Tumor Progression In Prostate Cancer Research Articles

Abstract 4619: Prolonged inhibition of androgen receptor signaling induces expression of Nuclear ErbB3 which renders prostate cancer cells susceptible to targeted inhibitors

Abstract Objective: Nuclear expression of the receptor tyrosine kinase (RTK) ErbB3/HER3 increases in highly aggressive prostate cancer cells, but its nuclear transport mechanism is currently unknown. Prostate tumors rely on the androgen receptor (AR), whose activation transcriptionally controls ErbB3 expression, but its role in ErbB3 nuclear localization had not previously been reported. Here, we investigated the mechanism by which subcellular localization of ErbB3 was altered in prostate cancer. Methods: ErbB3 localization was investigated in the human prostate cancer tumor progression model LNCaP, C4, C4-2 and C4-2B. ErbB3 was stimulated with heregulin-1β (HRG). Nuclear translocation was tested with a nucleocytoplasmic transport inhibitor panel (chlorpromazine, filipin III, amiloride and Leptomycin B) LNCaP and C4 cells were continuously cultured with the AR activation inhibitor abiraterone acetate (AbiAc) or ethanol (VEH) or treated with the AR inhibitors enzalutamide, darolutamide and apalutamide. ErbB3 activity and subcellular localization were analyzed using confocal microscopy/subcellular fractionation/immunoblot. Cell viability was determined by MTT assay. Proliferation and apoptosis were determined by flow cytometry. Invasive potential was investigated by crystal violet staining of cell colonies. ErbB3 expression was reduced by siRNA technology. Results: The ratio of nuclear to cytoplasmic ErbB3 increased in untreated cells from LNCaP<C4-2<C4<C4-2B. In all four lines, nuclear localization of ErbB3 peaked at 30 minutes after HRG treatment, with a rapid return to the cytoplasm in LNCaP cells (at 1 hour), slower in C4 cells (at 4 hours), and continued accumulation in the nucleus in C4-2 and C4-2B 8 hours following HRG treatment. Treatment with the transport inhibitor panel showed that nuclear accumulation of ErbB3 was prevented by the clathrin-dependent endocytosis inhibitor Chlorpromazine (CPZ) but not by the others. In LN-VEH and C4-VEH cells, HRG induced nuclear translocation of ErbB3 but not in LN-AbiAc or C4-AbiAc cells. Accordingly, HRG increased ErbB3 phosphorylation at Y1328 and nuclear Akt phosphorylation at S473 in VEH but not AbiAc cells. C4 cells, which had high baseline nuclear ErbB3, were more sensitive to the AR inhibitors and showed decreased viability and invasive potential compared to LNCaP cells. Conclusions: These results indicate that (1) ErbB3 nuclear localization required clathrin-dependent endocytosis (2) ligand binding of ErbB3 as well as the presence of an active AR is necessary for ErbB3 nuclear localization and (3) the presence of nuclear ErbB3 increases sensitivity to AR inhibitors. Citation Format: Maitreyee K. Jathal, Maria M. Mudryj, Paramita M. Ghosh. Prolonged inhibition of androgen receptor signaling induces expression of Nuclear ErbB3 which renders prostate cancer cells susceptible to targeted inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4619.

Abstract 2638: Sensitizing immunotherapy-refractory prostate cancer to immune checkpoint blockade therapy with optimized ketogenic diet regimens

Abstract The purpose of this study is to investigate strategies for enhancing immune checkpoint blockade (ICB) therapy in prostate cancer (PCa). To achieve this, we established a series of ICB-resistant PCa cell lines derived from a previously sensitive cell line, which originated from a murine model of metastatic PCa. Notably, we observed a reduction in major histocompatibility complex (MHC) class I protein levels in the resistant sublines. Our study demonstrated synergistic efficacy through the combination of anti-PD1 and anti-CTLA4 antibodies with histone deacetylase inhibitor (HDACi) vorinostat, a cyclic ketogenic diet (CKD), or β-hydroxybutyrate (BHB) supplementation. Importantly, these interventions significantly delayed PCa tumor progression. Furthermore, our analysis revealed that the effects of these interventions were contingent on both BHB and adaptive immunity. In-depth single-cell transcriptomic and proteomic profiling unveiled mechanisms involving cancer-cell-intrinsic changes (upregulated MHC class I molecules) and extrinsic factors, including CD8+ T cell chemoattraction, M1/M2 macrophage rebalancing, monocyte differentiation toward antigen-presenting cells, and reduced neutrophil presence. In conclusion, our findings provide valuable insights into the potential clinical utility of combining HDACi and optimized ketogenic diet regimens to enhance ICB therapy for PCa. Citation Format: Sean Murphy, Sharif Rahmy, Dailin Gan, Jun Li, Xuemin Lu, Xin Lu. Sensitizing immunotherapy-refractory prostate cancer to immune checkpoint blockade therapy with optimized ketogenic diet regimens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2638.

A novel circular RNA, circMAML3, promotes tumor progression of prostate cancer by regulating miR-665/MAPK8IP2 axis

Many studies have now demonstrated that circRNAs are aberrantly expressed in cancer and are involved in the regulation of malignant tumor progression. However, the role of circMAML3 (hsa_circ_0125392) in prostate cancer has not been reported. circMAML3 was selected from public data through screening. The circMAML3 circular characterization was performed using Sanger sequencing, agarose gel electrophoresis assay, RNase R assay and actinomycin D assay. The expression of circMAML3 in prostate cancer tissues and cells was detected by qRT-PCR. In vivo and in vitro experiments were conducted to investigate the biological functions of circMAML3 in prostate cancer. Finally, the underlying mechanism of circMAML3 was revealed by qRT-PCR, luciferase reporter assay, miRNA Pulldown, RNA immunoprecipitation, western blotting, and rescue assay. Compared to normal prostate tissue and prostate epithelial cells, circMAML3 is highly expressed in prostate cancer tissues and cell lines. CircMAML3 overexpression promotes prostate cancer proliferation and metastasis, while knockdown of circMAML3 exerts the opposite effect. Mechanistically, circMAML3 promotes prostate cancer progression by upregulating MAPK8IP2 expression through sponge miR-665. Our research indicates that circMAML3 promotes prostate cancer progression through the circMAML3/miR-665/MAPK8IP2 axis. circMAML3 and MAPK8IP2 are upregulated in prostate cancer expression and play an oncogenic role, whereas miR-665 is downregulated in prostate cancer and plays an oncogenic role. Therefore, CircMAML3 may be a potential biomarker for prostate cancer diagnosis, treatment and prognosis.

Exosome miRNA-203 promotes M1 macrophage polarization and inhibits prostate cancer tumor progression.

Prostate cancer (PCa) is a prevalent malignant neoplasm affecting the male reproductive system globally. However, the diagnostic and therapeutic approaches fall short of meeting the demands posed by PCa. Poor expression of miRNA-203 (miR-203) within PCa tissues and cells implies its potential utility as a diagnostic indicator for PCa. Exosomes (Exo), membranous vesicles released by various cells, are rich reservoirs of miRNAs. However, the presence of miR-203 presents within Exo derived from PCa cells remains unclarified. In this study, Exo was isolated from urine specimens collected from clinical PCa patients and LNCaP cells to detect miR-203 expression. Meanwhile, the impact of overexpressed miR-203 on M0 macrophages (mø) was analyzed. Subsequently, alterations in the proliferative, migratory, and invasive capacities of LNCaP cells were examined within a co-culture system featuring elevated miR-203 levels in both macrophages and LNCaP cells. Furthermore, the repercussions of miR-203 upregulation or inhibition were explored in a murine PCa tumor model. The results revealed that Exo manifested a circular or elliptical morphology, encapsulating a phospholipid bilayer approximately 100nm in diameter. Notably, Exo readily infiltrated, with both Exo and miR-203-overexpressing Exo prompting macrophage polarization toward the M1 subtype. In the co-culture system, miR-203 exhibited pronounced suppression of LNCaP cell proliferation, migration, and invasion, while concurrently fostering apoptosis as compared with the LNCaP group (Control). In vivo experiments further disclosed that miR-203 greatly inhibited the growth of PCa tumors in nude mice. Markedly heightened expression of M1 macrophage markers such as IL-1β, IL-6, IL-12, CXCL9, and CXCL10 was observed within the tumor microenvironment following miR-203 intervention, as opposed to the model group. However, the introduction of miR-203 antagomir led to a reversal in tumor growth trends. This investigation indicates the presence of miR-203 within the urine of PCa patients and Exo originating from cells, and that miR-203 exerted antitumor effect by facilitating M1 macrophage polarization. Our study furnishes valuable insights into the potential applicability of miR-203 as a diagnostic biomarker and therapeutic target for PCa.

Immune mechanisms shape the clonal landscape during early progression of prostate cancer.

Understanding the role of the immune microenvironment in modulating intratumor heterogeneity is essential for effective cancer therapies. Using multicolor lineage tracing in genetically engineered mouse models and single-cell transcriptomics, we show that slowly progressing tumors contain a multiclonal landscape of relatively homogeneous subpopulations within a well-organized tumor microenvironment. In more advanced and aggressive tumors, however, the multiclonal landscape develops into competing dominant and minor clones accompanied by a disordered microenvironment. We demonstrate that this dominant/minor landscape is associated with differential immunoediting, in which minor clones are marked by an increased expression of IFNγ-response genes and the Tcell-activating chemokines Cxcl9 and Cxcl11. Furthermore, immunomodulation of the IFNγ pathway can rescue minor clones from elimination. Notably, the immune-specific gene signature of minor clones exhibits a prognostic value for biochemical recurrence-free survival in human prostate cancer. These findings suggest new immunotherapy approaches for modulating clonal fitness and tumor progression in prostate cancer.

Mir-629 Repressed LATS2 Expression and Promoted the Proliferation of Prostate Cancer Cells.

MicroRNAs (miRNAs) have been discovered to play critical role in regulating prostate cancer (PC) progression. The function role of miR-629 in tumor progression of PC has not been studied. Here, we found that miR-629 was markedly upregulated in PC as determined using the cancer genome atlas (TCGA) dataset, clinical tissues, and cell lines. Functional analysis (MTT assays, colony formation assays, soft agar growth assay and BrdU incorporation assay) indicated that overexpression of miR-629 was drastically promoted, while miR-629-in significantly suppressed cell proliferation. LATS2 was predicted as a direct target of miR-629 and was confirmed by western blot and dual luciferase assay. Through downregulation of large tumor suppressor 2 (LATS2) by overexpression of miR-629, the p21 mRNA and protein were decreased while the Cyclin D3 mRNA and protein were enhanced, suggesting promoting of cell proliferation process. Additionally, knockdown of LATS2 reversed the inhibitory effect by miR-629-in in PC. Our study indicated that miR-629 might serve as a new promising target for PC treatment.

MiR-124-3p inhibits tumor progression in prostate cancer by targeting EZH2

Prostate cancer (PCa) is widespread cancer with significant morbidity and mortality rates. MicroRNAs (miRNAs) have been identified as important post-transcriptional modulators in various malignancies. This study investigated the miR-124-3p effect on PCa cell proliferation, infiltration, and apoptosis. EZH2 and miR-124-3p expression levels were measured in PCa tissues. PCa cell lines DU145 and PC3 were transfected with miR-124-3p inhibitors or analogs. EZH2 and miR-124-3p linkage was validated by conducting the luciferase enzyme reporter test. The cell viability and apoptosis were assessed by flow cytometry and MTT test. Cell movement was noted during infiltration using transwell assays. EZH2, AKT, and mTOR contents were assessed using qRT-PCR and western blotting. In clinical PCa specimens, miR-124-3p and EZH2 contents were inversely correlated. Further research has demonstrated that EZH2 is the miR-124-3p direct target. Furthermore, miR-124-3p overexpression reduced EZH2 levels and lowered cell viability, infiltration, and promoted cell death, whereas miR-124-3p silencing had the opposite effect. Overexpression of miR-124-3p decreased the phosphorylation level of AKT and mTOR, whereas miR-124-3p downregulation produced the opposite result. Our findings depict that miR-124-3p prevents PCa proliferative and invasive processes while promoting apoptosis by targeting EZH2.

The Circular RNA Circ_0085494 Regulates Prostate Cancer Progression Through NRBP1/miR-497-5p Axis.

Aberrant expression of circular RNA (circRNA) is closely linked to the progression of various human cancers, including prostate cancer (PCa). In this research, we aimed to investigate the biological role of and mechanisms of circ_0085494 in PCa. The quantitative real-time polymerase chain reaction (qRT-PCR) assay was used to detect circ_0085494, miR-497-5p, and nuclear receptor binding protein 1 (NRBP1) mRNA expression in PCa tissues and cell lines. Subsequently, colony formation, cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), and transwell assays were performed to evaluate PCa cell proliferation, migration, and invasion. Western blot assay was applied for assessing the protein levels. Dual-luciferase reporter and RNA pull-down assays were implemented for verifying the association between miR-497-5p and circ_0085494 or NRBP1. The role of circ_0085494 in vivo was measured by establishing a mice xenograft model. Circ_0085494 was highly expressed in PCa tissues and cells, and its absence suppressed PCa cell proliferation, migration, and invasion. Circ_0085494 impacts NRBP1 content by adsorbing miR-497-5p. Meanwhile, the repression of circ_0085494 absence on tumor growth in vivo was validated. Our finding revealed that circ_0085494 downregulation might repress PCa tumor progression through in part regulating the miR-497-5p/NRBP1 pathway.

Cancer-derived exosomal miR-375 targets DIP2C and promotes osteoblastic metastasis and prostate cancer progression by regulating the Wnt signaling pathway.

Bone metastasis is the most common complication responsible for most deaths in the advanced stages of prostate cancer (PCa). However, the exact mechanism of bone metastasis in PCa remains unelucidated. Herein, we explored the function and potential underlying mechanism of exosomal miR-375 in bone metastasis and tumor progression in PCa. This study revealed that miR-375 expression was markedly upregulated in advanced PCa with bone metastasis and metastatic PCa cell lines. Moreover, miR-375 showed high expression in PCa-derived exosomes and could be delivered to human mesenchymal stem cells (hMSCs) via exosomes. Mechanistically, miR-375 directly targeted DIP2C and upregulated the Wnt signaling pathway, thereby promoting osteoblastic differentiation in hMSCs. Furthermore, miR-375 promoted the proliferation, invasion, and migration of PCa cells in vitro and enhanced tumor progression and osteoblastic metastasis in vivo. Notably, the expression of miR-375, TCF-1, LEF-1, and β-catenin in was higher in PCa tissues with bone metastasis than in PCa tissues without bone metastasis and showed a continuous increase, whereas DIP2C, cyclin D1, and Axin2 showed an opposite expression pattern. In conclusion, our study suggests that cancer-derived exosomal miR-375 targets DIP2C, activates the Wnt signaling pathway, and promotes osteoblastic metastasis and PCa progression.

MAPK8IP2 is a potential prognostic biomarker and promote tumor progression in prostate cancer

BackgroundMAPK8IP2 is one of the JNK-interacting proteins (JIPs) family members, and is involved in the regulation of the JNK and P38 MAPK signaling pathways. MAPK8IP2 has been reported to be closely associated with several cancers. However, the biological function of MAPK8IP2 in prostate cancer (PCa) remains unclear.MethodsMAPK8IP2 expression in PCa and subgroups of PCa was analyzed by public databases. The prognostic role of MAPK8IP2 in prostate cancer was analyzed using the Cox regression method. The potential mechanism by which MAPK8IP2 affects PCa progression was investigated by utilizing public data, including genetic alteration, DNA methylation, m6A methylation, and immune infiltration data. We further performed in vitro assays to validate the effect of MAPK8IP2 on PCa cell proliferation, migration and invasion.ResultsMAPK8IP2 is highly expressed in PCa tissues. Overexpression of MAPK8IP2 is associated with adverse clinicopathological factors and a poor prognosis in PCa. Receiver operating curve analysis showed that MAPK8IP2 can distinguish PCa tissues from non-PCa tissues with a certain accuracy (AUC = 0.814). The MAPK8IP2 genetic alteration rate was 2.6% and MAPK8IP2 alterations correlated with a poor prognosis. We also found that CDK12 and TP53 mutations were associated with MAPK8IP2 expression. The DNA methylation level of MAPK8IP2 was higher in primary tumors than in normal tissues, and the high MAPK8IP2 DNA methylation group of PCa patients had poor survival. Enrichment analysis indicated that MAPK8IP2 was involved in the MAPK signaling pathway. In vitro, knockdown of MAPK8IP2 inhibited PCa cell proliferation, migration and invasion.ConclusionMAPK8IP2 is a potential target for PCa treatment and can serve as a novel biomarker for PCa diagnosis and prognosis evaluation.

LncRNA RHPN1-AS1 inhibition induces autophagy and apoptosis in prostate cancer cells via the miR-7-5p/EGFR/PI3K/AKT/mTOR signaling pathway.

LncRNA RHPN1-AS1 (RHPN1-AS1) has been confirmed to promote tumor progression in multiple cancers and is upregulated in prostate cancer (PCa), but whether it has an effect on PCa progression remains unclear. In this study, we found that PCa patients with high RHPN1-AS1 expression had a shorter survival time, and RHPN1-AS1 was significantly upregulated in PCa tissues and cells. Based on informatics analysis we predicted that miR-7-5p binds to 3'UTR of RHPN1-AS1 and epidermal growth factor receptor (EGFR) and verified it by luciferase reporter gene assay. Subsequently, we transfected PCa cells with RHPN1-AS1 overexpression vector (RHPN1-AS1), knockdown plasmids (sh-RHPN1-AS1) and/or miR-7-5p mimics or inhibitor and/or overexpression vector (EGFR) or small interfering RNA of EGFR (si-EGFR) or its control, and found that overexpression of RHPN1-AS1 inhibited miR-7-5p expression and promoted EGFR expression, silencing RHPN1-AS1 inhibited proliferation and invasion, and induced G2/M arrest, apoptosis and autophagy in PCa cells. 3MA (an inhibitor of autophagy)-mediated autophagy inhibition attenuated RHPN1-AS1 inhibition-induced apoptosis. Overexpression miR-7-5p or silencing EGFR promoted LC3-I to LC3-II conversion, enhanced autophagy activity, induced cleaved-caspase-3 expression and apoptosis in PCa cells. Furthermore, overexpression of RHPN1-AS1 promoted phosphorylation of phosphatidylinositol 3-kinase (PI3K), AKT and mTOR, inhibited LC3-I to LC3-II conversion and reduced apoptosis in PCa cells, while GSK2126458 (an inhibitor of PI3K) reversed the effect of RHPN1-AS1 on PCa cells. In summary, RHPN1-AS1 acted as a ceRNA of miR-7-5p to upregulate EGFR expression, silencing RHPN1-AS1 suppressed PCa tumor progression by inducing autophagy and apoptosis in PCa cells through the miR-7-5p/EGFR/PI3K/AKT/mTOR pathway.

Abstract 6125: ADAM9 secreted by cancer-associated stromal cells promotes metastasis of prostate cancer

Abstract Introduction: Cancer-associated fibroblasts (CAFs), the main cellular component of the tumor microenvironment (TME), contribute to tumor metastasis. Prostate cancer (PCa) is the most common malignancy among the elderly male population, and it is typically fatal when the disease progresses to metastasis and castration-resistant stage (CRPC). We have previously demonstrated that ADAM9 increased during PCa progression, and ROS is the critical mediator regulating ADAM9 expression in CRPC cells to promote tumor growth. In addition to membrane-anchored ADAM9, a secreted variant of ADAM9 (sADAM9) was recently identified in activated liver stromal cells that facilitated colon cancer liver metastasis. However, it is not clear whether sADAM9 is expressed by prostatic CAFs and, if so, whether it contributes to aggressive tumor phenotypes through tumor-stromal interactions. Objective: To investigate the sADAM9 variants in CAFs and to elucidate the effects of CAF-derived sADAM9 on the aggressiveness of PCa cells. Methods: Immunohistochemistry staining and ELISA of clinical samples were used to assess the expression of ADAM9 during PCa tumor progression. Laser microdissected tissues, primary cell lines, and experimental cell model of paired normal fibroblasts and CAF cells were analyzed. The effects of sADAM9 on CRPC progression were conducted in PC3 cells by WST-1 cell proliferation assay, transwell migration assay, and μ-slide chemotaxis assay. Immunoblot analyses of integrin β1 and migration-related kinases were used to verify the possible downstream signaling pathway of sADAM9. Results: Increased overall intensity of ADAM9 protein expression in PCa patients with metastasis compared. Elevated ADAM9 expression was observed in both cancerous epithelial cells and the tumor stroma. The average serum ADAM9 concentration measured by ELISA was 6.253 ± 0.448 ng/mL in BPH patient samples versus 10.89 ± 0.953 ng/mL in PCa patient samples. A smaller molecular weight (~50-kDa) of ADAM9 was detected in CM from cultured CAFs but not normal fibroblasts or PCa cell lines except for DU145 cells. Treatment of PC3 cells with sADAM9 induced a significant chemotactic migration while did not affect cell proliferation. Furthermore, a stark increase in activated integrin β1, phosphorylated focal adhesion kinase (p-FAK), and p-AKT, but not p-mTOR in PCa cells by sADAM9 suggested the induction of cell migration occurs through activation of the integrin β1/FAK signaling pathways. Conclusion: Herein, we have provided in vitro evidence to demonstrate the pro-metastatic function of sADAM9 from CAFs that facilitate the migration of CRPC cells through integrin receptor signaling, which could be partly responsible for the promotion of CRPC metastasis. Further molecular identification of controlling spliced ADAM9 expression in CAFs remains to investigate. Citation Format: Weichieh Chen, Le Thi Huynh Trang, Chia-Ling Hsieh, Katsumi Shigemura, Masato Fujisawa, Leland W.K. Chung, Shian-Ying Sung. ADAM9 secreted by cancer-associated stromal cells promotes metastasis of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6125.

Roles for CCN proteins in Lysophosphatidic Acid and Sphingosine 1‐Phosphate Signaling in Prostate Cancer Cells

IntroductionCysteine rich angiogenic factor (CCN1/Cyr61) and connective tissue growth factor (CCN2/CTGF) are members of the CCN family of matricellular proteins. These secreted proteins interact with extracellular matrix proteins and integrins to modulate cell signaling. CCN proteins have been implicated in survival and other mitogenic responses in various cancers. CCN1 and CCN2 are the two early inducible genes in this family. Lysophosphatidic acid (LPA) and sphingosine‐1‐phosphate (S1P) are lipid growth factors known to contribute to tumor progression in prostate cancer. In previous work, our group showed that LPA stimulates proliferation and migration in prostate cancer cells via the G protein‐coupled receptor LPAR1. Free fatty acid receptor 4 (FFAR4) agonists, inhibit LPA‐induced responses in prostate cancer cells. We also demonstrated that CCN1 expression is induced by LPA in human prostate cancer cells. However, the roles of CCN1 and CCN2 in cancer cell signaling have not been fully elucidated.HypothesisWe hypothesize that CCN proteins contribute to LPA‐ and S1P‐induced adhesion and mitogenic signaling in prostate cancer cells.Methods and ResultsPC‐3 human prostate cancer cells were used in the current study. For all experiments, cells were serum starved for 24 hours and then treated with and without 10 µM LPA or S1P. Immunoblotting showed that CCN1 and CCN2 were induced by LPA after 2‐5 hours in PC‐3 cells. The response was inhibited by the LPA receptor antagonist Ki1625. TUG‐891, an FFAR4 agonist, inhibited LPA‐induced CCN1 expression in PC‐3 cells. S1P also induced CCN1 expression in PC‐3 cells. In related experiments, whole cell extracts, and extracellular matrix were subjected to immunoblotting for CCN1, and immunofluorescence microscopy was used to localize the induced CCN1. CCN1 protein levels in extracellular matrix were increased 2‐5 hours after LPA treatment, as compared to untreated controls, and CCN1 was detected outside the cells by confocal microscopy. In cell adhesion assays, LPA‐treated PC‐3 cells exhibited an increase in adhesion to fibronectin‐coated 96‐well plates that was detected by 2 hours. Erk MAPK activation in response to LPA was biphasic, with the later phase occurring after CCN1/2 induction.ConclusionThe results of this study suggest that CCN proteins may play roles in LPA‐ and S1P‐induced signaling in prostate cancer cells and are thus potential therapeutic targets.

Editorial Comment to Tenascin C regulates cancer cell glycolysis and tumor progression in prostate cancer.

Editorial Comment to Tenascin C regulates cancer cell glycolysis and tumor progression in prostate cancer.

MiR-363-3p promotes prostate cancer tumor progression by targeting Dickkopf 3.

BackgroundProstate cancer (PCa) is a frequent malignant tumor worldwide with high morbidity along with mortality. MicroRNAs (miRNAs) have been identified as key posttranscriptional modulators in diverse cancers. Herein, we purposed to explore the impacts of miR‐363‐3p on PCa growth, migration, infiltration along with apoptosis.MethodsThe expressions of miR‐363‐3p along with Dickkopf 3 (DKK3) were assessed in clinical PCa specimens. We adopted the PCa cell line PC3 and transfected it using miR‐363‐3p repressors or mimic. The relationship of miR‐363‐3p with DKK3 was verified by a luciferase enzyme reporter assay. Cell viability along with apoptosis were determined by MTT assay coupled with flow cytometry analysis. Cell migration along infiltration were detected via wound healing, as well as Transwell assays. The contents of DKK3, E‐cadherin, vimentin along with N‐cadherin were analyzed via Western blotting accompanied with qRT–PCR.ResultsMiR‐363‐3p was found to be inversely associated with the content of DKK3 in clinical PCa specimens. Further investigations revealed that DKK3 was miR‐363‐3p's direct target. Besides, overexpression of miR‐363‐3p decreased the contents of DKK3, promoted cell viability, migration coupled with infiltration, and reduced cell apoptosis, while silencing of miR‐363‐3p resulted in opposite influence. Upregulation of miR‐363‐3p diminished E‐cadherin contents but increased vimentin along with N‐cadherin protein contents in PC3 cells; in contrast, miR‐363‐3p downregulation produced the opposite result.ConclusionOur study indicates that miR‐363‐3p promotes PCa growth, migration coupled with invasion while dampening apoptosis by targeting DKK3.

Tenascin C regulates cancer cell glycolysis and tumor progression in prostate cancer.

Tenascin C is a potential biomarker of cancer-associated fibroblasts and has been significantly associated with poor prognosis in patients with prostate cancer. However, the effects of Tenascin C in prostate cancer cell glycolysis largely remain unclear. Thus, this study aimed to investigate the Tenascin C expression in prostate cancer and its correlation to glycolysis-related protein and gene expression, clinicopathological parameters, and survival of patients. We performed immunohistochemical staining for Tenascin C in 141 cases of primary prostate cancer. Based on public data sets, we explored the association of Tenascin C with angiogenesis-related genes, M2 macrophage-related gene, androgen receptor levels, PI3K/AKT/NF-κB pathway genes, and glycolytic enzyme expression. The glucose uptake, lactate production, and glycolytic enzyme levels were detected by glycolysis assay and western blotting. Our results showed that Tenascin C expression is upregulated in prostate cancer tissues compared with benign prostatic hyperplasia tissues. High Tenascin C expression in prostate cancer cells was positively associated with lymph node metastasis, advanced clinical stage, the expression of CD105, CD206, and androgen receptor levels. The Kaplan-Meier curves showed a significant association of Tenascin C expression with the patient's overall survival. Tenascin C expression was positively associated with PI3K p85, pAKT-ser308, and NF-κB p65 protein expression in prostate cancer samples. Moreover, siRNA-mediated knockdown of Tenascin C expression inhibited cell glucose uptake, lactate production, and glycolytic-enzyme expression in prostate cancer cells invitro. Together, our findings suggest that Tenascin C is a prognostic marker for patients with prostate cancer and that its effects might be mediated via regulation of the glycolysis process of prostate cancer cells.

MiR-182-5p Modulates Prostate Cancer Aggressive Phenotypes by Targeting EMT Associated Pathways.

Prostate cancer (PCa) is a clinically heterogeneous disease, where deregulation of epigenetic events, such as miRNA expression alterations, are determinants for its development and progression. MiR-182-5p, a member of the miR-183 family, when overexpressed has been associated with PCa tumor progression and decreased patients’ survival rates. In this study, we determined the regulatory role of miR-182-5p in modulating aggressive tumor phenotypes in androgen-refractory PCa cell lines (PC3 and DU-145). The transient transfection of the cell lines with miR-182-5p inhibitor and mimic systems, significantly affected cell proliferation, adhesion, migration, and the viability of the cells to the chemotherapeutic agents, docetaxel, and abiraterone. It also affected the protein expression levels of the tumor progression marker pAKT. These changes, however, were differentially observed in the cell lines studied. A comprehensive biological and functional enrichment analysis and miRNA/mRNA interaction revealed its strong involvement in the epithelial-mesenchymal transition (EMT) process; expression analysis of EMT markers in the PCa transfected cells directly or indirectly modulated the analyzed tumor phenotypes. In conclusion, miR-182-5p differentially impacts tumorigenesis in androgen-refractory PCa cells, in a compatible oncomiR mode of action by targeting EMT-associated pathways.

Overexpression of claspin promotes docetaxel resistance and is associated with prostate-specific antigen recurrence in prostate cancer.

Although docetaxel (DTX) confers significant survival benefits in patients with castration‐resistant prostate cancer (CRPC), resistance to DTX inevitably occurs. Therefore, clarifying the mechanisms of DTX resistance may improve survival in patients with CRPC. Claspin plays a pivotal role in DNA replication stress and damage responses and is an essential regulator for the S‐phase checkpoint. CLSPN is an oncogenic gene that contributes to tumor proliferation in several human solid tumors. However, the clinical significance of claspin in prostate cancer (PCa) has not been examined. The present study aimed to elucidate the role of claspin and its relationship with DTX resistance in PCa. We immunohistochemically analyzed the expression of claspin in 89 PCa cases, of which 31 (35%) were positive for claspin. Claspin‐positive cases were associated with higher Gleason score, venous invasion, and perineural invasion. Kaplan–Meier analysis showed that high claspin expression was related to poor prostate‐specific antigen (PSA) relapse‐free prognosis. In a public database, high CLSPN expression was associated with poor PSA relapse‐free prognosis, Gleason score, T stage, lymph node metastasis, CRPC, and metastatic PCa. Claspin knockdown by siRNA decreased cell proliferation, upregulated DTX sensitivity, and suppressed the expression of Akt, Erk1/2, and CHK1 phosphorylation in DU145 and PC3 cell lines. Furthermore, claspin expression was much more upregulated in DTX‐resistant DU145 (DU145‐DR) than in parental DU145 cells. Claspin knockdown significantly upregulated the sensitivity to DTX in DU145‐DR cells. These results suggest that claspin plays an important role in PCa tumor progression and DTX resistance.

AR-mTOR-SRF Axis Regulates HMMR Expression in Human Prostate Cancer Cells.

The elevated expression of the hyaluronan-mediated motility receptor (HMMR) is known to be highly associated with tumor progression in prostate cancer, but the molecular mechanisms underlying the regulation of HMMR expression remain unclear. Here, we report that mammalian target of rapamycin (mTOR) is a key regulator of HMMR expression, for which its kinase activity is required. Pharmacological inhibitors of mTOR, such as rapamycin and Torin2, markedly suppressed the mRNA level as well as the protein level of HMMR in LNCaP and PC-3 cells. Our data demonstrate that such regulation occurs at the transcription level. HMMR promoter reporter assays revealed that the transcription factor SRF is responsible for the mTOR-mediated transcriptional regulation of HMMR gene. Consistently, the suppression of HMMR expression by Torin2 was noticeably reversed by the overexpression of SRF. Moreover, our findings suggest that the SRF binding sites responsible for the transcriptional regulation of HMMR through the mTOR-SRF axis are located in HMMR promoter sequences carrying the first intron, downstream of the translational start site. Furthermore, the upregulation of HMMR by DHT was abolished by stimulation with rapamycin, prior to DHT treatment, suggesting that mTOR activity is required for the induction of HMMR expression by androgen. Collectively, our study provides new mechanistic insights into the role of mTOR/SRF/AR signaling in HMMR regulation in prostate cancer cells.

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.