Role In Ovarian Carcinogenesis Research Articles

IMMUNOHISTOCHEMICAL EXPRESSION OF CD133 IN OVARIAN SURFACE EPITHELIAL TUMORS

Background: The identification of antigenic profile of the cancer stem cells might have relevant clinical implication as they're able to proliferate and self-renew, in turn sustaining tumor growth. CD133 is one of robust surface marker for cancer stem cells in various neoplastic human tissues, including ovaries. It is associated with the clinical outcome of patients. Objective: To assess the immunohistochemichal expression of cancer stem cell marker (CD133) in ovarian surface epithelial tumors. Methods: This study included a total of 100 ovarian tissue paraffin blocks, 70 tissue paraffin blocks included ovarian tumors obtained from patient who underwent total abdominal hysterectomy and bilateral salpingo-oophorectomy, while 30 tissue paraffin blocks assigned as the control group included normal ovarian tissue and fallopian tubes tissue. From each paraffin block, 2 sections were taken, one was stained with hematoxylin and eosin stain and the other section was stained immunohistochemically for CD133. Results: CD133 showed a high significant difference in its immunohistochemical expression between the control group and the case groups with the highest expression seen in malignant ovarian surface epithelial tumors (P < 0.001). CD133 expression was highly significantly associated with histopathological type (P < 0.001). CD133 also showed a significant relation to age (r = 0.254, P= 0.034), tumor grade (r = 0.794, P < 0.001), tumor stage (r = 0.543, P= 0.013) and presence of ascites (P < 0.001). Conclusion: CD133 expression revealed a highly significant differences in the ovarian surface epithelial tumors tissues compared to control group, which reflect its important role in ovarian carcinogenesis. Keywords: Ovarian surface epithelial tumors, ovarian carcinoma, cancer stem cells, cancer stem cell marker, CD133, immunohistochemistry Citation: Al-Shami SA, Qasim BJ, Hassan AM. Immunohistochemical expression of CD133 in ovarian surface epithelial tumors. Iraqi JMS. 2018; 16(3): 308-318. doi: 10.22578/IJMS.16.3.10

Abstract 4424: Micro-RNA mediated deregulation of hHB-EGF in ovarian cancer

Abstract Background: Aberrant expression of microRNAs (miRNAs) has been implicated in human carcinogenesis where they function as oncogenes or tumor suppressor genes. Overexpression of HB-EGF, member of the epidermal growth factor (EGF) family, plays pivotal role in ovarian carcinogenesis. However the molecular mechanisms underlying miRNA(s) mediated aberrant function of HB-EGF in ovarian carcinoma remain unresolved. Here, we investigated the miRNA(s) mediated deregulation of HB-EGF in human ovarian carcinogenesis. Methods: Web-based bioinformatics algorithms were used to identify candidate miRNAs with putative binding sites on target hHB-EGF mRNA. Expression of miRNAs and mRNA were checked by real-time PCR whereas protein expression was investigated by western blotting assay. Luciferase reporter assay was carried out to examine miR-27a target on 3´UTR of hHB-EGF mRNA. Changes in the oncogenic effects of HB-EGF were studied after gain/loss -of-function of the miR-27a using soft agar colony formation assay and 3D collagen culture. In vivo tumor growth potential of miR-27a was measured in BALB/c nude mice. Results: By using web-based bioinformatics algorithms 11 miRNAs ((miR-27a/b, -29a/b/c, -96, -128, -132, -182,-194, -212) having putative binding sites on the 3′UTR of hHB-EGF mRNA have been selected for further analysis. Among the putative candidates miR-27a showed maximum reciprocal expression with hHB-EGF and thus we selected miR-27a as promising candidate for hHB-EGF regulation. Overexpression of miR-27a significantly inhibited the hHB-EGF expression in SKOV3 cells by almost 50% and miR-27a inhibitors (anti-miR-27a) upregulated the expression of endogenous hHB-EGF mRNA whereas miR-27a overexpression decreased hHB-EGF protein level in MCAS cells suggesting that miR-27a modulate endogenous hHB-EGF expression both at mRNA and protein level. Luciferase reporter assay revealed that miR-27a directly targets 3´UTR of hHB-EGF mRNA at the predicted site. Ectopic expression of miR-27a suppresses in-vitro growth of SKOV3 ovarian cancer cells. In addition, in-vivo tumorigenecity of SKOV3 cells was also inhibited by miR-27a treatment. Conclusion: The current study has identified novel miR-27a that negatively regulates hHB-EGF expression by directly targeting 3`UTR of its mRNA. This miR-27a might act as oncosupressor in human epithelial ovarian cancer. Citation Format: Md. Mustafizur Rahman, Razia Sultana, Ryo Iwamoto, Eisuke Mekada. Micro-RNA mediated deregulation of hHB-EGF in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4424.

Dietary inflammatory index and ovarian cancer risk in a New Jersey case-control study.

Diet may influence the development of ovarian cancer. Although it has been shown that inflammation plays an important etiologic role in ovarian carcinogenesis, little is known about the influence of the inflammatory potential of food consumption. The aim of this study was to evaluate the effect of a proinflammatory diet, as indicated by a high dietary inflammatory index (DII®) score, on ovarian cancer risk, in a New Jersey population. Data from a case-control study conducted in New Jersey were used to estimate the relation between DII score and the risk for ovarian cancer. The study consisted of 205 cases with incident, histologically confirmed ovarian cancer, and 390 controls identified by random-digit dialing, based on Centers for Medicare & Medicaid Service lists, and area sampling. Computation of the DII was based on the intake of selected dietary factors assessed by a validated food frequency questionnaire (FFQ). Logistic regression models were fit to estimate odds ratios (ORs) and 95% confidence intervals (CIs) adjusted for potential covariates. Although there was no significant association observed in pre- and perimenopausal women, a significant association was observed between the most proinflammatory DII scores and ovarian cancer among postmenopausal women (ORQuartile4 vs1, 1.89; 95% CI, 1.02-3.52; Ptrend=0.03). Findings from the present study suggested that a proinflammatory diet may increase risk for ovarian cancer among postmenopausal women, and warrants further study to confirm this association.

Androgen receptor gene CAG repeat polymorphism and ovarian cancer risk: A meta-analysis.

Ovarian cancer is one of the common gynecological malignancies worldwide. It is usually diagnosed at a later stage, thus missing the best opportunity for treatment. Despite the advancement of ovarian cancer treatment, the prognosis is still poor. Androgen receptor (AR) may play a role in ovarian carcinogenesis. Previous studies regarding the association between AR CAG repeat length and ovarian cancer risk reported inconsistent results. Therefore, we conducted a meta-analysis to evaluate the association between AR CAG repeat length and ovarian cancer risk following the MOOSE guidelines. PubMed, Web of Science, EBSCO and other databases were searched up to September 15th 2016. Case control studies with clear definition of CAG repeat length and detailed genotype information were included. Two authors independently reviewed and extracted data. Pooled analysis and subgroup analysis stratified by ethnicity were performed for different genetic models. Begg's funnel plot and Egger's test were performed for publication bias estimation. Overall, there was no association between the AR CAG repeat polymorphism and ovarian cancer risk. However, short CAG repeat polymorphism was associated with increased ovarian cancer risk in African Americans and Chinese under the dominant model, whereas a reverse association was observed in Caucasians and Italians under the other three models. Our study results should be interpreted with caution. Further well-designed epidemiological and functional studies are needed to elucidate the role of AR in ovarian carcinogenesis.

The pro-inflammatory effect of obesity on high grade serous ovarian cancer

IntroductionObesity is a known generator of chronic inflammation but has an uncertain role in ovarian carcinogenesis and survival. Pro-inflammatory cytokines have previously been associated with poor outcomes. Given the established links, we sought to determine whether obesity and pro-inflammatory cytokines affect platinum sensitivity. MethodsA retrospective review was performed of patients undergoing primary debulking surgery (PDS) for high grade serous ovarian cancer (HGSC) who had available pre-operative serum. Oncologic and treatment characteristics were recorded and analyzed using SAS version 9.3. Bioplex reagent kit was used to measure serum cytokine concentrations. Results86 patients met study criteria. Most were Caucasian (88%) and non-diabetic (92%). All patients had advanced stage (III/IV) disease and received chemotherapy after PDS. In univariate analysis, lower VEGF (p=0.013) was associated with longer overall survival (OS). Low IL-8 level (p=0.053) was marginally associated with platinum resistant disease. After adjusting for covariates including residual disease and maintenance therapy, IL-8 was no longer associated with platinum sensitive status (p=0.13), VEGF remained associated with OS (low vs. high HR 0.3, 95% CI 0.1–0.8, p=0.018), and higher IL-12 was associated with longer PFS (HR 0.4, 95% CI 0.2–0.9, p=0.031). ConclusionIn HGSC, pro-inflammatory cytokines are influenced by obesity, as differing inter-cytokine correlations were observed based on BMI, possibly due to dysregulation between cytokines in the setting of obesity. Differences in survival and platinum sensitivity were not noted. Future studies are warranted to determine whether obesity may be a modifiable risk factor for poorer outcomes due to differing immune response.

Dietary inflammatory index and ovarian cancer risk in a large Italian case-control study.

While inflammation has been shown to play an important etiologic role in ovarian carcinogenesis, little is known about the association between inflammatory properties of diet and ovarian cancer risk. We explored the association between the dietary inflammatory index (DII) and ovarian cancer risk in a multicentric Italian case-control study conducted between 1992 and 1999. Cases were 1,031 women with incident, histologically confirmed ovarian cancer from four areas in Italy. Controls were 2,411 women admitted to the same network of hospitals as the cases for acute, non-malignant and non-gynecological conditions, unrelated to hormonal or digestive-tract diseases or committed to long-term modifications of diet. DII scores were computed based on 31 nutrients and food items assessed using a reproducible and validated 78-item food frequency questionnaire. Odds ratios (ORs) were estimated through logistic regression models adjusting for age, total energy intake and other recognized confounding factors. Subjects in the highest quartile of DII scores (i.e., with the most pro-inflammatory diets) had a higher risk of ovarian cancer compared to subjects in the lowest quartile (i.e., with an anti-inflammatory diet) (ORQuartile4vs1 1.47, 95% confidence interval, CI, 1.07, 2.01; p trend=0.009). When analyses were carried out using continuous DII, a significant positive association with ovarian cancer was observed: the OR for one-unit increment in DII score (corresponding to approximately 8% of its range in the current study, +6.0 to -6.20) was 1.08 (95% CI 1.02, 1.14). A pro-inflammatory diet as indicated by higher DII scores is associated with increased ovarian cancer risk.

Lésions épithéliales précoces dans les annexectomies prophylactiques chez des patientes à haut risque de cancer de l’ovaire : à propos d’une série de 93 cas

ObjectiveTubal lesions detected in specimen of risk reducing salpingo oophorectomy (RRSO) for mutation BRCA1/2 seems to play a role in ovarian carcinogenesis. The main objective of this study is to evaluate the prevalence of occult neoplasia, of Serous Tubal Intraepithelial Carcinoma (STIC), and signature P53 in a cohort of patients who underwent a risk reducing salpingo oophorectomy. MethodsFrom January 2010 to January 2014 unicentric, retrospective study on a consecutive cases cohort of RRSO for patients with a high risk of ovarian neoplasia (mutation BRCA 1/2 or family history). Pathological specimen should be analysed according to the SEE–FIM protocol. ResultsNinety-three RRSO were recorded. Among them, 44% of the patients had the germ line mutation BRCA1, 30.1% BRAC2 and 18.2% had no identified mutation. In all, 33.3% of the RRSO reveal a signature P53, in the fimbria for 93.9%, 7.9% of them were bilateral. 1,1‰ (n=1) of the patients presented a unilateral STIC. We obtained 4.3% of occult neplasia: 3 ovarian high-grade serous carcinomas and 1 tubal high-grade serous carcinoma. Only the tubal carcinoma coexists with STIC. Conclusion5,4% of the patients who underwent RRSO had a diagnostic of occult neoplasia. One percent of the patients had an isolated STIC. These results agree with recent data of the literature. Extensive examination of the Fallopian tube opens up a new way to understand ovarian carcinogenesis.

Association of androgen receptor CAG repeat polymorphism and risk of epithelial ovarian cancer

Biological and epidemiologic evidence suggested that androgen and its receptor may play an important role in ovarian carcinogenesis. However, results of previous association studies about ovarian cancer and AR CAG repeat polymorphism were inconsistent. Furthermore, none of these studies were conducted in Asians. We evaluated the relationship between AR CAG repeat length and epithelial ovarian cancer (EOC) risk among a Chinese population including 1800 pathologically confirmed EOC patients and 1800 frequency matched controls. Women with longer AR CAG repeats had a decreased EOC risk (OR=0.87 for per CAG_A increase, 95% CI: 0.81-0.95). Compared to those with shorter (<22) CAG_A repeat length, women with of longer (≥22) CAG_A repeats had a 34% decreased EOC risk (OR=0.66, 95% CI: 0.57-0.75). For CAG_S and CAG_L, the results remained consistent. Our findings suggest that androgen signaling contributes to the development of ovarian cancer.

SKP2/P27Kip1 pathway is associated with Advanced Ovarian Cancer in Saudi Patients.

Ovarian cancer is the most common gynecological malignancy and constitutes the fifth leading cause of female cancer death. Some biological parameters have prognostic roles in patients with advanced ovarian cancer and their expression may contribute to tumor progression. The aim of this study was to investigate the potential prognostic value of SKP2, genes P27Kip1, K-ras, c-Myc, COX2 and HER2 genes expression in ovarian cancer. This study was performed on two hundred formalin fixed paraffin embedded ovarian cancer and normal adjacent tissues (NAT). Gene expression levels were assessed using real time PCR and Western blotting. Elevated expression levels of SKP2, K-ras, c-Myc, HER2 and COX2 genes were observed in 61.5% (123/200), 92.5% (185/200), 74% (148/200), 96 % (192/200), 90% (180/200) and 78.5% (157/200) of cancer tissues, respectively. High expression of SKP2 and down-regulation of P27 was associated with advanced stages of cancer. The association between high expression of c-Myc and SKP2 with low expression of P27 suggested that the Skp2-P27 pathway may play an important role in ovarian carcinogenesis. Reduced expression of P27 is associated with advanced stage of cancer and can be used as a biological marker in clinical routine assessment and management of women with advanced ovarian cancer.

Fimbrial cells exposure to catalytic iron mimics carcinogenic changes.

Recent evidence strongly suggests that the fallopian tube is a site of origin of ovarian cancer. Although histological data show iron deposition in the fallopian tubes, its role remains unclear. To establish whether catalytic iron has a possible role in ovarian carcinogenesis, we isolated human fimbrial secretory epithelial cells (FSECs). Fimbrial secretory epithelial cells, isolated from women undergoing isteroannessiectomy, were treated with different doses of catalytic iron (0.05-100 mM) to study cell viability; NO production; p53, Ras, ERK/MAPK, PI3K/Akt, Ki67, and c-Myc protein expressions through Western blot analysis; and immunocytochemistry or immunofluorescence. In FSECs treated with catalytic iron for up to 6 days, we observed an increase in cell viability, NO production, and p53, pan-Ras, ERK/MAPK, PI3K/Akt, Ki67, and c-Myc activations (P < 0.05) in a dose-dependent and time-dependent manner. These same results were also observed in FSECs maintained for respectively 2 and 4 weeks in the absence of catalytic iron after 6 days of stimulation. Our model aimed at studying the main nongenetic risk factor for ovarian cancer, providing an alternative interpretation for the role of menstruation in increasing risk of this pathology. This in vitro model mimics several features of the precursor lesions and opens new scenarios for further investigations regarding the correlation between damages produced by repeated retrograde menstruation carcinogenic stimuli.

Hypermethylation of the TGF-β target, ABCA1 is associated with poor prognosis in ovarian cancer patients.

BackgroundThe dysregulation of transforming growth factor-β (TGF-β) signaling plays a crucial role in ovarian carcinogenesis and in maintaining cancer stem cell properties. Classified as a member of the ATP-binding cassette (ABC) family, ABCA1 was previously identified by methylated DNA immunoprecipitation microarray (mDIP-Chip) to be methylated in ovarian cancer cell lines, A2780 and CP70. By microarray, it was also found to be upregulated in immortalized ovarian surface epithelial (IOSE) cells following TGF-β treatment. Thus, we hypothesized that ABCA1 may be involved in ovarian cancer and its initiation.ResultsWe first compared the expression level of ABCA1 in IOSE cells and a panel of ovarian cancer cell lines and found that ABCA1 was expressed in HeyC2, SKOV3, MCP3, and MCP2 ovarian cancer cell lines but downregulated in A2780 and CP70 ovarian cancer cell lines. The reduced expression of ABCA1 in A2780 and CP70 cells was associated with promoter hypermethylation, as demonstrated by bisulfite pyro-sequencing. We also found that knockdown of ABCA1 increased the cholesterol level and promoted cell growth in vitro and in vivo. Further analysis of ABCA1 methylation in 76 ovarian cancer patient samples demonstrated that patients with higher ABCA1 methylation are associated with high stage (P = 0.0131) and grade (P = 0.0137). Kaplan-Meier analysis also found that patients with higher levels of methylation of ABCA1 have shorter overall survival (P = 0.019). Furthermore, tissue microarray using 55 ovarian cancer patient samples revealed that patients with a lower level of ABCA1 expression are associated with shorter progress-free survival (P = 0.038).ConclusionsABCA1 may be a tumor suppressor and is hypermethylated in a subset of ovarian cancer patients. Hypermethylation of ABCA1 is associated with poor prognosis in these patients.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-014-0036-2) contains supplementary material, which is available to authorized users.

Lesser-Known Molecules in Ovarian Carcinogenesis.

Currently, the deciphering of the signaling pathways brings about new advances in the understanding of the pathogenic mechanism of ovarian carcinogenesis, which is based on the interaction of several molecules with different biochemical structure that, consequently, intervene in cell metabolism, through their role as regulators in proliferation, differentiation, and cell death. Given that the ensemble of biomarkers in OC includes more than 50 molecules the interest of the researchers focuses on the possible validation of each one's potential as prognosis markers and/or therapeutic targets. Within this framework, this review presents three protein molecules: ALCAM, c-FLIP, and caveolin, motivated by the perspectives provided through the current limited knowledge on their role in ovarian carcinogenesis and on their potential as prognosis factors. Their structural stability, once altered, triggers the initiation of the sequences characteristic for ovarian carcinogenesis, through their role as modulators for several signaling pathways, contributing to the disruption of cellular junctions, disturbance of pro-/antiapoptotic equilibrium, and alteration of transmission of the signals specific for the molecular pathways. For each molecule, the text is built as follows: (i) general remarks, (ii) structural details, and (iii) particularities in expression, from different tumors to landmarks in ovarian carcinoma.

A novel serum microRNA panel to discriminate benign from malignant ovarian disease

Ovarian cancer is the seventh most common cancer in women and the most frequent cause of gynaecological malignancy-related mortality in women. Currently, no standardized reliable screening test exists. MicroRNA profiling has allowed the identification of signatures associated with diagnosis, prognosis and response to treatment of human tumours. The aim of this study was to determine if a microRNA signature could distinguish between malignant and benign ovarian disease. A training set of 5 serous ovarian carcinomas and 5 benign serous cystadenomas were selected for the initial experiments. The validation set included 20 serous ovarian carcinomas and 20 benign serous cystadenomas. The serum/plasma focus microRNA Exiqon panel was used for the training set. For the validation set a pick and mix Exiqon panel, which focuses on microRNAs of interest was used. A panel of 4 microRNAs (let-7i-5p, miR-122, miR-152-5p and miR-25-3p) was significantly down regulated in cancer patients. These microRNAs target WNT signalling, AKT/mTOR and TLR-4/MyD88, which have previously been found to play a role in ovarian carcinogenesis and chemoresistance. let-7i-5p, miR-122, miR-152-5p and miR-25-3p could act as diagnostic biomarkers in ovarian cancer.

Abstract 2369: Molecular biological characterization of newly established epithelial ovarian cancer (EOC) cell lines : The cyclin-dependent kinase 1 pathway

Abstract Background Gene expression pattern analysis comparing normal cells with cancer cells are performed by many competitors in the worldwide lab. Cancer has many different species in itself due to its individual differences, variations and heterogeneity. Therefore, it is necessary to spend more time in gene expression pattern analysis using microarray or whole genome sequencing. The purpose of this study was to identify genes and proteins that are highly differentially expressed in newly established epithelial ovarian cancer (EOC) cell lines, and to use this knowledge for the development of novel diagnostic and prognostic markers for EOC. We further performed the validation studies of several candidate genes. Results Comparison of gene expression patterns using microarray analysis enabled us to identify 84 genes that were commonly up-regulated and 132 genes that were down-regulated in cancer cell lines (up; &amp;gt; 2-fold, down; &amp;lt; 3-fold, P &amp;lt; 0.05). In 2-DE and MALDI-TOF/PMF, 31 up-regulated spots that had at least two-fold differences between the 3 EOC cell lines and HOSE cells used as controls were observed. 15 up-regulated genes in cell cycle related genes and 9 down-regulated genes in apoptosis related confirmed gene expression using quantitative RT-PCR. Pathway analysis was validated in cell cycle related genes in six EOC cell lines. Conclusions The data shows the aberrant expression of several cell cycle related genes in EOC and that Cyclin-dependent kinase 1 (CDK1) pathway may play a significant role in ovarian carcinogenesis, implying that CDK1 pathway could be used as a novel therapeutic target for ovarian cancer treatment. Citation Format: wookyeom yang, hanbyoul Cho, hayeon Shin, eunju Lee, hyunja Kwon, sol Kim, jaehoon Kim. Molecular biological characterization of newly established epithelial ovarian cancer (EOC) cell lines : The cyclin-dependent kinase 1 pathway. [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 2369. doi:10.1158/1538-7445.AM2014-2369

The role of RhoC in epithelial-to-mesenchymal transition of ovarian carcinoma cells

BackgroundRhoC is a small G protein/GTPase and involved in tumor mobility, invasion and metastasis. Previously, up-regulated RhoC expression is found to play an important role in ovarian carcinogenesis and subsequent progression by modulating proliferation, apoptosis, migration and invasion.MethodsWe transfected RhoC-expressing plasmid and RhoC siRNA into CAOV3 and OVCAR3 cells respectively. These cells and transfectants were exposed to vascular epithelial growth factor (VEGF), transforming growth factor (TGF)-β1 or their receptor inhibitors with the phenotypes and their related-molecules examined.ResultsTGF-β1R or VEGFR inhibitor suppressed the proliferation, migration, invasion and lamellipodia formation, the expression of N-cadherin, α-SMA, snail and Notch1 mRNA or protein, and enhanced E-cadherin mRNA and protein expression in CAOV3 and its RhoC-overexpressing transfectants, whereas both growth factors had the opposite effects in OVCAR3 cells and their RhoC-hypoexpressing transfectants. Ectopic RhoC expression enhanced migration, invasion, lamellipodia formation and the alteration in epithelial to mesenchymal transition (EMT) markers of CAOV3 cells regardless of the treatment of VEGFR or TGF-β1R inhibitor, whereas RhoC knockdown resulted in the converse in OVCAR3 cells even with the exposure to VEGF or TGF-β1.ConclusionRhoC expression might be involved in EMT of ovarian epithelial carcinoma cells, stimulated by TGF-β1 and VEGF.

Epigenetic alteration of p16 and retinoic acid receptor beta genes in the development of epithelial ovarian carcinoma.

Silencing of tumor suppressor and tumor-related genes by promoter hypermethylation is one of the major events in ovarian carcinogenesis. In this study, we analyzed aberrant promoter methylation of p16 and RAR-β genes in 134 epithelial ovarian carcinomas (EOCs), 23 low malignant potential (LMP) tumors, 26 benign cystadenomas, and 15 normal ovarian tissues. Methylation was investigated by methylation-specific PCR (MSP), and the results were confirmed by bisulfite DNA sequencing. Relative gene expression of p16 and RAR-β was done using quantitative reverse transcriptase PCR (qRT-PCR) on 51 EOC cases, 9 LMP tumors, and 7 benign cystadenomas with 5 normal ovarian tissues. Aberrant methylation for p16 and RAR-β was present in 43 % (58/134) and 31 % (41/134) in carcinoma cases, 22 % (05/23) and 52 % (12/23) in LMP tumors, and 42 % (11/26) and 69 % (18/26) in benign cystadenomas. No methylation was observed in any of the normal ovarian tissues. The mRNA expression level of p16 and RAR-β was significantly downregulated in EOC and LMP tumors than the corresponding normal tissues whereas the expression level was normal in benign cystadenomas for p16 and slightly reduced for RAR-β. A significant correlation of p16 promoter methylation was observed with reduced gene expression in EOC. For RAR-β, no significant correlation was observed between promoter methylation and gene expression. Our results suggest that epigenetic alterations of p16 and RAR-β have an important role in ovarian carcinogenesis and that mechanism along with methylation plays a significant role in downregulation of RAR-β gene in ovarian cancer.

Synergistic effects of combined platelet-activating factor receptor and epidermal growth factor receptor targeting in ovarian cancer cells

BackgroundGenetic alterations, including the overexpression of epidermal growth factor receptor (EGFR), play a crucial role in ovarian carcinogenesis. To date, EGFR targeting has shown limited antitumor effects in ovarian cancer when administered as monotherapy. We previously identified platelet-activating factor receptor (PAFR) as being overexpressed in ovarian cancer and found that its ligand PAF evoked EGFR phosphorylation. To determine whether PAFR targeting can enhance the antitumor efficacy of EGFR inhibition, we investigated the effects of a PAFR antagonist (WEB2086) in conjunction with an EGFR inhibitor (AG1478).MethodsThe expression of EGFR and PAFR in CAOV-3 and SKOV-3 ovarian cancer cell lines was measured by Western blot and immunocytochemistry. Synergy was determined using isobologram analysis. The effects of combined PAFR and EGFR targeting on both cells were assessed by using CCK-8, transwell, flow cytometry, western blot analysis. In vivo studies were conducted using CAOV-3 cells xenografted in nu/nu mice.ResultsTreatment with combination WEB2086 and AG1478 resulted in significantly greater inhibition of proliferation and invasion compared to either drug alone. When examining equipotent combinations of WEB2086 and AG1478 to determine potential synergy, a combination index (CI) of 0.49 was identified for CAOV-3 cells and a CI of 0.58 for SKOV-3 cells indicating synergy. This co-inhibition induced significantly more apoptosis and arrested the cells at G0/G1 phase in both cell lines. The activation of PAFR and/or EGFR induced phosphorylation of the mTOR, AKT, and MAPK pathways. Combined PAFR and EGFR targeting synergistically diminished the expression of PAFR and EGFR phosphorylation and downstream signaling. In vivo studies further verified the antitumor effects of combined PAFR and EGFR targeting in a CAOV-3 xenograft model.ConclusionsThese results suggest that WEB2086 and AG1478 are synergistic in ovarian cancer cells with high expression of both PAFR and EGFR. The presented approach may have important therapeutic implications in the treatment of ovarian cancer patients.

Exposure of fallopian tube epithelium to follicular fluid mimics carcinogenic changes in precursor lesions of serous papillary carcinoma

ObjectivesOvulation-related inflammation is suspected to have a causal role in ovarian carcinogenesis, but there are no human models to study the molecular pathways. Our aim is to develop such an ex-vivo model based on human fallopian tube (FT) epithelium exposed to human follicular fluid (FF). MethodsFT epithelium was dissociated from normal surgical specimens. FF was obtained from donors undergoing in-vitro fertilization. The cells were cultured on collagen-coated Transwells and incubated with FF for various periods of time. The transcriptomic changes resulting from FF treatment were profiled using Affymetrix expression arrays. Specific characteristics of the FT pre-cancerous lesions were studied using immunohistochemistry, immunofluorescence, RT-PCR and XTT assay. ResultsWe show that FF exposure causes up-regulation of inflammatory and DNA repair pathways. Double stranded DNA breaks are induced. There is a minor increase in cell proliferation. TP53, which is the hallmark of the precursor lesion in-vivo, is accumulated. Levels of expression and secretion of Interleukin-8 are significantly increased. ConclusionsOur model addresses the main non-genetic risk factor for ovarian cancer, namely the impact of ovulation. This study demonstrates the biological implications of in-vitro exposure of human FT epithelial cells to FF. The model replicates elements characterizing the precursor lesions of ovarian cancer, and warrants further investigation of the linkage between repeated exposure to ovulation-related damage and accumulation of neoplastic changes.

Human papillomavirus genotyping and integration in ovarian cancer Saudi patients

BackgroundHuman papillomavirus (HPV) is associated with different malignancies but its role in the pathogenesis of ovarian cancer is controversial. This study investigated the prevalence, genotyping and physical state of HPV in ovarian cancer Saudi patients.MethodsHundred formalin fixed paraffin embedded (FFPE) ovarian carcinoma tissues and their normal adjacent tissues (NAT) were included in the study. HPV was detected by nested polymerase chain reaction (PCR) using degenerated HPVL1 consensus primer pairs MY09/MY11 and GP5+/GP6 + to amplify a broad spectrum of HPV genotypes in a single reaction. The HPV positive samples were further genotyped using DNA sequencing. The physical state of the virus was identified using Amplification of Papillomavirus Oncogene Transcripts (APOT) assay in the samples positive for HPV16 and/or HPV18.ResultsHigh percentage of HPV (42%) was observed in ovarian carcinoma compared to 8% in the NAT. The high-risk HPV types 16, 18 and 45 were highly associated with the advanced stages of tumor, while low-risk types 6 and 11 were present in NAT. In malignant tissues, HPV-16 was the most predominant genotype followed by HPV-18 and -45. The percentage of viral integration into the host genome was significantly high (61.1%) compared to 38.9% episomal in HPV positive tumors tissues. In HPV18 genotype the percentage of viral integration was 54.5% compared to 45.5% episomal.ConclusionThe high risk HPV genotypes in ovarian cancer may indicate its role in ovarian carcinogenesis. The HPV vaccination is highly recommended to reduce this type of cancer.

Importance of promoter methylation of GATA4 gene in epithelial ovarian cancer

Ovarian cancer is the most lethal gynecological malignancy, with typically late diagnosis. Altered DNA methylation of tumor suppressor gene promoters probably plays a relevant role in ovarian carcinogenesis and frequently occurs as an early event in the development of different types of cancer including ovarian carcinoma. GATA4 methylation has been reported in a variety of human cancers. The aim of this study was to investigate promoter methylation of the GATA4 gene in ovarian cancer by comparison with that in normal ovarian tissue. To search for promoter methylation of the GATA4 gene we used MSP (methylation-specific PCR) to compare the methylation status in 67 tissue samples of ovarian cancer with that in 40 control samples. In our study, methylation-specific PCR revealed GATA4 promoter methylation in 21 of 67 specimens with ovarian cancer (31.3%), and in none of the control ovarian tissue samples. These results confirm that methylation in the GATA4 promoter region could play an important role in ovarian carcinogenesis, and show new loci which are highly methylated only in ovarian cancer samples and which are associated predominantly with the endometrioid type of ovarian carcinoma.