remains unclear. Using normal human prostatic epithelial cells, we examined the role of mitogen-activated protein kinase phosphatase 5 (MKP5) in mediating cancer preventive activities of vitamin D. Up-regulation of MKP5 mRNA by 1,25-dihydroxyvitamin-D3 (1,25D) was dependent on the vitamin D receptor. We also identified a putative positive vitamin D response element within the MKP5 promoter that associated with the vitamin D receptor following 1,25D treatment. MKP5 dephosphorylates/inactivates the stress-activated protein kinase p38. Treatment of prostate cells with 1,25D inhibited p38 phosphorylation, and MKP5 small interfering RNA blocked this effect. Activation of p38 and downstream production of interleukin 6 (IL-6) are proinflammatory. Inflammation and IL-6 overexpression have been implicated in the initiation and progression of prostate cancer. 1,25D pretreatment inhibited both UV- and tumor necrosis factor alpha-stimulated IL-6 production in normal cells via p38 inhibition. Consistent with inhibition of p38, 1,25D decreased UV-stimulated IL-6 mRNA stabilization. The ability of 1,25D to up-regulate MKP5 was maintained in primary prostatic adenocarcinoma cells but was absent in metastases-derived prostate cancer cell lines. The inability of 1,25D to regulate MKP5 in the metastasis-derived cancer cells suggests there may be selective pressure to eliminate key tumor suppressor functions of vitamin D during cancer progression. These studies reveal MKP5 as a mediator of p38 inactivation and decreased IL-6 expression by 1,25D in primary prostatic cultures of normal and adenocarcinoma cells, implicating decreased prostatic inflammation as a potential mechanism for prostate cancer prevention by 1,25D.

Calcitriol↗

PubMed2006

Expression of CCL5 (RANTES) and CCR5 in prostate cancer.

BACKGROUND: Expression of the inflammatory chemokine CCL5 (RANTES) by tumor cells is thought to correlate with the progression of several cancers. CCL5 was shown to induce breast cancer cell migration, mediated by the receptor CCR5. A CCR5 antagonist was demonstrated to inhibit experimental breast tumor growth. Recently, CCL5 and CCR5 mRNA expression was reported in prostate cancer (PCa) tissues. Herein, we characterized CCL5 and CCR5 expression in cultures of PCa cells and explored possible functions of CCL5 in PCa progression. METHODS: Quantitative RT-PCR, ELISA, and immunohistochemical staining were performed to examine CCL5 expression in prostate cell lines. CCR5 expression was measured by flow cytometry. Proliferation and invasion assays were performed to determine potential functions of CCL5 and CCR5 in PCa. RESULTS: Expression of CCL5 mRNA and protein was found in human PCa cell lines (PC-3; DU-145; LNCaP) and primary prostate adenocarcinoma cells. CCL5 and CCR5 were also detected in human PCa tissues. CCR5 expression was demonstrated on the cell surface of PCa cells, as well as in intracellular pools. Incubation with CCL5 (10-100 ng/ml) induced PCa cell proliferation, and the CCR5 antagonist TAK-779 inhibited CCL5-induced proliferation. CCL5 was found to stimulate PCa cell invasion, and TAK-779 blocked the effects of CCL5. CONCLUSIONS: In light of evidence that inflammation influences the pathogenesis of PCa, these results suggest that inflammatory chemokines, such as CCL5, expressed by prostate cells may act directly on the growth and survival of PCa cells. Chemokine receptor antagonists may thus block autocrine mechanisms of PCa progression.

Adenocarcinoma↗

PubMed2006

Aldo-keto reductase (AKR) 1C3: role in prostate disease and the development of specific inhibitors.

Human aldo-keto reductases (AKR) of the 1A, 1B, 1C and 1D subfamilies are involved in the pre-receptor regulation of nuclear (steroid hormone and orphan) receptors by regulating the local concentrations of their lipophilic ligands. AKR1C3 is one of the most interesting isoforms. It was cloned from human prostate and the recombinant protein was found to function as a 3-, 17- and 20-ketosteroid reductase with a preference for the conversion of Delta4-androstene-3,17-dione to testosterone implicating this enzyme in the local production of active androgens within the prostate. Using a validated isoform specific real-time RT-PCR procedure the AKR1C3 transcript was shown to be more abundant in primary cultures of epithelial cells than stromal cells, and its expression in stromal cells increased with benign and malignant disease. Using a validated isoform specific monoclonal Ab, AKR1C3 protein expression was also detected in prostate epithelial cells by immunoblot analysis. Immunohistochemical staining of prostate tissue showed that AKR1C3 was expressed in adenocarcinoma and surprisingly high expression was observed in the endothelial cells. These cells are a rich source of prostaglandin G/H synthase 2 (COX-2) and vasoactive prostaglandins (PG) and thus the ability of recombinant AKR1C enzymes to act as PGF synthases was compared. AKR1C3 had the highest catalytic efficiency (kcat/Km) for the 11-ketoreduction of PGD2 to yield 9alpha,11beta-PGF2 raising the prospect that AKR1C3 may govern ligand access to peroxisome proliferator activated receptor (PPARgamma). Activation of PPARgamma is often a pro-apoptotic signal and/or leads to terminal differentiation, while 9alpha,11beta-PGF2 is a pro-proliferative signal. AKR1C3 is potently inhibited by non-steroidal anti-inflammatory drugs suggesting that the cancer chemopreventive properties of these agents may be mediated either by inhibition of AKR1C3 or COX. To discriminate between these effects we developed potent AKR1C inhibitors based on N-phenylanthranilic acids that do not inhibit COX-1 or COX-2. These compounds can now be used to determine the role of AKR1C3 in producing two proliferative signals in the prostate namely testosterone and 9alpha,11beta-PGF2.

3-Hydroxysteroid Dehydrogenases↗

PubMed2005

Comparative gene and protein expression in primary cultures of epithelial cells from benign prostatic hyperplasia and prostate cancer.

Primary cultures are widely used to investigate the disease-specific biology of prostate cancer and benign prostatic hyperplasia (BPH). To identify genes differentially expressed between epithelial cells cultured from adenocarcinomas versus BPH tissues, we used probe array technology. Gene expression profiles were evaluated on Affymetrix Human Cancer G110 Array Chips containing approximately 1900 cancer-related genes. After defined statistical analysis, genes that were over-expressed in cancer cultures were identified. Protein expression of four of the differentially expressed genes was measured in immunoblots, and the expression of two other genes was measured by real-time reverse transcription-polymerase chain reaction (RT-PCR). While no gene or protein was consistently over-expressed in all cancer versus BPH cell cultures, cytokeratin 16 protein was highly elevated in several of the cancer cultures, suggesting that a hyperproliferative phenotype may be characteristic of prostate cancer cells.

Adenocarcinoma↗

PubMed2005

Genistein potentiates the growth inhibitory effects of 1,25-dihydroxyvitamin D3 in DU145 human prostate cancer cells: role of the direct inhibition of CYP24 enzyme activity.

In a search for improved therapies for prostate cancer, we investigated the effect of genistein in combination with 1alpha-25-dihydroxyvitamin D3 [1,25(OH)2D3], on the growth of DU145 human prostate cancer cells. DU145 cells were very resistant to the growth inhibitory action of 1,25(OH)2D3 or genistein when administered individually. However, the combination caused a significant growth inhibition seen at lower concentrations of both agents. 1,25(OH)2D3 induces the expression of the CYP24 gene, which codes for the enzyme that initiates the catabolism of 1,25(OH)2D3. We showed for the first time that genistein at low doses (50-100 nM) directly inhibited CYP24 at the enzyme level. Addition of genistein to mitochondrial preparations inhibited CYP24 enzyme activity in a noncompetitive manner. CYP24 inhibition by genistein increased the half-life of 1,25(OH)2D3 thereby augmenting the homologous up-regulation of the vitamin D receptor (VDR) both at the mRNA and protein levels. Genistein co-treatment enhanced 1,25(OH)2D3-mediated transactivation of the vitamin D responsive reporters OC-Luc and OP-Luc transfected into DU145 cells. Consistent with the growth inhibition due to the combination treatment, significant changes in the expression of genes involved in growth arrest and apoptosis were seen. We conclude that genistein potentiates the antiproliferative actions of 1,25(OH)2D3 in DU145 cells by two mechanisms: (i) an increase in the half-life of 1,25(OH)2D3 due to the direct inhibition of CYP24 enzyme activity and (ii) an amplification of the homologous up-regulation of VDR. Together these two effects lead to a substantial enhancement of the cellular responses to the growth inhibitory and pro-apoptotic signaling by 1,25(OH)2D3.

Anticarcinogenic Agents↗

PubMed2005

Identification of the major oxidative 3alpha-hydroxysteroid dehydrogenase in human prostate that converts 5alpha-androstane-3alpha,17beta-diol to 5alpha-dihydrotestosterone: a potential therapeutic target for androgen-dependent disease.

Androgen-dependent prostate diseases initially require 5alpha-dihydrotestosterone (DHT) for growth. The DHT product 5alpha-androstane-3alpha,17beta-diol (3alpha-diol), is inactive at the androgen receptor (AR), but induces prostate growth, suggesting that an oxidative 3alpha-hydroxysteroid dehydrogenase (HSD) exists. Candidate enzymes that posses 3alpha-HSD activity are type 3 3alpha-HSD (AKR1C2), 11-cis retinol dehydrogenase (RODH 5), L-3-hydroxyacyl coenzyme A dehydrogenase , RODH like 3alpha-HSD (RL-HSD), novel type of human microsomal 3alpha-HSD, and retinol dehydrogenase 4 (RODH 4). In mammalian transfection studies all enzymes except AKR1C2 oxidized 3alpha-diol back to DHT where RODH 5, RODH 4, and RL-HSD were the most efficient. AKR1C2 catalyzed the reduction of DHT to 3alpha-diol, suggesting that its role is to eliminate DHT. Steady-state kinetic parameters indicated that RODH 4 and RL-HSD were high-affinity, low-capacity enzymes whereas RODH 5 was a low-affinity, high-capacity enzyme. AR-dependent reporter gene assays showed that RL-HSD, RODH 5, and RODH 4 shifted the dose-response curve for 3alpha-diol a 100-fold, yielding EC(50) values of 2.5 x 10(-9) M, 1.5 x 10(-9) M, and 1.0 x 10(-9) M, respectively, when compared with the empty vector (EC(50) = 1.9 x 10(-7) M). Real-time RT-PCR indicated that L-3-hydroxyacyl coenzyme A dehydrogenase and RL-HSD were expressed more than 15-fold higher compared with the other candidate oxidative enzymes in human prostate and that RL-HSD and AR were colocalized in primary prostate stromal cells. The data show that the major oxidative 3alpha-HSD in normal human prostate is RL-HSD and may be a new therapeutic target for treating prostate diseases.

3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)↗

PubMed2005

Regulation of prostaglandin metabolism by calcitriol attenuates growth stimulation in prostate cancer cells.

Calcitriol exhibits antiproliferative and pro-differentiation effects in prostate cancer. Our goal is to further define the mechanisms underlying these actions. We studied established human prostate cancer cell lines and primary prostatic epithelial cells and showed that calcitriol regulated the expression of genes involved in the metabolism of prostaglandins (PGs), known stimulators of prostate cell growth. Calcitriol significantly repressed the mRNA and protein expression of prostaglandin endoperoxide synthase/cyclooxygenase-2 (COX-2), the key PG synthesis enzyme. Calcitriol also up-regulated the expression of 15-hydroxyprostaglandin dehydrogenase, the enzyme initiating PG catabolism. This dual action was associated with decreased prostaglandin E2 secretion into the conditioned media of prostate cancer cells exposed to calcitriol. Calcitriol also repressed the mRNA expression of the PG receptors EP2 and FP, providing a potential additional mechanism of suppression of the biological activity of PGs. Calcitriol treatment attenuated PG-mediated functional responses, including the stimulation of prostate cancer cell growth. The combination of calcitriol with nonsteroidal anti-inflammatory drugs (NSAIDs) synergistically acted to achieve significant prostate cancer cell growth inhibition at approximately 2 to 10 times lower concentrations of the drugs than when used alone. In conclusion, the regulation of PG metabolism and biological actions constitutes a novel pathway of calcitriol action that may contribute to its antiproliferative effects in prostate cells. We propose that a combination of calcitriol and nonselective NSAIDs might be a useful chemopreventive and/or therapeutic strategy in men with prostate cancer, as it would allow the use of lower concentrations of both drugs, thereby reducing their toxic side effects.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2005

Molecular targets of doxazosin in human prostatic stromal cells.

BACKGROUND: We used cDNA microarray analysis to obtain insights into the mechanisms of action of doxazosin, an alpha(1)-adrenergic receptor antagonist used to treat benign prostatic hyperplasia (BPH). METHODS: Hierarchical clustering analysis and significance analysis of microarray (SAM) were performed to identify genes differentially expressed between untreated stromal cells cultured from normal tissue and BPH, and changes in gene expression induced by doxazosin. Transcript levels of selected genes were validated by real-time reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: Hierarchical clustering analyses separated untreated normal and BPH cells. Sixty-seven genes whose expression varied at least twofold after doxazosin treatment in both normal and BPH cells were identified, as were 93 genes differentially regulated in normal versus BPH cells. Molecular targets consistent with tumor necrosis factor (TNF)-alpha-related activity were identified. CONCLUSIONS: Normal versus BPH stromal cells differ in global gene transcription. Doxazosin induced gene expression changes relevant to proliferation/apoptosis, immune defense, cell-cell signaling/signal transduction, and transcriptional regulation.

Adrenergic alpha-Antagonists↗

PubMed2005

Redox modulation of human prostate carcinoma cells by selenite increases radiation-induced cell killing.

Although selenium compounds have been extensively studied as chemopreventative agents for prostate cancer, little is known about the potential use of selenium compounds for chemotherapy. We have shown that selenite inhibits cell growth and induces apoptosis in androgen-dependent LAPC-4 prostate cancer cells. LAPC-4 cells were more sensitive to selenite-induced apoptosis than primary cultures of normal prostate cells. Selenite-induced apoptosis in LAPC-4 cells correlated with a decrease in the Bcl-2:Bax expression ratio. Selenite-induced oxidative stress and apoptosis are dependent upon its reaction with reduced GSH. LAPC-4 cells treated with selenite showed decreased levels of total GSH and increased concentrations of GSSG. Thus, selenite altered the intracellular redox status toward an oxidative state by decreasing the ratio of GSH:GSSG. Because increased levels of Bcl-2 and GSH are associated with radioresistance, we examined the ability of selenite to sensitize prostate cancer cells to gamma-irradiation. Both LAPC-4 and androgen-independent DU 145 cells pretreated with selenite showed increased sensitivity to gamma-irradiation as measured by clonogenic survival assays. Importantly, selenite-induced radiosensitization was observed in combination with a clinically relevant dose of 2 Gy. These data suggest that altering the redox environment of prostate cancer cells with selenite increases the apoptotic potential and sensitizes them to radiation-induced cell killing.

Apoptosis↗

PubMed2004

Interaction of nuclear receptor ligands with the Vitamin D signaling pathway in prostate cancer.

A number of hormonal ligands and/or the nuclear receptors that mediate their actions have been targeted for prostate cancer therapy. Androgens, the ligands for the androgen receptor (AR), are critical for the growth of prostate cancer. Inhibition of androgen production has been the mainstay of treatment for advanced prostate cancer for decades. Other more recently tested targets include retinoid receptors (RAR and RXR), glucocorticoid receptors (GR), estrogen receptors (ER) and peroxisome proliferator-activated receptors (PPAR). Calcitriol, acting through the Vitamin D receptor (VDR), has many tumor suppressive activities in the prostate, including inhibition of proliferation, induction of apoptosis and/or differentiation, and reduction of cellular invasion. Because of these properties, calcitriol and its less hypercalcemic analogs are being evaluated as agents to prevent or treat prostate cancer. Androgens, retinoids, glucocorticoids, estrogens and agonists of PPAR directly or indirectly impact Vitamin D signaling pathways, and vice versa. In order to design the most effective strategies to use calcitriol to prevent or treat prostate cancer, the interactions of other nuclear receptors and their ligands with the Vitamin D signaling pathway need to be considered.

Alitretinoin↗

PubMed2004

Expression of transforming growth factor-beta 1 and growth in soft agar differentiate prostate carcinoma-associated fibroblasts from normal prostate fibroblasts.

Carcinoma-associated fibroblasts (CAF) promote tumor progression of pre-neoplastic epithelial cells. To investigate the basis of this phenomenon, we compared the properties of fibroblasts cultured from normal human prostate (NHPF) to prostate CAF. NHPF and CAF were assayed for growth potential, cell death and proliferative capacity by measuring population doubling time, cell cycle distribution and capability to form colonies in soft agar. Resistance to genotoxic (UV radiation: 0-50 J/cm2) and chemotoxic (0-200 nM Taxol) agents were compared between CAF and NHPF by measuring cell viability and cell cycle analysis. Transforming growth factor beta1 (TGF-beta1) immunoreactivity was assessed in non-malignant and malignant prostatic tissue. No detectable differences were found when comparing CAF and NHPF with respect to population doubling time, cell cycle distribution and response to genotoxic and chemotoxic agents. The mean number of colonies in soft agar was 120.5 for CAF vs. 18.2 for NHPF (p < 0.05). Because TGF-beta1 and matrix metalloproteinase (MMP)-9 have been associated with growth of fibroblasts in soft agar and tumor promotion, we measured the expression of these factors in NHPF and CAF by ELISA. There was no difference in expression of MMP-9; however, TGF-beta1 was expressed in higher concentrations in CAF than in NHPF (p < 0.0014). Furthermore, TGF-beta1 expression was higher in the carcinoma-associated stroma of prostate cancer tissue than stroma of non-malignant prostatic tissue. Increased capability of CAF as compared to NHPF to form colonies in soft agar may be due to a higher expression of TGF-beta1 and correlates with the ability of CAF to promote malignant progression of prostate epithelial cells.

Agar↗

PubMed2004

Altered SMRT levels disrupt vitamin D3 receptor signalling in prostate cancer cells.

We hypothesized that key antiproliferative target genes for the vitamin D receptor (VDR) were repressed by an epigenetic mechanism in prostate cancer cells resulting in apparent hormonal insensitivity. To explore this possibility, we examined nuclear receptor corepressor expression in a panel of nonmalignant and malignant cell lines and primary cultures, and found frequently elevated SMRT corepressor mRNA expression often associated with reduced sensitivity to 1alpha,25-dihydroxyvitamin D(3) (1alpha,25(OH)2D3). For example, PC-3 and DU-145 prostate cancer cell lines had 1.8-fold and twofold increases in SMRT mRNA relative to normal PrEC cells (P<0.05). Similarly, 10/15 primary tumour cultures (including three matched to normal cells from the same donors) had elevated SMRT mRNA levels; generally NCoR1 and Alien were not as commonly elevated. Corepressor proteins often have associated histone deacetylases (HDAC) and reflectively the antiproliferative action of 1alpha,25(OH)2D3 can be 'restored' by cotreatment with low doses of HDAC inhibitors such as trichostatin A (TSA, 15 nM) to induce apoptosis in prostate cancer cell lines. To decipher the transcriptional events that lead to these cellular responses, we undertook gene expression studies in PC-3 cells after cotreatment of 1alpha,25(OH)2D3 plus TSA after 6 h. Examination of known VDR target genes and cDNA microarray analyses revealed cotreatment of 1alpha,25(OH)2D3 plus TSA cooperatively upregulated eight (out of 1176) genes, including MAPK-APK2 and GADD45alpha. MRNA and protein time courses and inhibitor studies confirmed these patterns of regulation. Subsequently, we knocked down SMRT levels in PC-3 cells using a small interfering RNA (siRNA) approach and found that GADD45alpha induction by 1alpha,25(OH)2D3 alone became very significantly enhanced. The same distortion of gene responsiveness, with repressed induction of GADD45alpha was found in primary tumour cultures compared and to matched peripheral zone (normal) cultures from the same donor. These data demonstrate that elevated SMRT levels are common in prostate cancer cells, resulting in suppression of target genes associated with antiproliferative action and apparent 1alpha,25(OH)2D3-insensitivity. This can be targeted therapeutically by combination treatments with HDAC inhibitors.

Base Sequence↗

PubMed2004

CXCR4 and CXCL12 (SDF-1) in prostate cancer: inhibitory effects of human single chain Fv antibodies.

PURPOSE: Metastasis is a major cause of morbidity in prostate cancer (PCa). Several studies have shown that the chemokine receptor CXCR4 and its ligand, CXCL12 (stromal cell-derived factor-1), regulate tumor cell metastasis to specific organs. Recently, it was demonstrated that CXCL12 enhances PCa cell adhesion, migration, and invasion, implicating CXCR4 in PCa metastasis. In this study, we examined the inhibitory effects of anti-CXCR4 antibodies on CXCL12-mediated PCa cell activities. EXPERIMENTAL DESIGN: We developed fully human single chain Fv antibodies (scFv), Ab124 and Ab125, against CXCR4 using the yeast two-hybrid system. We performed immunofluorescent staining, flow cytometry, and ELISA-binding assays to measure scFv binding to PCa cells. We also examined the effects of scFv on CXCL12-mediated calcium mobilization, cell migration, and invasion. RESULTS: Our results confirmed that PCa cell lines express cell-surface CXCR4. Real-time quantitative reverse transcription-PCR and immunohistochemical staining also verified that CXCR4 is expressed in primary cultures of prostate epithelial cells from adenocarcinomas and in human prostate tissues. Ab124 and Ab125 demonstrated specific binding to PCa cell lines by flow cytometry and in binding assays. Preincubation with scFv resulted in significant reduction of CXCL12-induced calcium mobilization in PC-3 and LNCaP cells. Ab124 and Ab125 also inhibited PCa cell migration toward CXCL12, as well as invasion through extracellular matrix gels. CONCLUSIONS: Ab124 and Ab125 inhibit CXCL12-mediated cellular activities by binding the receptor CXCR4. Recombinant scFv are an efficient mode of targeting tumor antigens. Considering the high incidence of PCa, the development of fully human scFv may be a useful therapeutic approach in the prevention and treatment of PCa metastasis.

Breast Neoplasms↗