Behavior Of Prostate Cancer Cells Research Articles

Comparative study of prostate cancer biophysical and migratory characteristics via iterative mechanoelectrical properties (iMEP) and standard migration assays

This study reveals a new microfluidic biosensor consisting of a multi-constriction microfluidic device with embedded electrodes for measuring the biophysical attributes of single cells. The biosensing platform called the iterative mechano-electrical properties (iMEP) analyzer captures electronic records of biomechanical and bioelectrical properties of cells. The iMEP assay is used in conjunction with standard migration assays, such as chemotaxis-based Boyden chamber and scratch wound healing assays, to evaluate the migratory behavior and biophysical properties of prostate cancer cells. The three cell lines evaluated in the study each represent a stage in the standard progression of prostate cancer, while the fourth cell line serves as a normal/healthy counterpart. Neither the scratch assay nor the chemotaxis assay could fully differentiate the four cell lines. Furthermore, there was not a direct correlation between wound healing rate or the migratory rate with the cells’ metastatic potential. However, the iMEP assay, through its multiparametric dataset, could distinguish between all four cell line populations with p-value < 0.05. Further studies are needed to determine if iMEP signatures can be used for a wider range of human cells to assess the tumorigenicity of a cell population or the metastatic potential of cancer cells.

CircFMN2 Sponges miR-1238 to Promote the Expression of LIM-Homeobox Gene 2 in Prostate Cancer Cells

Circular RNAs (circRNAs) regulate gene expression in different malignancies. However, the molecular mechanisms that link circRNAs with the tumorigenesis of prostate cancer (PCa) are not well understood. In the present study, we attempted to provide a novel basis for targeted therapy for PCa from the aspect of circRNA-microRNA (miRNA)-mRNA interaction. We investigated the expression of circRNAs in 5 paired PCa tissues and adjacent non-tumor tissues by microarray analysis. We focused on hsa_circ_0005100, which is located on chromosome 1 and derived from FMN2, and thus we named it circFMN2. The qRT-PCR was used to detect circFMN2 and target miRNA expression in PCa tissues and cell lines. Biological functional experiments were performed to detect the effects of circFMN2 on the biological behavior of PCa cells in vivo and in vitro. Bioinformatic analysis was utilized to predict potential miRNA target sites on circFMN2. High expression of circFMN2 was associated with PCa progression. Function assays revealed that knockdown of circFMN2 significantly reduced PCa cell growth in vitro and in vivo. Finally, we found that circFMN2 acts as a competing endogenous RNA (ceRNA) for miR-1238 to regulate LIM-homeobox gene 2 (LHX2) expression. circFMN2 regulates the miR-1238/LHX2 axis to promote PCa progression.

Breviscapine suppresses the growth and metastasis of prostate cancer through regulating PAQR4-mediated PI3K/Akt pathway

ObjectivesProstate cancer, one of the most frequently diagnosed tumors of men, leads to poor quality of life. Previous studies have shown that breviscapine (BRE) exerts therapeutic activity in malignant tumors. However, the role and mechanism of BRE exhibit an anti-tumor effect on prostate cancer are largely unknown. MethodsThe mRNA and protein levels in prostate cancer tissues and cell lines were measured using RT-qPCR, western blot, and immunohistochemical staining, respectively. Cell proliferation, invasion, and migration in both PC3 and DU145 cells were evaluated using CCK-8 and Transwell assay. The effect of BRE on cell proliferation and metastasis by regulating the PAQR4-mediated PI3K/Akt pathway in vitro and in vivo was determined. ResultsPAQR4 was significantly overexpressed in prostate cancer tissues and cell lines, which was positively correlated with poor prognosis. Knockdown of PAQR4 inhibited the proliferation, invasion, migration, and epithelial-mesenchymal transition (EMT) of both PC3 and DU145 cells. Mechanistically, BRE treatment significantly suppressed the malignant biological behavior of both prostate cancer cells by downregulating PAQR4 and blocking the PI3K/Akt pathway. In vivo experiments, BRE administration remarkably inhibited tumor growth and metastasis in a xenograft model of prostate cancer. ConclusionOur findings revealed that BRE exerts anti-tumor and anti-metastasis roles in prostate cancer by inhibiting PAQR4-mediated PI3K/Akt pathway, which provides a new therapeutic agent for prostate cancer clinical treatment.

Role of metabolic imaging in diagnosis of primary, metastatic, and recurrent prostate cancer.

The present review describes the current role of metabolic imaging techniques such as multiparametric MRI (mpMRI), magnetic resonance spectroscopic imaging (MRSI), hyperpolarized MRSI, and positron emission tomography (PET) in the diagnosis of primary prostate cancer, surveillance of low-grade disease, detection of metastases, and evaluation of biochemical recurrence after therapy. The natural history of prostate cancer ranges from indolent disease that is optimally monitored by active surveillance, to highly aggressive disease that can be lethal. Current diagnostic methods remain imperfect in noninvasively distinguishing between silent versus aggressive tumors. Hence, there is a high demand for noninvasive imaging techniques that offer insight into biological behavior of prostate cancer cells. Characterization of prostate cancer metabolism is a promising area to provide such insights. Metabolic imaging may allow for greater detection and ultimately characterization of tumor based on aggressiveness and spread. Hence, it has the potential to monitor tumor activity, predict prognostic outcomes, and guide individualized therapies.

&lt;p&gt;lncRNA HAND2-AS1 Regulates Prostate Cancer Cell Growth Through Targeting the miR-106a-5p/RBM24 Axis&lt;/p&gt;

IntroductionIncreasing evidence has shown that abnormally expressed long non-coding RNA (lncRNA) plays crucial roles in prostate cancer (PCa) progression.Materials and MethodsHere, we analyzed the expression level of lncRNA HAND2 antisense RNA 1 (HAND2-AS1) in PCa cells and tissues. Function assays were performed to investigate the biological roles of HAND2-AS1 in PCa cell behaviors. Bioinformatics methods, luciferase activity reporter assay, and RNA pull-down assay were performed to validate the connection of microRNA-106a-5p (miR-106a-5p) with HAND2-AS1. Also, the target of miR-106a-5p was explored using the same methods.ResultsOur results revealed HAND2-AS1 expression was decreased in both PCa cells and tissues. In vitro functional assays showed HAND2-AS1 could inhibit PCa cell proliferation and colony formation through promoting cell apoptosis. Dual-luciferase activity assays showed miR-106a-5p could directly bind with HAND2-AS1 and RNA binding motif protein 24 (RBM24). Mechanistically, we showed that HAND2-AS1 regulates PCa cell behaviors via targeting miR-106a-5p/RBM24 axis.ConclusionIn summary, our results illustrated that HAND2-AS1 functions as miR-106a-5p sponge to regulate RBM24 expression, and to influence PCa progression.

CMTM5-v1 inhibits cell proliferation and migration by downregulating oncogenic EGFR signaling in prostate cancer cells.

Anomalous epidermal growth factor receptor (EGFR) signaling plays an important role in the progression of prostate cancer (PCa) and the transformation to castration-resistant PCa (CRPC). A novel tumor suppressor CKLF-like MARVEL transmembrane domain-containing member 5(CMTM5) has a MARVEL domain and may regulate transmembrane signaling. Thus, we postulated that CMTM5 could regulate EGFR and its downstream molecules to affect the biological behaviors of PCa cells. In this study, we found that CMTM5 was expressed in benign prostatic hyperplasia (BPH) tissues but was undetectable in PCa cells. However, the EGFR was upregulated in PCa cells, especially in two metastatic CRPC cell lines, PC3 and DU145. Furthermore, ectopic expression of CMTM5-v1 suppressed cell proliferation and migration and p-EGFR levels. Further investigation revealed that restoration of CMTM5-v1 inhibited not only EGF-mediated proliferation but also chemotactic migration by EGF in PC3 and DU145 cells. Moreover, mechanistic studies showed that CMTM5-v1 attenuated EGF-induced receptor signaling by repressing EGFR and Akt phosphorylation in PCa cells, which were essential for malignant features. Finally, CMTM5-v1can promote the sensitivity of PC3 cells to Gefetinib, a tyrosine kinase inhibitor (TKI) targeting the EGFR. These observations indicate that CMTM5-v1 suppressed PCa cells through EGFR signaling. The loss of CMTM5 may participate in the progression of PCa resulting from deregulated EGFR, and CMTM5 might be associated with the efficacy of TKIs in terms of their potent inhibition of EGFR and human epidermal growth factor-2 (HER2) activation.

Cancer-Derived Exosomal miR-199b-5p Inhibits Distant Metastases of Prostate Cancer by Counteracting the DDR1-MAPK/ERK-EMT Pathway

Metastasis remains a primary challenge in treating prostate cancer (PCa). In this study, we analyzed the transcriptomes of localized PCa and bone metastatic PCa patients derived plasma exosomes by using high-throughput RNA sequencing. We found that miR-199b-5p was down-regulation in plasma exosomes of bone metastatic PCa, primary PCa tissues of metastatic patients, as well as bone-related metastatic PCa cells. Over-expressed miR-199b-5p could significantly suppress the proliferation, migration, and metastatic behavior of PCa cells. Clinical data form Taylor dataset indicated that low miR-199b-5p expression level was dramatically correlated with high Gleason score, advanced pathological stage, positive metastasis, and biochemical recurrence of PCa patients. DDR1 (discoid domain receptor 1), the direct target of miR-199b-5p, was responsible for miR-199b-5p down-regulation mediated PCa progression. This process might associate with the activation of MAPK/ERK signaling and the development of EMT. The clinical data from our hospital, TCGA dataset, and GEO database collectively suggested a positive relationship between DDR1 expression and the poor prognosis of PCa. Collectively, exosomal miR-199b-5p-DDR1 axis might act as the potential metastatic diagnostic biomarkers for PCa and provide a new therapeutic strategy for metastatic PCa. Funding Statement: This work was supported by the grants from National Natural Science Foundation of China (No. 81372774, No. 81572537 for Zhigang Zhao), The Key Program of Natural Science Foundation of Guangdong Province (No. 2015A030311007 for Zhigang Zhao), Science and Technology Program of Guangzhou (No. 201607010376 for Zhigang Zhao), and The Major Program of Department of Guangdong Education (No. 2017KZDXM067 for Zhigang Zhao). Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The present study was approved by the Human Study Ethics Committees of the First Affiliated Hospital of Guangzhou Medical University. All of the patients have signature informed consent. Human specimens were handled and made anonymous in adherence to the ethical and legal standards.

EHMT2 promotes the development of prostate cancer by inhibiting PI3K/AKT/mTOR pathway.

To investigate the expression characteristics and the potential role of euchromatic histone-lysine N-methyltransferase 2 (EHMT2) in the clinical pathology and prognosis of prostate cancer (PCa). Quantitative Real-time polymerase chain reaction (qRT-PCR) was performed to detect the expression of EHMT2 in 55 pairs of PCa tissues and adjacent normal tissues. The relationship between EHMT2 expression and the pathological features, as well as the prognosis of PCa patients was analyzed. EHMT2 expression in PCa cells was determined by qRT-PCR as well. In addition, EHMT2 knockdown model was constructed by transfection of the small interfering RNA in PCa cell lines PC-3 and DU-145. The regulatory effects of EHMT2 on the behaviors of PCa cells were evaluated through cell counting kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), colony formation assay and flow cytometry. Finally, we detected the protein levels of relative genes in PI3K/AKT/mTOR pathway through Western blot. QRT-PCR results showed that the expression level of EHMT2 in PCa tissues was markedly higher than that in adjacent normal tissues. Compared with PCa patients with low expression of EHMT2, those with high expression of EHMT2 had higher pathological grade and lower overall survival. EHMT2 was also highly expressed in PCa cell lines. Knockdown of EHMT2 inhibited the proliferative potential and induced apoptosis of PCa cells. Western blot results revealed that PCa cells with EHMT2 knockdown presented downregulated p-PI3K, p-AKT and p-mTOR. EHMT2 was highly expressed in PCa, and its expression is closely correlated with tumor stage and poor prognosis of PCa patients. EHMT2 promoted the malignant progression of PCa by inhibiting PI3K/AKT/mTOR pathway.

Cell Responses to Electrical Pulse Stimulation for Anticancer Drug Release.

Electrical stimulation is an attractive approach to tune on-demand drug release in the body as it relies on simple setups and requires typically 1 V or less. Although many studies have been focused on the development of potential smart materials for electrically controlled drug release, as well as on the exploration of different delivery mechanisms, progress in the field is slow because the response of cells exposed to external electrical stimulus is frequently omitted from such investigations. In this work, we monitor the behavior of prostate and breast cancer cells (PC-3 and MCF7, respectively) exposed to electroactive platforms loaded with curcumin, a hydrophobic anticancer drug. These consist in conducting polymer nanoparticles, which release drug molecules by altering their interactions with polymer, and electrospun polyester microfibres that contain electroactive nanoparticles able to alter the porosity of the matrix through an electro-mechanical actuation mechanism. The response of the cells against different operating conditions has been examined considering their viability, metabolism, spreading and shape. Results have allowed us to differentiate the damage induced in the cell by the electrical stimulation from other effects, as for example, the anticancer activity of curcumin and/or the presence of curcumin-loaded nanoparticles or fibres, demonstrating that these kinds of platforms can be effective when the dosage of the drug occurs under restricted conditions.

MiR-203 inhibits the malignant behavior of prostate cancer cells by targeting RGS17.

The aim of this study was to explore the role of microRNA-203 (miR-203) in Prostate Cancer (PCa), and to further verify its influence in PCa cell function. The expression level of miR-203 in 55 clinical PCa cases and cell lines was detected by qRT-PCR. Then, the target gene of miR-203 in PCa cells was predicted and verified by online prediction software and Luciferase reporter gene assay, respectively. Furthermore, the role of miR-203 in PCa cell proliferation, colony formation, cell cycle and metastasis capacities was detected through a series of in vitro experiments. The expression of miR-203 in PCa tissues and cells was significantly reduced when compared with that of normal tissues and cells. In searching for potential downstream targets of miR-203, a regulator of G-protein signaling 17 (RGS17) entered our sight due to its active role in a variety of malignant tumors. More importantly, the negative regulation of RGS17 by miR-203 was verified by Luciferase reporter gene assay. Functional experiments demonstrated that low expression of RGS17 in PCa cells induced by up-regulation of miR-203 could significantly restrain the proliferation, invasion and migration capacities of PCa cells. MiR-203 served as a tumor suppressor gene in PCa. Through targeting RGS17, miR-203 significantly controlled the malignant behavior of PCa cells. Our findings revealed that miR-203/RGS17 axis might be a potential therapeutic target for the treatment of PCa.

Abstract 2904: Intratumor δ-catenin heterogeneity driven by genomic rearrangement dictates growth factor dependent prostate cancer cell fate and behavior

Abstract Only a small number of genes are bona fide oncogenes and tumor suppressors such as Ras, Myc, p53 and PTEN. However, targeting these cancer drivers frequently fail to demonstrate sustained cancer remission. Tumor heterogeneity and evolution contribute to cancer resistance and pose challenges for cancer therapy due to differential gene expression profiles driving distinct tumor responses to treatments. Here we report that intratumor heterogeneity of Wnt/β-catenin/armadillo modulator δ-catenin controls individual cell behavior to promote cancer. The differential intratumor subcellular localization of δ-catenin mirrors its compartmentalization in prostate cancer xenograft cultures as a result of mutation-rendered δ-catenin truncations. Wildtype and δ-catenin mutants displayed non-overlapping protein interactomes that highlight rewiring of signal networks. Localization specific δ-catenin mutants influenced p120ctn-dependent Rho GTPase phosphorylation and shifted cells towards bFGF-responsive growth, a known signal to bypass androgen receptor dependence. Mutant δ-catenin promoted Myc-induced prostate tumorigenesis by increasing proliferation over apoptosis as well as β-catenin stabilization and HIF-1α expression. Therefore, intratumor δ-catenin heterogeneity by genetic remodeling promotes prostate cancer expansion towards androgen independence, supporting a neomorphism model paradigm for targeting tumor progression. Citation Format: Mingchuan Li, Jongdee Nopparat, Jiao Zhang, Byron J. Aguilar, Yan-Hua Chen, Jie Du, Xin Ai, Yong Luo, Yongguang Jiang, Christi Boykin, Qun Lu. Intratumor δ-catenin heterogeneity driven by genomic rearrangement dictates growth factor dependent prostate cancer cell fate and behavior [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 2904.

Abstract 40: Biomaterials and systems biology to guide breast cancer drug screening

Abstract Improved in vitro models are needed to better understand cancer progression and bridge the gap between in vitro proof-of-concept studies, in vivo validation, and clinical application. Many methods exist to create biomaterial platforms, including hydrogels, which we use to study cells in contexts more akin to what they experience in vivo. Our lab has multiple approaches to create such biomaterials, based on combinations of poly(ethylene glycol) (PEG) with peptides and zwitterions. In this presentation, I will discuss our findings in using these cell culture environments to understand the role of the extracellular matrix (ECM) in controlling cancer cell innate drug response via adaptive signaling. Specifically, I will present data comparing the behavior of breast, prostate, and ovarian cancer cells to chemotherapy and targeted drugs when cultured as 3D spheroids, on 2D gels, and as a function of the stiffness of the tumor microenvironment. This approach uncovered that cells on 2D hydrogels and spheroids encapsulated in 3D hydrogels were less responsive to receptor tyrosine kinase (RTK)-targeting drugs sorafenib and lapatinib, but not cytotoxic drugs, compared to single cells in hydrogels and cells on plastic. We found that transcriptomic differences between these in vitro models and tumor xenografts did not reveal mechanisms of ECM-mediated resistance to sorafenib. However, a systems biology analysis of phospho-kinome data uncovered that variation in MEK phosphorylation was associated with RTK-targeted drug resistance. Using sorafenib as a model drug, we found that co-administration with a MEK inhibitor decreased ECM-mediated resistance in vitro and reduced in vivo tumor burden compared to sorafenib alone. In sum, we provide a novel strategy for identifying and overcoming ECM-mediated resistance mechanisms by performing drug screening, phospho-kinome analysis, and systems biology across multiple biomaterial environments. Our work suggests that different model systems are important for evaluating cell response to receptor tyrosine kinase inhibitors, whose efficacy depends on cell-ECM interactions. Citation Format: Alyssa Schwartz, Lauren Barney, Lauren Jansen, Christopher Hall, Aaron Meyer, Shelly R. Peyton. Biomaterials and systems biology to guide breast cancer drug screening [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 40.

Abstract 66: MMP-7 cleavage of the Perlecan-Sema3A-Plexin(A1)-Neuropilin-1 complex promotes the progression of metastatic prostate cancer

Abstract Introduction: Perlecan/HSPG2 is a major component of the basement membrane and of reactive stroma. PCa relies on matrix metalloproteinase activity to pass through five perlecan-rich boundaries to metastasize to bone. Semaphorin 3A (Sema3A), a secreted factor, complexes with perlecan at the cell surface and binds to a plexin-neuropilin receptor, resulting in a cohesive tumor state. In prostate cancer (PCa) metastasis, cleavage of this complex and disruption of the perlecan-rich stroma promotes migration to secondary sites. Loss of the Perlecan-Sema3A-Plexin(A1)-Neuropilin-1 (PSPN)-complex by MMP-7 cleavage of perlecan disrupts PCa cohesion and enables the formation of circulating tumor cells (CTCs). Downstream FAK, Akt, and FOXM1 are activated by loss of the PSPN-complex in the tumor microenvironment, promoting growth and metastasis. The extent of MMP-7 cleavage of the other components of the PSPN-complex was unknown. Methods: Knockdowns using shRNA of various PSPN-complex components were used to identify the perlecan-Sema3A binding partners (plexin A1 and neuropilin-1) on the PCa cell surface. Both in silico and in vitro analysis of plexin A1 and neuropilin-1 revealed potential MMP-7 cleavage sites within the PSPN-complex. Silver staining was used to analyze the extent of cleavage of the PSPN-complex components by MMP-7. Western blot analysis was used to probe for any resulting molecular changes in downstream signaling from PSPN-complex components (FAK, Akt, and FOXM1). Changes in cell cohesion were monitored to associate changes in signaling with phenotypic changes in cell behavior. Results: Knockdowns of the plexin A1 component of the PSPN-complex prevented tumoroid-like formation of PCa cells, implicating this plexin as having a key role in maintaing PCa cohesion mediated through the PSPN-complex. MMP-7 cleavage of various PSPN-complex components resulted in complex destruction, PCa dyscohesion, and the activation of downstream metastasis-promoting factors FAK, Akt, and FOXM1. Conclusion: The dynamic interactions between the PSPN-complex and MMP-7 at the PCa cell surface governs the cohesion-dyscohesion behavior of PCa cells. Loss of the PSPN-complex by MMP-7 cleavage results in a dyscohesive and metastatic phenotype. Downstream factors FAK, Akt, and FOXM1 can be activated by loss of the PSPN-complex, further implicating the importance of this complex in the metastasis of PCa. These components offer a novel target to perturb the progression of metastatic disease and to prevent CTC formation in PCa patients. Citation Format: Tristen V. Tellman, Lissette A. Cruz, Brian J. Grindel, Leland W.K. Chung, Mary C. Farach-Carson. MMP-7 cleavage of the Perlecan-Sema3A-Plexin(A1)-Neuropilin-1 complex promotes the progression of metastatic prostate cancer [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 66.

MiR-506-3p acts as a novel tumor suppressor in prostate cancer through targeting GALNT4.

Researches have indicated that microRNA-506-3p (miR-506-3p) was downregulated and functioned as tumor suppressor in cancers. However, the biological role of miR-506-3p in prostate cancer (PCa) remains to be elucidated. Expression of miR-506-3p in PCa cell lines was measured by qRT-PCR. Effects of miR-506-3p expression on PCa cell behaviors were investigated with MTT assay, colony formation assay, and transwell invasion assay. Connection of miR-506-3p and N-Acetylgalactosaminyltransferase-4 (GALNT4) was analyzed with luciferase activity reporter assay and Western blot assay. miR-506-3p expression was downregulated in PCa cell lines. Function studies demonstrated that overexpression of miR-506-3p inhibits PCa tumor progression in vitro. Mechanistic investigations found GALNT4 was a direct target of miR-506-3p. Overexpression of GALNT4 reversed the tumor-suppressive effects of miR-506-3p on PCa cell. Our results elucidated genetic silencing of miR-506-3p enhances GALNT4 oncogene expression to accelerate PCa progression.

A Novel Monoclonal Antibody PC322 Inhibit Prostate Cancer Cell Autophagy

Autophagy is an important element in tumor progression. The clinical and commercial success of monoclonal antibodies (MAbs) for cancer has emerged as the fastest growing therapy. Prostate cancer is well appropriately suited for antibody therapy. PC322 is our newly prepared prostate cancer monoclonal antibody. In our current study, we described the impacts of PC322 on autophagy and biological behaviors in vitro and explored the possible mechanism of PC322 in prostate cancer cell autophagy. Role of PC322 on autophagy of LNCaP and C4-2 cells was observed with fluorescence microscopy, western blotting, and flow cytometry. Further, the function of PC322 on biological behaviors of prostate cancer cells was detected by CCK-8 kit, Hoechst 33342, wound healing assay and cell adhesion assay. Finally, we found that Autophagy-related genes3 (ATG3) was up-regulated with stimulation of Rapamycin (RAPA), while PC322 inhibited ATG3 expression. In summary, in vitro experiments demonstrated that the new monoclonal antibody PC322 inhibit cell autophagy, proliferation, and metastasis of LNCaP and C4-2 cells, and promote their apoptosis. PC322 reduces ATG3 expression. Thereby suppresses the formation of autophagosomes by inhibiting LC3-I converting to LC3-II.

Epithelial-To-Mesenchymal Transition Markers and CD44 Isoforms Are Differently Expressed in 2D and 3D Cell Cultures of Prostate Cancer Cells.

Three-dimensional (3D) cell cultures allow the mimic of functions of living tissues and provide key information encoded in tissue architecture. Considered the pivotal role of epithelial-to-mesenchymal transition (EMT) in carcinoma progression, including prostate cancer (PCa), we aimed at investigating the effect of the 3D arrangement on the expression of some key markers of EMT in cultured human prostate cancer (PCa) cells, to better understand PCa cell behavior. PC3 and DU145 PCa cells were cultured in RPMI cell culture medium either in 2D-monolayers or in 3D-spheroids. The main EMT markers E-cadherin, N-cadherin, α-smooth muscle actin (αSMA), vimentin, Snail, Slug, Twist and Zeb1 were evaluated by confocal microscopy, real-time PCR and Western blot. Confocal microscopy revealed that E-cadherin was similarly expressed at the cell boundaries on the plasma membrane of PCa cells grown in 2D-monolayers, as well as in 3D-spheroids, but resulted up-regulated in 3D-spheroids, compared to 2D-monolayers, at the mRNA and protein level. Moreover, markers of the mesenchymal phenotype were expressed at very low levels in 3D-spheroids, suggesting important differences in the phenotype of PCa cells grown in 3D-spheroids or in 2D-monolayers. Considered as a whole, our findings contribute to a clarification of the role of EMT in PCa and confirm that a 3D cell culture model could provide deeper insight into the understanding of the biology of PCa.

ZNF433 positively regulates the beta-catenin/ TCF pathway in prostate cancer and enhances the tumorigenicity of cancer cells.

BackgroundProstate cancer often shows the over-activation of beta-catenin/t-cell factor (TCF) signaling. It remains largely unknown how the beta-catenin/TCF transcriptional machinery is tightly controlled.MethodsThe ZNF433 mRNA and protein levels in the clinical tissues were examined using q-PCR, Western blot and immunohistochemistry. The phenotypes of prostate cancer cells were examined using MTT assay, Boyden chamber assay and anchorage-independent assay. The interaction between ZNF433 and beta-catenin was evaluated by immunoprecipitation.ResultsIn the present study, ZNF433 was upregulated in prostate cancer samples, and promoted the growth and migration of prostate cancer cells. Furthermore, ZNF433 was the binding partner of beta-catenin and activated beta-catenin/TCF signaling in prostate cancer. Moreover, ZNF433 enhanced the binding between beta-catenin and TCF4. In addition, NC043, small antagonist for beta-catenin/TCF complex, inhibited the malignant behaviors of prostate cancer cells driven by ZNF433.ConclusionIn summary, these studies demonstrate the tumor-promoting roles of ZNF433 in prostate cancer, and suggesting that ZNF433 was a potential target for the treatment.

USP44 Promotes the Tumorigenesis of Prostate Cancer Cells through EZH2 Protein Stabilization.

Ubiquitin-specific protease 44 (USP44) has been implicated in tumor progression and metastasis across various tumors. However, the function of USP44 in prostate cancers and regulatory mechanism of histone-modifying enzymes by USP44 in tumors is not well-understood. Here, we found that enhancer of zeste homolog 2 (EZH2), a histone H3 lysine 27 methyltransferase, is regulated by USP44. We showed that EZH2 is a novel target of USP44 and that the protein stability of EZH2 is upregulated by USP44-mediated deubiquitination. In USP44 knockdown prostate cancer cells, the EZH2 protein level and its gene silencing activity were decreased. Furthermore, USP44 knockdown inhibited the tumorigenic characteristics and cancer stem cell-like behaviors of prostate cancer cells. Inhibition of tumorigenesis caused by USP44 knockdown was recovered by ectopic introduction of EZH2. Additionally, USP44 regulates the protein stability of oncogenic EZH2 mutants. Taken together, our results suggest that USP44 promotes the tumorigenesis of prostate cancer cells partly by stabilizing EZH2 and that USP44 is a viable therapeutic target for treating EZH2-dependent cancers.

MiR-490-3p modulates the progression of prostate cancer through regulating histone deacetylase 2.

microRNAs (miRNAs) were regarded as critical participators for human cancers progression including prostate cancer (PCa) and have the potential to be used as treatment targets for cancers. Herein, we validated a tumor-suppressive miRNA, miR-490-3p, which may suppress PCa progression. Histone deacetylase 2 (HDAC2) is a protein that aberrantly expressed in several cancers. However, the role of HDAC2 in the progression of PCa has not been fully elucidated. Expression of miR-490-3p and HDAC2 in PCa was investigated. The effects of miR-490-3p or HDAC2 expression on PCa cell behaviors were analyzed. Association between miR-490-3p and HDAC2 was analyzed by luciferase activity reporter assay and Western blot assay. We demonstrated that miR-490-3p functioned as a tumor-suppressive role in PCa progression. We found miR-490-3p expression was decreased in PCa cell lines. Down-regulation of miR-490-3p promoted the growth, migration, invasion but inhibited apoptosis of PCa cells. HDAC2 was validated as a direct target of miR-490-3p and promoted the progression of PCa cells. Further studies showed that HDAC2 could reverse the effects of miR-490-3p on growth, migration, invasion and apoptosis of PCa cells. Our data highlighted the key role of miR-490-3p in the progression of PCa. Thus, miR-490-3p may be a novel cancer-specific therapeutic.

MicroRNA-425-5p promotes the development of prostate cancer via targeting forkhead box J3.

MicroRNAs (miRNAs) have critical roles in the progression of prostate cancer (PCa) and have the potential to be used as prognosis biomarkers. In this study, we aimed to investigate the role of miR-425-5p in the progression of PCa. miR-425-5p expression in PCa tumor tissues and cell lines was measured by Quantitative Real-time PCR (RT-qPCR). Effects of miR-425-5p expression on PCa cell proliferation, colony formation, cell migration, and cell invasion were measured. We found miR-425-5p expression was elevated in both PCa tissues and cell lines. Importantly, we found overexpression of miR-425-5p promoted proliferation, colony formation, migration and invasion of PCa cell lines in vitro. Forkhead box J3 (FOXJ3) was validated as a downstream target of miR-425-5p. Finally, we found the stimulation effects of miR-425-5p on PCa cell behaviors were fulfilled through directly regulating the expression of FOXJ3, which validated FOXJ3 as a functional target of miR-425-5p in PCa. Taken together, our results demonstrated miR-425-5p may contribute the malignancy progression of PCa in a mechanism involving FOXJ3, implicating miR-425-5p may be developed as therapeutic target for PCa in the future.