Genomic analysis of epithelial ovarian cancer

Ovarian cancer is a major health problem for women in the United States. Despite evidence of considerable heterogeneity, most cases of ovarian cancer are treated in a similar fashion. The molecular basis for the clinicopathologic characteristics of these tumors remains poorly defined. Whole genome expression profiling is a genomic tool, which can identify dysregulated genes and uncover unique sub-classes of tumors. The application of this technology to ovarian cancer has provided a solid molecular basis for differences in histology and grade of ovarian tumors. Differentially expressed genes identified pathways implicated in cell proliferation, invasion, motility, chromosomal instability, and gene silencing and provided new insights into the origin and potential treatment of these cancers. The added knowledge provided by global gene expression profiling should allow for a more rational treatment of ovarian cancers. These techniques are leading to a paradigm shift from empirical treatment to an individually tailored approach. This review summarizes the new genomic data on epithelial ovarian cancers of different histology and grade and the impact it will have on our understanding and treatment of this disease.     

Tissue quantification of CA 125 in epithelial ovarian cancer.

The objectives of this study were the determination of CA 125 in the cytosol of healthy and carcinomatous ovarian tissue by immunoanalysis, analysis of its correlation with the biological characteristics of ovarian carcinoma, determination of serum CA 125 levels, and study of the prognostic value of the marker in cytosol. The levels of the marker depend not only on the tumor's production rate, so its determination in tissue can indicate more accurately if the tumor is a producer of the marker and establish its value for the prognosis of the disease. Determination of CA 125 in tissue was performed by immunoanalysis in 50 ovarian epithelial cancer samples, 13 benign pathology samples and 32 healthy ovary samples. The presurgical serum level of the marker was also obtained. The correlation between the CA 125 level in the cytosol and the different biological characteristics of the ovarian carcinoma, the serum levels of the marker and survival were analyzed. The CA 125 level proved to be higher in malignant tissue (p < 0.0001). There was a significant association between the tissue marker and histological type (high CA 125 was associated with serous and endometrioid tumors) and between the marker and survival. No relation with stage was found. There was a correlation between the CA 125 level in the cytosol and serum both variables being dependent, with a correlation coefficient of 0.44. This good correlation speaks in favor of the usefulness of CA 125 determination in serum in the follow-up of ovarian cancer. Tumors having high tissue expression of CA 125 were found to have a double relative risk of death, independently of tumor stage.     

NuMA overexpression in epithelial ovarian cancer

Highly aneuploid tumours are common in epithelial ovarian cancers (EOC). We investigated whether NuMA expression was associated with this phenomenon.NuMA protein levels in normal and tumour tissues, ovarian cell lines and primary cultures of malignant cells derived from ovarian ascitic fluids were analysed by Affymetrix microarray analysis, immunoblotting, immunohistochemistry (IHC) and immunofluorescence (IF), with results correlated to associated clinical data. Aneuploidy status in primary cultures was determined by FACS analysis.Affymetrix microarray data indicated that NuMA was overexpressed in tumour tissue, primary cultures and cell lines compared to normal ovarian tissue. IHC revealed low to weak NuMA expression in normal tissues. Expression was upregulated in tumours, with a significant association with disease stage in mucinous EOC subtypes (p = 0.009), lymph node involvement (p = 0.03) and patient age (p = 0.04). Additional discontinuous data analysis revealed that high NuMA levels in tumours decreased with grade (p = 0.02) but increased with disease stage (p = 0.04) in serous EOC. NuMA expression decreased in late disease stage 4 endometrioid EOCs. High NuMA levels decreased with increased tumour invasion in all subtypes (p = 0.03). IF of primary cultures revealed that high NuMA levels at mitotic spindle poles were significantly associated with a decreased proportion of cells in cytokinesis (p = 0.05), increased binucleation (p = 0.021) and multinucleation (p = 0.007), and aneuploidy (p = 0.008).NuMA is highly expressed in EOC tumours and high NuMA levels correlate with increases in mitotic defects and aneuploidy in primary cultures.     

Comparative proteome analysis of human epithelial ovarian cancer

Background  

Epithelial ovarian cancer is a devastating disease associated with low survival prognosis mainly because of the lack of early detection markers and the asymptomatic nature of the cancer until late stage. Using two complementary proteomics approaches, a differential protein expression profile was carried out between low and highly transformed epithelial ovarian cancer cell lines which realistically mimic the phenotypic changes observed during evolution of a tumour metastasis. This investigation was aimed at a better understanding of the molecular mechanisms underlying differentiation, proliferation and neoplastic progression of ovarian cancer.     

Molecular pathogenesis and therapeutic targets in epithelial ovarian cancer

Ovarian cancer, the most aggressive gynecologic cancer, is the foremost cause of death from gynecologic malignancies in the developed world. Two primary reasons explain its aggressive behavior: most patients present with advanced disease at diagnosis, and die of recurrences from disease that has become resistant to conventional chemotherapies. In this paper on epithelial ovarian cancer (EOC), we will review molecular alterations associated with the few precursor lesions identified to date, followed by the more commonly recognized processes of de novo carcinogenesis, metastasis, and the development of chemoresistance. We will propose a unifying model of ovarian epithelial tumorigenesis that takes into account various hypotheses. We will also review novel approaches to overcome the major problem of chemoresistance in ovarian cancer. Finally, we will discuss advances and new challenges in the development of mouse model systems to investigate EOC precursor lesions, progression, metastasis, and chemoresistance.     

Lipid metabolism and Calcium signaling in epithelial ovarian cancer

Epithelial Ovarian cancer (EOC) is the deadliest gynecologic malignancy and represents the fifth leading cause of all cancer-related deaths in women. The majority of patients are diagnosed at an advanced stage of the disease that has spread beyond the ovaries to the peritoneum or to distant organs (stage FIGO III-IV) with a 5-year overall survival of about 29%. Consequently, it is necessary to understand the pathogenesis of this disease. Among the factors that contribute to cancer development, lipids and ion channels have been described to be associated to cancerous diseases particularly in breast, colorectal and prostate cancers. Here, we reviewed the literature data to determine how lipids or lipid metabolites may influence EOC risk or progression. We also highlighted the role and the expression of the calcium (Ca²⁺) and calcium-activated potassium (KCa) channels in EOC and how lipids might regulate them. Although lipids and some subclasses of nutritional lipids may be associated to EOC risk, lipid metabolism of LPA (lysophosphatidic acid) and AA (arachidonic acid) emerges as an important signaling network in EOC. Clinical data showed that they are found at high concentrations in EOC patients and in vitro and in vivo studies referred to them as triggers of the Ca²⁺entry in the cancer cells inducing their proliferation, migration or drug resistance. The cross-talk between lipid mediators and Ca²⁺ and/or KCa channels needs to be elucidated in EOC in order to facilitate the understanding of its outcomes and potentially suggest novel therapeutic strategies including treatment and prevention.     

The genetic epidemiology of early-onset epithelial ovarian cancer: a population-based study

We conducted a population-based study to determine the contribution of germline mutations in known candidate genes to ovarian cancer diagnosed at age <30 years. Women with epithelial ovarian cancer were identified through cancer registries. DNA samples were analyzed for mutations in BRCA1, the "ovarian cancer-cluster region" (nucleotides 3139-7069) of BRCA2, and the mismatch-repair genes hMSH2 and hMLH1. Probable germline mutations in hMLH1 were identified in 2 (2%; 95% confidence interval 1%-8%) of 101 women with invasive ovarian cancer diagnosed at age <30 years. No germline mutations were identified in any of the other genes analyzed. There were no striking pedigrees suggestive of families with either breast/ovarian cancer or hereditary nonpolyposis colorectal cancer (HNPCC). There was a significantly increased incidence of all cancers in first-degree relatives of women with invasive disease (relative risk [RR] = 1.6, P=.01) but not in second-degree relatives or in relatives of women with borderline cases. First-degree relatives of women with invasive disease had increased risks of ovarian cancer (RR = 4.8, P=.03), myeloma (RR = 10, P=.01), and non-Hodgkin lymphoma (RR = 7, P=.004). Germline mutations in BRCA1, BRCA2, msh2, and mlh1 contribute to only a minority of cases of early-onset epithelial ovarian cancer. Our data suggest that early-onset ovarian cancer is not associated with a greatly increased risk of cancer in close relatives.     

Primary culture of ovarian surface epithelial cells and ascites-derived ovarian cancer cells from patients

Our laboratory has refined the technique for isolating primary cultures of normal human ovarian surface epithelial (OSE) cells by combining two different protocols involving the enzymatic and mechanical removal of OSE cells from ovarian biopsies. A simple protocol of obtaining primary epithelial ovarian cancer (EOC) cells from the ascites fluid removed from patients with high-grade ovarian cancer is also described. These methods allow for the direct application of many molecular and cellular analyses of normal versus cancer cells isolated freshly from patients, with the added potential for retrospective analyses of archived cells and tissues. Thus, we have included optional steps for the immediate preparation of ascites-derived EOC cells to be used for subsequent cytological analyses. Initial isolation of OSE or EOC cells can be completed in 1 h, and primary cells are further expanded in culture for several weeks.     

Suppressive activities of mangiferin on human epithelial ovarian cancer

BackgroundEpithelial carcinoma is a subtype of ovarian cancers, with the highest lethality among all ovarian cancer subtypes. Hitherto surgical excision combined with chemotherapy has been the most extensively employed method in clinical treatment. However, the disease relapses very frequently, calling for more effective therapies. Mangiferin, a natural xanthone glucoside, has displayed promising anti-cancer activities by in vitro studies, but its therapeutic value in epithelial ovarian cancer treatment, either by in vivo or in vitro studies, remained to be known.PurposeThis study aimed to determine the suppressive activities of mangiferin on human epithelial ovarian cancer and elucidate the underlying molecular mechanisms.Study design and methodsWe employed the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the crystal violet assay to determine the half maximal inhibitory concentration (IC₅₀) values of mangiferin with paclitaxel as a positive control and the inhibitory effects of mangiferin on the proliferation of two human epithelial ovarian cancer cell lines. Wound healing and Transwell assays were used to determine anti-metastastic activities of mangiferin. ES-2 xenograft nude mouse model was used for the in vivo experiments. Western blotting, enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC) assays were carried out for evaluating the expression level of matrix metalloproteinase 2 (MMP2) and matrix metalloproteinase 9 (MMP9).ResultsIn the present study, we demonstrated by both in vitro and in vivo assays that mangiferin suppressed the progress of epithelial ovarian cancer in a dose-dependent manner. In the animals treated with mangiferin, the tumor volume and weight were reduced significantly. Analyses of involved molecular events demonstrated that mangiferin down-regulated the expression of metastasis-associated proteins MMP2 and MMP9.ConclusionMangiferin strongly inhibited the progression of human epithelial ovarian cancer by down-regulating MMP2 and MMP9.     

Pre-treatment neutrophil to lymphocyte ratio is elevated in epithelial ovarian cancer and predicts survival after treatment

Purpose  Inflammatory cells can both suppress and stimulate tumor growth, and the influence of inflammatory cells on clinical outcome has been the focus of many studies. The purpose of this study was to evaluate the effectiveness of the neutrophil to lymphocyte ratio (NLR), a measure of the systemic inflammatory response, as an additional discriminative biomarker in epithelial ovarian cancer and to determine whether it predicts survival and recurrence. Methods  We studied 192 patients with epithelial ovarian cancer, 173 with benign ovarian tumors, 229 with benign gynecologic disease, and 405 healthy controls. Serum CA125 levels and leukocyte counts according to subtypes were recorded prior to treatment in all study subjects. In epithelial ovarian cancer, the diagnostic usefulness of NLR, in combination with CA125, was evaluated. The correlation between NLR and overall and disease-free survival was analyzed using both univariate and multivariate analyses adjusting for the known prognostic factors (age, stage, cell type, and grade). Results  Preoperative NLR in ovarian cancer subjects (mean 6.02) was significantly higher than that in benign ovarian tumor subjects (mean 2.57), benign gynecologic disease subjects (mean 2.55), and healthy controls (mean 1.98) (P < 0.001). The sensitivity and specificity of NLR in detecting ovarian cancer was 66.1% (95% CI, 59.52–72.68%) and 82.7% (95% CI, 79.02–86.38%), respectively (cutoff value: 2.60). In early stage ovarian cancer, CA125 was not elevated in 19 out of 49 patients. Seven (36.8%) of these 19 patients were NLR positive. On Cox multivariate analysis, NLR positive, stage III/IV, and older age were independent poor prognostic factors, and being NLR positive was the most powerful predictive variable (Hazard Ratio = 8.42 [95% CI: 1.09–64.84], P = 0.041). Conclusions  Our findings provide evidence for the association between NLR and epithelial ovarian cancer. Preoperative NLR, in combination with CA125, may represent a simple and cost-effective method of identifying ovarian cancers, and an elevated NLR may predict an adverse outcome in ovarian cancer.     

BRCA1 regulates follistatin function in ovarian cancer and human ovarian surface epithelial cells

Follistatin (FST), a folliculogenesis regulating protein, is found in relatively high concentrations in female ovarian tissues. FST acts as an antagonist to Activin, which is often elevated in human ovarian carcinoma, and thus may serve as a potential target for therapeutic intervention against ovarian cancer. The breast cancer susceptibility gene 1 (BRCA1) is a known tumor suppressor gene in human breast cancer; however its role in ovarian cancer is not well understood. We performed microarray analysis on human ovarian carcinoma cell line SKOV3 that stably overexpress wild-type BRCA1 and compared with the corresponding empty vector-transfected clones. We found that stable expression of BRCA1 not only stimulates FST secretion but also simultaneously inhibits Activin expression. To determine the physiological importance of this phenomenon, we further investigated the effect of cellular BRCA1 on the FST secretion in immortalized ovarian surface epithelial (IOSE) cells derived from either normal human ovaries or ovaries of an ovarian cancer patient carrying a mutation in BRCA1 gene. Knock-down of BRCA1 in normal IOSE cells demonstrates down-regulation of FST secretion along with the simultaneous up-regulation of Activin expression. Furthermore, knock-down of FST in IOSE cell lines as well as SKOV3 cell line showed significantly reduced cell proliferation and decreased cell migration when compared with the respective controls. Thus, these findings suggest a novel function for BRCA1 as a regulator of FST expression and function in human ovarian cells.