Epithelial Ovarian Cancer Tissue Samples Research Articles

Regulation of ULK1 by WTAP/IGF2BP3 axis enhances mitophagy and progression in epithelial ovarian cancer

There is a pressing need for innovative therapeutic strategies for patients with epithelial ovarian cancer (EOC). Previous studies have shown that UNC-51-like kinase 1 (ULK1), a serine/threonine kinase, is crucial in regulating cellular autophagy and mitophagy across various tumor types. However, the clinical implications, biological functions, and potential mechanisms of ULK1 in EOC remain poorly understood. This study demonstrates that ULK1 expression is upregulated in EOC tissue samples and EOC cell lines, with increased ULK1 expression correlating with poor prognosis. Functionally, overexpressed ULK1 enhances the proliferation and migration abilities of EOC cells both in vitro and in vivo. Mechanistically, ULK1 was identified as an m6A target of WTAP. WTAP-mediated m6A modification of ULK1 enhanced its mRNA stability in an IGF2BP3-dependent manner, leading to elevated ULK1 expression and enhanced mitophagy in EOC. In summary, our research reveals that the WTAP/IGF2BP3-ULK1 axis significantly influences protective mitophagy in EOC, contributing to its progression. Therefore, the regulatory mechanisms and biological function of ULK1 identify it as a potential molecular target for therapeutic intervention in EOC.

Metabolic pathways of potential miRNA biomarkers derived from liquid biopsy in epithelial ovarian cancer.

Epithelial ovarian cancer (EOC) is the type of OC with the highest mortality rate. Due to the asymptomatic nature of the disease and few available diagnostic tests, it is mostly diagnosed at the advanced stage. Therefore, the present study aimed to discover predictive and/or early diagnostic novel circulating microRNAs (miRNAs or miRs) for EOC. Firstly, microarray analysis of miRNA expression levels was performed on 32 samples of female individuals: Eight plasma samples from patients with pathologically confirmed EOC (mean age, 45 (30-54) years), eight plasma samples from matched healthy individuals (HIs) (mean age, 44 (30-65) years), eight EOC tissue samples (mean age, 45 (30-54) years) and eight benign ovarian (mean age, 35 (17-70) years) neoplastic tissue samples A total of 31 significantly dysregulated miRNAs in serum and three miRNAs in tissue were identified by microarray. The results were validated using reverse transcription-quantitative PCR on samples from 10 patients with pathologically confirmed EOC (mean age, 47(30-54) years), 10 matched His (mean age, 40(26-65) years], 10 EOC tissue samples (mean age, 47(30-54) years) and 10 benign ovarian neoplastic tissue samples (mean age, 40(17-70) years). The 'Kyoto Encyclopedia of Genes and Genomes' (KEGG) database was used for target gene and pathway analysis. A total of three miRNAs from EOC serum (hsa-miR-1909-5p, hsa-miR-885-5p and hsa-let-7d-3p) and one microRNA from tissue samples (hsa-miR-200c-3p) were validated as significant to distinguish patients with EOC from HIs. KEGG pathway enrichment analysis showed seven significant pathways, which included 'prion diseases', 'proteoglycans in cancer', 'oxytocin signaling pathway', 'hippo signaling pathway', 'adrenergic signaling in cardiomyocytes', 'oocyte meiosis' and 'thyroid hormone signaling pathway', in which the validated miRNAs served a role. This supports the hypothesis that four validated miRNAs, have the potential to be a biomarker of EOC diagnosis and target for treatment.

A comparison of four technologies for detecting p53 aggregates in ovarian cancer.

The tumor suppressor protein p53 is mutated in half of all cancers and has been described to form amyloid-like structures, commonly known from key proteins in neurodegenerative diseases. Still, the clinical relevance of p53 aggregates remains largely unknown, which may be due to the lack of sensitive and specific detection methods. The aim of the present study was to compare the suitability of four different methodologies to specifically detect p53 aggregates: co-immunofluorescence (co-IF), proximity ligation assay (PLA), co-immunoprecipitation (co-IP), and the p53-Seprion-ELISA in cancer cell lines and epithelial ovarian cancer tissue samples. In 7 out of 10 (70%) cell lines, all applied techniques showed concordance. For the analysis of the tissue samples co-IF, co-IP, and p53-Seprion-ELISA were compared, resulting in 100% concordance in 23 out of 30 (76.7%) tissue samples. However, Co-IF lacked specificity as there were samples, which did not show p53 staining but abundant staining of amyloid proteins, highlighting that this method demonstrates that proteins share the same subcellular space, but does not specifically detect p53 aggregates. Overall, the PLA and the p53-Seprion-ELISA are the only two methods that allow the quantitative measurement of p53 aggregates. On the one hand, the PLA represents the ideal method for p53 aggregate detection in FFPE tissue, which is the gold-standard preservation method of clinical samples. On the other hand, when fresh-frozen tissue is available the p53-Seprion-ELISA should be preferred because of the shorter turnaround time and the possibility for high-throughput analysis. These methods may add to the understanding of amyloid-like p53 in cancer and could help stratify patients in future clinical trials targeting p53 aggregation.

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.

RNF144A suppresses ovarian cancer stem cell properties and tumor progression through regulation of LIN28B degradation via the ubiquitin-proteasome pathway.

Cancer stem cells (CSCs) are the main driving force of tumorigenesis, metastasis, recurrence, and drug resistance in epithelial ovarian cancer (EOC). The current study aimed to explore the regulatory effects of ring finger protein 144A (RNF144A), an E3 ubiquitin ligase, in the maintenance of CSC properties and tumor development in EOC. The expressions of RNF144A in EOC tissue samples and cells were examined. The knockdown or overexpression of a target gene was achieved by transfecting EOC cells with short hairpin RNA or adenoviral vectors. A mouse xenograft model was constructed by inoculating nude mice with EOC cells. Co-immunoprecipitation was used to determine the interaction between RNF144A and LIN28B. Downregulated RNF144A expression was observed in ovarian tumor tissues and EOC cells. Low RNF144A expression was positively associated with poor survival of EOC patients. RNF144A knockdown significantly enhanced sphere formation and upregulated stem cell markers in EOC cells, while RNF144A overexpression prevented EOC cells from acquiring stem cell properties. Also, the upregulation of RNF144A inhibited ovarian tumor growth and aggressiveness in cell culture and mouse xenografts. Further analysis revealed that RNF144A induced LIN28B degradation through ubiquitination in EOC cells. LIN28B upregulation restored the expressions of stem cell pluripotency-associated transcription factors in EOC cells overexpressing RNF144A. Taken together, our findings highlight the therapeutic potential of restoring RNF144A expression and thereby suppressing LIN28B-associated oncogenic signaling for EOC treatment. • Ring finger protein 144A (RNF144A) is downregulated in epithelial ovarian cancer (EOC) tissues and cell lines. • The overexpression of RNF144A prevents EOC cells from acquiring stem cell properties and inhibits ovarian tumor growth. • RNF144A induces LIN28B degradation through ubiquitination in EOC cells. • LIN28B upregulation restores the expressions of stem cell pluripotency-associated transcription factors in EOC cells overexpressing RNF144A.

CircRNA circ_0072995 promotes the progression of epithelial ovarian cancer by modulating miR-147a/CDK6 axis.

Background: Increasing evidence has indicated that circular RNAs (circRNAs) play vital roles in modulating tumor progression. However, regulatory roles and underlying mechanisms of circRNA circ_0072995 in epithelial ovarian cancer (EOC) are not well characterized.Results: Circ_0072995 was up regulated in EOC afflicted tissues and cell lines (HO8910 and A2780), and was mainly located in the cytoplasm. The expression of circ_0072995 was associated with the pathological grade of EOC for respective patients. Functional experiments revealed that circ_0072995 promoted EOC cell proliferation, migration, induced apoptosis, as well as enhanced tumorigenesis in vivo. Mechanistic analyses indicated that circ_0072995 may have acted as a sponge of miR-147a such as to relieve repressive effects of miR-147a upon its target CDK6.Conclusions: Our results revealed that circ_0072995 promoted EOC progression through the circ_0072995/miR-147a/CDK6 axis and may represent a strategy for treatment of EOC afflicted patients.Methods: Expression of circ_0072995 was evaluated in 40 EOC tissue samples and cell lines by qRT-PCR. The location of circ_0072995 was determined via nuclear-cytoplasmic fractionation. A series of functional experiments facilitated determinations of effects of circ_0072995 on EOC progression in vitro, and in vivo. Underlying mechanisms and influence of circ_0072995 on EOC were confirmed by bioinformatic analyses, luciferase reporter assays, qRT-PCR, and Western blotting.

High neuropilin and tolloid-like 1 expression associated with metastasis and poor survival in epithelial ovarian cancer via regulation of actin cytoskeleton.

Abnormal expression of neuropilin and tolloid‐like 1 (NETO1) has been detected in some human carcinomas. However, the expression of NETO1 and the underlying mechanism in epithelial ovarian cancer (EOC) remain unknown. In this study, we found that a higher NETO1 expression in EOC tissue samples compared to normal ovarian tissue samples was significantly correlated with worse overall survival. Additionally, Cox regression analysis suggested that NETO 1 was independently associated with overall survival. NETO1 overexpression enhanced the EOC cells’ migration and invasion capability in vitro via regulation of actin cytoskeleton. Mechanistically, silencing NETO1 reduced the expression of β‐tubulin, F‐actin and KIF2A. In conclusion, our results demonstrated the critical role of NETO1 in EOC invasion, and therapies aimed at inhibiting its expression or activity might significantly control EOC growth, invasion and metastatic dissemination.

The Wnt3a/β-catenin/TCF7L2 signaling axis reduces the sensitivity of HER2-positive epithelial ovarian cancer to trastuzumab

Epithelial ovarian cancer (EOC) is one of the most common and lethal gynecological cancers. Novel therapeutic agents have been developed for EOC, but patient survival remains poor. Trastuzumab has been approved for breast and gastric cancers with high expression of human epidermal growth factor receptor 2 (HER2), but it has not achieved any clinical success in EOC. Dysregulated Wnt/β-catenin signaling is involved in cancer development, but whether it plays a role in EOC resistance to trastuzumab remains largely unknown. Here, we observed that high expression of Wnt3a, β-catenin and TCF7L2, which can form a signaling axis in the Wnt/β-catenin pathway, commonly existed in HER2-positive EOC tissue samples and was correlated with a poor patient prognosis. Cell proliferation and migration assays and nude mouse xenograft model experiments demonstrated that the Wnt3a/β-catenin/TCF7L2 signaling axis promoted tumor cell growth and metastasis and reduced tumor sensitivity to trastuzumab. Analysis of downstream Akt signaling suggested that the function of the Wnt3a/β-catenin/TCF7L2 signaling axis was mediated, at least in part, through increasing Akt phosphorylation. Overall, this study reveals a crucial role for the Wnt3a/β-catenin/TCF7L2 signaling axis in EOC resistance to trastuzumab and the potential application of HER2-targeted drugs combined with inhibitors of this signaling axis for EOC treatment.

Annexin A2 and S100A10 as Candidate Prognostic Markers in Epithelial Ovarian Cancer.

Ovarian cancer (OC) is the 5th most common cancer among European women. Approximately 70-80% of OC is diagnosed at advanced stage resulting in an elevated mortality rate. The aim of this study was to examine whether Annexin A2 and S100A10 expression can be used as prognostic markers for epithelial ovarian cancer (EOC). Expression of Annexin A2 and S100A10 was evaluated in EOC tissue samples (n=303) by immunohistochemistry. The staining of the membrane, cytoplasmic and stroma was assessed according to intensity. The expression of both markers correlated to histological subtype, histological grading, International Federation of Gynecology and Obstetrics (FIGO) stage, and macro-radical surgery. Univariate Cox regression analysis showed that Annexin A2 and S100A10 in stromal tissue correlated with shorter overall survival (OS). Multivariate Cox regression analysis demonstrated no independent prognostic significance of stromal Annexin A2 expression. High expression of Annexin A2 and S100A10 in stromal tissue from EOC patients was associated with reduced OS; however, no independent prognostic value was found for any of the markers.

Upregulation of lncRNA AB073614 functions as a predictor of epithelial ovarian cancer prognosis and promotes tumor growth in vitro and in vivo.

Upregulation of lncRNA AB073614 is found in some cancer types and involved in tumor development and progression including ovarian cancer. However, the clinical value and functional role of lncRNA AB073614 in epithelial ovarian cancer (EOC) still needed to be investigated. We examined lncRNA AB073614 expression using quantitative real time polymerase chain reaction (qRT-PCR) in 75 paired of EOC tissue samples and adjacent normal tissues. Association of lncRNA AB073614 expression with overall survival (OS) was evaluated using Kaplan-Meier analysis. Univariate and multivariate analysis of factors associated with OS were assessed in EOC patients. After lncRNA AB073614 knockdown using siRNAs, the cell viability and cell colony forming assays were performed. Western blot analysis was used to assess relative protein expression. In present study, we demonstrated that lncRNA AB073614 was significantly upregulated in ovarian cancer tissues compared to adjacent normal tissues in patients. Higher lncRNA AB073614 expression significantly associated with tumor size, lymph node invasion, FIGO stage, and shorter OS rate of EOC patients. Furthermore, multivariate Cox regression analysis results showed that higher lncRNA AB0736141 was identified as an independent risk factor of OS in EOC patients. Moreover, we demonstrated that lncRNA AB0736141 knockdown suppressed EOC cell proliferation ability and cell colony formation in vitro. In vivo, we showed that AB0736141 knockdown suppressed tumor growth. We also revealed that lncRNA AB0736141 knockdown inhibited the PTEN/PI3K/AKT signaling pathway in EOC. Thus, these results indicated that LncRNA AB073614 may serve as a prognostic biomarker and potential target of treatment for EOC.

ITLNI identified by comprehensive bioinformatic analysis as a hub candidate biological target in human epithelial ovarian cancer.

BackgroundEpithelial ovarian cancer (EOC) is a female malignant tumor. Bioinformatics has been widely utilized to analyze genes related to cancer progression. Targeted therapy for specific biological factors has become more valuable.Materials and methodsGene expression profiles of GSE18520 and GSE27651 were downloaded from Gene Expression Omnibus. We used the “limma” package to screen differentially expressed genes (DEGs) between EOC and normal ovarian tissue samples and then used Clusterprofiler to do functional and pathway enrichment analyses. We utilized Search Tool for the Retrieval of Interacting Genes Database to assess protein–protein interaction (PPI) information and the plug-in Molecular Complex Detection to screen hub modules of PPI network in Cytoscape, and then performed functional analysis on the genes in the hub module. Next, we utilized the Weighted Gene Expression Network Analysis package to establish a co-expression network. Validation of the key genes in databases and Gene Expression Profiling Interactive Analysis (GEPIA) were completed. Finally, we used quantitative real-time PCR to validate hub gene expression in clinical tissue samples.ResultsWe analyzed the DEGs (96 samples of EOC tissue and 16 samples of normal ovarian tissue) for functional analysis, which showed that upregulated DEGs were strikingly enriched in phosphate ion binding and the downregulated DEGs were significantly enriched in glycosaminoglycan binding. In the PPI network, CDK1 was screened as the most relevant protein. In the co-expression network, one EOC-related module was identified. For survival analysis, database and clinical sample validation of genes in the turquoise module, we found that ITLN1 was positively correlated with EOC prognosis and had lower level in EOC than in normal tissues, which was consistent with the results predicted in GEPIA.ConclusionIn this study, we exhibited the key genes and pathways involved in EOC and speculated that ITLN1 was a tumor suppressor which could be used as a potential biomarker for treating EOC, Gene Expression Omnibus, prognosis.

AEG-1 Contributes to Metastasis in Hypoxia-Related Ovarian Cancer by Modulating the HIF-1alpha/NF-kappaB/VEGF Pathway.

Objective Ovarian carcinoma represents one of the deadliest malignancies among female cancer patients. Astrocyte-elevated gene-1 (AEG-1) participates in the ontogenesis of multiple human malignant diseases. Here we evaluated AEG-1, hypoxia-inducible factor- (HIF-) 1α, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and vascular endothelial growth factor (VEGF) amounts in hypoxia induced ovarian carcinoma cells. This study aimed to explore the mechanism by which AEG-1 regulates metastasis in hypoxia induced ovarian carcinoma. Patients and Methods AEG-1, HIF-1α, and VEGF protein amounts were evaluated by immunohistochemistry in 40 and 170 normal ovary and ovarian cancer tissue specimens, respectively. In addition, AEG-1, HIF-1α, NF-κB, and VEGF mRNA and protein levels were determined by reverse quantified RT-PCR and WB, respectively, at different time periods (0–24 h) in epithelial ovarian cancer (EOC) SKOV3 cells treated in a hypoxia incubator. Furthermore, NF-κB and VEGF gene and protein expression levels in AEG-1 knockdown EOC cells were quantitated by RT-PCR and WB, respectively. Results AEG-1, HIF-1α, and VEGF amounts were significantly elevated in EOC tissue samples compared with normal ovary specimens (p < 0.001). Positive expression of HIF-1α and AEG-1 was associated with higher metastatic rate (p < 0.01), lower FIGO stage (p < 0.001), and degree of differentiation (p < 0.001). Meanwhile, EOC SKOV3 cells grew upon exposure to hypoxia for 8 h (p < 0.001); at this time point, AEG-1, HIF-1α, NF-κB, and VEGF amounts peaked (p < 0.001), at both the gene and the protein levels. After AEG-1 knockdown, HIF-1α, NF-κB, and VEGF amounts were significantly decreased in EOC SKOV3 cells, also under hypoxic conditions (p < 0.01). Conclusions As an independent prognostic factor, AEG-1 was found to be significantly associated with hypoxia in ovarian cancer by regulating the HIF-1alpha/NF-kappaB/VEGF pathway. Therefore, AEG-1 may be useful in determining disease stage and prognosis in ovarian cancer.

Overexpression of trophoblast cell surface antigen 2 as an independent marker for a poor prognosis and as a potential therapeutic target in epithelial ovarian carcinoma.

Most patients with epithelial ovarian cancer (EOC) are diagnosed at an advanced stage, and therapeutic options for these patients are limited. The identification of suitable biomarkers could be helpful for patients with EOC, who might benefit from targeted therapies even in advanced stages of the disease. Trophoblast cell surface antigen 2 (TROP2) is highly expressed in various human malignant tumours; however, this has not been demonstrated clearly in EOC. In this study, we further evaluated whether TROP2 is a promising marker for EOC, and thus also a potential target for EOC immunotherapy. Quantitative real-time polymerase chain reaction (qPCR) and fluorescence-activated cell sorting (FACS) analysis were employed to determine TROP2 mRNA and protein expression in both human EOC and normal ovarian cell lines. Additionally, TROP2 protein expression was measured by immunohistochemistry in 128 EOC tissue samples, 21 normal ovarian tissues and 18 normal fallopiantubes. The correlations between TROP2 protein expression and patients' clinicopathological features were investigated, and survival outcomes were analysed. TROP2 mRNA and protein levels were upregulated significantly in EOC cell lines compared with normal cell lines. The protein of TROP2 was expressed in the majority of EOC tissue samples (90.6%) and overexpressed in 75 (58.6%) of the 128 tumour samples. TROP2 overexpression was correlated with relevant clinicopathological characteristics and was associated with significantly shortened overall survival and disease-free survival. Furthermore, TROP2 was an independent prognostic marker for EOCs as analysed by Cox regression. TROP2 was a potential biomarker for targeted therapy in patients with TROP2-overexpressing EOC.

MicroRNA-520g promotes epithelial ovarian cancer progression and chemoresistance via DAPK2 repression.

The lack of efficient tumor progression and chemoresistance indicators leads to high mortality in epithelial ovarian cancer (EOC) patients. Dysregulated miR-520g expression is involved in these processes in hepatic and colorectal cancers. In this study, we found that miR-520g expression gradually increased across normal, benign, borderline and EOC tissues. High miR-520g expression promoted tumor progression and chemoresistance to platinum-based chemotherapy, and reduced survival in EOC patients. miR-520g upregulation increased EOC cell proliferation, induced cell cycle transition and promoted cell invasion, while miR-520g downregulation inhibited tumor-related functions. In vivo, overexpression or downregulation of miR-520g respectively generated larger or smaller subcutaneous xenografts in nude mice. Death-associated protein kinase 2 (DAPK2) was a direct target of miR-520g. In 116 EOC tissue samples, miR-520g expression was significantly lower following DAPK2 overexpression. DAPK2 overexpression or miR-520g knockdown reduced EOC cell proliferation, invasion, wound healing and chemoresistance. This study suggests that miR-520g contributes to tumor progression and drug resistance by post-transcriptionally downregulating DAPK2, and that miR-520g may be a valuable therapeutic target in patients with EOC.

Promoter Hypermethylation of FANCF and Susceptibility and Prognosis of Epithelial Ovarian Cancer.

To assess the 5' CpG island methylation of Fanconi anemia, complementation group F (FANCF) gene in epithelial ovarian cancer (EOC) tissues and normal ovarian tissues and to investigate the relationship between FANCF methylation and clinicopathologic features and prognosis of EOC. The experiment was performed with 112 EOC tissue samples (case group) and 60 normal ovarian tissues (control group). With methylation-specific polymerase chain reaction (MSP), FANCF methylation status of cases and controls was assessed. And the association between FANCF methylation and the clinicopathological features of EOC was investigated with univariate survival analysis and Cox regression model analysis. The methylation-positive rate of the case group was significantly higher than that of the control group (P = 0.015). The FANCF promoter methylation rates showed significant differences in the comparisons stratified by age, International Federation of Gynecology and Obstetrics (FIGO) staging, histopathological classification, and lymph node metastasis (all P < .05). Univariate survival analysis showed there were significant differences in mean survival time between the groups based on FIGO staging, histopathological classification, lymph node metastasis, and FANCF methylation (all P < .05). Cox regression model analysis suggested that FIGO staging and FANCF methylation were independent risk factors for EOC prognosis. CpG island methylation of FANCF gene promoter region is strongly associated with the susceptibility and clinicopathologic features of EOC. The FIGO staging and FANCF methylation are independent risk factors for EOC prognosis.

Clinical implication of the serum galectin-1 expression in epithelial ovarian cancer patients.

Backgroundgalectin-1 has been implicated in tumor invasion and metastasis and is frequently over-expressed in epithelial ovarian cancer (EOC), but its potential as a biomarker remains unclear. In this novel study, we have explored the possible use of galectin-1 as a biomarker for EOC.Methodsgalectin-1 in sera was evaluated by ELISA in a pilot panel of EOC patients, healthy volunteers, patients with benign gynecologic tumors or other gynecologic malignancies. We examined galectin-1 expression in EOC tumor samples by Western Blot, qRT-PCR and immunohistochemistry. In vitro experiments were conducted to elucidate the biologic role of galectin-1 in EOC progression using over-expression of galectin-1 in OVCAR-3 cells. We also looked for the association of galectin-1 expression with clinic pathological variables and survival outcomes in EOC.ResultsA significant difference was detected in serum galectin-1 between EOC patients with non-metastatic and those with metastatic disease, but not between EOC patients and healthy volunteers. It increased in recurrent cases and decreased after debulking surgery. Both of galectin-1 mRNA and protein levels were increased in 90 % of the examined EOC tissue samples, compared with a wedge resection of a normal ovary. High galectin-1 in peritumor stroma was primarily detected in advanced stages of EOC. Over expression of galectin-1 significantly increased the ability of OVCAR-3 cells’ migration and invasion.ConclusionsOur results suggest that galectin-1 might play a role in tumor progression and be associated with poor outcome in EOC. It could be a novel prognostic and progression biomarker in EOC patients.

Abstract 4423: Chemoresistance regulation by COL11A1 in ovarian cancer

Abstract Epithelial ovarian carcinoma remains the most lethal gynecologic malignancy. While initial response to first-line therapy (consisting of surgical cytoreduction and combination platinum/taxane therapy) is usually effective, the majority of patients will ultimately recur with chemotherapy-resistant cancer and succumb to disease. Biomarkers that predict patient's responsiveness to chemotherapy might assist the development of better therapeutic strategies for aggressive cancers, such as ovarian cancer. Here, we investigated the molecular mechanisms of collagen type XI alpha 1 (COL11A1) in chemo-resistance of ovarian cancer. We analyzed the expression profiles of 60 epithelial ovarian cancer tissue samples obtained at first cytoreductive surgery to identify genes linked with chemotherapy response. Among the 47 selected genes, collagen type XI alpha 1 (COL11A1) was the most highly elevated in chemo-resistance tumors. Small interfering RNA-mediated specific reduction of COL11A1 protein levels increased the sensitivity of ovarian chemoresistant cells to cisplatin and paclitaxel. A combination of experimental approaches, including real-time RT-PCR, western blotting, luciferase assay and chromatin immunoprecipitation (ChIP) assays, showed that the c/EBPβ binding site on the COL11A1 promoter was identified as the major determinant of cisplatin and paclitaxel-dependent COL11A1 activation. Furthermore, COL11A1 silencing suppressed phosphorylated Akt expression, which was in concert with increased PDK1 degradation in chemoresistant cells. Analysis of 104 EOC patients showed that high COL11A1 mRNA levels are significantly associated with poor chemoresponse and poor clinical outcome. In conclusion, activation of COL11A1 by anticancer drugs confers chemoresistance on ovarian cancer cells through the activation of Akt/c/EBPβ pathway and PDK1 stabilization. Note: This abstract was not presented at the meeting. Citation Format: Yi-Hui Wu. Chemoresistance regulation by COL11A1 in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4423. doi:10.1158/1538-7445.AM2015-4423

CAP1 is overexpressed in human epithelial ovarian cancer and promotes cell proliferation.

Adenylate cyclase-associated protein 1 (CAP1) regulates both actin filaments and the Ras/cAMP pathway in yeast, and has been found play a role in cell motility and in the development of certain types of cancer. In the present study, we investigated CAP1 gene expression in human epithelial ovarian cancer (EOC). Western blot analysis and immunohistochemistry were performed using EOC tissue samples and the results revealed that CAP1 expression increased with the increasing grade of EOC. In the normal ovarian tissue samples however, CAP1 expression was barely detected. Using Pearson’s χ2 test, it was demonstrated that CAP1 expression was associated with the histological grade and Ki-67 expression. Kaplan-Meier analysis revealed that a higher CAP1 expression in patients with EOC was associated with a poorer prognosis. In in vitro experiments using HO-8910 EOC cells, the expression of CAP1 was knocked down using siRNA. The proliferation of the HO-8910 cells was then determined by cell cycle analysis and cell proliferation assay using the cell counting kit-8 and flow cytometry. The results revealed that the loss of CAP1 expression inhibited cell cycle progression. These findings suggest that a high expression of CAP1 is involved in the pathogenesis of EOC, and that the downregulation of CAP1 in tumor cells may be a therapeutic target for the treatment of patients with EOC.

Abstract 958: Activation of COL11A1 by anticancer drugs through IGF-1R/PI3K pathway confers chemoresistance in ovarian cancer via activating NF-κB-mediated Twist1 expression

Abstract Epithelial ovarian carcinoma remains the most lethal gynecologic malignancy. While initial response to first-line therapy (consisting of surgical cytoreduction and combination platinum/taxane therapy) is usually effective, the majority of patients will ultimately recur with chemotherapy-resistant cancer and succumb to disease. Biomarkers that predict patient's responsiveness to chemotherapy might assist the development of better therapeutic strategies for aggressive cancers, such as ovarian cancer. Here, we investigated the molecular mechanisms of collagen type XI alpha 1 (COL11A1) in chemo-resistance of ovarian cancer. We analyzed the expression profiles of 60 epithelial ovarian cancer tissue samples obtained at first cytoreductive surgery to identify genes linked with chemotherapy response. Among the 47 selected genes, collagen type XI alpha 1 (COL11A1) was the most highly elevated in chemo-resistance tumors. Small interference RNA-mediated specific reduction in COL11A1 protein levels increased the sensitivity to cisplatin and taxol in ovarian cancer cells. A combination of experimental approaches, including real-time RT-PCR, western blotting, luciferase assay and chromatin immunoprecipitation (ChIP) assays, showed that COL11A1 knockdown attenuated Twist1 expression and suppressed binding of NF-κB to its putative Twist1 promoter binding site, suggesting that the NF-κB/Twist1 axis is upregulated by COL11A1. Cisplatin and taxol treatment triggers the activation of IGF-1R/PI3K signaling cascades, leading to activation of COL11A1 and Twist1. Pharmacological inhibition of PI3K abrogated the cisplatin and taxol-triggered, COL11A1-dependent chemo-resistance. Furthermore, the c/EBPβ binding site on the COL11A1 promoter was identified as the major determinant of cisplatin and taxol-dependent COL11A1 activation. Analysis of 88 ovarian cancer patients indicated that high COL11A1 mRNA levels are significantly associated with high Twist1 mRNA levels and poor clinical outcome. We conclude that activation of COL11A1 by anti-cancer drugs through IGF-1R/PI3K pathway confers chemo-resistance in ovarian cancer cells via activating NF-κB-mediated Twist1 expression. Citation Format: Yi-Hui Wu, Yu-Fang Huang, Cheng-Yang Chou. Activation of COL11A1 by anticancer drugs through IGF-1R/PI3K pathway confers chemoresistance in ovarian cancer via activating NF-κB-mediated Twist1 expression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 958. doi:10.1158/1538-7445.AM2014-958

Deregulation of microcephalin and ASPM expression are correlated with epithelial ovarian cancer progression.

Mutations in the MCPH1 (Microcephalin) and ASPM (abnormal spindle-like microcephaly associated) genes cause primary microcephaly. Both are centrosomal associated proteins involved in mitosis. Microcephalin plays an important role in DNA damage response and ASPM is required for correct division of proliferative neuro-epithelial cells of the developing brain. Reduced MCPH1 mRNA expression and ASPM mRNA over-expression have been implicated in the development of human carcinomas. Epithelial ovarian cancer (EOC) is characterised by highly aneuploid tumours. Previously we have reported low Microcephalin and high ASPM protein levels and associations with clinico-pathological parameters in malignant cells from ascitic fluids. To confirm these previous findings on a larger scale Microcephalin and ASPM expression levels and localisations were evaluated by immunohistochemistry in two cohorts; a training set of 25 samples and a validation set of 322 EOC tissue samples. Results were correlated to the associated histopathological data. In normal ovarian tissues the Microcephalin nuclear staining pattern was consistently strong. In the cancer tissues, we identified low nuclear Microcephalin expression in high grade and advanced stage tumours (p<0.0001 and p = 0.0438 respectively). ASPM had moderate to high nuclear and low to moderate cytoplasmic expression in normal tissue. Cytoplasmic ASPM expression decreased with tumour grade and stage in the serous subtype of EOC (p = 0.023 and p = 0.011 respectively). Cytoplasmic ASPM increased with tumour stage in the endometrioid subtype (p = 0.023). Increasing tumour invasiveness (T3) and lymph node involvement (N1) also correlated with a decrease in cytoplasmic ASPM in EOC (p = 0.02 and p = 0.04 respectively). We have validated previous findings of deregulated expression of Microcephalin and ASPM in EOC by confirming associations for low nuclear Microcephalin levels and high cytoplasmic ASPM levels in a larger scale tumour tissue study. Microcephalin and ASPM may prove useful biomarkers in EOC.