Inhibit Prostate Cancer Proliferation Research Articles

Network pharmacology and experimental validation to explore the role and potential mechanism of Liuwei Dihuang Decoction in prostate cancer

ObjectiveTo evaluate the anti-tumor effector of Liuwei Dihuang Decoction (LWDHD) in prostate cancer (PCa) and explore the potential mechanism using experimental validation, network pharmacology, bioinformatics analysis, and molecular docking.MethodsCCK test, Clone formation assay and wound-healing assays were used to determine the effect of LWDHD on prostate cancer growth and metastasis. The active ingredients and targets of LWDHD were obtained from the TCMSP database, and the relevant targets were selected by GeneCards, OMIM and DisGeNET databases for PCa. The cross-targets of drugs and disease were imported into the STRING database to construct protein interactions. The network was also visualized using Cytoscape software and core targets are screened using the Network Analyzer plug-in. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment were analyzed using R software. TCGA database was used to analyze the correlation of bioinformatics genes. AutoDock vina was used to predict the molecular docking and binding ability of active ingredients to key targets. Through WB and q-PCR experiments, the above gene targets were detected to verify the effect of LWDHD on PCa.ResultsCCK and scratch tests confirmed that LWDHD could inhibit the proliferation, invasion and migration of prostate cancer cells. Clone formation experiments showed that LWDHD inhibited the long-term proliferative capacity of PC3 cells. LWDHD and PCa had a total of 99 common targets, establishing a “drug-ingredient-common target” network. Through GO and KEGG enrichment analysis, PI3K/AKT, MAPK, TP53 pathway, MYC, TNF pathway and other signaling pathways were found. Bioinformatics analysis showed that MYC gene was highly expressed and CCND1 and MAPK1 were low expressed in prostate cancer tissues. In addition, TP53, AKT1, MYC, TNF and CCND1 were positively correlated with MAPK1, among which AKT1 and CCND1 were most closely correlated with MAPK1. Molecular docking results showed that quercetin, kaempferol, β-sitosterol and other main active ingredients of LWDHD treatment for PCa were combined with core proteins MAPK1 and AKT1 well. WB and q-PCR results showed that LWDHD inhibited the expression of PI3K and AKT in PC3 cells.ConclusionThe mechanism of LWDHD therapy for PCa is a multi-target and multi-pathway complex process, which may be related to the biological processes mediated by MAPK1 and AKT1 pathways, such as cell proliferation and inhibition of metastasis, and the regulation of signaling pathways. The PI3K/AKT signaling pathway may be a central pathway of LWDHD to inhibit prostate cancer proliferation.

TRIM47 silencing inhibits the malignant biological behaviors of prostate cancer cells by regulating MDM2/p53 signaling.

Prostate cancer (PCa) is a prevalent male malignancy globally. Tripartite motif 47 (TRIM47) has been reported to be associated with PCa. However, how TRIM47 acts on PCa is still incompletely understood. Here, we explored the biological roles of TRIM47 in PCa cells and investigated its potential regulatory mechanism. TRIM47 expression in PCa cells was detected by qRT-PCR and western blot. After TRIM47 silencing, the viability of PCa cells was measured using CCK-8 method. Flow cytometry was employed to estimate cell cycle. Cell apoptotic level was subjected to appraisement with TUNEL assay. Additionally, wound healing- and transwell assays were adopted for evaluation of migration and invasion of PCa cells. Moreover, the Biogrid database and HDOCK SERVER predicated that TRIM47 could interact with mouse double minute 2 (MDM2), which was detected using the Co-immunoprecipitation (co-IP) assay and glutathione S-transferase (GST) pull-down assay. The expression of proteins in MDM2/p53 signaling was detected by western blot analysis. Results indicated that TRIM47 expression was highly expressed in PCa cells. TRIM47 knockdown inhibited PCa proliferation and cell cycle whereas promoted cell apoptosis. Besides, TRIM47 knockdown significantly inhibited the migration and invasion of PCa cells. In addition, TRIM47 was proved to bind to MDM2 and regulated MDM2/p53 expression. Importantly, MDM2 overexpression counteracted the impacts of TRIM47 knockdown on cell viability, cell cycle, apoptosis, migration and invasion by regulating the MDM2/p53 pathway. Collectively, our results suggested that TRIM47 silencing inhibits the malignant biological behaviors of prostate cancer cells by regulating MDM2/p53 signaling, which may provide a novel therapeutic target for PCa treatment.

Gastrodin inhibits prostate cancer proliferation by targeting canonical Wnt/β-catenin signaling pathway.

Prostate cancer is an epithelial malignant tumor occurring in the prostate and is the most common malignant tumor in the male genitourinary system. In recent years, the incidence of prostate cancer in China has shown a trend of sudden increase. The search for new and effective drugs to treat prostate cancer is therefore extremely important.The canonical Wnt/β-catenin signaling pathway has been shown to be involved in the regulation of tumor proliferation, migration and differentiation. Activation of the canonical Wnt/β-Catenin signaling pathway in the prostate has oncogenic effects. Drugs targeting the canonical Wnt/β-catenin signaling pathway have great potential in the treatment of prostate cancer. In this study, we found that Gastrodin could significantly inhibit the proliferation of prostate cancer cell line PC3 and DU145. Oral administration Gastrodin could significantly inhibit the tumor growth of PC3 cells subcutaneously injected. Gastrodin has an inhibitory effect on canonical Wnt/β-Catenin signaling pathway in Prostate cancer, and this inhibitory effect can be abolished by Wnt/β-Catenin agonist LiCl. These findings raise the possibility that Gastrodin can be used in the treatment of Prostate cancer by targeting canonical Wnt/β-Catenin signaling pathway.

LTBP2 inhibits prostate cancer progression and metastasis via the PI3K/AKT signaling pathway.

Biochemical recurrence (BCR) is a cause of concern in advanced prostate cancer (PCa). Thus, novel diagnostic biomarkers are required to improve clinical care. However, research on PCa immunotherapy is also scarce. Hence, the present study aimed to explore promising BCR-related diagnostic biomarkers, and their expression pattern, prognostic value, immune response effects, biological functions, and possible molecular mechanisms were evaluated. GEO datasets (GSE46602, GSE70768, and GSE116918) were downloaded and merged as the training cohort, and differential expression analysis was performed. Lasso regression and SVM-RFE algorithm, as well as PPI analysis and MCODE algorithm, were then applied to filter BCR-related biomarker genes. The CIBERSORT and estimation of stromal and immune cells in malignant tumor tissues using expression data (ESTIMATE) methods were used to calculate the fractions of tumor-infiltrating immune cells. GO/DO enrichment analyses were used to identify the biological functions. The expression of latent transforming growth factor β-binding protein 2 (LTBP2) was determined by RT-qPCR and western blotting. The role of LTBP2 in PCa was determined by CCK-8, Transwell, and the potential mechanism was investigated by KEGG and GSEA and confirmed by western blotting. In total, 44 BCR-related differentially expressed genes (DEGs) in the training cohort were screened. LTBP2 was found to be a diagnostic biomarker of BCR in PCa and was associated with CD4+ T-cell infiltration and response to anti-PD-1/PD-L1 immunotherapy. Subsequently, using the ESTIMATE algorithm, it was identified that LTBP2 was associated with the tumor microenvironment and could be a predictor of the clinical benefit of immune checkpoint blockade. Finally, the expression and biological function of LTBP2 were evaluated via cellular experiments. The results showed that LTBP2 was downregulated in PCa cells and inhibited PCa proliferation and metastasis via the PI3K/AKT signaling pathway in vitro. In conclusion, LTBP2 was a promising diagnostic biomarker of BCR of PCa and had an important role in CD4+ T-cell recruitment. Moreover, it was associated with immunotherapy in patients with PCa who developed BCR, and it inhibited PCa proliferation and metastasis via the PI3K/AKT signaling pathway in vitro.

FBXW2 inhibits prostate cancer proliferation and metastasis via promoting EGFR ubiquitylation and degradation

FBXW2 is a poorly characterized F-box protein, as a tumor suppressor that inhibits growth and metastasis of lung cancer by promoting ubiquitylation and degradation of oncogenic proteins, including SKP2 and β-catenin. However, what the biological functions of FBXW2 in prostate cancer cells and whether FBXW2 targets other substrates to involve in progression of prostate cancer is still unclear. Here, we reported that overexpression of FBXW2 attenuated proliferation and metastasis of PCa models both in vitro and in vivo, while FBXW2 depletion exhibited the opposite effects. Intriguingly, FBXW2 was an E3 ligase for EGFR in prostate cancer. EGFR protein level and its half-life were extended by FBXW2 depletion, while EGFR protein level was decreased, and its half-life was shortened upon overexpression of FBXW2, but not its dominant-negative mutant. Importantly, FBXW2 bond to EGFR via its consensus degron motif (TSNNST), and ubiquitylated and degraded EGFR, resulting in repression of EGF function. Thus, our data uncover a novel that FBXW2 as a tumor suppressor of prostate cancer, inhibits EGFR downstream by promoting EGFR ubiquitination and degradation, resulting in repression of cell proliferation and metastasis.

The lncRNA NEAT1/miRNA-766-5p/E2F3 Regulatory Axis Promotes Prostate Cancer Progression.

Background Prostate cancer (PCa) is one of the most common malignancies in men. Increasing evidence has demonstrated that dysregulation of long noncoding RNAs (lncRNAs) is closely related to carcinogenesis and cancer progression. lncRNA NEAT1 has recently been identified as a carcinogenic regulator of multiple cancers; however, the role of NEAT1 on PCa is still poorly understood. Methods Kaplan–Meier was conducted to determine the overall survival rate in PCa patients with aberrant NEAT1 levels. qRT-PCR analysis was performed to detect expressions of NEAT1 and miR-766-5p in tissues and cells. In addition, CCK-8, colony formation, flow cytometry analysis, wound healing, and transwell assay were conducted to determine cell proliferation, cell arrest, apoptosis, migration, and invasion. The western blot assay was utilized to determine E2F3 and cell growth-related proteins. The relationship between NEAT1 and miR-766-5p or miR-766-5p and E2F3 was verified by correlation analysis and dual-luciferase reporter assay. Results Here, we find that NEAT1 is overexpressed in PCa tissues and cell lines. Besides, silencing of NEAT1 inhibits cell proliferation, invasion, and migration and promotes cell apoptosis and cell cycle arrest. Further mechanistic studies find that NEAT1 sponges miR-766-5p, and miRNA-766-5p is negatively correlated with the expression of NEAT1. In addition, the functional experiment shows that upregulation of miRNA-766-5p inhibits PCa proliferation, migration, and invasion. Furthermore, E2F transcription factor 3 (E2F3) is testified to be the downstream target gene of miRNA-766-5p. Finally, the rescue experiment revealed that miRNA-766-5p inhibition largely restores NEAT1 downregulation-mediated function on PCa progression, while E2F3 knockdown partly removes the effects of miRNA-766-5p inhibitor. Conclusions In conclusion, NEAT1 facilitates PCa progression by targeting the miRNA-766-5p/E2F3 axis.

Abstract 1152: Targeted inhibition of SIRT6 by engineered exosomes suppressed tumorigenesis and metastasis in prostate cancer

Abstract The great majority of prostate cancer occurrence and development are driven by androgen receptor signaling, which has led to androgen dependent therapy (ADT) as the backbone of systemic therapy for advanced prostate cancer patients until now. Although ADT has improved clinical outcome in patients with castration-sensitive disease and the next-generation androgen receptor pathway inhibitors (ARPIs) like enzalutamide are commonly used in advanced prostate cancer patients, the cross-resistance between AR-targeted drugs significantly limits the therapeutic effect. Besides, early use and potentially long exposure to ARPIs preferentially lead to the evolvement of the alternative downstream signaling instead of AR pathway. Hence, targeting the AR fails to develop inescapable drug resistance, and no effective approach is available for targeting metastatic and castration-resistant prostate cancer (mCRPC), which is a major clinical challenge in prostate cancer treatment. Undoubtedly, new targets and therapeutic strategies are needed to improve the clinical outcomes. Sirt6 belongs to the Sirtuin family proteins, and as a nuclear protein, regulates diverse physiological functions like aging and metabolism. Silencing SIRT6 could inhibit prostate cancer proliferation, and in this study, we further exploited this phenomenon. We found that expression of SIRT6 was positively related with the Gleason score of the patients' prostate tissue sample and knockdown the expression of SIRT6 in prostate cancer cell lines significantly reduced migration and invasion both in vitro and in vivo. Furthermore, SIRT6 deficiency upregulated the expression of E-cadherin and downregulated the expression of N-cadherin and Vimentin, suggesting its role in regulation of epithelial-mesenchymal transition (EMT). Silencing SIRT6 inhibited both the proliferation and metastasis in prostate cancer, indicating the therapeutic value of targeting SIRT6 in the treatment of metastatic PCa. We then tested the therapeutic value of SIRT6 in prostate cancer in vivo by using aptamer-modified exosomes carrying SIRT6 siRNA. To achieve this goal, the cholesterol-covalent E3 aptamer was inserted into the membrane of 293T-derived exosomes to target specifically the prostate cancer cells, followed by SIRT6-siRNA loading into the modified exosomes with an electroporation apparatus. The engineered exosomes suppressed the proliferation and metastasis in subcutaneously and orthotopically xenograft mice tumor models, and confirmed that SIRT6 could be considered as a selective and novel target in high-grade prostate cancer. Citation Format: Qing Han, Qian Reuben Xie, Fan Li, Yirui Cheng, Tingyu Wu, Jianjun Sha, Weiliang Xia. Targeted inhibition of SIRT6 by engineered exosomes suppressed tumorigenesis and metastasis in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1152.

Abstract 1274: Metformin, not γ-tocotrienol, increases gene methylation and decreases expression of genes responsible for cell signal transduction in prostate cancer cells

Abstract Metformin and γ-tocotrienol (GT3) are promising chemopreventative agents with studies showing improved patient outcomes with metformin use in several cancer types, including prostate cancer. However, the cellular mechanisms underlying the improved survival rates in cancer patients treated with these drugs are incompletely understood. Inhibition of mammalian target of rapamycin (mTOR), alteration of p-c-JUN and p-ERK signaling pathways and inhibition of mitochondrial complex 1 have all been implicated as part of metformin's anti-tumorigenesis activities. More recently, metformin was shown to influence the activity of epigenetic modifying enzymes resulting in genome-wide alterations in DNA methylation patterns in breast cancer cell lines. In this study, we investigated whether GT3 and metformin treatment would alter methylation pathways in prostate cancer cells.The prostate cancer cell line PC-3 (androgen independent) and normal prostate control cell line RWPE-1 were treated with varying concentrations of either GT3, metformin, the methylation inhibitor azacitidine, or metformin plus azacitidine for 6 hours. Genomic DNA was isolated, and methylation assessed using the Illumina Infinium methylation EPIC BeadChip array. Analysis revealed treatment of PC-3 cells with metformin promoted hypermethylation of CC2D2B, GHDC, LINC01377, PLPP2, and PTK2 and decreased methylation of TSPY4 as compared to normal prostate controls. qRT-PCR experiments confirmed decreased expression of PTK2 mRNA, consistent with hypermethylation of PTK2. In contrast, GT3 did not affect methylation of any identifiable recognizable genes. This study suggests metformin, but not GT3, may inhibit prostate cancer proliferation partially through epigenetic changes which disrupt downstream focal adhesion kinase (FAK) signaling pathways. Metformin is a promising chemopreventive agent; studies investigating the impact of metformin on cellular adhesion and FAK pathways are ongoing in our laboratory. Citation Format: Christine Ann Moore, Janet Lightner, Michelle Duffourc, Koyamangalath Krishnan. Metformin, not γ-tocotrienol, increases gene methylation and decreases expression of genes responsible for cell signal transduction in prostate cancer cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1274.

A Novel Small Molecule Inhibits Tumor Growth and Synergizes Effects of Enzalutamide on Prostate Cancer.

Prostate cancer (PCa) is the second leading cause of cancer-related death for men in the United States. Approximately 35% of PCa recurs and is often transformed to castration-resistant prostate cancer (CRPCa), the most deadly and aggressive form of PCa. However, the CRPCa standard-of-care treatment (enzalutamide with abiraterone) usually has limited efficacy. Herein, we report a novel molecule (PAWI-2) that inhibits cellular proliferation of androgen-sensitive and androgen-insensitive cells (LNCaP and PC-3, respectively). In vivo studies in a PC-3 xenograft model showed that PAWI-2 (20 mg/kg per day i.p., 21 days) inhibited tumor growth by 49% compared with vehicle-treated mice. PAWI-2 synergized currently clinically used enzalutamide in in vitro inhibition of PCa cell viability and resensitized inhibition of in vivo PC-3 tumor growth. Compared with vehicle-treated mice, PC-3 xenograft studies also showed that PAWI-2 (20 mg/kg per day i.p., 21 days) and enzalutamide (5 mg/kg per day i.p., 21 days) inhibited tumor growth by 63%. Synergism was mainly controlled by the imbalance of prosurvival factors (e.g., Bcl-2, Bcl-xL, Mcl-1) and antisurvival factors (e.g., Bax, Bak) induced by affecting mitochondrial membrane potential/mitochondria dynamics. Thus, PAWI-2 utilizes a distinct mechanism of action to inhibit PCa growth independently of androgen receptor signaling and overcomes enzalutamide-resistant CRPCa. SIGNIFICANCE STATEMENT: Castration-resistant prostate cancer (CRPCa) is the most aggressive human prostate cancer (PCa) but standard chemotherapies for CRPCa are largely ineffective. PAWI-2 potently inhibits PCa proliferation in vitro and in vivo regardless of androgen receptor status and uses a distinct mechanism of action. PAWI-2 has greater utility in treating CRPCa than standard-of-care therapy. PAWI-2 possesses promising therapeutic potency in low-dose combination therapy with a clinically used drug (e.g., enzalutamide). This study describes a new approach to address the overarching challenge in clinical treatment of CRPCa.

A novel androgen-reduced prostate-specific lncRNA, PSLNR, inhibits prostate-cancer progression in part by regulating the p53-dependent pathway.

Prostate cancer (PCa) is one of the most common cancers in males in China. Long noncoding RNAs (lncRNAs) reportedly play crucial roles in human cancer progression in many studies. However, the molecular mechanisms underlying PCa progression remain unclear. We investigated the lncRNA transcriptome using publicly available RNA-sequencing data to identify prostate-specific lncRNAs. Then, the chromatin immunoprecipitation (ChIP) assay identified lncRNA with a direct binding to androgen receptor (AR), hereafter denoted as PSLNR. Quantitative real-time polymerase chain reaction analysis and Western blot analysis were performed to detect the expression of p53 signaling-related genes after overexpression PSLNR. The effects of overexpression of PSLNR on cell proliferation, cell cycle, and cell apoptosis were assessed by using CCK-8 and flow cytometric analysis. We then detected the expression of PSLNR in tissues. We reported a novel androgen-reduced prostate-specific lncRNA, PSLNR, that inhibited PCa progression via the p53-dependent pathway. By analyzing the NOCODE data set, we reported that PSLNR was specifically expressed in the prostate, suggesting the potential of PSLNR as a biomarker for PCa treatment. The AR pathway was also confirmed to be an upstream regulation signaling pathway of PSLNR by transcriptionally regulating its expression in androgen-dependent PCa cells. PSLNR also significantly inhibited PCa proliferation by inducing cell apoptosis in a p53-dependent manner. Thus, PSLNR may be a candidate diagnosis and therapeutic target for PCa. Our study revealed for the first time a novel androgen-reduced prostate-specific lncRNA, PSLNR, which inhibited PCa progression via the p53-dependent pathway, suggesting that PSLNR may be a candidate diagnosis and therapeutic target for PCa.

MiR-129-5p inhibits prostate cancer proliferation via targeting ETV1.

BackgroundProstate cancer is one of the most commonly diagnosed diseases in males.MethodsRT-qPCR was used to detect miR-129-5p expression in tumor tissues and adjacent normal tissues from patients with prostate cancer. The cell proliferation assay and colony forming assay were used to study the role of miR-129-5p in mediating prostate cancer cell growth. Bioinformatic analysis and dual luciferase assay were performed to predict and confirm ETV1 as a target gene of miR-129-5p.ResultsWe found that miR-129-5p levels were decreased significantly in human prostate cancer tissues compared with matched normal tissues from patients with prostate cancer. Overexpression of miR-129-5p suppressed prostate cancer cell growth while antagonist of miR-129-5p promoted cell proliferation in immortal prostate cell line RWPE-1. In addition, elevation of miR-129-5p decreased ETV1 expression in prostate cancer cells while downregulation of miR-129-5p increased ETV1 expression in RWPE-1. Mechanistically, ETV1 is confirmed a direct target of miR-129-5p in prostate cancer cells. Through repression of ETV1 expression, miR-129-5p could inactivate YAP signaling in prostate cancer cells. In addition, overexpression of ETV1 attenuated miR-129-5p induced cell proliferation in prostate cancer cells. Correlation analysis further revealed that there was a negative correlation between miR-129-5p levels and ETV1 mRNA levels in tumor tissues from patients with prostate cancer.ConclusionOur results identified miR-129-5p as a tumor suppressor in prostate cancer via repression of ETV1.

MicroRNA-588 is upregulated in human prostate cancer with prognostic and functional implications.

Epigenetic factors of microRNAs (miRNAs) are important biomarkers and modulators in human prostate cancer (PCa). In this work, we characterized the expression, biomarker-potential and functional regulation of miR-588 in PCa. Endogenous miR-588 levels were quantified by qRT-PCR in both prostate-specific antigen (PSA)-negative and PSA-positive PCa cell lines, as well as human PCa tumors. The associations between endogenous miR-588 and PCa patients' clinical outcomes and postoperative overall survival were statistically examined. Moreover, in both PSA-negative DU145 and PSA-positive LNCaP, downregulation of miR-588 was achived by transduction of miR-588 inhibitor lentivirus. The subsequent effects of miR-588 downregulation on PCa cell developments were investigated both in vitro and in vivo. MiR-588 was profoundly upregulated in both PSA-negative and PSA-positive PCa cells, as well as in PCa tumors. Significant miR-588 upregulation was found to be closely associated with PCa patients' poor clinical outcomes and shorter postoperative overall survivals. In DU145 and LNCaP cell lines, lentiviral transduction markedly downregulated endogenous miR-588 levels. MiR-588 downregulation was shown to profoundly inhibit PCa proliferation in vitro and xenograft in vivo. Aberrant upregulation of endogenous miR-588 in PCa patients may be a prognostic biomarker, indicative of their poor clinical outcomes. Inhibiting endogenous miR-588 may also serve as a therapeutic target for PCa treatment. This article is protected by copyright. All rights reserved.

Lobaplatin Inhibits Prostate Cancer Proliferation and Migration Through Regulation of BCL2 and BAX.

Lobaplatin is a diastereometric mixture of platinum (II) complexes, which contain a 1,2-bis (aminomethyl) cyclobutane stable ligand and lactic acid. Previous studies have showed that lobaplatin plays inhibiting roles in various types of tumors. However, the role of lobaplatin in prostate cancer remains unknown. Cell viability was detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay. Cell proliferation was detected by cell colony formation assay. Cell migration and invasion were determined by transwell migration and invasion assay. Cell apoptosis was detected by flow cytometry. The messenger RNA and protein expression levels were detected by quantitative real-time polymerase chain reaction and Western blot. Lobaplatin treatment inhibits cell viability, cell proliferation, cell migration, and invasion, while promotes cell apoptosis of prostate cancer cell lines DU145 and PC3. Meanwhile, lobaplatin treatment regulates apoptosis by downregulation of BCL2 expression and upregulation of BAX expression levels. Our study suggests lobaplatin inhibits prostate cancer proliferation and migration through regulation of BCL2 and BAX expression.

Long noncoding RNA SNHG7 accelerates prostate cancer proliferation and cycle progression through cyclin D1 by sponging miR-503

Increasing evidence has indicated the important roles of long non-coding RNAs (lncRNAs) in tumorigenesis and cellular progression, including prostate cancer. In this study, we aim to investigate the expression level of SNHG7 and its biological functions on prostate cancer cells. Results indicated that SNHG7 expression was significantly up-regulated in prostate cancer tissue and cell lines. Besides, the overexpression of SNHG7 was closely correlated with the poor prognosis. In vitro and in vivo, experiments demonstrated that SNHG7 knockdown markedly inhibited prostate cancer proliferation and cycle-related protein (CDK4, CDK6, Cyclin D1), induced cell cycle arrest at G0/G1 phase and suppressed tumor growth. Moreover, miR-503 was predicted by bioinformatics tools and validated using luciferase reporter assay to both directly inhibited SNHG7 and Cyclin D1 expression by targeting their RNA 3′-UTR. In conclusion, results present that SNHG7 regulates the cycle progression and acts as an oncogenic gene in the prostate cancer tumorigenesis via miR-503/Cyclin D1 pathway, revealing the vital role of lncRNA/miRNA/mRNA axis in prostate cancer carcinogenesis.

Inhibition of prostate cancer proliferation by Deferiprone.

Cancer growth and proliferation rely on intracellular iron availability. We studied the effects of Deferiprone (DFP), a chelator of intracellular iron, on three prostate cancer cell lines: murine, metastatic TRAMP-C2; murine, non-metastatic Myc-CaP; and human, non-metastatic 22rv1. The effects of DFP were evaluated at different cellular levels: cell culture proliferation and migration; metabolism of live cells (time-course multi-nuclear magnetic resonance spectroscopy cell perfusion studies, with 1-13 C-glucose, and extracellular flux analysis); and expression (Western blot) and activity of mitochondrial aconitase, an iron-dependent enzyme. The 50% and 90% inhibitory concentrations (IC50 and IC90 , respectively) of DFP for the three cell lines after 48h of incubation were within the ranges 51-67μM and 81-186μM, respectively. Exposure to 100μM DFP led to: (i) significant inhibition of cell migration after different exposure times, ranging from 12h (TRAMP-C2) to 48h (22rv1), in agreement with the respective cell doubling times; (ii) significantly decreased glucose consumption and glucose-driven tricarboxylic acid cycle activity in metastatic TRAMP-C2 cells, during the first 10h of exposure, and impaired cellular bioenergetics and membrane phospholipid turnover after 23h of exposure, consistent with a cytostatic effect of DFP. At this time point, all cell lines studied showed: (iii) significant decreases in mitochondrial functional parameters associated with the oxygen consumption rate, and (iv) significantly lower mitochondrial aconitase expression and activity. Our results indicate the potential of DFP to inhibit prostate cancer proliferation at clinically relevant doses and plasma concentrations.

MicroRNA-1297 inhibits prostate cancer cell proliferation and invasion by targeting the AEG-1/Wnt signaling pathway

MicroRNAs (miRNAs) have been known to be implicated in tumorigenic programs. miR-1297 has been reported to be dysregulated and involved in cancer progression in many types of human cancers. However, the expression level and the role of miR-1297 in prostate cancer remain unclear. Herein, we aimed to investigate the potential role and molecular mechanism of miR-1297 in prostate cancer progression. We found that miR-1297 was significantly downregulated in human prostate cancer specimens as well as in several prostate cancer cell lines. In addition, functional experiments demonstrated that overexpression of miR-1297 remarkably inhibited prostate cancer cell proliferation and invasion whereas miR-1297 suppression significantly promoted prostate cancer cell proliferation and invasion. Bioinformatics analysis showed that the Astrocyte elevated gene-1 (AEG-1), a well-known oncogene, is a predicted target of miR-1297. Dual-luciferase reporter assay showed that miR-1297 was able to directly target the 3’-untranslated region of AEG-1. In addition, RT-qPCR and Western blot analysis showed that miR-1297 regulated the mRNA and protein expression levels of AEG-1. We also showed that miR-1297 was able to regulate the Wnt signaling pathway. Moreover, rescue assays indicated that AEG-1 contributed to miR-1297-endowed effects on cell proliferation and invasion as well as Wnt signaling pathway. Taken together, these findings suggest that miR-1297 inhibits prostate cancer proliferation and invasion by targeting AEG-1, thereby providing novel insight into understanding the pathogenesis of prostate cancer. Thus, miR-1297 may be a novel potential therapeutic candidate to treat prostate cancer.

Epigenetics Reactivation of Nrf2 in Prostate TRAMP C1 Cells by Curcumin Analogue FN1.

It has previously been shown that curcumin can effectively inhibit prostate cancer proliferation and progression in TRAMP mice, potentially acting through the hypomethylation of the Nrf2 gene promoter and hence activation of the Nrf2 pathway to enhance cell antioxidative defense. FN1 is a synthetic curcumin analogue that shows stronger anticancer activity than curcumin in other reports. We aimed to explore the epigenetic modification of FN1 that restores Nrf2 expression in TRAMP-C1 cells. Stably transfected HepG2-C8 cells were used to investigate the effect of FN1 on the Nrf2- antioxidant response element (ARE) pathway. Real-time quantitative PCR and Western blotting were applied to study the influence of FN1 on endogenous Nrf2 and its downstream genes. Bisulfite genomic sequencing (BGS) and methylated DNA immunoprecipitation (MeDIP) were then performed to examine the methylation profile of the Nrf2 promoter. An anchorage-independent colony-formation analysis was conducted to examine the tumor inhibition activity of FN1. Epigenetic modification enzymes, including DNMTs and HDACs, were investigated by Western blotting. The luciferase reporter assay indicated that FN1 was more potent than curcumin in activating the Nrf2-ARE pathway. FN1 increased the expression of Nrf2 and its downstream detoxifying enzymes. FN1 significantly inhibited the colony formation of TRAMP-C1 cells. BGS and MeDIP assays revealed that FN1 treatment (250 nM for 3 days) reduced the percentage of CpG methylation of the Nrf2 promoter. FN1 also downregulated epigenetic modification enzymes. In conclusion, our results suggest that FN1 is a novel anticancer agent for prostate cancer. In the TRAMP-C1 cell line, FN1 can increase the level of Nrf2 and downstream genes via activating the Nrf2-ARE pathway and inhibit the colony formation potentially through the decreased expression of keap1 coupled with CpG demethylation of the Nrf2 promoter. This CpG demethylation effect may come from decreased epigenetic modification enzymes, such as DNMT1, DNMT3a, DNMT3b, and HDAC4.

251 Down-regulation of CDKN3 inhibited prostate cancer proliferation in vitro and in vivo via regulating cell cycle and DNA replication signaling

251 Down-regulation of CDKN3 inhibited prostate cancer proliferation in vitro and in vivo via regulating cell cycle and DNA replication signaling

Glucose-regulated protein 78 mediates the anticancer efficacy of shikonin in hormone-refractory prostate cancer cells.

Glucose-regulated protein 78 (GRP78) is a key modulator of prostate cancer progression and therapeutic resistance. Prostate cancer is a worldwide health problem, and therapeutic resistance is a critical obstacle for the treatment of hormone-refractory prostate cancer patients. Shikonin inhibits prostate cancer proliferation and metastasis. However, the role of GRP78 in the cytotoxic effect of shikonin in prostate cancer cells remains unclear. GRP78 expression was abolished using small interfering RNA (siRNA), and the anticancer effects of shikonin were assessed using MTT assays, the XCELLigence biosensor, flow cytometric cell cycle analysis, and Annexin V-PI apoptotic assays. PC-3 cells expressed more GRP78 than DU-145 cells, and the MTT assays revealed that DU-145 cells were more sensitive to shikonin than PC-3 cells. GRP78 knockdown (GRP78KD) PC-3 cells were more sensitive to shikonin treatment than scrambled siRNA control cells. Based on cell cycle analysis and AnnexinV-PI apoptotic assays, apoptosis dramatically increased in GRP78KD cells compared with the control PC-3 in response to shikonin. Finally, in response to shikonin treatment, Mcl-1 and Bcl-2 levels increased in the scrambled control cells treated with shikonin, whereas Bcl-2 decreased and Mcl-1 slightly increased in the GRP78KD PC-3 cells. The levels of Bax and Bad did not change in the scrambled control or GRP78KD cells after shikonin treatment. These results are consistent with the increased sensitivity to shikonin after knockdown of GRP78. GRP78 expression may determine the therapeutic efficacy of shikonin against prostate cancer cells.

Metadherin/astrocyte elevated gene-1-based DNA vaccine suppresses progression and metastasis in prostate cancer of mice

Objective To study the metadherin (MTDH)/astrocyte elevated gene-1 (AEG-1) as targets for gene vaccine inducing immunosuppression mice prostate cancer growth and metastasis of action research.Methods Male C57BL/6 mice (6-8 weekes old) were randomly divided into three groups (n =20),respectively,to phosphate buffered saline (PBS),Pub,Pub-MTDH/AEG-1 attenuated salmonella (1 × 10s cfu) of lavage servo immune; Immune once a week,a total of immune three times.Last one week after the immunization,all in three groups of mice prostate in situ (right and left dorsal lobe) vaccinated 10s RM-1 cell to build orthotopic transplantation tumor model of prostate,induced spontaneous pelvic cavity and retroperitoneal lymph node metastasis; Record each group mice with tumor size,pelvic cavity and retroperitoneal lymph node metastasis number [hematoxylin and eosin (HE) staining shall prevail],CD8 in through immunohistochemical method to detect in situ tumor,the expression of vascular endothelial growth factor (VEGF) and lymphatic vessel endothelial hyaluronic acid receptor-1 (LYVE-1) case,tumor cell apoptosis with TdT-mediated dUTP nick end labeling (TUNEL) method.Results pUb-MTDH/AEG-1 DNA vaccine group compared to the pUb,the PBS group could significantly stimulate CD8 + T cells and cytotoxic T lymphocyte (CTL) response; Effectively inhibit the growth of prostate cancer in situ in mice,MTDH group tumor volume (0.248 ± 0.102) compared with the Pub group (1.475 ± 0.314) (P ＜ 0.01),pelvic cavity and retroperitoneal lymph node metastasis 77.78％,35.7％ (P ＜ 0.05) ; Significantly enhanced the expression of CD8 + in the in situ tumor molecular:MTDH group (3.56 ± 0.85),Pub group (5.12 ± 0.90) (P ＜ 0.05),and promote tumor cell apoptosis:MTDH group (39.60 ± 3.28) ％,Pub group (16.18 ±2.52)％ (P＜0.01).Conclusion MTDH/targets AEG-1 gene vaccine in mice prevention immune models,can effectively induce the body specificity immune response,enhance cellular immunity and humoral immunity,inhibit prostate cancer proliferation,invasion,and promote tumor cell apoptosis,extend a tumor-burdened survival time in mice. Key words: Metadherin/astrocyte elevated gene-1 ; Gene vaccine; Prostate cancer; Metastasis