Human Prostatic Cells Research Articles

P66 - Androgens and TRIBs pseudokinases in modulating human prostate cells fate

P66 - Androgens and TRIBs pseudokinases in modulating human prostate cells fate

A basal-enriched microRNA is required for prostate tumorigenesis in a Pten knockout mouse model.

MicroRNAs (miRNAs) play important roles in prostate cancer development. However, it remains unclear how individual miRNAs contribute to the initiation and progression of prostate cancer. Here we show that a basal layer-enriched miRNA is required for prostate tumorigenesis. We identify miR-205 as the most highly expressed miRNA and enriched in the basal cells of the prostate. Although miR-205 is not required for normal prostate development and homeostasis, genetic deletion of miR-205 in a Pten null tumor model significantly compromises tumor progression and does not promote metastasis. In Pten null basal cells, loss of miR-205 attenuates pAkt levels and promotes cellular senescence. Furthermore, although overexpression of miR-205 in prostate cancer cells with luminal phenotypes inhibits cell growth in both human and mouse, miR-205 has a minimal effect on the growth of a normal human prostate cell line. Taken together, we have provided genetic evidence for a requirement of miR-205 in the progression of Pten null-induced prostate cancer.

Sweet Cherry Extract Targets the Hallmarks of Cancer in Prostate Cells: Diminished Viability, Increased Apoptosis and Suppressed Glycolytic Metabolism

The present work evaluated the anticancer properties of sweet cherry (Prunus avium) extract on human prostate cells. Several sweet cherry cultivars from Fundão (Portugal) were methanol-extracted and their phytochemical composition characterized. The Saco “late harvest” extract was highly-enriched in anthocyanins and selected for use in biological assays. Non-neoplastic (PNT1A) and neoplastic (LNCaP and PC3) human prostate cells were treated with 0–2,000 μg/ml of extract for 48–96 h. Cell viability was evaluated by the MTT assay. Apoptosis, oxidative stress, and glycolytic metabolism were assessed by Western blotting and enzymatic assays. Glucose consumption and lactate production were measured spectrophotometrically. Saco cherry extract diminished the viability of neoplastic and non-neoplastic cells, whereas enhancing apoptosis in LNCaP. Cherry extract-treatment also diminished oxidative damage and suppressed glycolytic metabolism in LNCaP cells. These findings widened the knowledge on the mechanisms by which cherry extract modulate cell physiology, demonstrating their broad action over the hallmarks of cancer.

The comparison between force volume and peakforce quantitative nanomechanical mode of atomic force microscope in detecting cell's mechanical properties.

Atomic force microscope (AFM) has been widely used in the biological field owing to its high sensitivity (subnanonewton), high spatial resolution (nanometer), and adaptability to physiological environments. Nowadays, force volume (FV) and peakforce quantitative nanomechanical (QNM) are two distinct modes of AFM used in biomechanical research. However, numerous studies have revealed an extremely confusing phenomenon that FV mode has a significant difference with QNM in determining the mechanical properties of the same samples. In this article, for the case of human benign prostatic hyperplasia cells (BPH) and two cancerous prostate cells with different grades of malignancy (PC3 and DU145), the differences were compared between FV and QNM modes in detecting mechanical properties. The results show measured Young's modulus of the same cells in FV mode was much lower than that obtained by QNM mode. Combining experimental results with working principles of two modes, it is indicated that surface adhesion is highly suspected to be a critical factor resulting in the measurement difference between two modes. To further confirm this conjecture, various weight ratios of polydimethylsiloxane (PDMS) were assessed by two modes, respectively. The results show that the difference of Young's modulus measured by two modes increases with the surface adhesion of PDMS, confirming that adhesion is one of the significant elements that lead to the measurement difference between FV and QNM modes.

Loss of the tumor suppressor, Tp53, enhances the androgen receptor-mediated oncogenic transformation and tumor development in the mouse prostate.

Recent genome analysis of human prostate cancers demonstrated that both AR gene amplification and TP53 mutation are among the most frequently observed alterations in advanced prostate cancer. However, the biological role of these dual genetic alterations in prostate tumorigenesis is largely unknown. In addition, there are no biologically relevant models that can be used to assess the molecular mechanisms for these genetic abnormalities. Here, we report a novel mouse model, in which elevated transgenic AR expression and Trp53 deletion occur simultaneously in mouse prostatic epithelium to mimic human prostate cancer cells. These compound mice developed an earlier onset of high-grade prostatic intraepithelial neoplasia and accelerated prostate tumors in comparison to mice harboring only the AR transgene. Histological analysis showed prostatic sarcomatoid and basaloid carcinomas with massive squamous differentiation in the above compound mice. RNA-sequencing analyses identified a robust enrichment of the signature genes for human prostatic basal cell carcinomas in the above prostate tumors. Master Regulator Analysis revealed SOX2 as a transcriptional regulator in prostatic basal cell tumors. Elevated expression of SOX2 and its downstream target genes were detected in prostatic tumors of the compound mice. Chromatin immunoprecipitation analyses implicate a co-regulatory role of AR and SOX2 in the expression of prostatic basal cell signature genes. Our data demonstrate a critical role of SOX2 in prostate tumorigenesis and provide mechanistic insight into prostate tumor aggressiveness and progression mediated by aberrant AR and p53 signaling pathways.

The effects of Cernitin® on inflammatory parameters and benign prostatic hyperplasia: An in vitro study.

The pollen extract Cernitin® is widely used for treatment of benign prostatic hyperplasia (BPH) and non-bacterial chronin prostatitis. However, little is known about the underlying molecular mechanisms to explain the clinical effects of Cernitin®. In this study, we sought to investigate the cellular mechanisms by which Cernitin® induces its effects on human prostatic cell lines BPH-1 and WPMY-1 and primary human peripheral blood mononuclear cells (hPBMCs) in vitro. We examined the effects of Cernitin® formulas T60 and GBX on the protein expression, proliferation, and cytokines production. Results revealed that Cernitin® upregulated antiinflammatory cytokine interleukin (IL)-10 and its receptors IL-10RA and IL-10B in addition to the upregulation of tumour necrosis factor-related apoptosis-inducing ligand in hPBMC. Interestingly, the levels of proinflammatory cytokines IL-6 and IL-8 were also increased. Furthermore, Cernitin® had significantly increased the level of IL-10 in BPH-1 and WPMY-1 cells. The level of IL-6 was also significantly increased in these cells although both T60 and GBX inhibited STAT-3 phosphorylation. Moreover, Cernitin® formulas had significantly reduced androgen receptor and prostate-specific antigen protein expression in stromal cells (p<.05). Treatment with GBX and T60 had significantly inhibited proliferation of BPH (p<.001) and stromal cells (p<.05), in a dose-dependent manner. Taken together, treatment with Cernitin® showed to regulate cytokines level in both prostatic cell lines and hPBMCs and it was associated with decreased androgen receptor and prostate-specific antigen levels WPMY-1 cells.

Detection of a prostate cancer cell line using a bioluminescent affiprobe: An attempt to develop a new molecular probe for ex vivo studies

Molecular probes have become an important tool to decipher cell and molecular events, as well as contributing to clinical diagnosis. In this study, we report on a novel bioluminescence affiprobe and evaluate its functionality by monitoring HER2 expression in the prostate cancer cells. The proposed bioluminescence affiprobe consists of a binding domain, an HER2 specific affibody molecule and a bioluminescent domain, recombinant Renilla luciferase. The experiments indicate that bioluminescence affiprobe can serve as a reliable and user-friendly probe for the detection of human HER2 positive prostate cells as well as ex vivo based detection of HER2-positive human prostate cancer specimens using luminometeric-based assays.

Abstract 1015: Molecular dissection of androgen receptor signaling in prostatic endothelial, epithelial, and stromal cells

Abstract Introduction and objectives: Androgen receptor (AR) is involved in development prostate cancer (CaP), and is a primary target for treatment of CaP. Androgen deprivation therapy (ADT) inhibits AR signaling by reducing AR ligands and/or blocking AR-ligand binding. ADT induces apoptosis of both endothelial cells and epithelial cells of human prostate, with apoptosis of endothelial cells preceding the apoptosis of epithelial cells. Further, the AR response to stimulation by androgen also differs between the 2 cell types. The current understanding of AR signaling was gained predominantly using CaP cell lines and samples prepared from whole tissue specimens. The present study sought to delineate AR signaling specifically in endothelial, epithelial, and stromal cells isolated from fresh clinical prostate tissue specimens and primary xenografts of human prostate tissue. Our results reveal the potentially different roles of AR signaling in different human prostate cell compartments, how AR regulates normal differentiated cell functions, and the biological consequences of ADT. Methods: Prostate tissue remnants were transplanted to male nude mice that were castrated and implanted with a sustained delivery device to maintain testosterone at human plasma levels. Endothelial and epithelial cells of both fresh prostate tissue and prostate tissue xenografts were isolated sequentially using magnetic beads conjugated with an antibody specific to epithelial cell or endothelial cell surface markers; the remaining cell fraction was defined as stromal cells. Transcriptomes were obtained using RNASeq, and were analyzed for differential expression of AR-regulated genes, AR co-regulators, and androgen metabolism enzymes. Results: Antibody-mediated cell type-specific enrichment isolated effectively the 3 cell types from the prostate from 16 patients. Among 1263 AR-regulated genes, 399, 313, and 223 were predominantly expressed in epithelial, endothelial or stromal cells, respectively. Among 179 AR co-regulators, 26, 26, and 18 were expressed predominantly in epithelial, endothelial or stromal cells, respectively. Among 128 potential androgen metabolic enzyme genes, 42, 15, and 12 were expressed predominantly in epithelial, endothelial or stromal cells, respectively. Conclusions: Differences in expression of genes associated with AR-mediated trans-regulation were apparent between the different cell types of prostate, whereas, the difference in expression of androgen metabolism genes was less striking. Therefore, the organ level outcome of ADT can only be understood by determining the response of the different cell types in CaP tissue. Citation Format: Nelson T. Gross, Jianming Wang, Gary Smith, Yue Wu. Molecular dissection of androgen receptor signaling in prostatic endothelial, epithelial, and stromal cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1015.

Synthesis and evaluation of anticancer properties of novel benzimidazole ligand and their cobalt(II) and zinc(II) complexes against cancer cell lines A-2780 and DU-145

Eighteen new cobalt(II) or zinc(II) complexes of benzimidazole bearing 1-benzyl and 2-phenyl moieties were synthesized from the reaction of appropriate benzimidazole ligands and CoCl2 or ZnCl2. Their structural characterizations were done by IR, NMR (1H, 13C) and UV–VIS spectrometers. Cytotoxic activities of eighteen new complexes and three benzimidazole ligands were determined using A-2780 (human ovarian) and DU-145 (human prostate) cell lines. Antitumor properties of all compounds were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell viability assay for the tested benzimidazole derivatives was performed and the LogIC50 values of the compounds were calculated after a 24-hour treatment. All tested benzimidazole derivatives showed higher or comparable antitumor activity against A-2780 cell lines compared to the standard drug docetaxel with a LogIC50 value of −0.81 µM (p < 0.05). Eight of the examined compounds (1, 3, 5, 6, 7, 9, 10 and 13) showed high cytotoxic activity against A-2780 compared to the standard drug docetaxel. While the LogIC50 of the docetaxel was −0.81 µM for A-2780 cells at 24 h, the IC50 values of compounds 1, 3, 5, 6, 7, 9, 10 and 13 were −0.97, −1.30, −0.22, 0.13, −0.16, −0.73 and −0.53 µM, respectively. Three of the compounds 1, 18 and V showed high cytotoxic activity against DU-145 compared to docetaxel (p < 0.05). While the LogIC50 of the docetaxel was −1.13 µM for DU-145 cells at 24 h, the LogIC50 values of compounds 1, 18 and V were 0.84, −0.38 and −0.66 µM, respectively.

Bakuchiol suppresses oestrogen/testosterone-induced Benign Prostatic Hyperplasia development through up-regulation of epithelial estrogen receptor β and down-regulation of stromal aromatase

Estrogens and androgens play critical roles during benign prostatic hyperplasia (BPH) development. Estrogen receptors (ERs), androgen receptor (AR) and aromatase, the key conversion enzyme of androgen to estrogen, are thought to be the effective targets for BPH treatment. Bakuchiol (Ba)-containing herb Psoralea corylifolia has been long-termed used for BPH patients in traditional Chinese medicine while the role and regulatory mechanism of Ba involved remain unclear. Human prostatic cell lines WPMY-1 and BPH-1 and oestrodial/testosterone-induced BPH rats were used as the in vitro and in vivo models. Ba significantly inhibited the proliferation of WPMY-1 and BPH-1 cells. In E2/T-induced BPH model, Ba treatment also significantly inhibited the enlargement of prostate, decreased PI values, reduced the thickness of periglanular smooth muscle layer, and down-regulated the expressions of PCNA and smooth muscle cell marker α-SMA, all of which were highly induced in BPH rats. Moreover, the basal and PGE2-induced expressions of aromatase were reduced in Ba-stimulated WPMY-1 cells, while the expression of ERβ was highly increased in Ba-stimulated BPH-1 cells, both of which are consistent with the findings in Ba group in vivo. Ba induced ERE activity in BPH-1 cells as E2 did; however, silence of ERβ not ERα, blocked Ba-induced ERE activity while E2 still exhibited the significant ERE activity, indicating the regulation of estrogen signaling by Ba is particularly via ERβ. In conclusion, by down-regulation of stromal aromatase and up-regulation of epithelial ERβ, Ba contributes to the balance of estrogen and androgen signaling and further inhibits BPH development.

Elemental distribution by synchrotron X‐ray microfluorescence of prostate 3D cell culture

Prostate cancer is a highly prevalent disease and ranks second among malignant neoplasms that affect men around the world, behind lung cancer alone. Trace elements are very important and are involved in many cellular processes. The X‐ray microfluorescence technique is an advanced tool of high spatial resolution, sensitivity, multielemental analysis, and nondestructiveness for trace element study. This study aimed to investigate the elemental distribution in spheroids obtained through the following human prostate cell lines using synchrotron X‐ray microfluorescence: tumor cell line androgen independent (DU145), tumor cell line androgen dependent (LNCaP), and normal cell line (RWPE‐1). The measurements were performed with a standard geometry of 45° of incidence, excited by a white beam using a pixel of 25 μm and an acquisition time of 300 ms/pixel at the X‐ray fluorescence beamline at the Synchrotron Light National Laboratory (Campinas, Brazil). The synchrotron X‐ray microfluorescence results showed differences between groups in all elements analyzed and suggested that further studies should be performed to understand the relationship of these trace elements with the progression and development of the disease.

Effects of isocorynoxeine, from Uncaria, on lower urinary tract dysfunction caused by benign prostatic hyperplasia via antagonism of α1A-adrenoceptors

Medical therapy of lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) targets smooth muscle contraction in the prostate, for which α1A-adrenoceptor (α1A-AR) antagonists have been considered to be the primary therapeutic method. We investigated the effects and underlying mechanisms of isocorynoxeine (ICN), one of indole alkaloids from Uncaria, on the treatment of LUTS secondary to BPH via α1A-ARs in mice. The effect of ICN on prostatic contractility was studied via myographic measurements in the prostates of rabbits. The effects of ICN on bladder function, serum-hormone levels, bladder histology, and prostate histology were determined in testosterone propionate-induced prostatic hyperplasic wild-type (WT) and α1A-AR knockout (α1A-KO) mice. The cytotoxicity of ICN in cultured human prostatic stromal cells (WPMY-1) was assessed by the following: a cell-counting kit, measuring the relaxant effect on WPMY-1 by a collagen gel contraction assay, intracellular Ca2+ mobilization indicated by Fluo-4, cytoskeletal organization by phalloidin staining, and expressions of α1A-AR-mediated key messengers by western blot analyses. ICN non-competitively antagonized the contractions of prostates induced by α1A-AR agonists. ICN treatment improved bladder functions in prostatic hyperplasic WT mice, whereas it failed to ameliorate bladder functions in prostatic hyperplasic α1A-KO mice. In WPMY-1, ICN relaxed cell contractions on collagen gels, disrupted F-actin organization, inhibited α1A-AR agonist-stimulated Ca2+ mobilization, and antagonized α1A-ARs via the RhoA/ROCK2/MLC signaling pathway. Our results suggest that ICN may be a promising therapeutic drug for targeting α1A-ARs in the treatment of BPH/LUTS.

Nonylphenol acts on prostate adenocarcinoma cells via estrogen molecular pathways

Estrogens play a role in the patho-physiology of the prostate. In the present work we studied the effects of nonylphenol (NP), a xenoestrogen, on human adenocarcinoma prostate cells (LNCaP). In order to understand molecular and cellular involvement, we observed the effects on cell cycle and we investigated the expression and the cellular localization of estrogen receptors and gene expression of cyclin D1, ki-67, c-myc, IL-8, IL-1β. We performed the same experiments with 17β-estradiol (E2), the most abundant estrogen circulating in nonpregnant humans in order to compare these two different substances. We demonstrated the ability of 1 × 10−10 M NP to induce proliferation of LNCaP, S-phase progression, increase of ERα expression and its translocation from the cytoplasm to the nucleus. Moreover, we observed an up-regulation of key target genes involved in cell cycle and inflammation process. Particularly, after NP treatment, IL-8 and IL-1β mRNA levels are increased more than 50% indicating a major NP involvement in inflammation processes than E2. These data suggest the proliferative effects of NP on prostate adenocarcinoma cells and highlight some aspects of molecular pathways involved in prostate responses to NP.

Genome-Wide Analysis of Low Dose Bisphenol-A (BPA) Exposure in Human Prostate Cells.

Endocrine disrupting compounds (EDCs) have the potential to cause adverse effects on wild-life and human health. Two important EDCs are the synthetic estrogen 17α-ethynylestradiol (EE2) and bisphenol-A (BPA) both of which are xenoestrogens (XEs) as they bind the estrogen receptor and dis-rupt estrogen physiology in mammals and other vertebrates. In the recent years the influence of XEs on oncogenes, specifically in relation to breast and prostate cancer has been the subject of considerable study.Methodology: In this study, healthy primary human prostate epithelial cells (PrECs) were exposed to environmentally relevant concentrations of BPA (5nM and 25nM BPA) and interrogated using a whole genome microarray.Results: Exposure to 5 and 25nM BPA resulted in 7,182 and 7,650 differentially expressed (DE) genes, respectively in treated PrECs. Exposure to EE2 had the greatest effect on the PrEC transcriptome (8,891 DE genes).Conclusion: We dissected and investigated the nature of the non-estrogenic gene signature associated with BPA with a focus on transcripts relevant to epigenetic modifications. The expression of transcripts encoding nuclear hormone receptors as well as histone and DNA methylation, modifying enzymes were significantly perturbed by exposure to BPA.

Low mRNA expression and activity of monoacylglycerol lipase in human SH-SY5Y neuroblastoma cells

Relatively little is known about the endocannabinoid system in human neuroblastoma cell lines. In the present study, we have investigated the expression of the genes coding for the enzymes involved in the synthesis and catabolism of endocannabinoids in the SH-SY5Y cell line. The expression of MGLL, the gene coding for the 2-arachidonoylglycerol hydrolytic enzyme monoacylglycerol lipase (MAGL), was found to be 85 and 340 fold lower than the expression levels for the genes coding for alpha/beta-hydrolase domain containing 6 and 12 (ABHD6, ABHD12), which are alternative hydrolytic enzymes for this endocannabinoid. In comparison, mRNA levels of MGLL were 1.5 fold higher than ABHD6 and 2 fold lower than the levels of ABHD12 in DU-145 human prostate cells. In functional assays, the hydrolysis of the 2-arachidonoylglycerol homologue 2-oleoylglycerol by intact SH-SY5Y cells was partially inhibited by the ABHD6 inhibitor WWL70, but not by the MAGL inhibitor JZL184, whereas the reverse was true in DU-145 cells. The combination of JZL184 + WWL70 did, however produce a significantly greater inhibition of 2-OG hydrolysis than seen with WWL70 alone in the SH-SY5Y cells. The low MGLL expression in the SH-SY5Y cells was not due to epigenetic silencing, since levels were not affected by treatment with the methylation inhibitor 5-aza-2′-deoxycytidine and/or the histone acetylase inhibitor trichostatin A. The low MGLL expression in SH-SY5Y cells should be taken into account when using these cells in experiments investigating the involvement of the endocannabinoid system in models of physiological and pathological processes.

Design, Synthesis, and Molecular Docking Study of Novel Heterocycles Incorporating 1,3,4-Thiadiazole Moiety as Potential Antimicrobial and Anticancer Agents.

A new series of 5-(3,5-dinitrophenyl)-1,3,4-thiadiazole derivatives were prepared and evaluated for their in vitro antimicrobial, antitumor, and DHFR inhibition activity. Compounds 9, 10, 13, and 16 showed strong and broad-spectrum antimicrobial activity comparable to Amoxicillin and Fluconazole as positive antibiotic and antifungal controls, respectively. Compounds 6, 14, and 15 exhibited antitumor activity against four human cancer cell lines, CCRF-CEM leukemia, HCT-15 colon, PC-3 prostate, and UACC-257 melanoma cell lines using Doxorubicin as a reference drug. Compounds 10, 13, 14, and 15 proved to be the most active DHFR inhibitors with an IC50 range of 0.04 ± 0.82–1.00 ± 0.85 µM, in comparison with Methotrexate (IC50 = 0.14 ± 1.38 µM). The highly potent DHFR inhibitors shared a similar molecular docking mode and made a critical hydrogen bond and arene‒arene interactions via Ser59 and Phe31 amino acid residues, respectively.

TUG1 promotes the development of prostate cancer by regulating RLIM.

The aim of this study was to investigate the specific role of Taurine up-regulated gene 1 (TUG1) in the development of prostate cancer (PCa), and to explore its underlying mechanism. The serum level of TUG1 in healthy subjects, benign prostatic hyperplasia (BPH) patients and PCa patients was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The correlation between TUG1 expression and clinical indexes of PCa patients was analyzed. TUG1 expression in PCa cells and human normal prostate cells was determined by qRT-PCR as well. Overexpression or knockdown of TUG1 was achieved by liposomal transfection. Subsequently, the regulatory effects of TUG1 on the proliferative and migratory capacities of DU145 cells were accessed by Cell Counting Kit-8 (CCK-8) assay, colony formation assay and transwell assay, respectively. An online software was used to predict whether RLIM could be regulated by TUG1, which was further verified by qRT-PCR. After RLIM knockdown in DU145 cells, the proliferative and migratory capacities were also determined. Finally, Western blot was conducted to determine relative protein expressions in the TGF-β1/Smad pathway after altering TUG1 expression in DU145 cells. TUG1 was highly expressed in serum samples of PCa patients when compared with healthy subjects and BPH patients. Besides, TUG1 expression in PCa patients with Gleason ≥ 7 was significantly higher than those with Gleason < 7. Meanwhile, TUG1 expression in PCa patients was remarkably higher than that of BPH patients at the PSA grey zone (4-10 ng/mg). ROC curves indicated that TUG1 might be a crucial hallmark to distinguish PCa patients from BPH patients and healthy subjects. The overexpression of TUG1 markedly promoted the proliferative and migratory capacities of DU145 cells. However, knockdown of TUG1 obtained the opposite results. QRT-PCR confirmed that TUG1 was positively correlated with RLIM at the mRNA level. RLIM knockdown significantly inhibited the proliferative and migratory capacities of DU145 cells. Furthermore, knockdown of TUG1 in DU145 cells markedly down-regulated TGF-β1 and p-Smad2, whereas up-regulated p-Smad7. TUG1 is highly expressed in peripheral blood of PCa patients, which can serve as a potential diagnostic marker for PCa. The overexpression of TUG1 promotes the proliferative and migratory capacities of PCa cells. Furthermore, TUG1 promotes the development of PCa by regulating RLIM through the TGF-β1/Smad pathway.

Bioassay-guided isolation and structure elucidation of cytotoxic stilbenes and flavonols from the leaves of Macaranga barteri

Bioassay-guided fractionation of the leaves of Macaranga barteri collected from Nigeria led to the isolation of three previously undescribed cytotoxic stilbenes, macabartebenes A–C (1–3), together with six known compounds including prenylated stilbenes: vedelianin (4), schweinfurthin G (5), and mappain (7), prenylated flavonols: 8-prenylkaempferol (6), and broussoflavonol F (8), and the geranylated flavonol, isomacarangin (9). The cytotoxicity of the compounds was evaluated against four human cancer cell lines, with vinblastine as the positive control and DMSO vehicle as the negative control. Vedelianin (IC50 = 0.32–0.54 μM) displayed the greatest antiproliferative activity across the panel of cancer cell lines amongst the compounds, while macabartebene A (IC50 = 0.60–0.79 μM) was the most potent of the previously unreported compounds. The compounds displayed varying selectivity towards the cancer cell lines compared to the normal human prostate cell line. The findings of this study revealed that M. barteri leaves contain several cytotoxic compounds.

Design, synthesis, cytotoxicity, and molecular modeling study of 2,4,6-trisubstituted pyrimidines with anthranilate ester moiety

A series of 2,4,6-trisubstituted pyrimidines with antharanilic acid ester moiety have been designed and synthesized by employing a one-pot multicomponent approach from methyl 5-(ethynyl)anthranilate, aroyl or cinnamoyl chlorides and various amidines. All the compounds were evaluated for their cytotoxic activity with respect to model cancer cell lines (CEM-13, U-937, MDA-MB-231, BT-474, DU-145) using conventional MTT assays. Structure-activity relationship analysis revealed that 4,6-diarylpyrimidines 13–17, substituted with a pyridine or a pyrimidine ring in the 2 position displayed an increasing of activity compared to 2-methyl or 2-phenyl substituted pyrimidines. The 2-amino substututed compound 9 showed selectivity on the human monocyte-like cells U-937. Trisubstituted pyrimidines 18–21, containing a (E)-styryl substituent in the 6 position of the pyrimidine core, demonstrated an increasing of activity against the breast cancer cell lines MDA-MB-231, BT-474, and also against the human prostate cell lines DU-145. Compounds 18 and 20 shown the best potency towards the cancer cell lines MDA-MB-231; theirs activity was comparable with the activity of Doxorubicin on this cell lines. These compounds were confirmed to be cyclin-dependent kinase 9 (CDK9) inhibitors through in silico molecular modeling studies for the mode of action. The newly synthesized compounds may serve as lead molecules for the future research regarding the identification of new anticancer agents in the pyrimidine series.

Dihydrotestosterone inhibits arylsulfatase B and Dickkopf Wnt signaling pathway inhibitor (DKK)-3 leading to enhanced Wnt signaling in prostate epithelium in response to stromal Wnt3A.

In tissue microarrays, immunostaining of the enzyme arylsulfatase B (ARSB; N-acetylgalactosamine-4-sulfatase) was less in recurrent prostate cancers and in cancers with higher Gleason scores. In cultured prostate stem cells, decline in ARSB increased Wnt signaling through effects on Dickkopf Wnt Signaling Pathway Inhibitor (DKK)3. The effects of androgen exposure on ARSB and the impact of decline in ARSB on Wnt signaling in prostate tissue were unknown. Epithelial and stromal tissues from malignant and normal human prostate were obtained by laser capture microdissection. mRNA expression of ARSB, galactose-6-sulfate-sulfatase (GALNS) and Wnt-signaling targets was determined by QPCR. Non-malignant human epithelial and stromal prostate cells were grown in tissue culture, including two-cell layer cultures. ARSB was silenced by specific siRNA, and epithelial cells were treated with stromal spent media following treatment with IWP-2, an inhibitor of Wnt secretion, and by exogenous recombinant human Wnt3A. Promoter methylation was detected using specific DKK3 and ARSB promoter primers. The effects of DHT and of ARSB overexpression on DKK expression were determined. Cell proliferation was assessed by BrdU incorporation. Normal stroma showed higher expression of vimentin, ARSB, and Wnt3A than epithelium. Normal epithelium had higher expression of E-cadherin, galactose 6-sulfate-sulfatase (GALNS), and DKK3 than stroma. In malignant epithelium, expression of ARSB and DKK3 declined, and expression of GALNS and Wnt signaling targets increased. In cultured prostate epithelial cells, Wnt-mediated signaling was greatest when ARSB was silenced and cells were exposed to exogenous Wnt3A. Exposure to 5α-dihydrotestosterone (DHT) increased ARSB and DKK3 promoter rmethylation, and effects of DHT on DKK3 expression were reversed when ARSB was overexpressed. Androgen-induced declines in ARSB and DKK3 may contribute to prostate carcinogenesis by sustained activation of Wnt signaling in prostate epithelium in response to stromal Wnt3A.