Epithelial Ovarian Cancer Tissues Research Articles

Mutation Landscape of Homologous Recombination Repair Genes in Epithelial Ovarian Cancer in China and Its Relationship With Clinicopathlological Characteristics.

ObjectiveThe status of homologous recombination repair (HRR) gene mutations and their impact on the survival of patients with Chinese epithelial ovarian cancer (EOC) are still unclear. In this study, we retrospectively analyzed the mutations of HRR genes in tumor tissues and evaluated their values for predicting the survival of Chinese EOC patients.MethodsA total of 273 primary EOC patients from five different hospitals between 2015 and 2016 were recruited. All patients received staging surgeries or debulking surgeries combined with systemic platinum-based chemotherapy. DNA was extracted from formalin-fixed, paraffin-embedded sections and analyzed for mutations using a 21-gene panel (including 13 well-known HRR genes) by next-generation sequencing.ResultsHigh-grade serous carcinoma (HGSOC) accounted for 76.2% of the cohort. A total of 34.1% (93/273) cases had 99 deleterious mutations in 9 HRR genes, namely, BRCA1 (56/273, 20.5%), BRCA2 (20/273, 7.3%), ATM (5/273, 1.8%), RAD51C (5/273, 1.8%), RAD51D (5/273, 1.8%), BRIP1 (2/273, 1.8%), CHEK2 (2/273, 0.7%), FANCI (2/273, 0.7%), and RAD54L (1/273, 0.4%). There is a strong mutual exclusion between HRR genes. The mutation landscape revealed several unappreciated deleterious variants in BRCA1/2 and other HRR genes reported previously. Estimated according to the mutation allele frequency, about 4.8% of the patients had potential somatic HRR gene mutations, which might be underestimated. Moreover, HRR mutations mainly exist in HGSOC (83/208, 39.9%), clear cell (2/30, 6.7%), and endometroid subtypes (8/20, 40%), but not seen in other rare subtypes. BRCA1 mutations tend to be present in younger patients with family history or multiple primary foci. Patients with BRCA1/2 mutations tend to have a longer progression-free survival and overall survival, while other HRR mutation carriers tend to have a shorter progression-free survival, but no significant difference in overall survival.ConclusionThis study revealed the distribution of HRR gene mutations in Chinese EOC tissues. BRCA1/2 account for the majority of HRR gene mutations and predict long prognosis in HGSOC. Non-BRCA HRR mutations also account for a very important proportion and might be associated with poor prognosis in HGSOC. It is suggested that HRR gene mutations need to be detected in EOC tissues and germline status be further clarified in clinical algorithm for potential targeted therapy, genetic screening, and prognosis prediction.

Knockdown of TRIM44 inhibits the progression of ovarian cancer and is related to the FOXM1-EZH2 signaling pathway.

BackgroundTripartite motif-containing protein 44 (TRIM44) was recently identified as a novel oncogene that is overexpressed in several types of human cancers. However, the biological functions of TRIM44 in epithelial ovarian cancer (EOC) remain unclear. Here, we aimed to investigate the role of TRIM44 in EOC and its clinical implications.MethodsTRIM44 was knocked down using shRNA transfection. In vitro proliferation, invasion, migration and apoptosis of ovarian cancer (OC) cells were detected by CCK8, colony formation assay, Transwell inserts and flow cytometry analysis. The growth ability of xenograft tumors was examined in vivo in a nude mouse metastatic tumor model. Finally, we performed gene chip analysis and ingenuity pathway analysis (IPA) to analyze the potential gene network.ResultsHigh expression of TRIM44 was observed in EOC tissues. Knockdown of TRIM44 expression substantially suppressed the proliferation, migration, invasion and colony-forming ability of EOC cells in vitro and attenuated tumor growth in vivo. Mechanistic studies revealed that silencing TRIM44 dramatically downregulated the expression of FOXM1, EZH2, CCNE2, CCND3 and BIRC5 in EOC cells, at least in part through inactivation of the FOXM1-EZH2 signaling pathway.ConclusionsCollectively, these data suggest that downregulation of TRIM44 inhibits the progression of EOC through suppression of the FOXM1-EZH2 signaling pathway. These results provide novel insight into the role of TRIM44 in tumorigenesis and suggest that it could be a potential therapeutic target for ovarian carcinoma.

KHDRBS3 promotes paclitaxel resistance and induces glycolysis through modulated MIR17HG/CLDN6 signaling in epithelial ovarian cancer

Paclitaxel (PTX) resistance contributes to mortality in epithelial ovarian cancer (EOC). Aerobic glycolysis is elevated in the tumor environment and may influence resistance to PTX in EOC. KH domain-containing, RNA-binding signal transduction-associated protein 3 (KHDRBS3) is an RNA binding protein that is up-regulated in EOC, but its underlying mechanism in EOC is unclear. Here, we investigate the role of KHDRBS3 in glycolysis and increased resistance to PTX. Expression of KHDRBS3 and Claudin (CLDN6) were measured in EOC tissue and cells by quantitative real-time PCR, western blotting and immunohistochemistry. The biological functions of KHDRBS3, MIR17HG and CLDN6 were examined using MTT, colony formation, apoptosis and seahorse assays in vitro. For in vivo experiments, a xenograft model was used to investigate the effects of KHDRBS3 and MIR17HG in EOC. Here, we investigate the role of KHDRBS3 in glycolysis and increased resistance to PTX. The expression of KHDRBS3 was up-regulated in PTX-resistant cells. KHDRBS3 knockdown restrained the IC50 of PTX, cell proliferation, colony formation and glycolysis in SKOV3-R and A2780-R cells in vitro and enhanced PTX sensitivity in a xenograft mouse model in vivo. KHDRBS3 interacts with lncRNA MIR17HG, which is down-regulated in EOC tissue and cells. The effect of KHDRBS3 overexpression on PTX resistance and glycolysis was rescued by MIR17HG overexpression. Additionally, MIR17HG interacts with the 3′UTR of CLDN6 and negatively regulates CLDN6 expression. MIR17HG overexpression suppressed the IC50 of PTX and glycolysis by targeting CLDN6. Our results reveal a KHDRBS3–MIR17HG–CLDN6 regulatory axis that contributes to enhanced glycolysis in EOC and represents a potential target for therapy.

LncRNA HCG18 upregulates TRAF4/TRAF5 to facilitate proliferation, migration and EMT of epithelial ovarian cancer by targeting miR-29a/b

BackgroundAlthough long noncoding RNA HLA complex group 18 (lncRNA HCG18) has been suggested to regulate cell growth in several tumours, the function of HCG18 in epithelial ovarian cancer (EOC) and its mechanism are still unclear.MethodsshRNAs were applied to reduce HCG18 and related genes. For overexpression of miRNA, a miRNA mimic was transfected into cells. Quantitative real-time PCR (qRT–PCR) was used to detect levels of HCG18, miR-29a/b, and mRNAs. MTT, colony formation, wound healing and Transwell assays were used to evaluate cell proliferation, migration and invasion, respectively. A luciferase reporter assay was utilized to evaluate NF-κB activity and the binding of miRNAs with HCG18 or TRAF4/5. BALB nude mice injected with cells stably expressing shHCG18 or shNC were used for in vivo modelling. Subcutaneous tumour growth was monitored in nude mice, and immunohistochemistry (IHC) was used to determine expression of the proliferation marker Ki67.ResultsAbnormal expression of HCG18 and miR-29a/b was observed in EOC tissues. Knockdown of HCG18 using shRNA inhibited proliferation, migration, EMT and the proinflammatory pathway in EOC cells. miR-29a/b mimics and TRAF4/5 knockdown exhibited effects similar to HCG18 knockdown. Further experiments suggested that HCG18 directly targets miR-29a/b and upregulates TRAF4/5 expression, which are inhibited by targeting miR-29a/b. Moreover, overexpression of TRAF4/5 antagonized the inhibitory effect of HCG18 knockdown, suggesting that they are involved in HCG18-mediated oncogenic effects. Silencing HCG18 reduced tumour size and levels of Ki67 and TRAF4/5 while increasing miR-29a/b levels in vivo.ConclusionsTaken together, our data revealed an oncogenic signalling pathway mediated by HCG18 in ovarian cell lines, which functions as a ceRNA of miR-29a/b and thus derepresses expression levels of TRAF4/5, facilitating NF-κB pathway-mediated promotion of EOC cell proliferation and migration.

Plasma circN4BP2L2 is a promising novel diagnostic biomarker for epithelial ovarian cancer

BackgroundCircular RNAs (circRNAs) are more stable than linear RNA molecules, which makes them promising diagnostic biomarkers for diseases. By circRNA-sequencing analysis, we previously found that circN4BP2L2 was significantly decreased in epithelial ovarian cancer (EOC) tissues, and was predictive of disease progression. The aim of this study was to evaluate the diagnostic value of plasma circN4BP2L2 in EOC.MethodsThree hundred seventy-eight plasma samples were acquired prior to surgery. Samples were obtained from 126 EOC patients, 126 benign ovarian cyst patients, and 126 healthy volunteers. CircN4BP2L2 was assessed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Cancer antigen 125 (CA125) and human epididymis protein 4 (HE4) were assessed using enzyme-linked immunosorbent assay (ELISA). EOC cells were transfected with small interference RNAs (siRNAs) and cell proliferation, migration, invasion, cell cycle and cell apoptosis were performed to assess the effect of circN4BP2L2 in EOC. Receiver operating curve (ROC), the area under the curve (AUC), sensitivity and specificity were estimated.ResultsPlasma circN4BP2L2 was significantly downregulated in EOC patients. Decreased circN4BP2L2 was significantly associated with advanced tumor stage, worse histological grade, lymph node metastasis and distant metastasis in EOC. CircN4BP2L2 inhibited tumor cell migration and invasion in vitro. CircN4BP2L2 could significantly separate EOC from benign (AUC = 0.82, P < 0.01) or normal (AUC = 0.90, P < 0.01) cohort. Early stage EOC vs benign (AUC = 0.81, P < 0.01) or normal (AUC = 0.90, P < 0.01) cohort could also be distinguished by circN4BP2L2. In discrimination between EOC cohort and benign or normal cohort, circN4BP2L2 performed equally well in both pre- and post-menopausal women. The combination of circN4BP2L2, CA125 and HE4 showed high sensitivity and specificity in detecting EOC cases.ConclusionsPlasma circN4BP2L2 is significantly downregulated in EOC and might serve as a promising novel diagnostic biomarker for EOC patients, especially in early stage EOC cases. CircN4BP2L2 might act as an adjunct to CA125 and HE4 in detecting EOC. Further large-scale studies are warranted to verify our results.

Diagnostic performance of microRNA-34a, let-7f and microRNA-31 in epithelial ovarian cancer prediction.

ObjectiveTo correlate the genome-wide methylation signature of microRNA genes with dysregulated expression of selected candidate microRNA in tissue and serum samples of epithelial ovarian cancer (EOC) and control using quantitative reverse transcription polymerase chain reaction (qRT-PCR), and evaluation of EOC predictive value of candidate microRNA at an early stage.MethodsWe performed Methylated DNA Immunoprecipitation coupled with NGS (MeDIP-NGS) sequencing of 6 EOC and 2 normal tissue samples of the ovary. Expression of selected microRNA from tissue (EOC=85, normal=30) and serum (EOC=50, normal=15) samples was evaluated using qRT-PCR. We conducted bioinformatics analysis to identify the candidate miRNA’s potential target and functional role.ResultsMeDIP-NGS sequencing revealed hypermethylation of several microRNAs gene promoters. Three candidate microRNAs were selected (microRNA-34a, let-7f, and microRNA-31) from MeDIP-NGS data analysis based on log2FC and P-value. The relative expression level of microRNA-34a, let-7f, and microRNA-31 was found to be significantly reduced in early-stage EOC tissues and serum samples (p<0.0001). The receiver operating characteristic analysis of microRNA-34a, let-7f and miR-31 showed improved diagnostic value with area under curve(AUC) of 92.0 (p<0.0001), 87.9 (p<0.0001), and 85.6 (p<0.0001) and AUC of 82.7 (p<0.0001), 82.0 (p<0.0001), and 81.0 (p<0.0001) in stage III-IV and stage I-II EOC serum samples respectively. The integrated diagnostic performance of microRNA panel (microRNA-34a+let-7f+microRNA-31) in late-stage and early-stage serum samples was 95.5 and 96.9 respectively.ConclusionOur data correlated hypermethylation-associated downregulation of microRNA in EOC. In addition, a combined microRNA panel from serum could predict the risk of EOC with greater AUC, sensitivity, and specificity.

EPDR1, Which Is Negatively Regulated by miR-429, Suppresses Epithelial Ovarian Cancer Progression via PI3K/AKT Signaling Pathway.

Epithelial ovarian cancer (EOC) is the main pathological type of ovarian cancer. In this study, we found that ependymin-related 1 (EPDR1) was remarkably downregulated in EOC tissues, and low EPDR1 expression was associated with International Federation of Gynecology and Obstetrics (FIGO) stage, metastasis, and poor prognosis. We confirmed that EPDR1 overexpression dramatically suppressed EOC cell proliferation, migration, and invasion in vitro and in vivo. Mechanistically, EPDR1 inhibited EOC tumorigenesis and progression, at least in part, through the repression of the PI3K (Phosphoinositide 3-kinase)/AKT (AKT Serine/Threonine Kinase 1) signaling pathway. Furthermore, the expression and function of EPDR1 were regulated by miR-429, as demonstrated by luciferase reporter assays and rescue experiments. In conclusion, our study validated that EPDR1, negatively regulated by miR-429, played an important role as a tumor-suppressor gene in EOC development via inhibition of the PI3K/AKT pathway. The miR-429/EPDR1 axis might provide novel therapeutic targets for individualized treatment of EOC patients in the future.

Exosome Component 4 Promotes Epithelial Ovarian Cancer Cell Proliferation, Migration, and Invasion via the Wnt Pathway.

BackgroundOf gynecologic malignancies, ovarian cancer is the leading cause of death, mainly due to the lack of sensitive tumor markers, which means it almost always presents at an advanced stage. Exosome Component 4 (EXOSC4) is involved in RNA degradation, but its role in epithelial ovarian cancer (EOC) is unclear.MethodsThe expression levels of EXOSC4 in EOC and normal ovarian tissue specimens were determined by immunohistochemical staining. The overall survival (OS) and progression-free survival (PFS) of patients with EOC were evaluated after patients were classified into high and low EXOSC4 expression groups, and the Cox regression model was established to identify independent predictors of patient prognosis. The effects of EXOSC4 on proliferation, colony formation, migration, and invasion were examined in the SKOV-3 and HO8910 cell lines by lentivirus-mediated shRNA knockdown. Flow cytometry was used to detect cell cycle changes. The mRNA levels of cyclin D1, CDK4, and c-myc were detected by RT-PCR. The protein expression levels of β-catenin, cyclin D1, CDK4, c-myc, vimentin, N-cadherin, and E-cadherin were assessed by western blot. Wnt/β-catenin activation was measured by TCF/LEF reporter assay.ResultsEXOSC4 was significantly elevated in EOC tissues and cell lines. High EXOSC4 expression was correlated with the International Federation of Gynecology and Obstetrics (FIGO) stage and pathological grade, and identified as an independent predictor of shorter OS and PFS. EXOSC4 knockdown suppressed proliferation, migration, and invasion in EOC cell lines. Cells were arrested at G0/G1 phase after EXOSC4 knockdown. The mRNA levels of cyclin D1, CDK4, and c-myc were decreased. β-catenin, cyclin D1, CDK4, c-myc, vimentin, and N-cadherin protein expression levels were reduced, while those of E-cadherin was increased. Wnt/β-catenin activity was suppressed after the EXOSC4 knockdown.ConclusionsEXOSC4 is involved in EOC. Knockdown of EXOSC4 can inhibit the proliferation, migration, and invasion ability of EOC by suppressing the Wnt pathway. EXOSC4 is expected to be a novel biomarker and molecular target in EOC.

Overexpression of miR-138-5p Sensitizes Taxol-Resistant Epithelial Ovarian Cancer Cells through Targeting Cyclin-Dependent Kinase 6

Background: Ovarian cancer, one of the most malignant diseases in female, is associated with poor diagnosis and low 5-year survival rate. Taxol is a widely used chemotherapeutic drug for the treatment of ovarian cancer by targeting the microtubules of the mitotic spindle to induce cancer cell death. However, with the widespread clinical applications of Taxol, a large fraction of ovarian cancer patients developed drug resistance. Results: Here, we report miR-138-5p is significantly downregulated in epithelial ovarian cancer tissues compared with their matched normal ovarian tissues. Overexpression of miR-138-5p effectively sensitized ovarian cancer cells to Taxol. By establishing Taxol-resistant cell line from the epithelial ovarian cancer cell line, HO-8910, we found miR-138-5p was significantly downregulated in Taxol-resistant cells. Furthermore, overexpression of miR-138-5p dramatically overcame the chemoresistance of Taxol-resistant cells. Intriguingly, bioinformatic analysis indicated miR-138-5p had putative binding sites for cyclin-dependent kinase 6 (CDK6). This negative regulation was further verified from epithelial ovarian cancer tissues. Luciferase assay demonstrated miR-138-5p could directly bind to 3′UTR of CDK6. Importantly, silencing CDK6 expression by siRNA successfully increased the sensitivity of both parental and Taxol-resistant ovarian cancer cells. Finally, rescue experiments clearly elucidated restoration of CDK6 in miR-138-5p-overexpressing ovarian cancer cells successfully recovered the Taxol resistance. Conclusion: In summary, these findings suggest important molecular mechanisms for the miR-138-5p-mediated Taxol sensitivity of ovarian cancer via directly targeting CDK6, suggesting miR-138-5p is an effective therapeutic target for the noncoding RNA-based anti-chemoresistance treatment.

STAT1-Induced Upregulation lncRNA LINC00958 Accelerates the Epithelial Ovarian Cancer Tumorigenesis by Regulating Wnt/β-Catenin Signaling.

Background Growing studies have demonstrated that long noncoding RNAs (lncRNAs) play important roles in tumor progression. In this study, we aimed to explore the potential roles of lncRNA LINC00958 (LINC00958) and its biological functions in epithelial ovarian cancer (EOC). Methods The expression of LINC00958 in 11 cases of EOC and adjacent nontumor specimens and five cell lines was detected by qRT-PCR. CCK-8, colony formation, and flow cytometry assays were conducted to study the cell viabilities of EOC cells. Wound scratch and transwell analyses were carried out for the examination of cell invasion and migration of EOC cells. The targeting associations between LINC00958 and STAT1 were demonstrated by ChIP analyses combined with luciferase reporter assays. The related proteins of Wnt/β-catenin signaling were determined using RT-PCR. Results Higher levels of LINC00958 were observed in EOC tissues and cell lines. Our data also revealed that high LINC00958 expression was partly induced by STAT1. Functionally, knockdown of LINC00958 suppressed the proliferation, migration, and invasion of EOC cells. Mechanistic investigation showed that the inhibitory effect of LINC00958 knockdown on EOC cells was mediated by the Wnt/β-catenin signaling. Conclusion Our findings suggested that STAT1-induced overexpression of LINC00958 promoted EOC progression by modulating Wnt/β-catenin signaling.

High Expression of MYL9 Indicates Poor Clinical Prognosis of Epithelial Ovarian Cancer.

The prognosis of Epithelial Ovarian Cancer (EOC) is poor, but the prognostic biomarkers are neither sensitive nor specific. Therefore, it is very important to search novel prognostic biomarkers for EOC. The present study aimed to investigate Myosin Light Chain 9(MYL9) expression in Epithelial Ovarian Cancer (EOC) tissues (including paraffin-embedded and fresh tissue samples) and its relationship with clinicopathological characteristics, as well as its potential prognostic value in patients with EOC. Between March 2009 and December 2018, all of 184 paraffin-embedded cancer tissues from patients with EOC and 41 paratumor tissues, pathologically confirmed at the Memorial Hospital of Sun Yat-sen University and Integrated Hospital of Traditional Chinese Medicine, Southern Medical University, were collected for the present study and were assessed for MYL9 protein expression patterns using Immunohistochemistry (IHC). Furthermore, from August 2013 to November 2019, 16 fresh EOC tissues and their paired paratumor tissues, pathologically confirmed at the Integrated Hospital of Traditional Chinese Medicine, Southern Medical University were analyzed using Reverse-Transcription Quantitative PCR (RT-qPCR) to detect MYL9 mRNA expression levels. The results showed that MYL9 expression was higher in cancer tissues compared with that in paratumor tissues, and MYL9 overexpression was associated with shorter Recurrence Free Survival (RFS) and Overall Survival (OS) of EOC patients. Furthermore, multivariate Cox model analysis indicated that MYL9 overexpression was an independent poor survival prediction in patients with EOC. MYL9 is upregulated in EOC and may serve as a useful patent of prognostic biomarker in EOC, and it may demonstrate an important value for the clinical treatment and supervision of patients with EOC.

Down-regulation of PSMD4 can attenuate autophagy, enhance the accumulation of intracellular ROS, and increase the sensitivity of epithelial ovarian cancer to carboplatin by inhibiting the NF-κB pathway.

BackgroundThe incidence of ovarian cancer ranks third among female gynecological cancers in the world, and more than 90% of patients are epithelial ovarian cancer (EOC). Carboplatin is the first-line chemotherapy drug for the treatment of EOC patients. However, patients who are resistant to carboplatin often do not benefit from it. Therefore, finding a key molecule that affects carboplatin sensitivity is expected to enhance the efficacy of carboplatin in EOC treatment.MethodsThe human EOC cell line SK-OV-3 and TOV-21G were used in this study. The second-generation sequencing technology was used to sequence the transcripts of carboplatin-resistant EOC cells and parental EOC cells. The bioinformatic analysis of the differentially expressed genes was performed by Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The EOC tissue chip in the Gene Expression Omnibus (GEO) database was also analyzed to screen the target gene. Flow cytometry, Hoechst staining, Western blot, MTS, mitochondrial specific reducing agent (Mito Tempo), and nuclear factor kappa-B (NF-κB) pathway inhibitor (BAY 11-7082) were used to explore the effect of proteasome 26S subunit, non-ATPase 4 (PSMD4) on the autophagy, apoptosis, and reactive oxygen species (ROS) accumulation of carboplatin-resistant EOC cells treated with carboplatin. In vivo, the carboplatin-resistant EOC cell lines treated with PSMD4 knockdown were injected subcutaneously into mice (twelve female BALB/c nude mice) to construct EOC subcutaneous xenograft tumor models.ResultsBased on second-generation sequencing technology and bioinformatics analysis, it was found that PSMD4 is the core molecule in the carboplatin resistance regulatory network in EOC, and its expression is up-regulated in EOC carboplatin-resistant tissues and cells. In vitro and in vivo experimental results show that the down-regulated expression of PSMD4 is closely related to the increase in sensitivity of EOC to carboplatin. Mechanically, we found that inhibiting PSMD4 expression may inhibit the activation of the NF-κB pathway, promote carboplatin-induced ROS accumulation in EOC cells, inhibit EOC cell autophagy, and enhance the sensitivity of EOC to carboplatin.ConclusionsDown-regulation of PSMD4 may inhibit the activation of the NF-κB pathway and autophagy, and up-regulate the level of intracellular ROS accumulation, thereby promoting carboplatin-mediated EOC cell apoptosis and enhancing carboplatin sensitivity.

Cysteine-Rich Intestinal Protein 1 Served as an Epithelial Ovarian Cancer Marker via Promoting Wnt/β-Catenin-Mediated EMT and Tumour Metastasis.

Objective To explore the expression, functions, and the possible mechanisms of cysteine-rich intestinal protein 1 (CRIP1) in epithelial ovarian cancer. Methods Using open microarray datasets from The Cancer Genome Atlas (TCGA), we identified the tumorigenic genes in ovarian cancer. Then, we detected CRIP1 expression in 26 pairs of epithelial ovarian cancer tissue samples by immunohistochemistry (IHC) and performed a correlation analysis between CRIP1 and the clinicopathological features. In addition, epithelial ovarian cancer cell lines A2780 and OVCAR3 were used to examine CRIP1 expression by western blot and qRT-PCR. Various cell function experiments related to tumorigenesis were performed including the CCK8 assay, EdU, Annexin V-FITC/PI apoptosis assay, wound healing, and Transwell assay. In addition, the expression of epithelial-mesenchymal transition (EMT) markers was detected by western blot to illustrate the relationship between CRIP1 and EMT. Furthermore, KEGG pathway enrichment analysis and western blot were conducted to reveal the signaling pathways in which CRIP1 is involved in ovarian cancer pathogenesis. Results CRIP1 was identified as an oncogene from the TCGA database. The IHC score demonstrated that the CRIP1 protein was expressed at a higher level in tumours than in tumour-adjacent tissues and was associated with a higher pathological stage, grade, and positive lymphatic metastasis. In cell models, CRIP1 was overexpressed in serous epithelial ovarian cancer. Cell function experiments showed that the knockdown of CRIP1 did not significantly affect cell proliferation or apoptosis but could exert an inhibitory effect on cell migration and invasion, and also induce changes in EMT markers. Furthermore, KEGG pathway enrichment analysis and western blot showed that CRIP1 could induce ovarian cancer cell metastasis through activation of the Wnt/β-catenin pathway. Conclusion This study is the first to demonstrate that CRIP1 acts as an oncogene and may promote tumour metastasis by regulating the EMT-related Wnt/β-catenin signaling pathway, suggesting that CRIP1 may be an important biomarker for ovarian cancer metastasis and progression.

Identification of solute-carrier family 27A molecules (SCL27As) as a potential biomarker of ovarian cancer based on bioinformatics and experiments

BackgroundOvarian cancer is one of the 3 major gynecological malignancies with high mortality, poor prognosis, and lack of specific diagnostic and prognostic markers. Solute-carrier family 27A molecules (SCL27As) play a crucial role in multiple malignant tumors via the regulation of long-chain fatty acid uptake and subsequent regulation of lipid metabolism. To date, the specific mechanisms and roles of SCL27As in epithelial ovarian cancer (EOC) have remained unclear.MethodsThe Oncomine and Gene Expression Profiling Interactive Analysis (GEPIA) databases and the Kaplan-Meier plotter were used to explore the differential expression and the prognostic value of SCL27As in EOC. The expression of SCL27A6 in 20 normal ovarian tissues and 120 ovarian cancer tissues was detected by immunohistochemistry (IHC). Cell Counting Kit-8 (CCK8) assay and colony-forming experiments were conducted to evaluate the role of SCL27A6 in the proliferation of ovarian cancer cells, so as to verify the clinical application value of SCL27A6 in the diagnosis and prognosis of ovarian cancer. We extracted the data of SCL27A6 for multiple bioinformatics analysis to identify the potential regulatory mechanism of SLC27A6.ResultsThe expression levels of SLC27A1 and SLC27A6 were significantly decreased in ovarian cancer tissues. Prognostic analysis showed that SLC27A2, SLC27A4, SLC27A5, and SLC27A6 expression levels were significantly correlated with overall survival (OS) in EOC patients. Moreover, the expression of the SLC27A6 protein was decreased in EOC tissues, which was related to the prognosis. Additionally, knocking down the expression of SLC27A6 could significantly enhance the malignant biological behavior of ovarian cancer cells. The SLC27A6 gene may be involved in the proteasome, cell cycle, Hippo signaling pathway, and so on.ConclusionsThis study revealed the abnormal expression and prognostic value of SLC27As in EOC. In addition, it was highlighted that SLC27A6 may be a novel biomarker for the diagnosis and prognostication of EOC patients.

Phosphorylation of Cofilin-1 Enhances Paclitaxel Resistance of Epithelial Ovarian Cancer Cells by Inhibiting Apoptosis

ObjectiveTo investigate the molecular mechanism of high phosphorylation levels of cofilin-1 (p-CFL-1) associated with paclitaxel resistance in epithelial ovarian cancer (EOC) cells. MethodsCells displaying varying levels of p-CFL-1 and CFL-1 were created by plasmid transfection and shRNA interference. Cell inhibition rate indicating paclitaxel efficacy was assessed by Cell Counting Kit-8 (CCK-8) assay. Apoptosis was assessed by flow cytometry and protein levels were detected by western blotting. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to measure the expression levels of phosphokinases and phosphatases of CFL-1. Survival analysis evaluated the correlation between the prognosis of EOC patients and the levels of p-CFL-1 and slingshot-1 (SSH-1). ResultsHigh levels of p-CFL-1 were observed in EOC cells that survived treatment with high doses of paclitaxel. SKOV3 cell mutants with upregulated p-CFL-1 showed impaired paclitaxel efficacy, as well as decreased apoptosis rates and pro-survival patterns of apoptosis-specific protein expression. Cytoplasmic accumulation of p-CFL-1 inhibited paclitaxel-induced mitochondrial apoptosis. SSH-1 silencing mediated CFL-1 phosphorylation in paclitaxel-resistant SKOV3 cells. Clinically, the high level of p-CFL-1 and the low level of SSH-1 in EOC tissues were closely related to chemotherapy resistance and poor prognosis in EOC patients. ConclusionThe SSH-1/p-CFL-1 signaling pathway mediates paclitaxel resistance by apoptosis inhibition in EOC and is expected to be a potential prognostic predictor.

Deregulation of Exo70 Facilitates Innate and Acquired Cisplatin Resistance in Epithelial Ovarian Cancer by Promoting Cisplatin Efflux.

Simple SummaryInnate and acquired platinum resistance are the leading causes of epithelial ovarian cancer (EOC) mortality. However, the mechanisms remain elusive. Here we found that Exo70, a key subunit of the exocyst, is upregulated in EOC and promotes cisplatin efflux to facilitate innate resistance. More interestingly, cisplatin could downregulate Exo70 to sustain cell sensitivity. However, this function was hampered during prolonged cisplatin treatment, which in turn stabilized Exo70 to facilitate the acquired cisplatin resistance of EOC cells. Our study potentiates Exo70 as a promising target to overcome cisplatin resistance in EOC.Whilst researches elucidating a diversity of intracellular mechanisms, platinum-resistant epithelial ovarian cancer (EOC) remains a major challenge in the treatment of ovarian cancer. Here we report that Exo70, a key subunit of the exocyst complex, contributes to both innate and acquired cisplatin resistance of EOC. Upregulation of Exo70 is observed in EOC tissues and is related to platinum resistance and progression-free survival of EOC patients. Exo70 suppressed the cisplatin sensitivity of EOC cells through promoting exocytosis-mediated efflux of cisplatin. Moreover, cisplatin-induced autophagy-lysosomal degradation of Exo70 protein by modulating phosphorylation of AMPK and mTOR, thereby reducing the cellular resistance. However, the function was hampered during prolonged cisplatin treatment, which in turn stabilized Exo70 to facilitate the acquired cisplatin resistance of EOC cells. Knockdown of Exo70, or inhibiting exocytosis by Exo70 inhibitor Endosidin2, reversed the cisplatin resistance of EOC cells both in vitro and in vivo. Our results suggest that Exo70 overexpression and excessive stability contribute to innate and acquired cisplatin resistance through the increase in cisplatin efflux, and targeting Exo70 might be an approach to overcome cisplatin resistance in EOC treatment.

Identification and analysis of genes associated with epithelial ovarian cancer by integrated bioinformatics methods.

BackgroundThough considerable efforts have been made to improve the treatment of epithelial ovarian cancer (EOC), the prognosis of patients has remained poor. Identifying differentially expressed genes (DEGs) involved in EOC progression and exploiting them as novel biomarkers or therapeutic targets is of great value.MethodsOverlapping DEGs were screened out from three independent gene expression omnibus (GEO) datasets and were subjected to Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses. The protein-protein interactions (PPI) network of DEGs was constructed based on the STRING database. The expression of hub genes was validated in GEPIA and GEO. The relationship of hub genes expression with tumor stage and overall survival and progression-free survival of EOC patients was investigated using the cancer genome atlas data.ResultsA total of 306 DEGs were identified, including 265 up-regulated and 41 down-regulated. Through PPI network analysis, the top 20 genes were screened out, among which 4 hub genes, which were not researched in depth so far, were selected after literature retrieval, including CDC45, CDCA5, KIF4A, ESPL1. The four genes were up-regulated in EOC tissues compared with normal tissues, but their expression decreased gradually with the continuous progression of EOC. Survival curves illustrated that patients with a lower level of CDCA5 and ESPL1 had better overall survival and progression-free survival statistically.ConclusionTwo hub genes, CDCA5 and ESPL1, identified as probably playing tumor-promotive roles, have great potential to be utilized as novel therapeutic targets for EOC treatment.

LncRNA CDKN2A-AS1 facilitates tumorigenesis and progression of epithelial ovarian cancer via modulating the SOSTDC1-mediated BMP-SMAD signaling pathway

ABSTRACT Ovarian cancer (OC) is the fifth most common female malignant tumor and the leading cause of cancer-related death in women worldwide. Epithelial ovarian cancer (EOC) is the predominant type of OC. Investigating the mechanism underlying tumorigenesis and progression of EOC is urgent. Our previous research has shown that long non-coding RNAs (lncRNAs) CDKN2A-AS1 is upregulated in EOC tissues and cells. Furthermore, we have predicted that CDKN2A-AS1 is associated with the bone morphogenetic protein (BMP)-SMAD signaling pathway, which is negatively regulated by the sclerostin domain containing 1 (SOSTDC1). Therefore, we conjecture that the CDKN2A-AS1 regulate BMP-SMAD signaling pathway via interacting with SOSTDC1, which need more investigation. Moreover, the functions of the BMP-SMAD signaling pathway and the SOSTDC1 on EOC are still unclear. Herein, we unearthed that CDKN2A-AS1, BMP2/4/7, SMAD1/5/9 and phosphorylation of SMAD1/5/9 (p-SMAD1/5/9) were upregulated in EOC tissues and cells, whereas SOSTDC1 was downregulated in EOC tissues and cells. We firstly demonstrated that CDKN2A-AS1 bound directly with the SOSTDC1. CDKN2A-AS1 downregulated the expression of SOSTDC1, but upregulated the expression of BMP2/4/7, SMAD1/5/9, and p-SMAD1/5/9. CDKN2A-AS1 promoted the proliferation, migration, invasion of EOC cells and tumor growth in vivo, whereas SOSTDC1 inhibited the proliferation, migration, invasion of EOC cells. Knockdown SOSTDC1 rescued the inhibitory effect of si-lncRNA CDKN2A-AS1 on the EOC cells proliferation, migration and invasion. These results demonstrated that CDKN2A-AS1activated the BMP-SMAD signaling pathway by directly bind with SOSTDC1 to promote EOC tumor growth. CDKN2A-AS1/SOSTDC1 axis may provide a novel therapeutic strategy for EOC treatment.

The Sonic Hedgehog signaling pathway regulates autophagy and migration in ovarian cancer.

BackgroundThe Sonic Hedgehog (SHH) signaling pathway plays an important role in various types of human cancers including ovarian cancer; however, its function and underlying mechanism in ovarian cancer are still not entirely understood.MethodsWe detected the expressions of SHH and SQSTM1 in borderline ovarian tumor tissues, epithelial ovarian cancer (EOC) tissues and benign ovarian tumor tissues. Cyclopamine (Cyp, a well‐known inhibitor of SHH signaling pathway) and chloroquine (CQ, the pharmaceutical inhibitor of autophagy) were used in vivo and in vitro (autophagic flux, CCK‐8 assay, wound healing assay, transwell assay, tumor xenograft model). The mechanism of action was explored through Quantitative RT‐PCR and Western Blot.ResultsWe found up‐regulation of SHH and accumulation of SQSTM1/P62 in epithelial ovarian cancer. Cyp induced autophagy through the PI3K/AKT signaling pathway. Moreover, low‐dose Cyp and chloroquine (CQ) significantly promoted the migratory ability of SKOV3 cells.ConclusionsOur findings suggest that inhibition of the SHH pathway and autophagy may be a potential and effective therapy for the treatment of ovarian cancer.

Prolyl-4-Hydroxylase α Subunit 2 as a Novel Potential Biomarker for Predicting the Prognosis of Epithelial Ovarian Carcinoma.

ObjectiveEpithelial ovarian cancer (EOC) is one of the leading causes of death worldwide. The aim of this study was to explore the prognostic significance of prolyl-4-hydroxylase α subunit 2 (P4HA2) in patients with EOC.Patients and MethodsA total of 217 clinical samples (EOC tissues, 167 cases; normal ovarian, 50 cases) were collected and pathologically confirmed using hematoxylin and eosin (H&E) staining. P4HA2 expression in clinical samples was stained by immunohistochemistry (IHC). Relationship between P4HA2 expression and clinicopathological characteristics of EOC patients were analyzed using chi-square test. The differential expression of targets was analyzed in Oncomine database. The prognostic value of P4HA2 was investigated in clinical EOC patients and Kaplan–Meier (KM) Plotter database.ResultsIHC staining showed that P4HA2 was significantly up-regulated in EOC tissues, compared to the normal tissues. Two databases retrieved from Oncomine database further confirmed the up-regulation P4HA2 in EOC. Chi-square test demonstrated that P4HA2 expression was associated with clinical stage (p=0.036), tumor grade (p<0.001), and residual disease (p=0.022). Both in clinical samples and KM Plotter database, high P4HA2 expression was significantly associated with worse progression-free survival (PFS) and overall survival (OS). Cox’s proportional hazards regression analysis suggested that high P4HA2 expression were independent risk factors for the survival of EOC patients. Besides, we confirmed the positive correlation between P4HA2 and COL1A1 expression. Moreover, COL1A1 was found to be up-regulated in EOC and also associated with short PFS and OS.ConclusionThe present study preliminarily proved that P4HA2 expression was associated with clinical outcome in EOC patients. P4HA2 might be a prognostic factor for EOC progression, and has the potential to be a valuable therapeutic target for EOC.