紫杉醇抑制前列腺癌增殖的体内实验研究

目的探讨紫杉醇对人前列腺癌细胞PC-3增殖的体内抑制作用。方法建立体内绿色荧光蛋白（GFP）标记的人雄激素非依赖性前列腺癌细胞PC-3裸鼠原位移植瘤模型,观察紫杉醇对裸鼠前列腺癌原位移植瘤的体积、重量的影响。结果裸鼠模型体内实验显示,与对照组（100μL生理盐水）相比,紫杉醇处理组（0.5 mg/kg）在给药第18天后能显著抑制前列腺肿瘤的体积（P〈0.05）;紫杉醇处理组在抑制前列腺肿瘤重量方面与对照组相比亦有明显抑制作用（P〈0.05）。与对照组相比G31P处理组VEGF（P〈0.05）的表达差异具有统计学意义（免疫组化法）。结论紫杉醇在体内实验中能明显抑制人雄激素非依赖性前列腺癌细胞系PC-3的增殖。     

Connexins,important players in the dissemination of prostate cancer cells

Over the past 50 years, increasing experimental evidences have established that connexins (Cxs) and gap junctional intercellular communication (GJIC) ensure an important role in both the onset and development of cancerous processes. In the present review, we focus on the impact of Cxs and GJIC during the development of prostate cancer (PCa), from the primary growth mainly localized in acinar glands and ducts to the distant metastasis mainly concentrated in bone. As observed in several other types of solid tumours, Cxs and especially Cx43 exhibit an ambivalent role with a tumour suppressor effect in the early stages and, conversely, a rather pro-tumoural profile for most of invasion and dissemination steps to secondary sites. We report here the current knowledge on the function of Cxs during PCa cells migration, cytoskeletal dynamics, proteinases activities and the cross talk with the surrounding stromal cells in the microenvironment of the tumour and the bones. In addition, we discuss the role of Cxs in the bone tropism even if the prostate model is rarely used to study the complete sequence of cancer dissemination compared to breast cancer or melanoma. Even if not yet fully understood, these recent findings on Cxs provide new insights into their molecular mechanisms associated with progression and bone targeted behaviour of PCa. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve.     

Adiponectin as a growth inhibitor in prostate cancer cells

Prostate cancer is associated with obesity. However, the molecular basis of this association is not well known. Adiponectin is a major adipose cytokine that decreases in circulation in obesity and ameliorates obesity. Here, we identify adiponectin as a novel inhibitor in prostate cancer cell growth. Adiponectin occurs in non-proteolytic (full-length adiponectin: f-adiponectin) and proteolytic (globular adiponectin) forms in various oligomeric states (trimer, hexamer, and high molecular weight complex). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay demonstrates that f-adiponectin inhibits prostate cancer cell growth drastically at subphysiological concentrations. Furthermore, velocity sedimentation analysis shows that the high molecular weight complex of f-adiponectin is the inhibitory form. Moreover, f-adiponectin suppresses leptin- and/or insulin-like growth factor-I (IGF-I)-stimulated, androgen-independent DU145 cell growth, and dihydrotestosterone-stimulated, androgen-dependent LNCaP-FGC cell growth. In addition, f-adiponectin enhances doxorubicin inhibition of prostate cancer cell growth. Therefore, f-adiponectin is a molecular mediator between prostate cancer and obesity, and may be therapeutic to prostate cancer.     

Genistein inhibits the contact-stimulated migration of prostate cancer cells

The results of several epidemiological studies have suggested that a soybean-based diet is associated with a lower risk of prostate cancer. We investigated the effect of the soy isoflavone genistein on the proliferation and contact-stimulated migration of rat prostatic carcinoma MAT-LyLu and AT-2 cell lines. Genistein almost completely inhibited the growth of both MAT-LyLu and AT-2 cells in the concentration range from 25 to 100 μM, but the addition of 1 μM genistein to the medium significantly stimulated the proliferation of both cell lines. Additionally, at concentrations above 25 μM, genistein showed a potent cytotoxic effect. However, the central finding of this study is that at physiologically relevant concentrations (1 μM and 10 μM), genistein inhibits the motility of prostate cancer cells stimulated by homo-and heterotypic contacts. These results show that at physiological concentrations, genistein exerts an inhibitory effect on the migration of prostate cancer cells and suggest that it may be one of the factors responsible for the anti-metastatic activity of plant isoflavonoids     

Proteomic analysis of cancer stem cells in human prostate cancer cells

Results from recent studies support the hypothesis that cancer stem cells (CSCs) are responsible for tumor initiation and formation. Here, we applied a proteome profiling approach to investigate the mechanisms of CSCs and to identify potential biomarkers in the prostate cancer cell line DU145. Using MACS, the DU145 prostate cancer cell line was isolated into CD44+ or CD44− cells. In sphere culture, CD44+ cells possessed stem cell characteristics and highly expressed genes known to be important in stem cell maintenance. In addition, they showed strong tumorigenic potential in the clonogenic assay and soft agar colony formation assay. We then analyzed and identified proteins that were differentially expressed between CD44+ and CD44− using two-dimensional gel electrophoresis and LC-MS/MS. Cofilin and Annexin A5, which are associated with proliferation or metastasis in cancer, were found to be positively correlated with CD44 expression. These results provide information that will be important to the development of new cancer diagnostic tools and understanding the mechanisms of CSCs although a more detailed study is necessary to investigate the roles of Cofilin and Annexin A5 in CSCs.     

p66Shc regulates migration of castration-resistant prostate cancer cells

Metastatic castration-resistant (CR) prostate cancer (PCa) is a lethal disease for which no effective treatment is currently available. p66Shc is an oxidase previously shown to promote androgen-independent cell growth through generation of reactive oxygen species (ROS) and is elevated in clinical PCa and multiple CR PCa cell lines. We hypothesize p66Shc also increases the migratory activity of PCa cells through ROS and investigate the associated mechanism. Using the transwell assay, our study reveals that the level of p66Shc protein correlates with cell migratory ability across several PCa cell lines. Furthermore, we show hydrogen peroxide treatment induces migration of PCa cells that express low levels of p66Shc in a dose-dependent manner, while antioxidants inhibit migration. Conversely, PCa cells that express high levels of endogenous p66Shc or by cDNA transfection possess increased cell migration which is mitigated upon p66Shc shRNA transfection or expression of oxidase-deficient dominant-negative p66Shc W134F mutant. Protein microarray and immunoblot analyses reveal multiple proteins, including ErbB-2, AKT, mTOR, ERK, FOXM1, PYK2 and Rac1, are activated in p66Shc-elevated cells. Their involvement in PCa migration was examined using respective small-molecule inhibitors. The role of Rac1 was further validated using cDNA transfection and, significantly, p66Shc is found to promote lamellipodia formation through Rac1 activation. In summary, the results of our current studies clearly indicate p66Shc also regulates PCa cell migration through ROS-mediated activation of migration-associated proteins, notably Rac1.     

Mitostatin is down-regulated in human prostate cancer and suppresses the invasive phenotype of prostate cancer cells

MITOSTATIN, a novel putative tumor suppressor gene induced by decorin overexpression, is expressed in most normal human tissues but is markedly down-regulated in advanced stages of mammary and bladder carcinomas. Mitostatin negatively affects cell growth, induces cell death and regulates the expression and activation levels of Hsp27. In this study, we demonstrated that ectopic expression of Mitostatin in PC3, DU145, and LNCaP prostate cancer cells not only induced a significant reduction in cell growth, but also inhibited migration and invasion. Moreover, Mitostatin inhibited colony formation in soft-agar of PC3 and LNCaP cells as well as tumorigenicity of LNCaP cells in nude mice. Conversely, targeting endogenous Mitostatin by siRNA and anti-sense strategies in PC3 and DU145 prostate cancer cells enhanced the malignant phenotype in both cell lines. In agreement of these anti-oncogenic roles, we discovered that Mitostatin was absent in ∼35% (n = 124) of prostate tumor samples and its overall reduction was associated with advanced cancer stages. Collectively, our findings indicate that MITOSTATIN may acts as a tumor suppressor gene in prostate cancer and provide a novel cellular and molecular mechanism to be further exploited and deciphered in our understanding of prostate cancer progression.     

Amyloid precursor protein regulates migration and metalloproteinase gene expression in prostate cancer cells

Amyloid precursor protein (APP) is a type I transmembrane protein, and one of its processed forms, β-amyloid, is considered to play a central role in the development of Alzheimer’s disease. We previously showed that APP is a primary androgen-responsive gene in prostate cancer and that its increased expression is correlated with poor prognosis for patients with prostate cancer. APP has also been implicated in several human malignancies. Nevertheless, the mechanism underlying the pro-proliferative effects of APP on cancers is still not well-understood. In the present study, we explored a pathophysiological role for APP in prostate cancer cells using siRNA targeting APP (siAPP). The proliferation and migration of LNCaP and DU145 prostate cancer cells were significantly suppressed by siAPP. Differentially expressed genes in siAPP-treated cells compared to control siRNA-treated cells were identified by microarray analysis. Notably, several metalloproteinase genes, such as ADAM10 and ADAM17, and epithelial–mesenchymal transition (EMT)-related genes, such as VIM, and SNAI2, were downregulated in siAPP-treated cells as compared to control cells. The expression of these genes was upregulated in LNCaP cells stably expressing APP when compared with control cells. APP-overexpressing LNCaP cells exhibited enhanced migration in comparison to control cells. These results suggest that APP may contribute to the proliferation and migration of prostate cancer cells by modulating the expression of metalloproteinase and EMT-related genes.     

Unripe Rubus coreanus Miquel suppresses migration and invasion of human prostate cancer cells by reducing matrix metalloproteinase expression

Rubus coreanus Miquel (RCM) is used to promote prostate health and has been shown to have anti-oxidant and anti-carcinogenic activities. However, the effects and mechanisms of RCM on prostate cancer metastasis remain unclear. PC-3 and DU 145 cells were treated with ethanol or water extract of unripe or ripe RCM and examined for cell invasion, migration, and matrix metalloproteinases (MMPs) activity and expression. Phosphoinositide 3-kinase (PI3K) and Akt activities were examined. Unripe RCM extracts exerted significant inhibitory effects on cell migration, invasion, and MMPs activities. A significant reduction in MMPs activities by unripe RCM ethanol extract treatment (UE) was associated with reduction of MMPs expression and induction of tissue inhibitors of metalloproteinases (TIMPs) expression. Furthermore, PI3K/Akt activity was diminished by UE treatment. In this study, we demonstrated that UE decreased metastatic potential of prostate cancer cells by reducing MMPs expression through the suppression of PI3K/Akt phosphorylation, thereby decreasing MMP activity and enhancing TIMPs expression.     

A new proteasome inhibitor, TP-110, induces apoptosis in human prostate cancer PC-3 cells

Proteasome inhibitors are useful in the treatment of cancer. Recently, we found a new proteasome inhibitor, TP-110, derived from tyropeptin A produced by Kitasatospora sp. Here we report that TP-110 induces apoptosis in human prostate cancer PC-3 cells. TP-110 showed strong cytotoxicity to PC-3 cells (IC(50)=0.05 muM). It increased the number of cells in the G(2)-M phase and increased the accumulated amounts of the p21 and p27 proteins, which are negative regulators of cell cycle progression. Furthermore, it induced apoptosis along with chromatin condensation and DNA fragmentation in PC-3 cells, and TP-110-induced apoptosis appeared to be associated with caspase activation. Additionally, TP-110 inhibited not only the degradation of IkappaB and the nuclear translocation of nuclear factor-kappaB (NF-kappaB), but also the DNA binding activity of NF-kappaB. These results indicate that TP-110 shows a strong growth inhibition and apoptosis in PC-3 cells.     

Continuous exposure of nicotine and cotinine retards human primary pterygium cell proliferation and migration

Pterygium is a triangular-shaped hyperplastic growth, characterized by conjunctivalization, inflammation, and connective tissue remodeling. Our previous meta-analysis found that cigarette smoking is associated with a reduced risk of pterygium. Yet, the biological effect of cigarette smoke components on pterygium has not been studied. Here we reported the proliferation and migration properties of human primary pterygium cells with continuous exposure to nicotine and cotinine. Human primary pterygium cells predominantly expressed the α5, β1, and γ subunits of the nicotinic acetylcholine receptor. Continuous exposure to the mixture of 0.15 μM nicotine and 2 μM cotinine retarded pterygium cell proliferation by 16.04% (P = 0.009) and hindered their migration by 11.93% ( P = 0.039), without affecting cell apoptosis. SNAIL and α-smooth muscle actin protein expression was significantly downregulated in pterygium cells treated with 0.15 μM nicotine-2 μM cotinine mixture by 1.33- ( P = 0.036) and 1.31-fold ( P = 0.001), respectively. Besides, the 0.15 μM nicotine-2 μM cotinine mixture also reduced matrix metalloproteinase (MMP)-1 and MMP-9 expressions in pterygium cells by 1.56- ( P = 0.043) and 1.27-fold ( P = 0.012), respectively. In summary, this study revealed that continuous exposure of nicotine and cotinine inhibited human primary pterygium cell proliferation and migration in vitro by reducing epithelial-to-mesenchymal transition and MMP protein expression, partially explaining the lower incidence of pterygium in cigarette smokers.     

Luteolin suppresses growth and migration of human lung cancer cells

Luteolin, 3′,4′,5,7-tetrahydroxyflavone, has been shown to possess antioxidant, anti-inflammation and anti-cancer properties. However, its role in lung cancer remains poorly understood. Here we examined the anti-tumorigenic role of luteolin in a commonly used lung cancer cell line. Luteolin inhibited the growth of A549 cells by inducing G1 phase cell cycle arrest and apoptosis. Furthermore, stress fiber assembly and cell migration in A549 cells was markedly suppressed by luteolin.     

RKIP inhibits the migration and invasion of human prostate cancer PC-3M cells through regulation of extracellular matrix

Raf kinase inhibitor protein (RKIP) plays a pivotal role in several intracellular signaling cascades and has been implicated as a metastasis suppressor in multiple cancer cells including prostate cancer cells, but the mechanism is not very clear. In this study, we investigated the effect of RKIP on cell proliferation, migration and invasion using human prostate cancer PC-3M cells as a model system. Our results indicate that RKIP does not effect cell proliferation in PC-3M cells, but inhibits both cell migration and cell invasion. In association with this inhibitory effect, RKIP down-regulates matrix metalloproteinases (MMP-2 and MMP-9), cathepsin B and urinary plasminogen activator (uPA). Also RKIP has the ability to regulate the expression of E-cadherin. But ectopic expression of RKIP does not affect the level of the Snail protein. As it has been indicated here, RKIP inhibits the migration and invasion ability of human prostate cancer cells through regulation of the extracellular matrix. These findings provide new mechanistic insight how RKIP suppresses metastasis in vitro.     

Antiandrogen-induced cell death in LNCaP human prostate cancer cells

Antiandrogens such as Casodex (Bicalutamide) are designed to treat advance stage prostate cancer by interfering with androgen receptor-mediated cell survival and by initiating cell death. Treatment of androgen sensitive, non-metastatic LNCaP human prostate cancer cells with 0-100 microM Casodex or 0-10 ng/ml TNF-alpha induces cell death in 20-60% of the cells by 48 h in a dose-dependent manner. In cells treated with TNF-alpha, this is accompanied by the loss of mitochondrial membrane potential (DeltaPsim) and cell adhesion. In contrast, cells treated with Casodex display loss of cell adhesion, but sustained mitochondrial dehydrogenase activity. Overexpression of Bcl-2 in LNCaP cells attenuates the induction of cell death by TNF-alpha but not Casodex, suggesting that mitochondria depolarization is not required for the induction of cell death by Casodex. While both TNF-alpha and Casodex-induced release of cytochrome c in LNCaP cell is predominantely associated with the translocation and cleavage of Bax, our data also suggest that Casodex induces cell death by acting on components downstream of decline of DeltaPsim and upstream of cytochrome c release. Furthermore, while induction of both caspase-3 and caspase-8 activities are observed in TNF-alpha and Casodex-treated cells, a novel cleavage product of procaspase-8 is seen in Casodex-treated cells. Taken together, these data support the hypothesis that Casodex induces cell death by a pathway that is independent of changes in DeltaPsim and Bcl-2 actions and results in an extended lag phase of cell survival that may promote the induction of an invasive phenotype after treatment.     

Swainsonine activates mitochondria-mediated apoptotic pathway in human lung cancer A549 cells and retards the growth of lung cancer xenografts

Swainsonine (1, 2, 8-trihyroxyindolizidine, SW), a natural alkaloid, has been reported to exhibit anti-cancer activity on several mouse models of human cancer and human cancers in vivo. However, the mechanisms of SW-mediated tumor regression are not clear. In this study, we investigated the effects of SW on several human lung cancer cell lines in vitro. The results showed that SW significantly inhibited these cells growth through induction of apoptosis in different extent in vitro. Further studies showed that SW treatment up-regulated Bax, down-regulated Bcl-2 expression, promoted Bax translocation to mitochondria, activated mitochondria-mediated apoptotic pathway, which in turn caused the release of cytochrome c, the activation of caspase-9 and caspase-3, and the cleavage of poly (ADP-ribose) polymerase (PARP), resulting in A549 cell apoptosis. However, the expression of Fas, Fas ligand (FasL) or caspase-8 activity did not appear significant changes in the process of SW-induced apoptosis. Moreover, SW treatment inhibited Bcl-2 expression, promoted Bax translocation, cytochrome c release and caspase-3 activity in xenograft tumor cells, resulting in a significant decrease of tumor volume and tumor weight in the SW-treated xenograft mice groups in comparison to the control group. Taken together, this study demonstrated for the first time that SW inhibited A549 cancer cells growth through a mitochondria-mediated, caspase-dependent apoptotic pathway in vitro and in vivo.     

Crk II silencing down-regulates IGF-IR and inhibits migration and invasion of prostate cancer cells

Crk (C10 regulator of kinase) adaptor proteins are highly expressed in many types of human cancers and often contribute to aggressive cancer phenotypes. Crk II, a member of CRK family, has been reported to regulate cell migration and metastasis in breast cancer cells. However, its role in other cancer types has not been reported. In this study, we investigated the molecular function of Crk II in prostate cancer (PCa) cells (CWR-22rv1) in vitro and using a mouse tumor model. Results showed that Crk II knockdown by shRNA-mediated silencing (Crk II-shRNA) in the PCa cells significantly inhibited both cancer cell migration and invasion in cell culture study. Crk II-shRNA cancer cells also significantly decreased colony formation in vitro, but had no significant reduction of tumor volume after 4 weeks of cancer cell xenografting in vivo when compared to the scramble control. Interestingly, Crk II-shRNA cancer cells showed a greatly reduced level of insulin-like growth factor 1 receptor (IGF-1R) and decreased signaling of the IGF-1R/PI3K/Akt axis upon IGF-1 ligand stimulation. A close interaction between Crk II and IGF-1R was demonstrated upon co-immunoprecipitation of IGF-1R with Crk II protein. Further, treatment of cells with either proteosomal degradation or protein synthesis inhibitor showed higher proportion of ubiquitin-associated IGF-1R and faster degradation of IGF-1R in Crk II-shRNA cells in comparison with that in the control cancer cells. Taken together, these data suggest that Crk II plays an important role in the regulation of IGF-1R protein stability and affects downstream of IGF-1R signaling pathways. Therefore, targeting Crk-II can block IGF-1R growth signaling and suppress cancer cell invasion and progression.