High-grade Serous Ovarian Tumors Research Articles

Individuality in FGF1 expression significantly influences platinum resistance and progression-free survival in ovarian cancer

Background:Ovarian cancer is frequently advanced at presentation when treatment is rarely curative. Response to first-line platinum-based chemotherapy significantly influences survival, but clinical response is unpredictable and is frequently limited by the development of drug-resistant disease.Methods:We used qRT–PCR analysis to assess intertumour differences in the expression of fibroblast growth factor 1 (FGF1) and additional candidate genes in human ovarian tumours (n=187), and correlated individuality in gene expression with tumour histology, chemotherapy response and survival. We used MTT assays to assess platinum chemosensitivity in drug-sensitive and drug-resistant ovarian cell lines.Results:Marked intertumour differences in gene expression were observed, with each tumour having a unique gene expression profile. Nine genes, including FGF1 (P=1.7 × 10−5) and FGFR2 (P=0.003), were differentially expressed in serous and nonserous tumours. MDM2 (P=0.032) and ERBB2 (P=0.064) expression was increased in platinum-sensitive patients, and FGF1 (adjusted log-rank test P=0.006), FGFR2 (P=0.04) and PDRFRB expression (P=0.037) significantly inversely influenced progression-free survival. Stable FGF1 gene knockdown in platinum-resistant A2780DPP cells re-sensitised cells to both cisplatin and carboplatin.Conclusion:We show for the first time that FGF1 is differentially expressed in high-grade serous ovarian tumours, and that individuality in FGF1 expression significantly influences progression-free survival and response to platinum-based chemotherapy.

CD20+ Tumor-Infiltrating Lymphocytes Have an Atypical CD27− Memory Phenotype and Together with CD8+ T Cells Promote Favorable Prognosis in Ovarian Cancer

Tumor-infiltrating lymphocytes (TIL), in particular CD8(+) T cells and CD20(+) B cells, are strongly associated with survival in ovarian cancer and other carcinomas. Although CD8(+) TIL can mediate direct cytolytic activity against tumors, the role of CD20(+) TIL is poorly understood. Here, we investigate the possible contributions of CD20(+) TIL to humoral and cellular tumor immunity. Tumor and serum specimens were obtained from patients with high-grade serous ovarian cancer. CD8(+) and CD20(+) TIL were analyzed by immunohistochemistry and flow cytometry. Immunoglobulin molecules were evaluated by DNA sequencing. Serum autoantibody responses to the tumor antigens p53 and NY-ESO-1 were measured by ELISA. The vast majority of CD20(+) TIL were antigen experienced, as evidenced by class-switching, somatic hypermutation, and oligoclonality, yet they failed to express the canonical memory marker CD27. CD20(+) TIL showed no correlation with serum autoantibodies to p53 or NY-ESO-1. Instead, they colocalized with activated CD8(+) TIL and expressed markers of antigen presentation, including MHC class I, MHC class II, CD40, CD80, and CD86. The presence of both CD20(+) and CD8(+) TIL correlated with increased patient survival compared with CD8(+) TIL alone. In high-grade serous ovarian tumors, CD20(+) TIL have an antigen-experienced but atypical CD27(-) memory B-cell phenotype. They are uncoupled from serum autoantibodies, express markers of antigen-presenting cells, and colocalize with CD8(+) T cells. We propose that the association between CD20(+) TIL and patient survival may reflect a supportive role in cytolytic immune responses.

Abstract 4270: Novel stromal angiogenic factor MFAP5 in ovarian cancer progression

Abstract Using transcriptome profiling analysis, we identified a gene signature in the fibroblastic stromal component of high grade serous ovarian tumor tissue. One of the most differentially expressed proteins was microfibrillar associated protein 5 (MFAP5). This study aims at validating the differential expression patterns of MFAP5 in ovarian cancer stroma, evaluating the clinical significance of stromal MFAP5 over-expression and delineating the functional role and mechanism of MFAP5 in modulating angiogenesis and tumor progression. Quantitative RT-PCR analysis was first performed on RNA isolated from microdissected normal and malignant ovarian tissue samples and immunolocalization of MFAP5 protein was performed on paraffin tissue sections (n=102) to validate differential stromal MFAP5 expression. Both demonstrated significant higher levels of MFAP5 expression in the fibroblastic stromal component of cancer tissue samples. Kaplan Meier analysis demonstrated a significant correlation between stromal MFAP5 expression and poor overall patient survival (p<0.001). Cox regression analysis showed a hazard ratio of 2.603 (p<0.001). Further correlation studies showed that MFAP5 expression correlated with CD34 positive microvessel densities, suggested a pro-angiogenic role of MFAP5. Thus, functional studies of exogenous MFAP5 on angiogenesis modulation and ovarian cancer cell growth were performed. Motility assay and tube formation assay were conducted on human microvascular endothelial cell 1 (hMEC-1) and telomerase immortalized microvascular endothelial cells (TIME). Increased motility and tube formation were observed in the presence of exogenous MFAP5 in both cell lines. In addition, both anti-αVα3 integrin antibody and cell permeant calcium chelator, BAPTA, AM, attenuated MFAP5 stimulated motility. These suggested that MFAP5 exerts its biological activity through the αVα3 integrin receptor and calcium signal transduction. The hypothesis is further supported by the successful detection of calcium influx and increased amplitude and change of distribution of traction force exerted by endothelial cells after treatment with MFAP5. In vivo, extensive CD34 positive tubular network was formed in the matrigel mixed with recombinant MFAP5 injected subcutaneously into mice when compared to the control. Co-injection of MFAP5 overexpressing ovarian fibroblasts and cancer cells subcutaneously into mice led to increased tumor size and increased number of CD34 positive microvessels within the tumor tissue. Finally, in vivo silencing of stromal MFAP5 using MFAP5 specific siRNA delivered in chitosan nanoparticle significantly decreased tumor sizes and intratumor microvessel densities. To conclude, stromal MFAP5 is a novel angiogenic factor that acts on αVα3 integrin receptor. It modulates endothelial cell behavior through calcium signaling which may lead to tumor progression and subsequent poor patient survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4270. doi:1538-7445.AM2012-4270

Abstract 3004: Biomarker identification through integrative bioinformatics analysis of serous epithelial ovarian cancer tumor samples

Abstract Ovarian cancer is the leading cause of death from gynecological malignancies and the fifth major cancer in women in the world. Once diagnosed, ovarian cancer is usually treated by cytoreductive surgery followed by platinum and taxane-based chemotherapeutic drugs. However, resistance to chemotherapy is a major impediment in management of serous epithelial ovarian cancer (SEOC). We hypothesize that a multifaceted view of the alterations taking place at multiple cellular levels using molecular profiling technologies will offer insight into the mechanisms which play key roles in drug resistant ovarian carcinomas. Also, the application of appropriate bioinformatic and statistical data processing and analysis is of utmost importance in identification of key drug resistance pathways. Current study is performed on 25 high-grade serous epithelial ovarian tumor tissue samples from patients that demonstrated favorable, or unfavorable response to chemotherapy treatment. Four different microarray platforms were used for molecular profiling of the full sample cohort at different molecular levels, namely: Single Nucleotide Polymorphisms (SNP), mRNA expression, miRNA expression and promotor tiling arrays (methylation)., Integrative and systematic analyses using up-to-date statistical approaches, such as empirical Bayes, AUC, SAM, permuted t-test and lassoed PCA, among others, have been employed on these large datasets obtained through the various high-throughput platforms. Preliminary mRNA expression analysis identified an enrichment of upregulated genes involved in cellular growth and proliferation, cellular development as well as differential gene expression changes in the TGFB1, TNF, PI3K, IFNG networks, between the chemotherapy responsive and unresponsive groups. The major molecular and cellular functions associated were cell-to-cell signaling, molecular transport and cellular movement. Differences were also seen in the, CTNNB1, LH and FSH networks as analysed by Ingenuity Pathway Analysis. Additionally, genes involved in activation of NFκB pathway showed differential expression in the two groups. Furthermore, our ongoing development of a streamlined database in which the multiple data types obtained from our statistical analyses are stored, will allow for localized or genome wide querying across the multiple levels of biological data. These approaches and software will potentially elucidate the synergistic roles that the various biological levels play in the deregulation of pathways involved in primary chemoresistance. Our research findings will lead to the determination of putative candidates for diagnostic and prognostic biomarkers that can be targeted for development of treatment regimens for the treatment of SEOC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3004. doi:1538-7445.AM2012-3004

The anti-adhesive mucin podocalyxin may help initiate the transperitoneal metastasis of high grade serous ovarian carcinoma

High grade serous ovarian tumors often metastasize transperitoneally, a process that begins when small tumor nodules de-adhere and are released into the fluid of the abdominal cavity where they float freely to reach new sites on the peritoneal wall. Podocalyxin, a small anti-adhesive sialomucin, has been shown to contribute to non-adhesive membrane domain formation in some epithelia and is overexpressed in a variety of cancers. We therefore assessed podocalyxin expression on a previously characterized tissue microarray and found that 87% (169/194) of high grade serous epithelial ovarian carcinomas were positive for podocalyxin. In addition, cell surface localization of podocalyxin was associated with a significant decrease in disease-free survival in these tumors. When podocalyxin was force-expressed in serous ovarian carcinoma-derived OVCAR-3 cells it was targeted to the cell surface and it decreased the adhesion of these cells to mesothelial monolayers, fibronectin and immobilized β1 integrin-binding antibodies. This decrease in adhesion was associated with a modest decrease in cell surface β1 integrin. In monolayer culture, podocalyxin was targeted to the free, apical domains of OVCAR-3 cells and it appeared to decrease β1 integrin levels on the attached basolateral domains of the same cells. Furthermore, in 3-dimensional basement membrane gel culture, the cells formed small, cohesive nodules and podocalyxin localized to membrane domains at the cell-basement membrane interface. Therefore, podocalyxin's ability to facilitate the formation of non-adhesive membrane domains may contribute to the formation of free-floating high grade serous tumor nodules during the initial steps of transperitoneal metastasis.

Abstract PL05-04: Epigenetic marks as biomarkers and their mechanisms of maintenance as targets in ovarian cancer.

Abstract PL05-04: Epigenetic marks as biomarkers and their mechanisms of maintenance as targets in ovarian cancer.

Increased tissue immunoexpression of YKL-40 protein in high grade serous ovarian cancers

YKL-40 is a glycoprotein secreted by numerous human cells, such as cartilage, synovial, and endothelial cells. The biological role of YKL-40 has not yet been fully unveiled, however, its participation is perceived in angiogenesis, growth, proliferation, differentiation, and remodeling processes. The primary goal of our study was to evaluate possible differences in tissue immunoexpression of YKL-40, assumed between high grade and low-grade ovarian cancers and between the above-mentioned cancer types and benign lesions. Another purpose was to find out whether immunoexpression of the studied protein could correlate with the tumor proliferation process, evaluated by Ki-67 immunoexpression. The analysis comprised 45 women, diagnosed and treated for epithelial ovarian tumors at the Medical University of Lodz between 1997 and 2002. YKL-40 protein immunoexpression was semiquantitatively assessed, whereas immunoexpression of Ki-67 was evaluated using a computer image analysis system. Significantly higher immunoexpression values of both examined proteins were observed in high-grade serous ovarian cancers vs. low-grade and benign tumors. Moreover, a significant positive correlation was identified between the immunoexpressions of YKL-40 and Ki-67 proteins in the studied groups of tumors. In conclusion, the obtained data suggest an overt prominence of TKL-40 tissue immunoexpression of YKL-40 in high-grade serous ovarian tumors, which could then be approached as a helpful, additional marker to identify more aggressive ovarian cancers.

Newly diagnosed and relapsed epithelial ovarian carcinoma: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up

Newly diagnosed and relapsed epithelial ovarian carcinoma: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up

Abstract 2054: A microRNA hypothesis for the precursor lesions of ovarian cancer

Abstract Ovarian cancer is the leading cause of death among gynecologic diseases in Western countries. Ovarian surface epithelial (OSE) cells were previously considered as the cells of origin of ovarian cancer. The ovarian surface epithelium is derived from the coelomic layer, which is consisted of peritoneal mesothelial cells. Malignant ovarian cancer cells, however, are found to be associated with epithelial phenotypes resembling cells from fallopian tube, uterus, and cervix. Recent studies have suggested that significant percentages of familial and sporadic high-grade serous ovarian carcinomas could be explained by an origin in the distal fallopian tube. We and others have identified overexpression of miR-200 family members in ovarian caner cells. In order to investigate whether the miR-200 pathway is involved in the early stages of ovarian pathogenesis, we have studied the expression levels of the pathway components in a panel of samples isolated from primary cultures and laser microdissection. The ovarian samples included OSEs, epithelial tumor cells from benign tumors, borderline tumors, nonserous invasive tumors, serous invasive tumors, ovarian cancer cell lines, and tumor-associated stromal fibroblasts. Fallopian tube samples included normal fallopian tubal epithelial (FTE) cells, tumor cells and stromal fibroblasts from frozen tubal intraepithelial carcinomas (TICs) of fallopian tube. Gene expression was determined using quantitative real-time polymerase chain reactions (qRT-PCRs). The reactions for the 4 target genes of miR-200s (ZEB1, ZEB2, TGFb1 and TGFb2), and 21 effector genes that were regulated by the target genes, were performed using a high-throughput BioMark array platform (Fluidigm). qRT-PCR of epithelial tumor cells from ovarian tumors and fallopian TICs showed concurrent up-regulation of miR-200s, down-regulation of the target genes, and up-regulation of several effector genes. To confirm the role of miR-200 family in regulating gene expression, synthetic miRNA precursors were transfected into normal OSE and FTE cells and qRT-PCRs were performed to determine the expression of target and effector genes. The results showed similar reduced expression of target genes and elevated levels of CDH1 and EpCAM effector genes in both normal OSE and FTE cells. However, the most interesting finding was the specific induction of tumor marker, CA125, in the normal FTE cells. Immunocytochemistry confirmed the induced CA125 proteins in FTE cells. In conclusion, the activation of the miR-200 pathway may be an early lesion that renders the OSE and FTE cells more susceptible to oncogenic mutations and histologic differentiation. As high-grade serous ovarian tumors usually express higher levels of CA125, the greater capacity of FTE cells to express CA125 by miR-200 induction is consistent with the notion that high-grade serous ovarian tumors have an origin in the distal fallopian tube. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2054.

Whole Genome Oligonucleotide-Based Array Comparative Genomic Hybridization Analysis Identified Fibroblast Growth Factor 1 As a Prognostic Marker for Advanced-Stage Serous Ovarian Adenocarcinomas

To identify markers that can predict overall survival in patients with high-grade advanced stage serous adenocarcinomas. Oligonucleotide array comparative genomic hybridization (aCGH) was performed on 42 microdissected high-grade serous ovarian tumor samples. aCGH segments were obtained and a prediction Cox model was built and validated by the standard leave one out analysis. Both DNA and mRNA copy numbers of selected genes located on the candidate aCGH segments were determined by quantitative polymerase chain reaction (qPCR) and quantitative reverse transcriptase PCR (qRT-PCR) analyses. The gene that showed the highest correlation was further validated on an independent set of specimens and was selected for further functional studies. Two chromosomal regions, 4p16.3 and 5q31-5q35.3, exhibited the strongest correlation with overall survival (P < .01). From the 5q31 region, fibroblast growth factor 1 (FGF-1) was selected for further validation study. FGF-1 mRNA copy number was significantly correlated with DNA copy number and protein expression levels (P = .021 and < .001), and both FGF-1 mRNA and protein levels were significantly associated with overall survival (P = .018 and .042). This association was validated for protein expression on an independent set of 81 samples, significant to P = .006. Further studies showed significant correlation between FGF-1 protein expression and CD31+ staining in the tumor stroma (P = .024). Finally, both cancer cells and endothelial cells treated with exogenous FGF-1 showed a significant increase in cell motility and survival. Amplification of FGF-1 at 5q31 in ovarian cancer tissues leads to increased angiogenesis, and autocrine stimulation of cancer cells, which may result in poorer overall survival in patents with high-grade advanced stage serous ovarian cancer.