Proteomic analysis of a preneoplastic phenotype in ovarian surface epithelial cells derived from prophylactic oophorectomies
Introduction
The epithelial ovarian carcinomas, which originate from the ovarian surface epithelium (OSE), are the prime cause of death from gynecological malignancies in European and North American women. One reason for the high mortality of ovarian cancer is that, unlike many other cancer types, ovarian cancer is notorious for its insidious properties in early stages. Close to 70% of patients present with the disease spread beyond the pelvis, resulting in a long-term survival rate of only 29% [1]. Indeed, if a woman is diagnosed with an early stage (stage I) ovarian cancer, the survival rate is close to 90% without altering current therapeutic approaches. Thus, it is urgently needed to understand the early events and etiology of the disease. Although there are many genetic and environmental factors which can influence a woman's risk of getting breast and ovarian cancer, a strong family history is by far the most important and best-defined epidemiological risk factor.
Recently, cancer-prone women with an inherited predisposition to ovarian cancer, often BRCA1 (a candidate tumor suppressor in breast and ovarian carcinomas) mutation carriers, have undergone prophylactic oophorectomies as a preventive approach [2], [3]. The high prevalence of ovarian cancer in cancer-prone women provides an excellent model to uncover new players in early ovarian carcinogenesis and, perhaps, means of defining and detecting risk factors at an early, curable stage. Importantly, several studies have identified microscopic benign-to-malignant morphologic features in these ovarian specimens, suggesting the existence of preneoplastic phenotypes in the cells [4], [5], [6], [7]. However, little is known about the molecular changes that are associated with or account for the preneoplastic morphologic changes.
Over 90% of ovarian cancers are thought to arise from the ovarian surface epithelium (OSE), which is a simple epithelial layer covering the ovaries. Since OSE is a minute part of the intact ovary, only limited amounts of tissue can be obtained from a single specimen. Thus, the ability to culture OSE provides an opportunity of obtaining large enough quantities of relatively pure populations of ovarian epithelial cells for in vitro studies [8], [9]. Although several previous studies have identified genes differentially expressed in ovarian cancer, our analysis represents a better approximation of the preneoplastic stage of ovarian cancer development. The majority of previous analyses were performed in ovarian cancer cells and compared with normal OSE cells [10], [11], [12], whereas we chose to compare overtly normal OSE cells of high-risk individuals to OSE cells of the general population. We and others have revealed the presence of premalignant histologic and/or biologic alterations in OSE cells from prophylactically removed ovaries of high-risk individuals [13], [14], [15], [16], [17].
This study has used a proteomic approach to compare the protein profiles of OSE. This powerful analytical technology, in contrast to array methodologies as in previous comparative gene studies, is able to provide an unbiased and comprehensive expression profiling without prior knowledge of the expressed proteins in the starting material [10], [11], [12]. It also offers the advantages of detection at the functional level of protein expression and the ability to also detect posttranslational modifications of proteins, which can easily be missed by transcriptional profiling. Differential expression of selected genes observed in this study was further confirmed by Western blotting and real-time reverse transcription polymerase chain reaction.
Section snippets
Cell cultures
Experimentation with human tissues was approved by the university's ethics committee prior to this study and the normal human OSE samples collected were made anonymous. Briefly, NFH-OSE (OSE-29, OSE-80 and OSE-398) cells were obtained from women in the general population with no family history of breast/ovarian cancer, having surgery for non-malignant gynecological diseases. FH-OSE (OSE-229F, OSE-261F and OSE-267F) were obtained from women who underwent prophylactic oophorectomy because of
Proteomic analysis
Fig. 1 shows typical silver-stained 2-D gel images for the FH IOSE-267F and the NFH IOSE-80 cell lines side by side. Around 1500 spots were detected on each gel, ranging from 6 to 200 kDa in size and 4 to 10 in pI. Normalized spot-volume comparison was made with the assistance of image analysis software (ImageMaster). Six areas where significant and consistent spot changes occurred in all cases of FH-OSE when compared to the gel images of control NFH-OSE cell lines were circled. In some cases,
Discussion
Ovaries from cancer-prone women who underwent prophylactic surgery provide an excellent opportunity to identify preneoplastic alterations predisposing to ovarian carcinogenesis. The findings here represent one of the most comprehensive studies to date, confirming and extending previous results indicating potentially preneoplastic features in ovaries from women with an inherited predisposition for ovarian cancer [4], [5], [6], [7], [21]. Since the ovaries or OSE cells removed prophylactically
Acknowledgments
We thank the B.C. Hereditary Cancer Program for providing data on BRCA1 mutation analysis in female patients from high-risk families. This work was supported by the Science Faculty Collaborative Seed Grant and Hong Kong Research Grants Council Grants HKU 7484/04M (to A.S.T.W.), HKU 7227/02M (to Q.Y.H.), HKU 7218/02M and HKU 7395/03M (to J.F.C.), the Department of Chemistry and the Areas of Excellence scheme of Hong Kong University Grants Committee.
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Ovarian surface epithelium as a source of ovarian cancers: Unwarranted speculation or evidence-based hypothesis?
2013, Gynecologic OncologyCitation Excerpt :Histologic evidence includes dysplastic lesions and in situ carcinomas in OSE-lined inclusion cysts [17,25,26], confirming that at least some carcinomas do arise from OSE. Functional and molecular preneoplastic or cancer-related changes in OSE include altered signal transduction including constitutive expression and upregulation of Akt [27,28], loss of basement membranes and the tumor suppressor Dab2 with overexpression of COX-2 in morphologically altered and adjacent morphologically normal OSE [25], augmented antiapoptotic chaperone proteins [29], altered responses to hormones and growth factors [30,31], transformative changes in response to oxidative stress [17] and, importantly, p53 overexpression with and without TP53 mutations in morphologically normal OSE, sometimes adjacent to dysplasias and in situ carcinomas [17], which is analogous to the “p53 signatures”, that signify serous carcinoma precursor lesions in fimbrial epithelium [1]. At present, perhaps the most convincing argument against OSE being a significant source of HGSOCs is that very early stage HGSOCs arising in the ovary have been observed only very rarely, though HGSOCs are by far the most common ovarian malignancies.
Application of proteomics in ovarian cancer: Which sample should be used?
2009, Gynecologic OncologyCitation Excerpt :Thirty-seven differentially expressed proteins were present and identified mainly as proteins involved in cytoskeleton organization, cell motility and adhesion. In a study of He et al. [49] ovarian surface epithelium of patients with hereditary ovarian cancer (HOC) undergoing prophylactic surgery and of patients with benign gynaecological disease were cultured and subsequently the protein profiles were compared. They found eight proteins which were altered in HOC including proteins involved in protein synthesis and processing which could protect cells against cell death.
Proteomic identification of tumor-associated protein in ovarian serous cystadenocarinoma
2009, Cancer LettersCitation Excerpt :In their study, proteome profiles of OSE from women with or without a history of familial ovarian cancer (FH-OSE/NFH-OSE) were compared. Mitochondrial short-chain enoyl-CoA hydratase was found to have a higher expression in FH-OSE [21]. Taken these observations together, we propose mitochondrial short-chain enoyl-CoA hydratase was not only associated with malignancy disease, but might have indicative value for premalignant changes.
Proteomics in Cancer
2007, Advances in Clinical ChemistryCitation Excerpt :For example, a 2D‐PAGE study determined protein patterns associated with a predisposition to develop ovarian cancer in ovarian surface epithelium obtained during prophylactic oophorectomy in high‐risk female patients. Eight proteins altered in high‐risk patients were identified: three were already known as ovarian tumor‐associated proteins, providing a proof of concept, whereas five were novel findings, representing potential early markers for the evaluation of the risk of developing ovarian cancer [85]. Numerous groups are pursuing similar serum‐based approaches to ovarian cancer diagnosis (reviewed in [86]): characteristic SELDI‐TOF and MALDI‐TOF mass spectral patterns could be identified in sera of patients that may yield a sensitive and specific signature for ovarian cancer [87].