Androgen-independent PC-3 Human Prostate Cancer Research Articles

Autophagy inhibition improves the chemotherapeutic efficacy of cruciferous vegetable-derived diindolymethane in a murine prostate cancer xenograft model.

Prostate cancer is the second leading cause of cancer-related deaths in men in North America and there is an urgent need for development of more effective therapeutic treatments against this disease. We have recently shown that diindolylmethane (DIM) and several of its halogenated derivatives (ring-DIMs) induce death and protective autophagy in human prostate cancer cells. However, the in vivo efficacy of ring-DIMs and the use of autophagy inhibitors as adjuvant therapy have not yet been studied in vivo. The objective of this study was to determine these effects on tumor growth in nude CD-1 mice bearing bioluminescent androgen-independent PC-3 human prostate cancer cells. We found that chloroquine (CQ) significantly sensitized PC-3 cells to death in the presence of sub-toxic concentrations of DIM or 4,4'-Br2DIM in vitro. Moreover, a combination of DIM (10mg/kg) and CQ (60mg/kg), 3× per week, significantly decreased PC-3 tumor growth in vivo after 3 and 4weeks of treatment. Furthermore, 4,4'-Br2DIM at 10mg/kg (3× per week) significantly inhibited tumour growth after 4weeks of treatment. Tissues microarray analysis showed that DIM alone or combined with CQ induced apoptosis marker TUNEL; the combination also significantly inhibited the cell proliferation marker Ki67. In conclusion, we have confirmed that DIM and 4,4'-Br2DIM are effective agents against prostate cancer in vivo and shown that inhibition of autophagy with CQ enhances the anticancer efficacy of DIM. Our results suggest that including selective autophagy inhibitors as adjuvants may improve the efficacy of existing and novel drug therapies against prostate cancer.

Lithocholic acid induces endoplasmic reticulum stress, autophagy and mitochondrial dysfunction in human prostate cancer cells.

Lithocholic acid (LCA) is a secondary bile acid that is selectively toxic to human neuroblastoma, breast and prostate cancer cells, whilst sparing normal cells. We previously reported that LCA inhibited cell viability and proliferation and induced apoptosis and necrosis of androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cells. In the present study, we investigated the roles of endoplasmic reticulum (ER) stress, autophagy and mitochondrial dysfunction in the toxicity of LCA in PC-3 and autophagy deficient, androgen-independent DU-145 cells. LCA induced ER stress-related proteins, such as CCAAT-enhancer-binding protein homologous protein (CHOP), and the phosphorylation of eukaryotic initiation factor 2-alpha (p-eIF2α) and c-Jun N-terminal kinases (p-JNK) in both cancer cell-types. The p53 upregulated modulator of apoptosis (PUMA) and B cell lymphoma-like protein 11 (BIM) levels were decreased at overtly toxic LCA concentrations, although PUMA levels increased at lower LCA concentrations in both cell lines. LCA induced autophagy-related conversion of microtubule-associated proteins 1A/1B light chain 3B (LC3BI–LC3BII), and autophagy-related protein ATG5 in PC-3 cells, but not in autophagy-deficient DU-145 cells. LCA (>10 µM) increased levels of reactive oxygen species (ROS) concentration-dependently in PC-3 cells, whereas ROS levels were not affected in DU-145 cells. Salubrinal, an inhibitor of eIF2α dephosphorylation and ER stress, reduced LCA-induced CHOP levels slightly in PC-3, but not DU-145 cells. Salubrinal pre-treatment increased the cytotoxicity of LCA in PC-3 and DU-145 cells and resulted in a statistically significant loss of cell viability at normally non-toxic concentrations of LCA. The late-stage autophagy inhibitor bafilomycin A1 exacerbated LCA toxicity at subtoxic LCA concentrations in PC-3 cells. The antioxidant α-tocotrienol strongly inhibited the toxicity of LCA in PC-3 cells, but not in DU-145 cells. Collectively, although LCA induces autophagy and ER stress in PC-3 cells, these processes appear to be initially of protective nature and subsequently consequential to, but not critical for the ROS-mediated mitochondrial dysfunction and cytotoxicity of LCA. The full mechanism of LCA-induced mitochondrial dysfunction and cytotoxicity in the similarly sensitive DU-145 cells remains to be elucidated.

Effect of 2-methoxyestradiol combined with quercetin on prostate cancer in vitro

To detect the effects of quercetin (Que) combined with 2-methoxyestradiol (2-ME) on the proliferation of androgen-dependent LNCaP human prostate cancer cells line and androgen-independent PC-3 human prostate cancer cells line, and to evaluate the antitumor effects of different combos of the two drugs. After LNCaP and PC-3 cells were treated with different concentration of quercetin (0, 3.125, 6.25, 12.5, 25, 50, 100, 200 μmol/L) or 2-ME (0, 0.312 5, 0.625, 1.25, 2.5, 5, 10 μmol/L) for 48 h, the inhibitory rates of cell growth were tested using trypan blue staining method respectively. Then the concentration-effect curves were drawn and IC(50) values were calculated. According to the fitted dose-effect curves and IC(50) values, appropriate concentrations of quercetin and 2-ME were selected to compose 16 different combos. Then cells were treated with different combos of Que and 2-ME for 48 h, and then the growth inhibitory rates of cell growth were detected. According to the equation and median-effect principle, the CI values of 16 different combos of Que and 2-ME were calculated to evaluate their antitumor effects. The inhibition rate of LNCaP or PC-3 cell growth treated with varying doses of quercetin or 2-ME alone showed a dose-dependent increase respectively. The IC(50) values of quercetin and 2-ME were 23.29 μmol/L and 1.89 μmol/L for LNCaP cells; and 22.12 μmol/L and 1.74 μmol/L for PC-3 cells respectively. After treated with 16 combos of Que (5, 10, 20, 40 μmol/L) and 2-ME (0.5, 1, 3, 5 μmol/L) for 48 h, for LNCaP cells, lower dose of Que (5 and 10 μmol/L) with higher dose of 2-ME (3 and 5 μmol/L) showed synergistic activity, whereas for PC-3 cells, besides the above combination of Que 10 μmol/L and 2-ME 3 μmol/L, higher dose of quercetin (20 and 40 μmol/L) with higher dose of 2-ME (3 and 5 μmol/L) also showed synergistic activity. Both quercetin and 2-methoxyestradiol could inhibit the growth of LNCaP and PC-3 human prostate cancer cells in a dose dependent manner. We confirmed that combinations of quercetin and 2-ME at appropriate concentrations have the potential for synergetic antiproliferative activity in vitro.

Combination of Quercetin and 2-Methoxyestradiol Enhances Inhibition of Human Prostate Cancer LNCaP and PC-3 Cells Xenograft Tumor Growth.

Quercetin and 2-Methoxyestradiol (2-ME) are promising anti-cancer substances. Our previous in vitro study showed that quercetin synergized with 2-Methoxyestradiol exhibiting increased antiproliferative and proapoptotic activity in both androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cell lines. In the present study, we determined whether their combination could inhibit LNCaP and PC-3 xenograft tumor growth in vivo and explored the underlying mechanism. Human prostate cancer LNCaP and PC-3 cells were inoculated subcutaneously in male BALB/c nude mice. When xenograft tumors reached about 100 mm3, mice were randomly allocated to vehicle control, quercetin or 2-Methoxyestradiol singly treated and combination treatment groups. After therapeutic intervention for 4 weeks, combination treatment of quercetin and 2-ME i) significantly inhibited prostate cancer xenograft tumor growth by 46.8% for LNCaP and 51.3% for PC-3 as compared to vehicle control group, more effective than quercetin (28.4% for LNCaP, 24.8% for PC3) or 2-ME (32.1% for LNCaP, 28.9% for PC3) alone; ii) was well tolerated by BALB/c mice and no obvious toxic reactions were observed; iii) led to higher Bax/Bcl-2 ratio, cleaved caspase-3 protein expression and apoptosis rate; and iv) resulted in lower phosphorylated AKT (pAKT) protein level, vascular endothelial growth factor protein and mRNA expression, microvascular density and proliferation rate than single drug treatment. These effects were more remarkable compared to vehicle group. Therefore, combination of quercetin and 2-ME can serve as a novel clinical treatment regimen owning the potential of enhancing antitumor effect on prostate cancer in vivo and lessening the dose and side effects of either quercetin or 2-ME alone. These in vivo results will lay a further solid basis for subsequent researches on this novel therapeutic regimen in human prostate cancer.

Alpha-tomatine synergises with paclitaxel to enhance apoptosis of androgen-independent human prostate cancer PC-3 cells in vitro and in vivo

Alpha (α)-tomatine, a major saponin found in tomato has been shown to inhibit the growth of androgen-independent prostate cancer PC-3 cells. The effects of α-tomatine in combination with the chemotherapeutic agent paclitaxel against PC-3 cells were investigated in the present study. Combined treatment with a sub-toxic dose of α-tomatine and paclitaxel significantly decreased cell viability with concomitant increase in the percentage of apoptotic PC-3 cells. The combined treatment, however, had no cytotoxic effect on the non-neoplastic prostate RWPE-1 cells. Apoptosis of PC-3 cells was accompanied by the inhibition of PI3K/Akt pro-survival signaling, an increase in the expression of the pro-apoptotic protein BAD but a decrease in the expressions of anti-apoptotic proteins, Bcl-2 and Bcl-xL. Results from a mouse xenograft model showed the combined treatment completely suppressed subcutaneous tumor growth without significant side effects. Consistent with its in vitro anti-cancer effects, tumor materials from mice showed increased apoptosis of tumor cells with reduced protein expression of activated PI3K/Akt. These results suggest that the synergistic anti-cancer effects of paclitaxel and α-tomatine may be beneficial for refractory prostate cancer treatment.

Treatment of PC-3 cells with ultrasound combined with microbubbles induces distinct alterations in the expression of Bcl-2 and Bax

The objective of the present study was to investigate whether ultrasound combined with microbubbles induces apoptotic cell death in androgen-independent prostate cancer cells and to identify the probable mechanism. We used ultrasound in continuous wave mode with a frequency of 21 kHz and a spatial-average temporal-average intensity of 46 mW/cm2. Ultrasound combined with microbubbles (200 μL SonoVue™) was used to treat androgen-independent human prostate cancer PC-3 cells for 30 s. PC-3 cells were divided into three groups: the control group, the ultrasound group and the ultrasound combined with microbubbles group. Immediately after treatment, trypan blue exclusion was used to assess cell viability. Cell apoptosis at 24 h after treatment was measured using transmission electron microscopy and flow cytometry. Western blotting was used to evaluate the expression of the apoptosis-related proteins, Bcl-2 and Bax. Ultrasound combined with microbubbles had a minimal effect on the viability of PC-3 cells and induced minimal levels of cell lysis. The level of apoptosis in PC-3 cells induced by this modality was significantly higher than in controls (12.77 ± 0.31% vs. 2.56 ± 0.22%, P<0.01). Treatment with ultrasound combined with microbubbles increased the expression of Bax, a pro-apoptotic protein, and decreased the expression of Bcl-2, an anti-apoptotic protein. It was concluded that ultrasound combined with microbubbles induces apoptotic cell death in human prostate cancer PC-3 cells through down-regulation of Bcl-2 and up-regulation of Bax.

Quercetin synergizes with 2-methoxyestradiol inhibiting cell growth and inducing apoptosis in human prostate cancer cells

Lack of effective treatment options for castration-resistant prostate cancer reinforces the great need to develop novel drug therapies. Quercetin is a plant-derived flavonoid that can induce apoptosis in prostate cancer cells. 2-Methoxyestradiol (2-ME) is an endogenous estrogenic metabolite that also has antineoplastic activity. However, these two agents have limited bioavailability. Herein, we explored the antiproliferative and proapoptotic activities of quercetin combined with 2-ME in both androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cell lines. Compared to quercetin and 2-ME alone, combining quercetin with 2-ME at appropriate concentrations i) showed synergistic antiproliferative and proapoptotic activities; ii) increased G2/M phase population of cells; iii) decreased the ratio of Bcl-2/Bax significantly. The combination of quercetin and 2-ME is a new clinically relevant treatment regimen which has the potential of enhancing the antitumor effect on prostate cancer and lessening the side effect of either quercetin or 2-ME alone.

Monascuspiloin Induces Apoptosis and Autophagic Cell Death in Human Prostate Cancer Cells via the Akt and AMPK Signaling Pathways

Monascus pigments have been reported to possess anticancer effects in various cancer cells; however, the molecular mechanisms of their anticancer properties remain largely unknown. Monascuspiloin is an analogue of the Monascus pigment monascin, and its anticancer growth activity against human prostate cancer cells was evaluated using in vitro and in vivo models. Monascuspiloin effectively inhibits the growth of both androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cells. Monascuspiloin preferentially induces apoptosis in LNCaP cells by attenuating the PI3K/Akt/mTOR pathway. In androgen-independent PC-3 cells, monascuspiloin induces G2/M arrest and autophagic cell death by an AMPK-dependent pathway. Induction of autophagy in PC-3 cells further sensitizes cells to apoptosis induced by monascuspiloin. Monascuspiloin inhibits tumor growth in nude mice bearing PC-3 xenografts through induction of apoptosis and autophagy. This study is the first to demonstrate that monascuspiloin has therapeutic potential for the treatment of both androgen-dependent and -independent human prostate cancers.

Flavonoid ampelopsin inhibits the growth and metastasis of prostate cancer in vitro and in mice.

The objective of this study was to evaluate the chemopreventive effect of a novel flavonoid, ampelopsin (AMP) on the growth and metastasis of prostate cancer cells. AMP showed the more potent activity in inhibiting the proliferation of androgen-sensitive LNCaP and, to less extent, androgen-independent PC-3 human prostate cancer cell lines in vitro, primarily by induction of apoptosis associated with down-regulation of bcl-2. On the other hand, AMP showed much less activity in inhibiting the proliferation of normal prostate epithelial cells than that of prostate cancer cell lines. AMP also inhibited the migration and invasion of PC-3 cells in vitro associated with down-regulation of CXCR4 expression. In the animal study using an orthotopic prostate tumor model, AMP (150 and 300 mg/kg body weight) inhibited the growth of PC-3 tumors and lymph node and lung metastases in a dose-dependent manner. Compared to the control mice, mice treated with AMP at 300 mg/kg BW had reduced final tumor weight by 49.2% (P<0.05), lymph node metastases by 54.5% (P = 0.3) and lung metastases by 93% (P<0.05), but had no apparent alteration on food intake or body weight. The in vivo anti-growth and anti-metastasis activities of AMP were associated with induction of apoptosis and inhibition of proliferation of prostate cancer cells, reduction of prostate tumor angiogenesis, and reduction of CXCR4 expression. Our results provide supporting evidence to warrant further investigation to develop AMP as a novel efficacious and safe candidate agent against progression and metastasis of prostate cancer.

Carnosic acid modulates Akt/IKK/NF-κB signaling by PP2A and induces intrinsic and extrinsic pathway mediated apoptosis in human prostate carcinoma PC-3 cells

This study investigates the efficacy of carnosic acid (CA), a polyphenolic diterpene, isolated from the plant rosemary (Rosemarinus officinalis), on androgen-independent human prostate cancer PC-3 cells. CA induced anti-proliferative effects in PC-3 cells in a concentration- and time-dependent manner, which was due to apoptotic induction as evident from flow-cytometry, DNA laddering and TUNEL assay. Apoptosis was associated with the activation of caspase-8, -9, -3 and -7, increase in Bax:Bcl-2 ratio, release of cytochrome-c and decrease in expression of inhibitor of apoptosis (IAP) family of proteins. Apoptosis was attenuated upon pretreatment with specific inhibitors of caspase-8 (Z-IETD-fmk) and caspase-9 (Z-LEHD-fmk) suggesting the involvement of both intrinsic and extrinsic apoptotic cascades. Further, apoptosis resulted from the inhibition of IKK/NF-κB pathway as evident from decreased DNA binding activity, nuclear translocation of p50 and p65 and IκBα phosphorylation. The down-regulation of IKK/NF-κB was associated with inhibition of Akt phosphorylation and its kinase activity with a concomitant increase in the serine/threonine protein phosphatase 2A (PP2A) activity. Pharmacologic inhibition of PP2A by okadaic acid and calyculin A, significantly reversed CA-mediated apoptotic events in PC-3 cells indicating that CA induced apoptosis by activation of PP2A through modulation of Akt/IKK/NF-κB pathway. In addition, CA induced apoptosis in another androgen refractory prostate cancer DU145 cells via intrinsic pathway as evidenced from the activation of caspase 3, cleavage of PARP, increase in Bax:Bcl-2 ratio and cytochrome-c release. Carnosic acid, therefore, may have the potential for use in the prevention and/or treatment of prostate cancer.

Inhibition of tumor growth and sensitization to chemotherapy by RNA interference targeting interleukin‐6 in the androgen‐independent human prostate cancer PC3 model

The objective of the present study was to investigate the inhibitory effects of interleukin-6 (IL-6) secretion by androgen-independent human prostate cancer PC3 cells on their growth and chemosensitivity. In this study, we established PC3 in which the expression vector containing short hairpin RNA (shRNA) targeting IL-6 was introduced (PC3/sh-IL6). Changes in the growth and sensitivity to docetaxel in PC3/sh-IL6 were compared with those in PC3 transfected with control vector alone (PC3/Co). Concentration of IL-6 in the culture supernatant from PC3/sh-IL6 was approximately 20% of that from PC3/Co. Both in vitro and in vivo, the growth of PC3/sh-IL-6 was significantly inferior to that of PC3/Co, accompanying downregulation of Bcl-2, Bcl-xL, phosphorylated Akt, p44/42 mitogen-activated protein kinase, and signal transducers and activation of transcription 3 in PC3/sh-IL-6 compared with that in PC3/Co. Despite the higher sensitivity of PC3/sh-IL6 to docetaxel than that of PC3/Co, the secretion of IL-6 by both cell lines was increased after treatment with docetaxel due to the formation of positive autocrine loops between these cell lines and NFκB signaling pathways. Furthermore, combined treatment with the proteasome inhibitor bortezomib, which completely inhibited the docetaxel-induced IL-6 secretion via the inactivation of NFκB signaling, resulted in the marked sensitization of these cell lines to docetaxel both in vitro and in vivo. These findings suggest that suppressed IL-6 secretion using shRNA, either alone or in combination with docetaxel and bortezomib, could be a useful therapeutic strategy against androgen-independent prostate cancer.

Oncolytic targeting of androgen-sensitive prostate tumor by the respiratory syncytial virus (RSV): consequences of deficient interferon-dependent antiviral defense

BackgroundOncolytic virotherapy for cancer treatment utilizes viruses for selective infection and death of cancer cells without any adverse effect on normal cells. We previously reported that the human respiratory syncytial virus (RSV) is a novel oncolytic virus against androgen-independent PC-3 human prostate cancer cells. The present study extends the result to androgen-dependent prostate cancer, and explores the underlying mechanism that triggers RSV-induced oncolysis of prostate cancer cells.MethodsThe oncolytic effect of RSV on androgen-sensitive LNCaP human prostate cancer cells and on androgen-independent RM1 murine prostate cancer cells was studied in vitro in culture and in vivo in a xenograft or allograft tumor model. In vitro, cell viability, infectivity and apoptosis were monitored by MTT assay, viral plaque assay and annexin V staining, respectively. In vivo studies involved virus administration to prostate tumors grown in immune compromised nude mice and in syngeneic immune competent C57BL/6J mice. Anti-tumorogenic oncolytic activity was monitored by measuring tumor volume, imaging bioluminescent tumors in live animals and performing histopathological analysis and TUNEL assay with tumorsResultsWe show that RSV imposes a potent oncolytic effect on LNCaP prostate cancer cells. RSV infectivity was markedly higher in LNCaP cells compared to the non-tumorigenic RWPE-1 human prostate cells. The enhanced viral burden led to LNCaP cell apoptosis and growth inhibition of LNCaP xenograft tumors in nude mice. A functional host immune response did not interfere with RSV-induced oncolysis, since growth of xenograft tumors in syngeneic C57BL/6J mice from murine RM1 cells was inhibited upon RSV administration. LNCaP cells failed to activate the type-I interferon (IFNα/β)-induced transcription factor STAT-1, which is required for antiviral gene expression, although these cells could produce IFN in response to RSV infection. The essential role of IFN in restricting infection was further borne out by our finding that neutralizing IFN activity resulted in enhanced RSV infection in non-tumorigenic RWPE-1 prostate cells.ConclusionsWe demonstrated that RSV is potentially a useful therapeutic tool in the treatment of androgen-sensitive and androgen-independent prostate cancer. Moreover, impaired IFN-mediated antiviral response is the likely cause of higher viral burden and resulting oncolysis of androgen-sensitive prostate cancer cells.

Abstract B203: Mitochondria-targeted superoxide spin-traps for treatment of cancers with high oxidative stress

Abstract Many human malignancies such as prostate cancer (CaP) and primary brain tumor (GBM) exhibit high oxidative stress. Developing a successful therapeutic drug to treat advanced metastatic CaP and GBM is a major unmet medical need. It has been demonstrated that in certain chemo- and/or radiation resistant cancer cells with high oxidative stress, cells depend on reactive oxygen species (ROS) for survival and proliferation. Therefore, reducing ROS can be a successful strategy for developing chemotherapeutic drugs for use against chemo- and/or radiation-resistant tumors. The mitochondria are a major source of superoxide and other ROS. We focused on targeting spin-trapping nitroxide analogs at the mitochondrial interstitial space with appropriate linker length in order to remove the superoxide in order to reduce the cellular ROS levels without interfering with the mitochondrial electron transport chain. Our lead compound in this new class of drugs is Mito-tempol-C10. The drug successfully blocked ROS production in human CaP and GBM cell lines as determined by DCF dye oxidation assay. It exhibited marked growth inhibitory effect against androgen-dependent as well as androgen-independent cultured human CaP cells (IC50 &amp;lt; 1 µM) and similar growth inhibitory activity against several human GBM cells lines as well as cultured primary brain tumor stem cells as determined by DNA fluorescence measurement assay. Preliminary pharmacokinetic (PK) study show that the drug has serum half-life of approximately 45 minutes, but is retained by the tumor and other animal tissue for more than 24 hours. The drug is tolerated by ICR white mice, nude mice and NOD-SCID mice at a dose of 15 mg/kg i.p. given once every week for four weeks without any overt systemic toxicity. Preliminary efficacy study at this treatment condition showed marked effect on the growth of intracranially implanted, radiation-resistant U-87 MG human GBM cells in NOD-SCID mice and a small, but significant effect on the growth of androgen-independent PC-3 human prostate cancer cell xenografts growing on the flanks of nude mice. These results confirm that mitochondria-targeted spin-traps that reduce cellular ROS can be therapeutically effective in treatment of advanced, hard-to-treat human tumors with high oxidative stress. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B203.

Detection of Intracellular Granularity Induction in Prostate Cancer Cell Lines by Small Molecules Using the HyperCyt® High-Throughput Flow Cytometry System

Prostate cancer is a leading cause of death among men due to the limited number of treatment strategies available for advanced disease. Discovery of effective chemotherapeutics involves the identification of agents that inhibit cancer cell growth. Increases in intracellular granularity have been observed during physiological processes that include senescence, apoptosis, and autophagy, making this phenotypic change a useful marker for identifying small molecules that induce cellular growth arrest or death. In this regard, epithelial-derived cancer cell lines appear uniquely susceptible to increased intracellular granularity following exposure to chemotherapeutics. We have established a novel flow cytometry approach that detects increases in side light scatter in response to morphological changes associated with intracellular granularity in the androgen-sensitive LNCaP and androgen-independent PC3 human prostate cancer cell lines. A cell-based assay was developed to screen for small molecule inducers of intracellular granularity using the HyperCyt high-throughput flow cytometry platform. Validation was performed using the Prestwick Chemical Library, where known modulators of LNCaP intracellular granularity, such as testosterone, were identified. Nonandrogenic inducers of granularity were also detected. A further screen of approximately 25,000 small molecules led to the identification of a class of aryl-oxazoles that increased intracellular granularity in both cell lines, often leading to cell death. The most potent agents exhibited submicromolar efficacy in LNCaP and PC3 cells.

Triterpenoids and Alkaloids from the Roots of Peganum Nigellastrum

Four lupane-type triterpenoids (1–4) and seven alkaloids (5–11) were isolated from the roots of Peganum nigellastrum. On the basis of spectroscopic and chemical evidence, the structures of the compounds were elucidated as 3α-hydroxy-27- trans-caffeoyloxylup-20(29)-en-28-oic acid methyl ester (1), 3β-hydroxy-27- trans-caffeoyloxylup-20(29)-en-28-oic acid methyl ester (2), 3α-acetoxy-27- trans-caffeoyloxylup-20(29)-en-28-oic acid methyl ester (3), 3β-acetoxy-27- trans-caffeoyloxylup-20(29)-en-28-oic acid methyl ester (4), luotonin C (5), luotonin D (6), harmine (7), harmol (8), harmaline (9), deoxyvasicinone (10) and vasicinone (11). Compounds 1, 3 and 4 are novel triterpenoids, and these pentacyclic triterpenoids were evaluated for their cytotoxicity against the androgen-sensitive LNCaP and androgen-independent PC-3 human prostate cancer cells.

Krüppel-Like Factor 4 Is a Novel Mediator of Selenium in Growth Inhibition

A previous prevention trial showed that selenium supplementation was effective in reducing (by 50%) the incidence of prostate cancer. Selenium has been reported to inhibit the growth of prostate cancer cells in vitro. Multiple mechanisms are likely to be operative in the underlying effect of selenium. Here, we report that Krüppel-like factor 4 (KLF4), a transcription factor of the KLF family, is an important target of selenium. We found that selenium up-regulates KLF4 expression and increases the DNA-binding activity of KLF4 in both the androgen-dependent LNCaP and the androgen-independent PC-3 human prostate cancer cells. The increase of KLF4 mRNA is accounted for primarily by enhanced transcription, although the contribution of a slight abatement in mRNA degradation cannot be ruled out. KLF4 knockdown using short interference RNA significantly weakens the effects of selenium on DNA synthesis inhibition, apoptosis induction, and the expression of three KLF4 target genes, cyclin D1, p21/WAF1, and p27/Kip1. In addition, the overexpression of KLF4 not only leads to an induction of apoptosis in the control cells, but also enhances the DNA synthesis-suppressive and-proapoptotic activities of selenium. Taken together, our results suggest that KLF4 plays a key role in mediating the growth-inhibitory effect of selenium in prostate cancer cells.

MP-17.08: Roles of vinexinβ in growth and paclitaxel resistance in androgen-independent human prostate cancer PC-3 cells

MP-17.08: Roles of vinexinβ in growth and paclitaxel resistance in androgen-independent human prostate cancer PC-3 cells

A Novel Antisense Oligonucleotide Inhibiting Several Antiapoptotic Bcl-2 Family Members Induces Apoptosis and Enhances Chemosensitivity in Androgen-Independent Human Prostate Cancer PC3 Cells

A Novel Antisense Oligonucleotide Inhibiting Several Antiapoptotic Bcl-2 Family Members Induces Apoptosis and Enhances Chemosensitivity in Androgen-Independent Human Prostate Cancer PC3 Cells

Soy Isoflavones Alter Expression of Genes Associated with Cancer Progression, Including Interleukin-8, in Androgen-Independent PC-3 Human Prostate Cancer Cells

High consumption of soy isoflavones in Asian diets has been correlated with a lower incidence of clinically important cases of prostate cancer. The chemopreventive properties of these diets may result from an interaction of several types of isoflavones, including genistein and daidzein. The present study investigated the effects of a soy isoflavone concentrate (ISF) on growth and gene expression profiles of PC-3 human prostate cancer cells. Trypan blue exclusion and [3H]-thymidine incorporation assays showed that ISF decreased cell viability and caused a dose-dependent inhibition of DNA synthesis, respectively, with 50% inhibition (IC50) of DNA synthesis at 52 mg/L (P = 0.05). The glucoside conjugates of genistein and daidzein in ISF were converted to bioactive free aglycones in cell culture in association with the inhibition of DNA synthesis. Flow cytometry and Western immunoblot analyses showed that ISF at 200 mg/L caused an accumulation of cells in the G2/M phase of the cell cycle (P < 0.05) and decreased cyclin A by 20% (P < 0.05), respectively. The effect of ISF on the gene expression profile of PC-3 cells was analyzed using Affymetrix oligonucleotide DNA microarrays that interrogate approximately 17,000 human genes. Of the 75 genes altered by ISF, 28 were upregulated and 47 were downregulated (P < 0.05). Further analysis showed that IL-8, matrix metalloproteinase 13, inhibin beta A, follistatin, and fibronectin mRNA levels were significantly reduced, whereas the expression of p21(CIP1), a major cell cycle inhibitory protein, was increased. The effects of ISF on the expression of IL-8 and p21(CIP1) mRNA and protein were validated at high and low ISF concentrations. Our data show that ISF inhibits the growth of PC-3 cells through modulation of cell cycle progression and the expression of genes involved in cell cycle regulation, metastasis, and angiogenesis.

A novel antisense oligonucleotide inhibiting several antiapoptotic Bcl-2 family members induces apoptosis and enhances chemosensitivity in androgen-independent human prostate cancer PC3 cells

Bcl-2 and Bcl-xL are associated with treatment resistance and progression in many cancers, including prostate cancer. The objective of this study was to determine whether a novel bispecific antisense oligonucleotide targeting both Bcl-2 and Bcl-xL induces apoptosis and enhances chemosensitivity in androgen-independent PC3 prostate cancer cells. An antisense oligonucleotide with complete sequence identity to Bcl-2 and three-base mismatches to Bcl-xL selected from five antisense oligonucleotides targeting various regions with high homology between Bcl-2 and Bcl-xL was found to be the most potent inhibitor of both Bcl-2 and Bcl-xL expression in PC3 cells. This selected Bcl-2/Bcl-xL bispecific antisense oligonucleotide reduced mRNA and protein levels in a dose-dependent manner, reducing Bcl-2 and Bcl-xL protein levels to 12% and 19%, respectively. Interestingly, Mcl-1 was down-regulated as well, although levels of Bax, Bad, or Bak were not altered after treatment with this bispecific antisense oligonucleotide. Indirect down-regulation of inhibitor of apoptosis (IAP) family, including XIAP, cIAP-1 and cIAP-2, via second mitochondria-derived activator of caspases was also observed after bispecific antisense oligonucleotide treatment. Executioner caspase-3, caspase-6, and caspase-7 were shown to be involved in apoptosis induced by bispecific antisense oligonucleotide. This Bcl-2/Bcl-xL bispecific antisense oligonucleotide also enhanced paclitaxel chemosensitivity in PC3 cells, reducing the IC50 of paclitaxel by >90%. These findings illustrate that combined suppression of antiapoptotic Bcl-2 family members using this antisense oligonucleotide could be an attractive strategy for inhibiting cancer progression through alteration of the apoptotic rheostat in androgen-independent prostate cancer.