TOPAS-Tissue: A Framework for the Simulation of the Biological Response to Ionizing Radiation at the Multi-Cellular Level.

This work aims to develop and validate a framework for the multiscale simulation of the biological response to ionizing radiation in a population of cells forming a tissue. We present TOPAS-Tissue, a framework to allow coupling two Monte Carlo (MC) codes: TOPAS with the TOPAS-nBio extension, capable of handling the track-structure simulation and subsequent chemistry, and CompuCell3D, an agent-based model simulator for biological and environmental behavior of a population of cells. We verified the implementation by simulating the experimental conditions for a clonogenic survival assay of a 2-D PC-3 cell culture model (10 cells in 10,000 µm2) irradiated by MV X-rays at several absorbed dose values from 0-8 Gy. The simulation considered cell growth and division, irradiation, DSB induction, DNA repair, and cellular response. The survival was obtained by counting the number of colonies, defined as a surviving primary (or seeded) cell with progeny, at 2.7 simulated days after irradiation. DNA repair was simulated with an MC implementation of the two-lesion kinetic model and the cell response with a p53 protein-pulse model. The simulated survival curve followed the theoretical linear-quadratic response with dose. The fitted coefficients α = 0.280 ± 0.025/Gy and ß = 0.042 ± 0.006/Gy2 agreed with published experimental data within two standard deviations. TOPAS-Tissue extends previous works by simulating in an end-to-end way the effects of radiation in a cell population, from irradiation and DNA damage leading to the cell fate. In conclusion, TOPAS-Tissue offers an extensible all-in-one simulation framework that successfully couples Compucell3D and TOPAS for multiscale simulation of the biological response to radiation.

Solid Lipid Nanoparticles Based on Babassu Oil and Copaiba Oleoresin: A Promising Approach for Prostate Cancer Therapy.

Solid lipid nanoparticles (SLNs) represent promising nanostructures for drug delivery systems. This study successfully synthesized SLNs containing different proportions of babassu oil (BBS) and copaiba oleoresin (COPA) via the emulsification-ultrasonication method. Before SLN synthesis, the identification and quantification of methyl esters, such as lauric acid and ß-caryophyllene, were performed via GC-MS analysis. These methyl esters were used as chemical markers and assisted in encapsulation efficiency experiments. A 22 factorial design with a center point was employed to assess the impact of stearic acid and Tween 80 on particle hydrodynamic diameter (HD) and polydispersity index (PDI). Additionally, the effects of temperature (8 ± 0.5 °C and 25 ± 1.0 °C) and time (0, 7, 15, 30, 40, and 60 days) on HD and PDI values were investigated. Zeta potential (ZP) measurements were utilized to evaluate nanoparticle stability, while transmission electron microscopy provided insights into the morphology and nanometric dimensions of the SLNs. The in vitro cytotoxic activity of the SLNs (10 µg/mL, 30 µg/mL, 40 µg/mL, and 80 µg/mL) was evaluated using the MTT assay with PC-3 and DU-145 prostate cancer cell lines. Results demonstrated that SLNs containing BBS and COPA in a 1:1 ratio exhibited a promising cytotoxic effect against prostate cancer cells, with a percentage of viable cells of 68.5% for PC-3 at a concentration of 30 µg/mL and 48% for DU-145 at a concentration of 80 µg/mL. These findings underscore the potential therapeutic applications of SLNs loaded with BBS and COPA for prostate cancer treatment.

Thymoquinone Potentially Modulates the Expression of Key Onco- and Tumor Suppressor miRNAs in Prostate and Colon Cancer Cell Lines: Insights from PC3 and HCT-15 Cells.

Prostate cancer (PC) and colon cancer significantly contribute to global cancer-related morbidity and mortality. Thymoquinone (TQ), a naturally occurring phytochemical found in black cumin, has shown potential as an anticancer compound. This study aimed to investigate the effects of TQ on the expression profile of key tumor suppressor and onco-suppressor miRNAs in PC3 prostate cancer cells and HCT-15 colon cancer cells. Cell viability assays revealed that TQ inhibited the growth of both cell lines in a dose-dependent manner, with IC50 values of approximately 82.59 µM for HCT-15 and 55.83 µM for PC3 cells. Following TQ treatment at the IC50 concentrations, miRNA expression analysis demonstrated that TQ significantly downregulated miR-21-5p expression in HCT-15 cells and upregulated miR-34a-5p, miR-221-5p, miR-17-5p, and miR-21-5p expression in PC3 cells. However, no significant changes were observed in the expression levels of miR-34a-5p and miR-200a-5p in HCT-15 cells. The current findings suggest that TQ might exert its antiproliferative effects by modulating specific tumor suppressor and onco-suppressor miRNAs in prostate and colon cancer cells. Further investigations are warranted to elucidate the precise underlying mechanisms and to explore the therapeutic potential of TQ in cancer treatment. To the best of our knowledge, this is the first report regarding the effect of TQ on the miRNA expression profile in colon and prostate cancer cell lines.

A Triazaspirane Derivative Inhibits Migration and Invasion in PC3 Prostate Cancer Cells.

Cancer is a serious health problem due to the complexity of establishing an effective treatment. The purpose of this work was to evaluate the activity of a triazaspirane as a migration and invasion inhibitor in PC3 prostatic tumor cells through a possible negative regulation of the FAK/Src signal transduction pathway and decreased secretion of metalloproteinases 2 and 9. Molecular docking analysis was performed using Moe 2008.10 software. Migration (wound-healing assay) and invasion (Boyden chamber assay) assays were performed. In addition, the Western blot technique was used to quantify protein expression, and the zymography technique was used to observe the secretion of metalloproteinases. Molecular docking showed interactions in regions of interest of the FAK and Src proteins. Moreover, the biological activity assays demonstrated an inhibitory effect on cell migration and invasion, an important suppression of metalloproteinase secretion, and a decrease in the expression of p-FAK and p-Src proteins in treated PC3 cells. Triazaspirane-type molecules have important inhibitory effects on the mechanisms associated with metastasis in PC3 tumor cells.

Tempol differential effect on prostate cancer inflammation: In vitro and in vivo evaluation.

BACKGROUND: Tempol is a redox-cycling nitroxide that acts directly on inflammation. However, few studies have reported the use of tempol in prostate cancer (PCa). The present study investigated the effects of tempol on inflammation related to NF-κB signaling, using hormone-dependent or hormone-independent cell lines and the transgenic adenocarcinoma of the mouse prostate PCa animal model in the early and late stages of cancer progression. METHODS: PC-3 and LnCaP cells were exposed to different tempol doses in vitro, and cell viability assays were performed. The optimal treatment dose was chosen for subsequent analysis using western blotting. Five experimental groups were evaluated in vivo to test for tempol effects in the early (CT12 and TPL12 groups) and late stages (CT20, TPL20-I, and TLP20-II) of PCa development. The TPL groups were treated with 50 or 100 mg/kg tempol. All control groups received water as the vehicle. The ventral lobe of the prostate was collected and subjected to immunohistochemical and western blot analysis. RESULTS: Tempol treatment reduced cellular proliferation in vitro and improved prostatic morphology in vivo, thereby decreasing tumor progression. Tempol reduced inflammation in preclinical models, and downregulated the initial inflammatory signaling through toll-like receptors, not always mediated by the MyD88 pathway. In addition, it upregulated iκB-α and iκB -ß levels, leading to a decrease in NF-κB, TNF-α, and other inflammatory markers. Tempol also influenced cell survival markers. CONCLUSIONS: Tempol can be considered a beneficial therapy for PCa treatment owing to its anti-inflammatory and antiproliferative effects. Nevertheless, the action of tempol was different depending on the degree of the prostatic lesion in vivo and hormone reliance in vitro. This indicates that tempol plays a multifaceted role in the prostatic tissue environment.

Severe Congenital Neutropenia Type 4: A Rare Disease Harboring a G6pc3 Gene Pathogenic Variant Particular to the Mexican Population.

Background: Severe congenital neutropenia type 4 (SCN4) is a rare autosomal recessive granulopoiesis disorder caused by G6PC3 gene pathogenic variants. The estimated prevalence is 1/10,000,000 people. Over 90% of patients present a syndromic form with variable multisystemic involvement, including congenital heart defects, increased visibility of superficial veins (IVSV), inflammatory bowel disease, and congenital urogenital defects as prominent symptoms. Objectives: The objective of the study was to study non-hematological phenotypic findings that suggest a clinical diagnosis of SCN4. Methods: We examined medical records of patients diagnosed with neutropenia from January 2000 to December 2020, selecting cases with non-hematologic manifestations for phenotypic description and G6PC3 gene sequencing. Results: We found 11 cases with non-hematologic features: congenital heart defects in 8, IVSV in 6, inflammatory bowel disease in 4, urogenital defects in 4, and similar facial appearance. In addition, Sanger sequencing confirmed 3 homozygous cases for the c.210delC variant, a compound heterozygous harboring this variant, and a c.199_218+1 deletion. Conclusions: Our findings of the c.210delC variant in very close geographical settings, to date, have only been reported among Mexicans, and a mutual uncommon surname in two families strongly supports a founder effect for the variant in the studied population. Furthermore, the described non-hematologic symptoms in patients with severe primary neutropenia should be explored, confirming SCN4 by investigating G6PC3 gene mutations.

Severe Congenital Neutropenia Type 4: A Rare Disease Harboring a G6pc3 Gene Pathogenic Variant Particular to the Mexican Population

ABSTRACT Background: Severe congenital neutropenia type 4 (SCN4) is a rare autosomal recessive granulopoiesis disorder caused by G6PC3 gene pathogenic variants. The estimated prevalence is 1/10,000,000 people. Over 90% of patients present a syndromic form with variable multisystemic involvement, including congenital heart defects, increased visibility of superficial veins (IVSV), inflammatory bowel disease, and congenital urogenital defects as prominent symptoms. Objectives: The objective of the study was to study non-hematological phenotypic findings that suggest a clinical diagnosis of SCN4. Methods: We examined medical records of patients diagnosed with neutropenia from January 2000 to December 2020, selecting cases with non-hematologic manifestations for phenotypic description and G6PC3 gene sequencing. Results: We found 11 cases with non-hematologic features: congenital heart defects in 8, IVSV in 6, inflammatory bowel disease in 4, urogenital defects in 4, and similar facial appearance. In addition, Sanger sequencing confirmed 3 homozygous cases for the c.210delC variant, a compound heterozygous harboring this variant, and a c.199_218+1 deletion. Conclusions: Our findings of the c.210delC variant in very close geographical settings, to date, have only been reported among Mexicans, and a mutual uncommon surname in two families strongly supports a founder effect for the variant in the studied population. Furthermore, the described non-hematologic symptoms in patients with severe primary neutropenia should be explored, confirming SCN4 by investigating G6PC3 gene mutations.

Fibronectin Modulates the Expression of miRNAs in Prostate Cancer Cell Lines.

Prostate cancer (PCa) is a significant cause of cancer-related deaths among men and companion animals, such as dogs. However, despite its high mortality and incidence rates, the molecular mechanisms underlying this disease remain to be fully elucidated. Among the many factors involved in prostate carcinogenesis, the extracellular matrix (ECM) plays a crucial role. This ECM in the prostate is composed mainly of collagen fibers, reticular fibers, elastic fibers, proteoglycans and glycoproteins, such as fibronectin. Fibronectin is a glycoprotein whose dysregulation has been implicated in the development of multiple types of cancer, and it has been associated with cell migration, invasion, and metastasis. Furthermore, our research group has previously shown that fibronectin induces transcriptional changes by modulating the expression of protein coding genes in LNCaP cells. However, potential changes at the post-transcriptional level are still not well understood. This study investigated the impact of exposure to fibronectin on the expression of a key class of regulatory RNAs, the microRNAs (miRNAs), in prostate cancer cell lines LNCaP and PC-3. Five mammalian miRNAs (miR-21, miR-29b, miR-125b, miR-221, and miR-222) were differentially expressed after fibronectin exposure in prostate cell lines. The expression profile of hundreds of mRNAs predicted to be targeted by these miRNAs was analyzed using publicly available RNA-Sequencing data (GSE64025, GSE68645, GSE29155). Also, protein-protein interaction networks and enrichment analysis were performed to gain insights into miRNA biological functions. Altogether, these functional analyzes revealed that fibronectin exposure impacts the expression of miRNAs potentially involved in PCa causing changes in critical signaling pathways such as PI3K-AKT, and response to cell division, death, proliferation, and migration. The relationship here demonstrated between fibronectin exposure and altered miRNA expression improves the comprehension of PCa in both men and other animals, such as dogs, which naturally develop prostate cancer.

The Isoflavanoid (+)-PTC Regulates Cell-Cycle Progression and Mitotic Spindle Assembly in a Prostate Cancer Cell Line.

Prostate cancer is the second most common malignancy in men and the development of effective therapeutic strategies remains challenging when more advanced, androgen-independent or insensitive forms are involved. Accordingly, we have evaluated, using flow cytometry, confocal microscopy and image analysis, the anti-proliferative effects of (+)-2,3,9-trimethoxypterocarpan [(+)-PTC, 1] on relevant human prostate cancer cells as well as its capacity to control mitosis within them. In particular, the studies reported herein reveal that (+)-PTC exerts anti-proliferative activity against the PC-3 cell lines by regulating cell-cycle progression with mitosis being arrested in the prophase or prometaphase. Furthermore, it emerges that treatment of the target cells with this compound results in the formation of monopolar spindles, disorganized centrosomes and extensively disrupted γ-tubulin distributions while centriole replication remains unaffected. Such effects suggest (+)-PTC should be considered as a possible therapy for androgen-insensitive/independent prostate cancer.

Additional activation of the AR gene may be involved in the development of the castration resistance phenotype in prostate cancer.

INTRODUCTION: Several studies have already shown that changes in the AR gene may be associated with a more aggressive disease phenotype and even castration-resistant prostate cancer. Thus, we investigated cytogenetic and molecular alterations linked to AR. MATERIALS AND METHODS: To evaluate AR methylation, we performed a cytogenetic-molecular analysis using fluorescence in situ hybridization that uses specific probes for the AR gene (Xq11.12) and the X chromosome centromere. For AR activity, we performed a qualitative analysis of human androgen receptor activity. To analyze the expression of AR in PC3 and LNCaP cell lines, we used qPCR assays. RESULTS: In the qPCR assay, we found downregulation of AR in the PC3 cell line compared with the LNCaP. We found the presence of X chromosome polysomy in PC-3 and LNCaP cell lines by FISH assay. In the HUMARA-Q assay, we found two X chromosomes/cell and the activity of both AR in the PC-3 cell line. In LNCaP cells, we found two X chromosomes/cell and methylation of only one AR. CONCLUSION: Castration-resistant prostate cancer phenotype represents a significant challenge in the setting of urological management. The X chromosomes and AR-linked alterations may contribute to a better understanding of the disease. However, further studies should be performed in an attempt to elucidate as much as possible the role of AR in the castration-resistant prostate cancer phenotype.

Severe congenital neutropenia due to G6PC3 deficiency: Case series of five patients and literature review.

BACKGROUND AND OBJECTIVES: Glucose-6-phosphate catalytic subunit 3 (G6PC3) deficiency is characterized by severe congenital neutropenia with recurrent pyogenic infections, a prominent superficial venous pattern and cardiovascular and urogenital malformations caused by an alteration of glucose homeostasis, with increased endoplasmic reticulum stress and cell apoptosis. METHODS: We reviewed our patients with G6PC3 deficiency diagnosed along the last decade in Mexico; we also searched the PubMed/Medline database for the terms ('G6PC3 deficiency' OR 'Dursun syndrome' OR 'Severe congenital neutropenia type 4'), and selected articles published in English from 2009 to 2020. RESULTS: We found 89 patients reported from at least 14 countries in 4 continents. We describe five new cases from Mexico. Of the 94 patients, 56% are male, 48% from Middle East countries and none of them had adverse reactions to live vaccines; all presented with at least 1 severe infection prior to age 2. Seventy-five per cent had syndromic features, mainly atrial septal defect in 55% and prominent superficial veins in 62%. CONCLUSIONS: With a total of 94 patients reported in the past decade, we delineate the most frequent laboratory and genetic features, their treatment and outcomes, and to expand the knowledge of syndromic and non-syndromic phenotypes in these patients.

Exposure to Bacteriophages T4 and M13 Increases Integrin Gene Expression and Impairs Migration of Human PC-3 Prostate Cancer Cells.

The interaction between bacteriophages and integrins has been reported in different cancer cell lines, and efforts have been undertaken to understand these interactions in tumor cells along with their possible role in gene alterations, with the aim to develop new cancer therapies. Here, we report that the non-specific interaction of T4 and M13 bacteriophages with human PC-3 cells results in differential migration and varied expression of different integrins. PC-3 tumor cells (at 70% confluence) were exposed to 1 × 107 pfu/mL of either lytic T4 bacteriophage or filamentous M13 bacteriophage. After 24 h of exposure, cells were processed for a histochemical analysis, wound-healing migration assay, and gene expression profile using quantitative real-time PCR (qPCR). qPCR was performed to analyze the expression profiles of integrins ITGAV, ITGA5, ITGB1, ITGB3, and ITGB5. Our findings revealed that PC-3 cells interacted with T4 and M13 bacteriophages, with significant upregulation of ITGAV, ITGA5, ITGB3, ITGB5 genes after phage exposure. PC-3 cells also exhibited reduced migration activity when exposed to either T4 or M13 phages. These results suggest that wildtype bacteriophages interact non-specifically with PC-3 cells, thereby modulating the expression of integrin genes and affecting cell migration. Therefore, bacteriophages have future potential applications in anticancer therapies.

Caryophyllene Oxide, the Active Compound Isolated from Leaves of Hymenaea courbaril L. (Fabaceae) with Antiproliferative and Apoptotic Effects on PC-3 Androgen-Independent Prostate Cancer Cell Line.

Cancer treatment frequently carries side effects, therefore, the search for new selective and effective molecules is indispensable. Hymenaea courbaril L. has been used in traditional medicine in South America to treat several diseases, including prostate cancer. Leaves' extracts from different polarities were evaluated using the 3-(4,5-methyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) cell viability assay to determine the cytotoxicity in prostate p53-null cells, followed by bio-guided fractionations to obtain the most cytotoxic fraction considering the selectivity index. The most cytotoxic fraction was analyzed by GC/MS to identify the active compounds. The majority compound, caryophyllene oxide, induced early and late apoptosis, depolarized the mitochondrial membrane, leading to several morphological changes and shifts in apoptotic proteins, and caspases were evidenced. Depolarization of the mitochondrial membrane releases the pro-apoptotic protein Bax from Bcl-xL. The apoptosis process is caspase-7 activation-dependent. Caryophyllene oxide is a safe anti-proliferative agent against PC-3 cells, inducing apoptosis with low toxicity towards normal cells.

The Glypican proteoglycans show intrinsic interactions with Wnt-3a in human prostate cancer cells that are not always associated with cascade activation.

BACKGROUND: Prostate cancer occurs through multiple steps until advanced metastasis. Signaling pathways studies can result in the identification of targets to interrupt cancer progression. Glypicans are cell surface proteoglycans linked to the membrane through glycosylphosphatidylinositol. Their interaction with specific ligands has been reported to trigger diverse signaling, including Wnt. In this study, prostate cancer cell lines PC-3, DU-145, and LNCaP were compared to normal prostate RWPE-1 cell line to investigate glypican family members and the activation of the Wnt signaling pathway. RESULTS: Glypican-1 (GPC1) was highly expressed in all the examined cell lines, except for LNCaP, which expressed glypican-5 (GPC5). The subcellular localization of GPC1 was detected on the cell surface of RWPE-1, PC-3, and DU-145 cell lines, while GPC5 suggested cytoplasm localization in LNCaP cells. Besides glypican, flow cytometry analysis in these prostate cell lines confirmed the expression of Wnt-3a and unphosphorylated ß-catenin. The co-immunoprecipitation assay revealed increased levels of binding between Wnt-3a and glypicans in cancer cells, suggesting a relationship between these proteoglycans in this pathway. A marked increase in nuclear ß-catenin was observed in tumor cells. However, only PC-3 cells demonstrated activation of canonical Wnt signaling, according to the TOPFLASH assay. CONCLUSIONS: GPC1 was the majorly expressed gene in all the studied cell lines, except for LNCaP, which expressed GPC5. We assessed by co-immunoprecipitation that these GPCs could interact with Wnt-3a. However, even though nuclear ß-catenin was found increased in the prostate cancer cells (i.e., PC-3, DU-145 and LNCaP), activation of Wnt pathway was only found in PC-3 cells. In these PC-3 cells, GPC1 and Wnt-3a revealed high levels of colocalization, as assessed by confocal microscopy studies. This suggests a localization at the cellular surface, where Frizzled receptor is required for downstream activation. The interaction of Wnt-3a with GPCs in DU-145 and LNCaP cells, which occurs in absence of Wnt signaling activation, requires further studies. Once non-TCF-LEF proteins can also bind ß-catenin, another signaling pathway may be involved in these cells with regulatory function.

Bioassay-Guided Identification of the Antiproliferative Compounds of Cissus trifoliata and the Transcriptomic Effect of Resveratrol in Prostate Cancer Pc3 Cells.

The bioassay-guided fractionation of a CHCl3-MeOH extract from the stems of Cissus trifoliata identified an active fraction against PC3 prostate cancer cells. The treatment for 24 h showed an 80% reduction in cell viability (p ≤ 0.05) by a WST-1 assay at a concentration of 100 µg/mL. The HPLC-QTOF-MS analysis of the fraction showed the presence of coumaric and isoferulic acids, apigenin, kaempferol, chrysoeriol, naringenin, ursolic and betulinic acids, hexadecadienoic and octadecadienoic fatty acids, and the stilbene resveratrol. The exposure of PC3 cells to resveratrol (IC25 = 23 µg/mL) for 24 h induced significant changes in 847 genes (Z-score ≥ ±2). The functional classification tool of the DAVID v6.8 platform indicates that the underlying molecular mechanisms against the proliferation of PC3 cells were associated (p ≤ 0.05) with the process of differentiation and metabolism. These findings provide experimental evidence suggesting the potential of C. trifoliata as a promising natural source of anticancer compounds.

Bioactive compounds, antioxidant activity and antiproliferative effects in prostate cancer cells of green and roasted coffee extracts obtained by microwave-assisted extraction (MAE).

Coffee consumption has been investigated as a protective factor against prostate cancer. Coffee may be related to prostate cancer risk reduction due to its phytochemical compounds, such as caffeine, chlorogenic acids, and trigonelline. The roasting process affects the content of the phytochemicals and undesired compounds can be formed. Microwave-assisted extraction is an alternative to conventional extraction techniques since it preserves more bioactive compounds. Therefore, this study aimed to evaluate the phytochemical composition and the putative preventive effects in prostate cancer development of coffee beans submitted to four different coffee-roasting degrees extracted using microwave-assisted extraction. Coffea arabica green beans (1) were roasted into light (2), medium (3) and dark (4) and these four coffee samples were submitted to microwave-assisted extraction. The antioxidant capacity of these samples was evaluated by five different methods. Caffeine, chlorogenic acid and caffeic acid were measured through HPLC. Samples were tested against PC-3 and DU-145 metastatic prostate cancer cell lines regarding their effects on cell viability, cell cycle progression and apoptotic cell death. We found that green and light roasted coffee extracts had the highest antioxidant activity. Caffeine content was not affected by roasting, chlorogenic acid was degraded due to the temperature, and caffeic acid increased in light roasted and decreased in medium and dark roasted. Green and light roasted coffee extracts promoted higher inhibition of cell viability, caused greater cell cycle arrest in S and G2/M and induced apoptosis more compared to medium and dark roasted coffee extracts and the control samples. Coffee extracts were more effective against DU-145 than in PC-3 cells. Our data provide initial evidence that among the four tested samples, the consumption of green and light coffee extracts contributes to inhibit prostate cancer tumor progression features, potentially preventing aspects related to advanced prostate cancer subtypes.

Estrogen Receptor Signaling Pathways Involved in Invasion and Colony Formation of Androgen-Independent Prostate Cancer Cells PC-3.

Castration-resistant prostate cancer (CRPC) is an advanced and androgen-independent form of prostate cancer. Recent studies of rapid actions mediated by estrogen in the prostate and its relationship with CRPC are emerging. We have previously shown that estrogen receptor (ER) promotes migration and invasion of the androgen-independent prostate cancer cells PC-3, but the signaling pathways involved in these events remain to be elucidated. Therefore, this study aimed to analyze the role of ERα and ERß in the activation of SRC, and the involvement of SRC and PI3K/AKT on invasion and colony formation of the PC-3 cells. Our results showed that the activation of ERα (using ERα-selective agonist PPT) and ERß (using ERß-selective agonist DPN) increased phosphorylation of SRC in PC-3 cells. In the presence of the selective inhibitor for SRC-family kinases PP2, the effects of DPN and PPT on transmigration and soft agar colony formation assays were decreased. Furthermore, SRC is involved in the expression of the non-phosphorylated ß-catenin. Finally, using PI3K specific inhibitor Wortmannin and AKT inhibitor MK2206, we showed that PI3K/AKT are also required for invasion and colony formation of PC-3 cells simulated by ER. This study provides novel insights into molecular mechanisms of ER in PC-3 cells by demonstrating that ER, located outside the cell nucleus, activates rapid responses molecules, including SRC and PI3K/AKT, which enhance the tumorigenic potential of prostate cancer cells, increasing cell proliferation, migration, invasion, and tumor formation.

Effects of RhoA and RhoC upon the sensitivity of prostate cancer cells to glutamine deprivation.

RhoA and RhoC contribute to the regulation of glutamine metabolism, which is a crucial determinant of cell growth in some types of cancer. Here we investigated the participation of RhoA and RhoC in the response of prostate cancer cells to glutamine deprivation. We found that RhoA and RhoC activities were up- or downregulated by glutamine reduction in PC3 and LNCaP cell lines, which was concomitant to a reduction in cell number and proliferation. Stable overexpression of wild type RhoA or RhoC did not alter the sensitivity to glutamine deprivation. However, PC3 cells expressing dominant negative RhoAN19 or RhoCN19 mutants were more resistant to glutamine deprivation. Our results indicate that RhoA and RhoC activities could affect cancer treatments targeting the glutamine pathway.

Cytotoxic effects of a triterpene-enriched fraction of Cecropia pachystachya on the human hormone-refractory prostate cancer PC3 cell line.

BACKGROUND: Prostate cancer (PCa) is the most diagnosed invasive cancer and a leading cause of death in men in western countries. Most patients initially respond to androgen deprivation but finally develop hormone-refractory disease, which results in advanced clinical failure and death. Since hormone-refractory disease is highly radiotherapy and chemotherapy resistant, increasing interest has been placed on finding novel therapies for this advanced type of Pca. PURPOSE: The potential cytotoxic effects of the crude extract and fractions obtained from the leaves of Cecropia pachystachya Trécul on different human cancer cell lines were investigated. Additionally, the mechanism of cell death induction of the most active sample (triterpene-enriched fraction, TEF) on the human hormone-refractory prostate PC3 cell line was examined. METHODS: Sulforhodamine B assay was used to measure the viability of human tumor and non-tumor cell lines. To elucidate the mechanism of PC3 cells death induced by TEF, different methodological approaches were used: cell cycle analysis and annexin V/PI staining, nuclear morphological analysis, and senescence-associated-ß-galactosidase assay. Moreover, the mitochondrial membrane potential was measured, and the long-term effects of TEF on PC3 cells were evaluated. RESULTS: TEF exerted cytotoxic effects on PC3 cells but not on human non-tumor cells. The analysis of nuclear morphology of PC3 cells treated with TEF increased the number of cells with large and regular nuclei suggesting senescence induction, which was supported by ß-galactosidase overexpression. Regarding PC3 cells cycle, TEF reduced the number of cells in G1 phase and increased that in sub G0/G1. Apoptosis was not involved in PC3 cell death. However, there was a decrease in mitochondrial membrane potential without the participation of reactive oxygen species (ROS) in the cytotoxic effects detected. Furthermore, there was a decrease in the number of viable cells able to duplicate after long-term TEF treatment. CONCLUSIONS: The results showed the in vitro cytotoxic potential of the triterpene-enriched fraction obtained from the leaves of C. pachystachya on human prostate cancer PC3 cell line.

Estrogen Receptors Promote Migration, Invasion and Colony Formation of the Androgen-Independent Prostate Cancer Cells PC-3 Through ß-Catenin Pathway.

Prostate cancer is initially dependent on the androgen, gradually evolves into an androgen-independent form of the disease, also known as castration-resistant prostate cancer (CRPC). At this stage, current therapies scantily improve survival of the patient. Androgens and estrogens are involved in normal prostate and prostate cancer development. The mechanisms by which estrogens/estrogen receptors (ERs) induce prostate cancer and promote prostate cancer progression have not yet been fully identified. Our laboratory has shown that androgen-independent prostate cancer cells PC-3 express both ERα and ERß. The activation of ERß increases the expression of ß-catenin and proliferation of PC-3 cells. We now report that the activation of ERß promotes the increase of migration, invasion and anchorage-independent growth of PC-3 cells. Furthermore, the activation of ERα also plays a role in invasion and anchorage-independent growth of PC-3 cells. These effects are blocked by pretreatment with PKF 118-310, compound that disrupts the complex ß-catenin/TCF/LEF, suggesting that ERs/ß-catenin are involved in all cellular characteristics of tumor development in vitro. Furthermore, PKF 118-310 also inhibited the upregulation of vascular endothelial growth factor A (VEGFA) induced by activation of ERs. VEGF also is involved on invasion of PC-3 cells. In conclusion, this study provides novel insights into the signatures and molecular mechanisms of ERß in androgen-independent prostate cancer cells PC-3. ERα also plays a role on invasion and colony formation of PC-3 cells.