Regional localization of activin-βA,activin-βC,follistatin, proliferation,and apoptosis in adult and developing mouse prostate ducts

Activins and inhibins, members of the TGF-β superfamily, are growth and differentiation factors involved in the regulation of several biological processes, including reproduction, development, and fertility. Previous studies have shown that the activin-β_A subunit plays a pivotal role in prostate development. Activin-A inhibits branching morphogenesis in the developing prostate, and its expression is associated with increased apoptosis in the adult prostate. Follistatin, a structurally unrelated protein to activins, is an antagonist of activin-A. A balance between endogenous activin-A and follistatin is required to maintain prostatic branching morphogenesis. Deregulation of this balance leads to branching inhibition or excessive branching and increased maturation of the stroma surrounding the differentiating epithelial ducts. Recent work identified another member of the TGF-β superfamily, the activin-β_C subunit, as a novel antagonist of activin-A. Over-expression of activin-C (β_C-β_C) alters prostate homeostasis, by interfering with the activin-A signaling. The current study characterized the spatiotemporal localization of activin-A, activin-C and follistatin in the adult and developing mouse prostate using immunohistochemical analysis. Results showed activin-C and follistatin are differentially expressed during prostate development and suggested that the antagonistic property of follistatin is secondary to the action of activin-C. In conclusion, the present study provides evidence to support a role of activin-C in prostate development and provides new insights in the spatiotemporal localization of activins and their antagonists during mouse prostate development.     

Interstitial cells of Cajal mediate excitatory sympathetic neurotransmission in guinea pig prostate

Morphological and functional studies have confirmed that interstitial cells of Cajal (ICCs) are involved in many enteric motor neurotransmission pathways. Recent investigations have demonstrated that human and guinea pig prostate glands possess a distinct cell type with morphological and immunological similarities to ICCs. These prostate ICCs have a close relationship with nerve bundles and smooth muscle cells. Prostate smooth muscle tone is largely induced by stimulation from the sympathetic nervous system, which releases excitatory norepinephrine (NE) to act on the α1-adrenoceptor. We have performed morphological and functional experiments to determine the role of ICCs in sympathetic neurotransmission in the guinea pig prostate based on the hypothesis that prostate ICCs act as mediators of sympathetic neurotransmission. Immunohistochemistry revealed many close points of contact between ICCs and sympathetic nerve bundles and smooth muscle cells. Double-labeled sections revealed that α1-adrenoceptor and the gap junction protein connexin 43 were expressed in prostate ICCs. Surprisingly, prostate ICCs co-expressed tyrosine hydroxylase and dopamine β-hydroxylase, two markers of sympathetic neurons. Functionally, the application of NE evoked a large single inward current in isolated prostate ICCs in a dose-dependent manner. The inward current evoked by NE was mediated via the activation of α1-adrenoceptors, because it was abolished by the non-specific α-adrenoceptor antagonist, phentolamine and the specific α1-adrenoceptor antagonist, prazosin. Thus, ICCs in the guinea pig prostate are target cells for prostate sympathetic nerves and possess the morphological and functional characteristics required to mediate sympathetic signals.     

Administration of an LHRH-antagonist to male mice: effects on in vivo secretion of hormones and on the growth of a transplantable human prostatic carcinoma

The potent luteinizing hormone releasing hormone (LHRH) antagonist [N-Ac-D-p-Cl-Phe1,2,D-Trp3,D-Arg6,D-Ala10]-LHRH was chronically administered to male nude mice bearing the transplantable human hormone-dependent prostatic adenocarcinoma PC-82. Treatment of tumor-bearing male mice with a daily dose of 100 micrograms (4 mg/kg b w.) for 21 days did not significantly affect the growth of the PC-82 tumor tissue, or the weights of ventral prostate, seminal vesicles and testes. At 24 hours after the last dose of the antagonist the mean plasma-testosterone (T) value in these animals was not different from the control level. Administration of similar doses of the antagonist to intact normal immunocompetent male mice significantly reduced plasma LH concentrations and suppressed plasma-T to near-castrate levels, when blood was taken 2 hours after the last injection. At 24 hours after the last dose, however, plasma concentrations of LH and T had returned to control levels. This time-dependent pattern of T suppression by the antagonist was confirmed by a time-course experiment in animals receiving a single dose of the compound. These data demonstrate that a daily high dose of this antagonist cannot effectively suppress plasma-T in male mice. Therefore, the mouse may not be a suitable model for the investigation of the "castration-like" effect of LHRH-antagonists on androgen-dependent prostate xenografts.     

Similarities and Distinctions in Actions of Surface-Directed and Classic Androgen Receptor Antagonists

The androgen receptor (AR) surface-directed antagonist MJC13 inhibits AR function and proliferation of prostate cancer (PC) cells. These effects are related to arrest of an AR/chaperone complex in the cytoplasm. Here, we compared MJC13 and classic AR antagonists such as flutamide and bicalutamide. Microarray analysis and confirmatory qRT-PCR reveals that MJC13 and flutamide inhibit dihydrotestosterone (DHT)-dependent genes in LNCaP PC cells. Both compounds are equally effective on a genome wide basis and as effective as second generation AR antagonists (MDV3100, ARN-509) at selected genes. MJC13 inhibits AR binding to the prostate specific antigen (PSA) promoter more strongly than flutamide, consistent with different mechanisms of action. Examination of efficacy of MJC13 in conditions that reflect aspects castrate resistant prostate cancer (CRPC) reveals that it inhibits flutamide activation of an AR mutant (ART877A) that emerges during flutamide withdrawal syndrome, but displays greatly restricted gene-specific activity in 22Rv1 cells that express a constitutively active truncated AR and is inactive against glucocorticoid receptor (GR), which can co-opt androgen-dependent signaling networks in CRPC. Importantly, MJC13 inhibits AR interactions with SRC2 and β-catenin in the nucleus and, unlike flutamide, strongly inhibits amplification of AR activity obtained with transfected SRC2 and β-catenin. MJC13 also inhibits DHT and β-catenin-enhanced cell division in LNCaP cells. Thus, a surface-directed antagonist can block AR activity in some conditions in which a classic antagonist fails and may display utility in particular forms of CRPC.     

非那雄胺联合M受体拮抗剂治疗前列腺增生合并膀胱过度活动症的疗效

目的:探讨非那雄胺联合M受体拮抗剂对前列腺增生合并膀胱过度活动症临床疗效的影响。方法:回顾性研究我院前列腺增生合并膀胱过度活动症的患者60例,随机分为实验组和对照组,每组30例。对照组患者给予M受体拮抗剂,实验组患者在对照组的基础上给予非那雄胺。观察并比较治疗前后两组患者国际前列腺症状评分(IPSS)、膀胱过度活动症评分(OABSS)、残余尿量、最大尿流率、尿急次数以及治疗期间不良反应。结果:与对照组相比,实验组患者IPSS及OABSS较低(P0.05);残余尿量较少,最大尿流率较大,尿急次数较少(P0.05);不良反应发生率较低(P0.05)。结论:非那雄胺联合M受体拮抗剂能够降低前列腺增生合并膀胱过度活动症的不良反应发生率,提高临床疗效,值得在临床上推广应用。     

Design,syntheses, and characterization of piperazine based chemokine receptor CCR5 antagonists as anti prostate cancer agents

Chemokine receptor CCR5 plays an important role in the pro-inflammatory environment that aids in the proliferation of prostate cancer cells. Previously, a series of CCR5 antagonists containing a piperidine ring core skeleton were designed based upon the proposed CCR5 antagonist pharmacophore from molecular modeling studies. The developed CCR5 antagonists were able to antagonize CCR5 at a micromolar level and inhibit the proliferation of metastatic prostate cancer cell lines. In order to further explore the structure–activity-relationship of the pharmacophore identified, the molecular scaffold was expanded to contain a piperazine ring as the core. A number of compounds that were synthesized showed promising anti prostate cancer activity and reasonable cytotoxicity profiles based on the biological characterization.     

Anibamine,a natural product CCR5 antagonist,as a novel lead for the development of anti-prostate cancer agents

Accumulating evidence indicates that the chemokine receptor CCR5 and the chemokine CCL5 may be involved in the proliferation and metastasis of prostate cancer. Consequently, chemokine receptor CCR5 antagonists could potentially act as anti-prostate cancer agents. As the first natural product CCR5 antagonist, anibamine provides a novel chemical structural skeleton compared with other known antagonists identified through high-throughput screening. Our studies demonstrate that anibamine produces significant inhibition of prostate cancer cell proliferation at micromolar to submicromolar concentrations as well as suppressing adhesion and invasion of the highly metastatic M12 prostate cancer cell line. Preliminary in vivo studies indicate that anibamine also inhibits prostate tumor growth in mice. These findings indicate that anibamine may prove to be a novel lead compound for the development of prostate cancer therapeutic agents.     

Up-regulation of Bcl-2 is required for the progression of prostate cancer cells from an androgen-dependent to an androgen-independent growth stage

Bcl-2 is an anti-apoptotic oncoprotein and its protein levels are inversely correlated with prognosis in many cancers.However, the role of Bcl-2 in the progression of prostate cancer is not clear. Here we report that Bcl-2 is required for theprogression of LNCaP prostate cancer cells from an androgen-dependent to an androgen-independent growth stage. ThemRNA and protein levels of Bcl-2 are significantly increased in androgen-independent prostate cancer cells. shRNA-medi-ated gene silencing of Bcl-2 in androgen-independent prostate cancer cells promotes UV-induced apoptosis and suppressesthe growth of prostate tumors in vivo. Growing androgen-dependent cells under androgen-deprivation conditions resultsin formation of androgen-independent colonies; and the transition from androgen-dependent to androgen-independentgrowth is blocked by ectopic expression of the Bcl-2 antagonist Bax or Bcl-2 shRNA. Thus, our results demonstratethat Bcl-2 is not only critical for the survival of androgen-independent prostate cancer cells, but is also required for theprogression of prostate cancer cells from an androgen-dependent to an androgen-independent growth stage.     

CXCR4 expression in prostate cancer progenitor cells

Tumor progenitor cells represent a population of drug-resistant cells that can survive conventional chemotherapy and lead to tumor relapse. However, little is known of the role of tumor progenitors in prostate cancer metastasis. The studies reported herein show that the CXCR4/CXCL12 axis, a key regulator of tumor dissemination, plays a role in the maintenance of prostate cancer stem-like cells. The CXCL4/CXCR12 pathway is activated in the CD44(+)/CD133(+) prostate progenitor population and affects differentiation potential, cell adhesion, clonal growth and tumorigenicity. Furthermore, prostate tumor xenograft studies in mice showed that a combination of the CXCR4 receptor antagonist AMD3100, which targets prostate cancer stem-like cells, and the conventional chemotherapeutic drug Taxotere, which targets the bulk tumor, is significantly more effective in eradicating tumors as compared to monotherapy.     

Pharmacological characterization of [3H]-JTH-601, a novel alpha1-adrenoceptor antagonist: binding to recombinant human alpha1-adrenoceptors and human prostates

Several alpha1-adrenoceptor (AR) selective antagonists are now widely used to improve lower urinary tract symptoms in benign prostatic hyperplasia patients. However, these drugs often result in orthostatic hypotension, because of their poor uroselectivity; the blockade of alpha1-AR not only in prostate but also in vasculature. Here we have investigated uroselectivity of JTH-601, a newly developed antagonist, in radioligand binding experiment using recombinant human alpha1-AR subtypes and human prostate. In saturation experiments, [3H]-JTH-601 showed subtype selectivity: high affinity to alpha1a-AR (pKd; 9.88+/-0.09), lower affinity to alpha1b-AR (pKd; 8.96+/-0.17) and no specific binding at concentrations up to 3000 pM to alpha1d-AR. In competition experiments, JTH-601 and its metabolic compound (JTH-601-G1) also showed alpha1a-AR selectivity, exhibiting approximately 5 times higher affinity for alpha1a-AR than for alpha1b-AR, 10 to 20 times higher affinity than for alpha1d-AR, respectively. [3H]-JTH-601 also bound to human prostate membranes in monophasic manner with high affinity constant (pKd; 9.89+/-0.12, Bmax=123.6+/-16 fmol/mg protein). JTH-601 is a unique alpha1-AR antagonist that shows high affinity and selectivity for human recombinant alpha1a- and human prostate. This new compound is useful for understanding alpha1-AR pharmacology and may have a therapeutic value.     

Calpain activation through galectin-3 inhibition sensitizes prostate cancer cells to cisplatin treatment

Prostate cancer will develop chemoresistance following a period of chemotherapy. This is due, in part, to the acquisition of antiapoptotic properties by the cancer cells and, therefore, development of novel strategies for treatment is of critical need. Here, we attempt to clarify the role of the antiapoptotic molecule galectin-3 in prostate cancer cells using siRNA and antagonist approaches. The data showed that Gal-3 inhibition by siRNA or its antagonist GCS-100/modified citrus pectin (MCP) increased cisplatin-induced apoptosis of PC3 cells. Recent studies have indicated that cisplatin-induced apoptosis may be mediated by calpain, a calcium-dependent protease, as its activation leads to cleavage of androgen receptor into an androgen-independent isoform in prostate cancer cells. Thus, we examined whether calpain activation is associated with the Gal-3 function of regulating apoptosis. Here, we report that Gal-3 inhibition by siRNA or GCS-100/MCP enhances calpain activation, whereas Gal-3 overexpression inhibits it. Inhibition of calpain using its inhibitor and/or siRNA attenuated the proapoptotic effect of Gal-3 inhibition, suggesting that calpain activation may be a novel mechanism for the proapoptotic effect of Gal-3 inhibition. Thus, a paradigm shift for treating prostate cancer is suggested whereby a combination of a non-toxic anti-Gal-3 drug together with a toxic chemotherapeutic agent could serve as a novel therapeutic modality for chemoresistant prostate cancers.     

17β-Estradiol regulates oxidative stress in prostate cancer cell lines according to ERalpha/ERbeta ratio

Estrogen action is mediated by the two receptor isoforms: estrogen receptor alpha and beta. Both receptors are expressed in human prostate tissue and have different action profiles. ERalpha is positively correlated with the malignancy of prostate cancer, while ERbeta may protect against abnormal prostate cell growth. 17β-Estradiol (E2), at least in part, induces cancerous transformations by causing deleterious mutations through the formation of reactive oxygen species (ROS). The aim was to study the effect of E2 on oxidative stress and the expression of uncoupling proteins (UCPs) and antioxidant enzymes in several prostate cancer cell lines with different ERalpha/ERbeta ratios. The cell prostate lines with a lower ERalpha/ERbeta ratio had lower oxidative stress, which could be partially explained by the increased expression of antioxidant enzymes and UCPs. Moreover, the action of E2 on the expression of antioxidant enzymes and UCPs was dual and dependent on the ERalpha/ERbeta ratio. Treatments with 0.1 nM E2 in cell lines with high ERalpha/ERbeta ratio produced a decrease in antioxidant enzymes and UCPs levels, with an increase in ROS production. These effects disappeared when the treatment was done in the presence of an ERalpha antagonist (MPP). In the cell lines with greatest levels of ERbeta and the lowest ERalpha/ERbeta ratio, E2 treatment caused the up-regulation of antioxidant enzymes and UCPs with a look-up decrease in ROS production. These effects were reversed when the cells were treated with E2 in the presence of an ERbeta antagonist (R,R-THC). On the whole, our results suggest a dual E2 effect; increasing or decreasing oxidative stress in part by modulation of UCPs and antioxidant enzymes according to the abundance ERbeta and ERalpha/ERbeta ratio in prostate cancer cell lines.     

Aryl hydrocarbon receptor signaling involved in the invasiveness of LNCaP cells

There is now mounting evidence that the aryl hydrocarbon receptor (AhR) plays an important role in physiologic responses such as development, cell cycle regulation, immune function and also malignant transformation in various tissues. The strong nuclear AhR expression is observed in the invasive phenotype, and an elevated nuclear AhR expression is associated with a poor prognosis of human prostate cancer. On the other hand, there are conflicting results that the AhR deficiency results in increased susceptibility to prostate tumors in mouse model. In the present study, we investigated AhR expression and its role in the growth and invasiveness of human prostate cancer cells. The AhR protein expression was detected in prostate cancer cell lines and human prostate cancer tissues. A small interfering RNA targeting AhR, constitutive active AhR expression vector, and AhR agonist and antagonist were used to moderate its expression and signaling. The induction of AhR signaling attenuated invasiveness of prostate cancer cells without affecting the cellular growth rate. These results suggest that AhR signaling in prostate cancer cells facilitates invasion of these cells, and modulation with this signaling can be a potential therapeutic target of invasive tumors.     

Epigenetic regulation of Wnt signaling pathway in urological cancer

Constitutive activation of the Wnt signaling pathway is a common feature of solid tumors and contributes to uncontrolled cell-growth and impaired differentiation. We hypothesized that gene silencing mediated through aberrant promoter methylation of upstream Wnt antagonist genes might result in β-catenin accumulation, resulting in constitutive Wnt activation. Wnt antagonist genes (SFRP1, WIF1, APC and CDH1) and CTNNB1 promoter methylation was examined in genomic DNA extracted from 12 urological cancer cell lines and correlated with CTNNB1 mRNA expression. Promoter methylation status was then assessed in 36 BCa, 30 PCa, 31 RCT, and normal bladder mucosa (15), prostate (10) and renal (5) tissue samples. Finally, CTNNB1 mRNA relative expression levels were correlated with Wnt antagonist gene methylation status in RCT. Methylation was found in at least one Wnt antagonist gene and the CTNNB1 promoter was unmethylated in all cancer cell lines tested. When gene methylation levels were compared between cancer cell lines with high and low CTNNB1 mRNA expression, a trend was found for increased CDH1 promoter methylation levels in the former. BCa and PCa tumors demonstrated high frequency of promoter methylation at all tested genes. In RCT, CTNNB1 was unmethylated in all cases and the overall frequency of promoter methylation at the remainder genes was lower. Interestingly, median CTNNB1 mRNA expression levels were significantly higher in RCTs methylated in at least one Wnt antagonist gene promoter. We concluded that epigenetic deregulation of Wnt pathway inhibitors may contribute to aberrant activation of Wnt signaling pathway in bladder, prostate and renal tumors.