Progression Of Epithelial Ovarian Carcinoma Research Articles

Downregulation of INPP4B is Associated with Poor Prognosis in Epithelial Ovarian Carcinoma.

INPP4B is a tyrosine-specific phosphatase in the human body, which plays an important role in the developing process of carcinogenesis. However, The correlation between INPP4B and epithelial ovarian cancer is rarely explored. In this study, the expression of INPP4B in human epithelial ovarian carcinoma and normal ovaries was detected, to explore the correlation between INPP4B expression and clinicopathological risk factors of epithelial ovarian carcinoma and to clarify its significance in the developing process of and prognosis of epithelial ovarian carcinoma. The expression of INPP4B in various tumors was detected by bioinformatics method, and the expression in epithelial ovarian cancer and normal control group was detected by Elisa. The immunohistochemical method was used in this experiment to analyze the expression of INPP4B in specimens of 100 cases of epithelial ovarian carcinoma and 20 cases of normal ovaries. Analysis of clinicopathological risk factors and related survival analysis was carried out on the expression of INPP4B in 100 cases of epithelial ovarian carcinoma. The results showed that the positive expressed INPP4B protein in epithelial ovarian carcinoma was significantly less, compared with that in normal ovaries (P < 0.05). The expression of INPP4B was significantly associated with many clinicopathologic factors, such as tumor differentiation (P < 0.001), FIGO stage (P < 0.001), lymph node metastasis (P < 0.001) and distant metastasis at recurrence (P=0. 009), but not with age, pathologic type of tumor, serum CA125 at recurrence and chemotherapy sensitivity. In epithelial ovarian carcinoma, there is a downregulation of INPP4B expression, which may be related to poor tumor differentiation, late FIGO stage, lymph node metastasis, distant metastasis at recurrence and insensitivity to chemotherapy. Under-expression of INPP4B, lymph node metastasis, FIGO stage, and distant metastasis at recurrence are factors of poor prognostic. The under-expression level of INPP4B may be involved in the progression of epithelial ovarian carcinoma.

KLF5-mediated CDCA5 expression promotes tumor development and progression of epithelial ovarian carcinoma

Cell division cycle associated 5 (CDCA5) is correlated with the development and progression of many malignant tumors. However, little is known about its role in epithelial ovarian cancer (EOC) progression. In this study, the clinical value, biological function and underlying mechanisms of CDCA5 in EOC were evaluated. CDCA5 mRNA and protein levels were substantially upregulated in EOC and had a significant positive correlation with adverse clinicopathological characteristics and a poor prognosis. CDCA5 facilitated proliferation, invasion, and metastasis and disrupted mitochondrial-mediated endogenous apoptosis by activating the cell cycle pathway and inhibiting the P53 pathway in EOC cells. Conversely, knockdown of CDCA5 expression blocked the malignant activities of EOC cells and suppressed the growth of xenograft tumors in vivo. Mechanistically, the transcription factor KLF5 bound to a specific site in the CDCA5 promoter and promoted CDCA5 expression. Moreover, KLF5 overexpression rescued the negative regulation of inhibited CDCA5 expression on EOC cell proliferation. In conclusion, our findings revealed that CDCA5 promoted tumor progression of EOC via the KLF5/CDCA5/cell cycle and P53 axes, which might provide new insights into the roles of CDCA5 in EOC.

FOXA1 can be modulated by HDAC3 in the progression of epithelial ovarian carcinoma

FOXA1 is associated with malignant tumors, but the function of FOXA1 in EOC is unclear. HDAC3 can influence the proliferation, migration and invasion ability of EOC. In this study, we wanted to explore the function of FOXA1 in ovarian cancer and the relationship between HDAC3 and FOXA1.The expression of HDAC3 and FOXA1 was detected by immunohistochemical staining of primary lesions from 127 epithelial ovarian carcinoma patients. A proliferation assay, a Transwell assay, an apoptosis assay and animal experiments were used to assess the proliferation, invasion and apoptosis abilities of ovarian cancer cells before and after transfection with FOXA1. The relevance of the in vitro findings was confirmed in xenografts. The H-scores for FOXA1 and HDAC3 staining in FIGO stage III-IV were noticeably higher and predicted adverse clinical outcomes in patients with ovarian cancer. The expression level of HDAC3 was significantly correlated with the expression level of FOXA1. Invasion, proliferation and apoptosis capacity and tumor formation were decreased in the FOXA1-knockdown cells. Experiments in xenografts confirmed that HDAC3 mediated tumor formation. In conclusion, FOXA1 can be modulated by HDAC3 through the Wnt/β-catenin signaling pathway, and FOXA1 plays essential roles in the proliferation, apoptosis and invasion of EOC cell lines and xenograft experiments.

Identification of key molecular markers in epithelial ovarian cancer by integrated bioinformatics analysis

ObjectiveThe current research was aimed to identify candidate genes associated with development and progression of epithelial ovarian carcinoma using bioinformatics analysis. Materials and methodsWe screened and validated candidate genes associated with carcinogenesis and development of epithelial ovarian carcinoma via bioinformatic analysis in three microarray datasets (GSE14407, GSE29450, and GSE54388) downloaded from the Gene Expression Omnibus (GEO) database. ResultsOur bioinformatic analysis identified 514 differentially expressed genes (DEGs) and nine candidate hub genes (CCNB1, CDK1, BUB1, CDC20, CCNA2, BUB1B, AURKA, RRM2, and TTK). Survival analysis using the Kaplan–Meier plotter showed that high expression levels of seven candidate genes (CCNB1, RRM2, BUB1, CCNA2, AURKA, CDK1, and BUB1B) were associated with poor overall survival (OS). Gene Expression Profiling Interactive Analysis (GEPIA) revealed a higher expression level of these seven candidate genes in ovarian carcinoma samples than in normal ovarian samples. Immunostaining results from the Human Protein Atlas (HPA) database suggested that the protein expression levels of CCNB1, CCNA2, AURKA, and CDK1 were increased in ovarian cancer tissues. No difference was observed in RRM2 protein expression level between normal ovarian and ovarian cancer samples. Oncomine analysis revealed an association between the expression patterns of BUB1B, CCNA2, AURKA, CCNB1, CDK1, and BUB1 and patient clinicopathological information. Finally, six genes, namely CCNB1, CCNA2, AURKA, BUB1, BUB1B, and CDK1, were identified as hub genes and a transcription factor (TF)-gene regulatory network was constructed to identify TFs, including POLR2A, ZBTB11, KLF9, and ELF1, that were implicated in regulating these hub genes. ConclusionSix significant hub DEGs associated with a poor prognosis in epithelial ovarian cancer were identified. These could be potential biomarkers for ovarian cancer patients.

SP1-Regulated Non-Coding RNA SNHG22 Promotes Ovarian Cancer Growth and Glycolysis.

ObjectivesLong non-coding RNAs (lncRNAs) play a crucial part in cancer progression. However, in epithelial ovarian carcinoma (EOC), the role of SNHG22 needs to be further explained.MethodsQuantitative real-time PCR was used to detect the expression of SNHG22. EOC cells were stably transfected with lentivirus approach and cell proliferation, glycolysis and cell apoptosis, as well as tumorigenesis in animal were performed to assess the effects of SNHG22 in EOC. Chromatin immunoprecipitation (ChIP) and luciferase reporter assay were conducted to confirm the relationship between SP1 and SNHG22.ResultsHigher expressed SNHG22 was associated with a poor prognosis in EOC tissues. SNHG22 facilitated glycolysis and proliferation. Mechanistically, LDHA deficiency and glycolysis inhibitor (2-DG, 3-BG) partly rescued proliferation. SP1 mediated SNHG22 expression at the transcriptional level and the SNHG22 promoter region (−900~ −600) was necessary for SP1 binding. Hypoxia and HIF-1α also upregulated SNHG22 expression.ConclusionSNHG22 is an independent prognostic biomarker for EOC. SNHG22 promotes EOC progression and is a prospective therapeutic target.

MicroRNA‑195 suppresses cell proliferation, migration and invasion in epithelial ovarian carcinoma via inhibition of the CDC42/CCND1 pathway

Epithelial ovarian carcinoma (EOC) is the most common cause of gynecological cancer mortality, and poses a threat to women. MicroRNA-195 (miR-195) has been reported to induce apoptosis of human OVCAR-3 cells by inhibiting the VEGFR2/AKT pathway. However, the role of miR-195 in EOC remains unknown. A previous study reported that cell division cycle 42 (CDC42) can serve as a target gene of miR-195 and mediate malignant progression of esophageal squamous cell carcinoma (ESCC). The aim of the present study was to investigate the role of miR-195 in EOC and the regulation in CDC42/CCND1 pathway. Tissues samples and clinical materials were collected from 78 enrolled patients with EOC to analyze the expression and clinical significance of miR-195, CDC42 and cyclin D1 (CCND1). Human EOC cell lines OVCA420, OVCAR-3, A2780 and SKOV3 cell lines were used to assess the expression and function of miR-195, CDC42 and CCND1 in vitro. Cell proliferation, the cell cycle and apoptosis, as well as the cell migratory and invasive abilities were detected in vitro using BrdU incorporation, colony formation, wound healing and Transwell invasion assays, along with flow cytometry. miR-195 was downregulated, while CDC42 and CCND1 were upregulated in human EOC tissues and cells, and the aberrant expression of both was associated with increased EOC malignancy. Moreover, miR-195 expression was negatively correlated with CDC42 and CCND1 expression levels, and negatively regulated these expression levels. Thus, it was suggested that miR-195 functions as a tumor suppressor, but CDC42 and CCND1 act as tumor promoters based their abilities to enhance cell proliferation, cell cycle entry, migration and invasion, as well as decrease apoptosis in OVCAR-3 cells. the present results demonstrated that miR-195 inhibited human EOC progression by downregulating CDC42 and CCND1 expression. Furthermore, it was identified that miR-195, CDC42 and CCND1 may be effective biomarkers for EOC diagnosis and treatment.

Abstract GMM-032: UNRAVELLING THE STEROID METABOLOME IN EPITHELIAL OVARIAN CANCER

Abstract INTRODUCTION: Ovarian tissue possesses the capacity for steroidogenesis from as early as seven weeks gestation. Perturbations in the steroidogenic pathways have been implicated in the development and progression of epithelial ovarian carcinoma in adulthood but the evidence has been limited. Analysis of RNA-seq data from ovarian tissue now allows the genes involved in these pathways to be studied comprehensively helping direct future investigations and highlighting possible mechanisms of the disease. METHODS: RNA-seq data from 293 serous epithelial ovarian carcinomas from The Cancer Genome Atlas were extracted and the genes involved in steroidogenesis were systematically interrogated for high and low expression. Pathway analysis was employed to confirm significant findings. Comparison was made with novel RNA-seq data from first trimester fetal ovary and healthy adult ovary. RESULTS: Genes involved in the synthesis of androgens (HSD17B1, SRD5A1, AKR1C3, AR and STS) were all significantly upregulated in the high grade serous ovarian carcinoma group when compared to normal healthy ovary (p&amp;lt;0.05). Expression of several steroidogenic genes in malignancy appeared to mimic increased fetal expression with relative quiescence noted in the healthy ovary group. CONCLUSIONS: Transcriptome analysis shows upregulated gene expression of the androgen synthesis pathway is associated with epithelial ovarian cancer. This is supported by immunohistochemistry findings in recent studies. Investigation of the androgen pathway does remain a valid opportunity for mechanistic insight into this disease and potentially diagnostic and therapeutic exploitation. Ovarian steroidogenic gene expression in the first trimester appears to correlate with neoplastic growth in serous ovarian cancer but the significance of this is uncertain. Citation Format: David N. Jeevan, Wiebke Arlt, Paul A. Foster and Sudha Sundar. UNRAVELLING THE STEROID METABOLOME IN EPITHELIAL OVARIAN CANCER [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr GMM-032.

SNHG22 overexpression indicates poor prognosis and induces chemotherapy resistance via the miR-2467/Gal-1 signaling pathway in epithelial ovarian carcinoma.

Recently, an increasing number of studies have reported that dysregulation of long noncoding RNAs (lncRNAs) plays an important role in cancer initiation and progression, including in epithelial ovarian carcinoma (EOC). However, little is known about the detailed biological functions of the lncRNA small nucleolar RNA host gene 22 (SNHG22) during the progression of EOC. Here, we found that SNHG22 was significantly increased in EOC tissues and was significantly associated with a low level of differentiation. Forced SNHG22 expression promoted chemotherapy resistance in EOC cells. Knockdown of SNHG22 expression increased the sensitivity of EOC cells to cisplatin and paclitaxel. Importantly, we found that SNHG22 could directly interact with miR-2467 and lead to the release of miR-2467-targeted Gal-1 mRNA. Moreover, SNHG22 overexpression induced EOC cell resistance to chemotherapy agents via PI3K/AKT and ERK cascade activation. In summary, our findings demonstrate that SNHG22 plays a critical role in the chemotherapy resistance of EOC by mediating the miR-2467/Gal-1 regulatory axis.

CD44 Regulates Formation of Spheroids and Controls Organ-Specific Metastatic Colonization in Epithelial Ovarian Carcinoma.

Disseminating epithelial ovarian cancer cells often become assembled into spheroids prior to their arrival at metastatic sites within the peritoneal cavity. Although epithelial ovarian carcinoma (EOC) is the deadliest gynecologic malignancy, the mechanisms regulating formation and metastatic potential of spheroids are poorly understood. We show that expression of a cell surface glycoprotein CD44 is an important contributing factor for spheroid formation and spheroid adhesion to mesothelial cells, and its loss impairs mesenteric metastasis. In contrast, loss of CD44 resulted in significant increase of tumor burden at several locoregional sites, including liver, and unleashed distant metastases to the thoracic cavity. Altogether our studies suggest that CD44 regulates metastatic progression of EOC in an organ-specific manner. IMPLICATIONS: Expression of CD44 promotes spheroid formation, mesothelial adhesion, and formation of mesenteric metastasis, but it suppresses development of metastasis to several peritoneal sites, including liver, and the thoracic cavity.

The role of transforming growth factor-β (TGF-β) in epithelial ovarian cancer progression via immunosuppression

Objective: Increased TGF-β signaling is associated with poorer prognosis in advanced-stage epithelial ovarian carcinoma (EOC). We sought to demonstrate the role of the TGF-β-mediated immune suppression in the progression of EOC.

Expression of zinc finger transcription factors (ZNF143 and ZNF281) in serous borderline ovarian tumors and low-grade ovarian cancers

Low-grade ovarian cancers represent up to 8% of all epithelial ovarian carcinomas (EOCs). Recent studies demonstrated that epithelial-mesenchymal transition (EMT) is crucial for the progression of EOCs. EMT plays a key role in cancer invasion, metastasis formation and chemotherapy resistance. An array of novel EMT transcription factors from the zinc finger protein family have been described recently, among them zinc finger protein 143 (ZNF143) and zinc finger protein 281 (ZNF281). The study included tissue specimens from 42 patients. Based on histopathological examination of surgical specimens, eight lesions were classified as serous borderline ovarian tumors (sBOTs) and 34 as low-grade EOCs. The proportions of the ovarian tumors that tested positively for ZNF143 and ZNF281 were 90 and 57%, respectively. No statistically significant differences were found in the expressions of ZNF143 and ZNF281 transcription factors in SBOTs and low-grade EOCs. Considering the expression patterns for ZNF143 and ZNF281 identified in this study, both sBOTs and low-grade EOCs might undergo a dynamic epithelial-mesenchymal interconversion. The lack of statistically significant differences in the expressions of the zinc finger proteins in sBOTs and low-grade serous EOCs might constitute an evidence for common origin of these two tumor types.

Cancer-associated fibroblasts in epithelial ovarian carcinoma: relation to clinicopathologic characteristics and microvessel density

Background Epithelial ovarian carcinoma (EOC) is the leading cause of mortality among all gynecologic cancers. Recent studies have emphasized the role of the tumor microenvironment in tumor progression. Cancer-associated fibroblasts (CAFs) are the most frequent component in the tumor microenvironment, and α-smooth muscle actin (α-SMA) is the most commonly used marker to identify CAF. This study aimed to investigate the potential role of CAFs in the progression of EOC. Furthermore, we analyzed the relation between the presence of CAF in EOC and clinicopathologic characteristics and tumor microvessel density (MVD). Methods Immunohistochemical staining was performed on 75 cases of epithelial ovarian neoplasms using α-SMA (as a marker of CAFs) and CD34 (for assessment of MVD in EOC cases). Results The presence of CAFs, as assessed by α-SMA intensity and percentage of positive stained area, was significantly more prominent in malignant cases, as compared with borderline and benign cases. It was found that abundant CAFs were significantly associated with high-grade tumors, stage III tumors and tumors with vascular invasion and omental metastasis. A significant positive correlation was detected between the presence of CAFs and tumor MVD in EOC. Conclusion CAFs significantly increase from benign through borderline to malignant ovarian neoplasms. CAFs are significantly higher in high-grade tumors, advanced stage tumors and in tumors with vascular invasion or omental metastasis. CAFs are significantly correlated with tumor MVD in EOC. Thus, CAFs may serve as a poor prognostic indicator in patients with EOC.

Long non-coding RNA CCAT2 promotes epithelial-mesenchymal transition involving Wnt/β-catenin pathway in epithelial ovarian carcinoma cells.

Long non-coding RNA colon cancer-associated transcript 2 (CCAT2) is dysregulated in a variety of types of human cancer. However, the role of CCAT2 in epithelial ovarian carcinoma (EOC) remains largely unknown. The aim of this study is to investigate the effect of CCAT2 on epithelial-mesenchymal transition (EMT) and related molecular mechanisms in epithelial ovarian cancer cells. In the current paper, we found that CCAT2 was significantly upregulated in EOC SKOV3, A2780 and HO8910 cell lines compared with the normal ovarian epithelial HUM-CELL-0088 cell line. Functional assays demonstrated that the knockdown of CCAT2 inhibited migration and invasion of EOC cells in vitro. Moreover, our results showed that silencing CCAT2 inhibited EMT by the upregulation of epithelial cadherin and downregulation of neural cadherin, zinc finger protein SNAI and Twist-related protein 1 in SKOV3 and A2780 cell lines. But, that was reversed by the treatment with lithium chloride (LiCl), by which the canonical Wnt/β-catenin pathway could be activated. In addition, we further investigated the role of CCAT2 in the modulation of Wnt/β-catenin signaling pathway. Our results revealed that knockdown of CCAT2 inhibited the expression of β-catenin and the activity of T-cell factor/lymphoid enhancer factor, acting as a key transcription factor of Wnt signaling pathway. Collectively, these results indicate that CCAT2 may promote EMT, at least partly through Wnt/β-catenin signaling pathway in EOC cells. Thus, CCAT2 might play a critical role in EOC progression and serve as a valuable target for the treatment of ovarian cancer.

The role of the long non-coding RNA TDRG1 in epithelial ovarian carcinoma tumorigenesis and progression through miR-93/RhoC pathway.

As one of the most frequently diagnosed cancers in women, the development and progression of epithelial ovarian carcinoma (EOC) remains an open area of research. The role of long non-coding RNAs (lncRNAs) in EOC is an emerging field of study. We found that LncRNA TDRG1 (human testis development-related gene 1) was highly expressed in EOC tissues than in normal ovarian tissues, and expression differed significantly with differentiation. LncRNA TDRG1 downregulation suppressed EOC cell proliferation, migration, and invasion, while its overexpression had the opposite effect. Bioinformatic predictions and dual-luciferase reporter assays showed that LncRNA TDRG1 has possible miRNA-93 (miR-93) binding sites. LncRNA TDRG1 downregulation upregulated miR-93 expression, while its overexpression reduced miR-93 expression. In addition, TDRG1 downregulation reduced the expression of Ras homolog gene family member C (RhoC), P70 ribosomal S6 kinase (P70S6 K), Bcl-xL, and matrix metalloproteinase 2 (MMP2) protein, which are regulated by miR-93, while its upregulation induced RhoC, P70S6 K, Bcl-xL, and MMP2 protein expression. In vivo, LncRNA TDRG1 overexpression induced tumor development and RhoC expression. Taken together, our results demonstrated for the first time that LncRNA TDRG1 may be a new and important diagnostic and therapeutic target in EOC.

DLEU1 contributes to ovarian carcinoma tumourigenesis and development by interacting with miR-490-3p and altering CDK1 expression.

Recently, a large number of studies have focused on the important role of long non‐coding RNAs (lncRNAs) in metabolism and development and have found that abnormal lncRNA expression is associated with the pathogenesis and development of many diseases. The lncRNA DLEU1 is involved in many solid tumours and haematological malignancies. However, its role in epithelial ovarian carcinoma (EOC) and the associated molecular mechanisms has not been reported. In this study, quantitative reverse transcription–PCR (qRT–PCR) demonstrated higher lncRNA DLEU1 expression in EOC tissues than in normal tissues. Plasmid transfection of DLEU1 to up‐regulate its expression in the ovarian cancer cell lines A2780 and OVCAR3 increased cell proliferation, migration, and invasion, while inhibited apoptosis. Nude mouse xenograft assay demonstrated that DLEU1 overexpression promoted tumour growth in vivo. QRT–PCR showed decreased miR‐490‐3p expression, while Western blotting demonstrated increased its target genes CDK1, cyclinD1 and SMARCD1, as well as matrix metalloproteinase‐2 (MMP2), Bcl‐xL and P70S6K protein expression, respectively. Short interfering RNA silencing of DLEU1 produced opposite results, where qRT–PCR showed increased miR‐490‐3p expression. The dual‐luciferase reporter assay revealed a direct interaction between DLEU1 and miR‐490‐3p. MiR‐490‐3p plays a tumour suppressor role in epithelial ovarian cancer by targeting CDK1 regulation and influencing SMARCD1 and cyclin D1 (CCND1) expressions. Therefore, we suggest that through interaction with miR‐490‐3p, DLEU1 may influence the expression of CDK1, CCND1 and SMARCD1 protein, subsequently promoting the development and progression of EOC.

Upregulation of the lncRNA Meg3 induces autophagy to inhibit tumorigenesis and progression of epithelial ovarian carcinoma by regulating activity of ATG3.

Maternally expressed gene 3 (Meg3), a long non-coding RNA, has been reported to be associated with the pathogenesis of multiple malignancies. However, little is known regarding the role of Meg3 in epithelial ovarian cancer (EOC). In this study, we found that the expression of Meg3 was lower in epithelial ovarian carcinoma, and has potential to be considered as a biomarker for ovarian cancer. After transfecting the ovarian cancer cell lines OVCAR3 and A2780 with Meg3, phenotypic changes and autophagy-related molecules were examined. Upregulation of Meg3 inhibited cell proliferation, plate colony formation, induced cell cycle arrest in G2 phases, and promoted apoptosis. Observation of autophagosomes was performed by transmission electron microscopy. The expression levels of LC3-II, ATG3, LAMP1 were elevated, while SQSTM1/p62 expression declined. Upregulated expression of Meg3 also suppressed tumorigenesis in vivo in a xenograft mouse model through upregulating ATG3 expression. RIP (ribonucleoprotein immunoprecipitation) and RNA pull-down assays showed that Meg3 was co-immunoprecipitated with ATG3. In addition, Meg3 protected ATG3 mRNA from degradation following treatment with actinomycin D. Overall, our results suggest that the lncRNA Meg3 acts as a tumor suppressor in EOC by regulating ATG3 activity and inducing autophagy.

Capn4 Enhances Osteopontin Expression through Activation of the Wnt/β-Catenin Pathway to Promote Epithelial Ovarian Carcinoma Metastasis

Background and Aim: Increasing evidence shows that the calpain regulatory subunit Capn4 can modulate the proliferation and metastasis of cancer cells, and plays an important role in the development of malignant tumors. However, there is no information on the clinical significance of Capn4 in epithelial ovarian carcinoma (EOC) or the molecular mechanisms by which Capn4 promotes the growth and metastasis of EOC. Therefore, the aim of this study was to clarify the role of Capn4 in EOC. Methods: We evaluated Capn4 and osteopontin (OPN) expression in EOC cell lines and tissues from patients with ovarian cancer by western blotting and immunohistochemical analysis. We then created cell lines with downregulated and upregulated Capn4 expression, using Capn4-targeting small interfering RNA and a pcDNA3.1-Capn4 overexpression vector, respectively, to investigate its function in EOC in vitro. In addition, we investigated the potential mechanism underlying the function of Capn4 by examining the effect of modifying Capn4 expression on Wnt/β-catenin signaling pathway-related genes by western blotting. Results: Capn4 was overexpressed in clinical EOC tissues compared with that in normal ovarian epithelial tissue, and was associated with poor clinical outcomes. Upon silencing or overexpressing Capn4 in EOC cells, we concluded that Capn4 promotes cell proliferation and migration in vitro. Furthermore, Capn4 promoted EOC metastasis by interacting with the Wnt/β-catenin signaling pathway to upregulate OPN expression. Conclusion: Our study indicates that Capn4 plays a critical role in the progression and metastasis of EOC, and could be a potential therapeutic target for EOC management.

METCAM/MUC18 is a novel tumor and metastasis suppressor for the human ovarian cancer SKOV3 cells.

BackgroundIncreased expression of METCAM/MUC18, a trans-membrane cell adhesion molecule in the Ig-like gene superfamily, has been associated with the malignant progression of epithelial ovarian carcinomas. To investigate if this is a fortuitous correlation or if METCAM/MUC18 actually plays a role in the progression of the cancer, we tested effects of enforced expression of METCAM/MUC18 on in vitro behaviors, in vivo tumorigenesis, and in vivo malignant progression of human ovarian cancer SK-OV-3 cells, which minimally expressed this protein.MethodsFor in vitro and in vivo tests, we transfected human METCAM/MUC18 cDNA gene into SK-OV-3 cells in a mammalian expression vector pcDNA3.1+ and obtained G418-resistant (G418R) clones, which expressed various levels of human METCAM/MUC18. To mimic physiological situations, we used pooled METCAM/MUC18-expressing and control (vector) clones for testing effects of human METCAM/MUC18 over-expression on in vitro motility and invasiveness, and on in vivo tumor formation and metastasis in female athymic nude mice. Effects of METCAM/MUC18 on the expression of various downstream key factors related to tumorigenesis were also evaluated by Western blot analyses.ResultsThe over-expression of METCAM/MUC18 inhibited in vitro motility and invasiveness of SK-OV-3 cells. SK-OV-3 cells of the control (vector) clone (3D), which did not express human METCAM/MUC18, supported the formation of a solid tumor after SC injection of the cells at dorsal or ventral sites and also formation of solid tumor and ascites after IP injection in the intraperitoneal cavity of nude mice. In contrast, SK-OV-3 cells from the METCAM/MUC18-expressing clone (2D), which expressed a high level of METCAM/MUC18, did not support the formation of a solid tumor at SC sites, or formation of ascites in the intraperitoneal cavity of nude mice. Expression levels of downstream key factors, which may affect tumor proliferation and angiogenesis, were reduced in tumors induced by the METCAM/MUC18-expressing clone (2D).ConclusionsWe conclude that increased human METCAM/MUC18 expression in ovarian cancer SK-OV-3 cells suppressed tumorigenesis and ascites formation in nude mice, suggesting that human METCAM/MUC18 plays a suppressor role in the progression of ovarian cancer, perhaps by reducing proliferation and angiogenesis.

MiR-26b/KPNA2 axis inhibits epithelial ovarian carcinoma proliferation and metastasis through downregulating OCT4.

Karyopherin alpha 2 (KPNA2) is a nuclear transport protein upregulated in many cancers. Our previous study has identified KPNA2 overexpression in epithelial ovarian carcinoma (EOC) tissues, which predicts poor prognosis. However, the mechanism of KPNA2 overexpression in EOC remains unclear. This study aimed to examine the role of miRNA in KPNA2 dysregulation. Our results showed that miR-26b was downregulated in EOC samples, and correlated inversely with KPNA2 expression. Low expression of miR-26b was associated with advanced FIGO stage, poor differentiation, higher risk of distant metastasis and recurrence. Downregulation of miR-26b predicted poor disease-free survival and overall survival in EOC patients. KPNA2 was validated as a direct target of miR-26b. Knockdown of KPNA2 or ectopic expression of miR-26b could downregulate OCT4, vimentin and upregulate E-cadherin. Reintroduction of KPNA2 partially abrogated the suppression effect induced by miR-26b. We further verified that miR-26b/KPNA2/OCT4 axis inhibited EOC cell viability, migratory ability and sphere-forming capacity in vitro and in vivo. In conclusion, our results reveal that miR-26b is downregulated in EOC, and directly targets KPNA2. miR-26b/KPNA2 axis suppresses tumor proliferation and metastasis through decreasing OCT4 expression, which is indicative of the important role of miR-26b/KPNA2/OCT4 axis in EOC carcinogenesis and progression.

RhoC is a major target of microRNA-93-5P in epithelial ovarian carcinoma tumorigenesis and progression.

BackgroundAn increasing amount of evidence has revealed that microRNAs regulate various biological processes, including cell differentiation, cell proliferation, apoptosis, drug resistance, and fat metabolism. Studies have shown that miR-93’s targetome in cancer has not been fully defined. Moreover, the role of miR-93 in epithelial ovarian carcinoma (EOC) remains largely unknown.MethodsMIR-93 mRNA expression in normal ovarian tissue, benign tumors, borderline tumors, primary ovarian carcinomas, and metastatic omentum was quantified. The ovarian carcinoma cell lines OVCAR3, SKOV3/DDP, and HO8910-PM were transfected with miR-93-5P, after which cell phenotype and expression of relevant molecules were assayed. Dual-luciferase reporter assay and a xenograft mouse model were used to examine miR-93 and its target gene RHOC (Ras homolog gene family member C).ResultsMIR-93 mRNA expression was significantly lower in ovarian carcinomas and borderline tumors than in normal ovarian tissues (p < 0.05), and was lower in metastatic omentum than in relative primary ovarian carcinomas (p < 0.05). MIR-93 mRNA expression was also negatively associated with differentiation (well vs. poor and moderate) and International Federation of Gynecology and Obstetrics staging (FIGO stage I/II vs. stage III/IV) in ovarian carcinoma (p < 0.05), besides, miR-93 was higher expressed in mucinous adenocarcinoma than the other types (p < 0.05). MiR-93-5P overexpression reduced proliferation (p < 0.05); promoted G1 or S arrest and apoptosis (p < 0.05); suppressed migration and invasion (p < 0.05); and reduced RhoC, P70S6 kinase, Bcl-xL, matrix metalloproteinase 9 (MMP9) mRNA or protein expression; conversely, it induced P53 and cleaved PARP expression (p < 0.05). Dual-luciferase reporter assay indicated that miR-93 directly targeted RhoC by binding its 3′ untranslated region. MiR-93-5P transfection also suppressed tumor development and RhoC expression (determined by immunohistochemistry) in vivo in the xenograft mouse model (p < 0.05).ConclusionsThis is the first demonstration that miR-93-5P may inhibit EOC tumorigenesis and progression by targeting RhoC. These findings indicate that miR-93-5P is a potential suppressor of ovarian cellular proliferation. The involvement of miR-93-5P–mediated RhoC downregulation in inhibiting EOC aggressiveness may provide extended insight into the molecular mechanisms underlying cancer aggressiveness.