A selective alpha1D-adrenoreceptor antagonist inhibits human prostate cancer cell proliferation and motility "in vitro"

The progression of prostate cancer (PC) to a metastatic hormone refractory disease is the major contributor to the overall cancer mortality in men, mainly because the conventional therapies are generally ineffective at this stage. Thus, other therapeutic options are needed as alternatives or in addition to the classic approaches to prevent or delay tumor progression. Catecholamines participate to the control of prostate cell functions by the activation of alpha1-adrenoreceptors (alpha1-AR) and increased sympathetic activity has been linked to PC development and evolution. Molecular and pharmacological studies identified three alpha1-AR subtypes (A, B and D), which differ in tissue distribution, cell signaling, pharmacology and physiological role. Within the prostate, alpha1A-ARs mainly control stromal cell functions, while alpha1B- and alpha1D- subtypes seem to modulate glandular epithelial cell growth. The possible direct contribution of alpha1D-ARs in tumor biology is supported by their overexpression in PC. The studies here presented investigate the "in vitro" antitumor action of A175, a selective alpha1D-AR antagonist we have recently obtained by modifying the potent, but not subtype-selective alpha1-AR antagonist (S)-WB4101, in the hormone-refractory PC3 and DU145 PC cell lines. The results indicate that A175 has an alpha1D-AR-mediated significant and dose-dependent antiproliferative action that possibly involves the induction of G0/G1 cell cycle arrest, but not apoptosis. In addition, A175 reduces cell migration and adhesiveness to culture plates. In conclusion, our work clarified some cellular aspects promoted by alpha1D-AR activity modulation and supports a further pharmacological approach in the cure of hormone-refractory PC, by targeting specifically this AR subtype

PBCs environmental pollution and epigenome: a new role for androgen receptor-dependent modulation?

Epigenome is an important target of environmental effects, modulating disease susceptibility during the whole life. In our previous studies we have demonstrated that prenatal exposure to polychlorinated biphenyls (PCBs), an important class of endocrine disruptors, alters in liver some histone post-translational modifications (H3K4me3/H4K16Ac) and the expression of the corresponding modulating enzymes (Jarid1b/SirtT1) and reduces the androgen receptor (AR) levels1. Furthermore, it is known that steroid receptors could act also as co-regulator of histone modification enzymes. It is also remarkable that AR and Jarid1b (demethylating enzyme) interact each other and that Jarid1b is able to potentiate the transcriptional activity of AR2. The AR down-regulation, shown by our data, is not directly related to the reduction of H3K4me3 levels in the AR promoter, as our ChIP experiments have indicated1. The observed AR reduction might be related to the down-regulation of AR induced by its own activation3. The aim of this work was to characterize the complex scenario of AR involvement in regulating histone modifications. First of all, we investigated if a reconstituted mixture of PCBs is able to modulate AR transcriptional activity; we observed that PCBs are able to induce AR mediated transcription in a dose dependent way and that Jarid1b presence potentiates the PCB effect on AR transcriptional activity. Furthermore we showed that the localization of PCB- activated AR is influenced by Jarid1b presence. Finally, using AR promoter fused with a reporter gene, we found that PCBs are able to auto-downregulate AR, especially at the presence of Jarid1b. In conclusion it is possible to hypothesize that AR modulation exerted by PCB pass through chromatin structure remodelling. It remains to clarify if AR is involved in mediating PCB induced disruption of Jarid1b

Effects of Developmental Exposure to 2,2′,4,4′,5-Pentabromodiphenyl Ether (PBDE-99) on Sex Steroids, Sexual Development, and Sexually Dimorphic Behavior in Rats

Increasing concentrations of polybrominated flame retardants, including polybrominated diphenyl ethers (PBDEs), in breast milk cause concern about possible developmental effects in nursed babies. Because previous studies in rats have indicated effects on sex steroids and sexually dimorphic behavior after maternal exposure to polychlorinated biphenyls (PCBs), our goal in the present study was to determine if developmental exposure to 2,2′,4,4′,5-pentabromodiphenyl ether (PBDE-99) induces similar endocrine-mediated effects. Pregnant rats were exposed to vehicle or PBDE-99 (1 or 10 mg/kg body weight, daily during gestational days 10–18). For comparison, we also included a group exposed to the technical PCB mixture Aroclor 1254 (30 mg/kg body weight, daily). PBDE exposure resulted in pronounced decreases in circulating sex steroids in male offspring at weaning and in adulthood. Female offspring were less affected. Anogenital distance was reduced in male offspring. Puberty onset was delayed in female offspring at the higher dose level, whereas a slight acceleration was detected in low-dose males. The number of primordial/primary ovarian follicles was reduced in females at the lower dose, whereas decline of secondary follicles was more pronounced at the higher dose. Sweet preference was dose-dependently increased in PBDE-exposed adult males, indicating a feminization of this sexually dimorphic behavior. Aroclor 1254 did not alter sweet preference and numbers of primordial/primary and secondary follicles but it did affect steroid concentrations in males and sexual development in both sexes. PBDE concentrations in tissues of dams and offspring were highest on gestational day 19. These results support the hypothesis that PBDEs are endocrine-active compounds and interfere with sexual development and sexually dimorphic behavior

Transient expression of the 5alpha-reductase type 2 isozyme in the rat brain in late fetal and early postnatal life

The enzyme 5alpha-reductase plays a key role on several brain functions controlling the formation of anxiolytic/anesthetic steroids derived from progesterone and deoxycorticosterone, the conversion of testosterone to dihydrotestosterone, and the removal of excess of potentially neurotoxic steroids. Two 5alpha-reductase isoforms have been cloned: 5alpha-reductase type 1 is widely distributed in the body, and 5alpha-reductase type 2 is confined to androgen-dependent structures. In this study, the gene expression of the two 5alpha-reductase isozymes has been analyzed in fetal, postnatal, and adult rat brains by RT-PCR followed by Southern analysis. 5Alpha-reductase type 1 messenger RNA is always detectable in the rat brain [from gestational day 14 (GD14) to adulthood]. 5Alpha-reductase type 2 messenger RNA expression is undetectable on GD14, increases after GD18, peaks on postnatal day 2, then decreases gradually, becoming low in adulthood. This pattern of expression appears to be correlated with the rate of production of testosterone by the testis. The possible control by androgens of gene expression of the two isozymes has been studied in brain tissues of animals exposed in utero to the androgen antagonist flutamide; the sex of the animals was determined by genetic sex screening of the SRY gene located on the Y-chromosome. In the brain of male embryos, flutamide treatment inhibited the expression of 5alpha-reductase type 2; this effect was much less pronounced in females. Moreover, 5alpha-reductase type 2 gene expression in cultured hypothalamic neurons is highly induced by testosterone and by the phorbol ester 12-O-tetradecanoyl-phorbol-13 acetate. The transient, androgen-regulated, expression of 5alpha-reductase type 2 overlaps the critical period of development, which may be important for sexual differentiation of the brain and for the formation of anxiolytic/anesthetic steroids involved in the stress responses associated with parturition

Epigenome and Environment: effects of a PCB exposure on epigenome during early development in the rat

Introduction: Epigenetic represents the genome programming to express appropriate set of genes in a temporal and spatial way during life. Epigenetic signature is dynamic and responsive to environment, especially in critical periods of development. Deregulation of epigenetic mechanisms may be responsible for incorrect transcription patterns, resulting in stable modifications of key physiological processes. Exposure to PCBs (polychlorinated biphenyls) during gestation and lactation, modifies the gene expression of important mediators of sex steroid action and of their receptors. Androgen receptor (AR) could also act as co-regulator of histone modifications enzymes. Aim: to evaluate whether prenatal administration of a reconstituted PCB mixture is able to influence epigenome Results: PCBs reduce histone post-translational modifications (H3K4me3 and H4K16Ac) in a dimorphic way, possibly as the result of a decreased expression of the histone modification enzymes, Jarid1b and SirtT1. AR gene and protein expression is also reduced, especially in females. ChIP performed against H3K4me3 shows a correlation between the expression of some genes involved in embryonic differentiation (e.g., TSHR and GabaRr3) and histone post- translational gene modifications. Conclusion: the influence of PCB exposure during differentiation on epigenome is dimorphic and affects chromatin packaging, probably through an impairment of interaction between androgen receptor and histone modification enzymes

In vitro chronic administration of ERbeta selective ligands and prostate cancer cell growth : hypotheses on the selective role of 3beta-adiol in AR-positive RV1 cells

Prostate cancer (PC) progression from androgen-dependent (AD) to castration-resistant (CR) disease is a process caused by modifications of different signal transduction pathways within tumor microenvironment. Reducing cell proliferation, estrogen receptor beta (ERbeta) is emerging as a potential target in PC chemoprevention. Among the known selective ERbeta ligands, 3beta-Adiol, the endogenous ligand in the prostate, has been proved to counteract PC progression. This study compares the effects of chronic exposure (1-12 weeks) to different ERbeta selective ligands (DPN, 8beta-VE2, 3beta-Adiol) on proliferation of human androgen-responsive CWR22Rv1 cells, representing an intermediate phenotype between the AD- and CR-PC. 3beta-Adiol (10 nM) is the sole ligand decreasing cell proliferation and increasing p21 levels. In vitro transcriptional activity assays were performed to elucidate different behavior between 3beta-Adiol and the other ligands; in these experiments the endogenous and the main ERbeta subtype activation were considered. It is concluded that ERbeta activation has positive effects also in androgen-responsive PC. The underlying mechanisms are still to be clarified and may include the interplay among different ERbeta subtypes and the specific PC microenvironment. ERbeta agonists might be useful in counteracting PC progression, although the final outcome may depend upon the molecular pattern specific to each PC lesion

Novel Molecular Targets of Azadirachta indica Associated with Inhibition of Tumor Growth in Prostate Cancer

Advanced prostate cancer has significant long-term morbidity, and there is a growing interest in alternative and complimentary forms of therapy that will improve the outcomes of patients. Azadirachta indica (common name: neem) contains multiple active compounds that have potent anti-inflammatory and anticancer properties. The present study investigates the novel targets of the anticancer activity of ethanol extract of neem leaves (EENL) in vitro and evaluates the in vivo efficacy in the prostate cancer models. Analysis of the components in the EENL by mass spectrometry suggests the presence of 2′,3′-dehydrosalannol, 6-desacetyl nimbinene, and nimolinone. Treatment of C4-2B and PC-3M-luc2 prostate cancer cells with EENL inhibited the cell proliferation. Genome-wide expression profiling, using oligonucleotide microarrays, revealed genes differentially expressed with EENL treatment in prostate cancer cells. Functional analysis unveiled that most of the up-regulated genes were associated with cell death, and drug metabolism, and the down-regulated genes were associated with cell cycle, DNA replication, recombination, and repair functions. Quantitative PCR confirmed significant up-regulation of 40 genes and immunoblotting revealed increase in the protein expression levels of HMOX1, AKR1C2, AKR1C3, and AKR1B10. EENL treatment inhibited the growth of C4-2B and PC-3M-luc2 prostate cancer xenografts in nude mice. The suppression of tumor growth is associated with the formation of hyalinized fibrous tumor tissue and the induction of cell death by apoptosis. These results suggest that EENL-containing natural bioactive compounds could have potent anticancer property and the regulation of multiple cellular pathways could exert pleiotrophic effects in prevention and treatment of prostate cancer

Gestational Exposure to Low Dose Bisphenol A Alters Social Behavior in Juvenile Mice

Bisphenol A (BPA) is a man-made compound used to make polycarbonate plastics and epoxy resins; public health concerns have been fueled by findings that BPA exposure can reduce sex differences in brain and some behaviors. We asked if a low BPA dose, within the range measured in humans, ingested during pregnancy, would affect social behaviors in prepubertal mice. We noted sex differences in social interactions whereby females spent more time sitting side-by-side, while males engaged in more exploring and sitting alone. In addition BPA increased display of nose-to-nose contacts, play solicitations and approaches in both sexes. Interactions between sex and diet were found for self grooming, social interactions while sitting side-by-side and following the other mouse. In all these cases interactions were produced by differences between control and BPA females. We examined brains from embryos during late gestation to determine if gene expression differences might be correlated with some of the sexually dimorphic or BPA affected behaviors we observed. Because BPA treatments ended at birth we took the brains during embryogenesis to increase the probability of discovering BPA mediated effects. We also selected this embryonic age (E18.5) because it coincides with the onset of sexual differentiation of the brain. Interestingly, mRNA for the glutamate transporter, Slc1a1, was enhanced by exposure to BPA in female brains. Also we noted that BPA changed the expression of two of the three DNA methyltransferase genes, Dnmt1 and Dnmt3a. We propose that BPA affects DNA methylation of Sc1a1 during neural development. Sex differences in juvenile social interactions are affected by BPA and in particular this compound modifies behavior in females