Normal Ovarian Epithelium Research Articles

Screening for ovarian cancer: what are the optimal surrogate endpoints for clinical trials?

The inability to identify relevant markers for presymptomatic screening in early stage or "preinvasive" ovarian cancer has plagued investigators and clinicians facing the problems of early detection. The characteristic late stage of disease at initial presentation has hindered our understanding of the biologic progression and stepwise molecular alterations that result in ovarian carcinoma. To date, most screening studies have focused on identifying early anatomic changes using ultrasound or fluctuations in serum biomarkers such as CA-125. These screening methodologies have proven inadequate in both sensitivity and specificity for early stage ovarian cancer detection. Molecular analysis of ovarian carcinomas has revealed alterations in oncogenes and tumor suppressor genes associated with these tumors. The HER-2/neu oncogene, a member of the epidermal growth factor family, is amplified or overexpressed in approximately 25-30% of ovarian carcinomas. Significant data substantiate an important role for HER-2/neu in the pathophysiology of ovarian cancer. While potentially an attractive surrogate endpoint biomarker (SEB), serum HER-2/neu levels have not proven to be a useful screening modality. In response to the urgent need for improved early detection for ovarian cancer, our current research efforts include differential hybridization studies between normal and malignant ovarian epithelium to define potentially unique ovarian cancer antigens which may ultimately have utility; defining physical alterations that occur in malignant ovarian tissues using implanted telemetry systems; studies using positron emission tomography to detect changes in glucose metabolism between normal and malignant ovarian tissues; and screening studies using a 3-dimensional ultrasound unit to improve the accuracy of this technique in recognizing early neoplastic changes. By taking diverse approaches to tackle this problem, an improved understanding of ovarian carcinogenesis should translate into the identification of appropriate SEBs for early detection.

Coordinate expression of urinary-type plasminogen activator and its receptor accompanies malignant transformation of the ovarian surface epithelium

OBJECTIVE: Because elevated expression and cell surface association of urinary-type plasminogen activator have been linked to invasive potential in certain tumor types, we examined the expression of urinary-type plasminogen activator and urinary-type plasminogen activator receptor in ovarian epithelial carcinoma tissues and cells as compared with normal ovarian epithelium. STUDY DESIGN: Monoclonal antibodies specific for urinary-type plasminogen activator and urinary-type plasminogen activator receptor were used for immunohistochemical staining of tissues and cells to assess expression of these antigens in frozen sections of normal and tumor tissue. Substrate zymography was used to detect plasminogen activator activity in ovarian carcinoma ascites and in conditioned media of cultured cells, whereas a Western blot assay was used to identify urinary-type plasminogen activator receptor in cultured cells. RESULTS: Normal ovarian epithelium expressed urinary-type plasminogen activator receptor (4/4 positive) but little or no urinary-type plasminogen activator (0/4 positive), whereas epithelial ovarian carcinomas frequently expressed urinary-type plasminogen activator (4/8 positive) in conjunction with urinary-type plasminogen activator receptor (7/9 positive). High levels of urinary-type plasminogen activator were detected in 15 of 19 samples of ascites. DOV 13, OVCA 420, OVCA 429, OVCA 432, and OVCA 433 cell lines secreted urinary-type plasminogen activator in variable quantities, whereas normal ovarian epithelial cells did not secrete any detectable plasminogen activator. Urinary-type plasminogen activator receptor had similar levels of expression in all cancer cell lines and normal ovarian epithelium CONCLUSION: Overexpression of urinary-type plasminogen activator is associated with malignant transformation of the ovarian epithelium. Increased cell surface proteolysis mediated by urinary-type plasminogen activator bound to cell surface urinary-type plasminogen activator receptor may contribute to metastatic behavior in ovarian carcinoma. OBJECTIVE: Because elevated expression and cell surface association of urinary-type plasminogen activator have been linked to invasive potential in certain tumor types, we examined the expression of urinary-type plasminogen activator and urinary-type plasminogen activator receptor in ovarian epithelial carcinoma tissues and cells as compared with normal ovarian epithelium. STUDY DESIGN: Monoclonal antibodies specific for urinary-type plasminogen activator and urinary-type plasminogen activator receptor were used for immunohistochemical staining of tissues and cells to assess expression of these antigens in frozen sections of normal and tumor tissue. Substrate zymography was used to detect plasminogen activator activity in ovarian carcinoma ascites and in conditioned media of cultured cells, whereas a Western blot assay was used to identify urinary-type plasminogen activator receptor in cultured cells. RESULTS: Normal ovarian epithelium expressed urinary-type plasminogen activator receptor (4/4 positive) but little or no urinary-type plasminogen activator (0/4 positive), whereas epithelial ovarian carcinomas frequently expressed urinary-type plasminogen activator (4/8 positive) in conjunction with urinary-type plasminogen activator receptor (7/9 positive). High levels of urinary-type plasminogen activator were detected in 15 of 19 samples of ascites. DOV 13, OVCA 420, OVCA 429, OVCA 432, and OVCA 433 cell lines secreted urinary-type plasminogen activator in variable quantities, whereas normal ovarian epithelial cells did not secrete any detectable plasminogen activator. Urinary-type plasminogen activator receptor had similar levels of expression in all cancer cell lines and normal ovarian epithelium CONCLUSION: Overexpression of urinary-type plasminogen activator is associated with malignant transformation of the ovarian epithelium. Increased cell surface proteolysis mediated by urinary-type plasminogen activator bound to cell surface urinary-type plasminogen activator receptor may contribute to metastatic behavior in ovarian carcinoma.

Coordinate expression of urinary-type plasminogen activator and its receptor accompanies malignant transformation of the ovarian surface epithelium

OBJECTIVE: Because elevated expression and cell surface association of urinary-type plasminogen activator have been linked to invasive potential in certain tumor types, we examined the expression of urinary-type plasminogen activator and urinary-type plasminogen activator receptor in ovarian epithelial carcinoma tissues and cells as compared with normal ovarian epithelium. STUDY DESIGN: Monoclonal antibodies specific for urinary-type plasminogen activator and urinary-type plasminogen activator receptor were used for immunohistochemical staining of tissues and cells to assess expression of these antigens in frozen sections of normal and tumor tissue. Substrate zymography was used to detect plasminogen activator activity in ovarian carcinoma ascites and in conditioned media of cultured cells, whereas a Western blot assay was used to identify urinary-type plasminogen activator receptor in cultured cells. RESULTS: Normal ovarian epithelium expressed urinary-type plasminogen activator receptor (4/4 positive) but little or no urinary-type plasminogen activator (0/4 positive), whereas epithelial ovarian carcinomas frequently expressed urinary-type plasminogen activator (4/8 positive) in conjunction with urinary-type plasminogen activator receptor (7/9 positive). High levels of urinary-type plasminogen activator were detected in 15 of 19 samples of ascites. DOV 13, OVCA 420, OVCA 429, OVCA 432, and OVCA 433 cell lines secreted urinary-type plasminogen activator in variable quantities, whereas normal ovarian epithelial cells did not secrete any detectable plasminogen activator. Urinary-type plasminogen activator receptor had similar levels of expression in all cancer cell lines and normal ovarian epithelium. CONCLUSION: Overexpression of urinary-type plasminogen activator is associated with malignant transformation of the ovarian epithelium. Increased cell surface proteolysis mediated by urinary-type plasminogen activator bound to cell surface urinary-type plasminogen activator receptor may contribute to metastatic behavior in ovarian carcinoma. (AM J OBSTET GYNECOL 1994;170:1285-96.)

Overexpression of the tyrosine phosphatase PTP1B is associated with human ovarian carcinomas

OBJECTIVE: Our purpose was to determine whether protein tyrosine phosphatase 1B is overexpressed in ovarian cancers, possibly altering the balance of intracellular tyrosine phosphorylation. STUDY DESIGN: The expression of protein tyrosine phosphatase 1B was assayed in frozen sections from 54 human ovarian carcinomas and seven normal ovaries by immunochemical staining with monoclonal antibody AE4-2J, which is specific for protein tyrosine phosphatase 1B. The expression of protein tyrosine phosphatase 1B - specific messenger ribonucleic acid in tumors was determined by Northern analysis. The results were analyzed statistically by means of Fisher's exact test. RESULTS: Minimal staining was observed in normal ovarian epithelium. In contrast, 43 of 54 (79.6%) tumors displayed increased protein tyrosine phosphatase 1B expression, which is statistically associated with malignancy. Overexpression was associated with the expression of the p185c-erbB-2, p170EGFR, and p165mCSFR growth factor receptor protein tyrosine kinases. Protein tyrosine phosphatase 1B messenger ribonucleic acid expression was inconsistently increased in tumor cells. CONCLUSION: Increased expression of protein tyrosine phosphatase 1B in ovarian cancers that also express protein tyrosine kinases suggests that protein tyrosine phosphatase 1B may play a role in the growth regulation of ovarian cancers. (AM J OBSTET GYNECOL 1994;170:1177-83.)

Prevalence and significance of HER-2/neu expression in early epithelial ovarian cancer.

Although expression of the HER-2/neu oncogene may be of some prognostic importance in advanced ovarian cancer, its role in early-stage disease has not been established. The current study examined the prevalence and significance of HER-2/neu expression in early epithelial ovarian cancer. The authors analyzed the expression of HER-2/neu on frozen tumor specimens from 40 patients with early epithelial ovarian cancer using the indirect immunoperoxidase technique with monoclonal antibodies that detect epitopes on the extracellular domain of the HER-2/neu protein. All patients underwent comprehensive surgical staging. HER-2/neu expression was graded as negative, weak, moderate (1+ to 2+), or strong (3+). Complete clinical data and long-term follow up were available for all patients. The distribution of patients by stage was as follows: Stage IA, 6; IB, 0; IC, 14; IIA, 4; IIB, 6; IIC, 10. The mean patient age was 53 years. Fourteen patients had serous tumors; nine, endometrioid; eight, clear cell; eight, mucinous; and one, undifferentiated. Intratumoral heterogeneity of HER-2/neu expression was observed with most specimens. In eight specimens (20%), some areas of the tumor showed strong (3+) expression, beyond the level that can be seen in normal ovarian epithelium. Twenty-eight specimens (70%) showed moderate (1+ to 2+) staining, whereas four specimens (10%) showed negative or weak staining. At a mean follow-up time among surviving patients of 32 months, 15 patients (37%) have had cancer recurrence. No statistically significant relationship was found between HER-2/neu expression and survival, disease-free survival, stage, or grade. A significant increase was found in 3+ expression of HER-2/neu in clear cell tumors. Consistent HER-2/neu overexpression occurs infrequently in early ovarian cancer, making it unlikely that such overexpression is a general early event in ovarian carcinogenesis. HER-2/neu expression does not appear to be a strong prognostic marker in early epithelial ovarian cancer.

Secretion of extracellular matrix‐degrading proteinases is increased in epithelial ovarian carcinoma

The biochemical events associated with tumor invasion involve localized degradation of the basement membrane by tumor-associated proteinases. In this study, we have characterized the proteinase secretion profiles of 5 ovarian epithelial carcinoma cell lines (DOV 13, OVCA 420, OVCA 429, OVCA 432, OVCA 433) as well as normal ovarian epithelial cells. Immunocapture assays demonstrated that all 5 carcinoma cell lines produce both secreted and surface-associated plasminogen activator. Urinary-type plasminogen activator (u-PA) production was one order of magnitude greater than production of tissue-type plasminogen activator (t-PA). Furthermore, t-PA secretion by normal ovarian epithelial cells was not detectable, whereas u-PA production was 17- to 38-fold lower than in ovarian carcinoma cells. Western-blotting analysis demonstrated that u-PA was secreted as the single chain form (scu-PA) when cells were cultured in serum-free medium. Incubation of plasminogen with ovarian carcinoma cell-conditioned medium resulted in direct activation of the zymogen to plasmin. Furthermore, following incubation of cells with plasminogen, plasmin was eluted from the cell surface, indicating that ovarian carcinoma cells contain binding sites for plasminogen/plasmin which are accessible to surface-associated plasminogen activators. In addition to plasminogen activators, metalloproteinases were also produced by DOV 13, OVCA 429 and OVCA 433 cells. DOV 13 cells produce a 68-kDa metalloproteinase similar to matrix metalloproteinase 2 (MMP-2) whereas a 92-kDa enzyme similar to MMP-9 is secreted by OVCA 429 and 433. Together, ovarian carcinoma-associated plasminogen activators and metalloproteinases catalyze the hydrolysis of the major basement membrane protein components, type-IV collagen, type-IV gelatin, laminin and fibronectin. The enhanced proteolytic capability of ovarian carcinoma cells relative to normal ovarian epithelium suggests a biochemical mechanism by which invasion and spread of ovarian epithelial carcinoma may be mediated.

Prognostic significance of HER-2/ neu expression in advanced epithelial ovarian cancer: A multivariate analysis

We tested the hypothesis that there is prognostic significance to the level of expression of the protooncogene HER-2/neu in advanced ovarian cancer, as prior studies have suggested. We determined expression of HER-2/neu by immunohistochemistry, with monoclonal antibody 9G6 and the indirect immunoperoxidase technique, on frozen tumor specimens from 105 patients with stage III or IV epithelial ovarian cancer. All patients were treated at Memorial Sloan-Kettering Cancer Center, and no patient was lost to follow-up. Median follow-up among surviving patients is 34 months. HER-2/neu expression was scored as negative, weak, 1+, 2+, or 3+. The staining pattern of normal ovarian epithelium was scored negative to 1+. Multivariate analysis was performed to evaluate the prognostic significance of HER-2/neu expression. Twenty-five of the 105 patients (24%) showed strong membrane staining (3+); the other tumor specimens showed weaker membrane staining or no immunoreactivity. There was no correlation of HER-2/neu expression with any of a variety of clinical factors, including stage, grade, cell type, and residual tumor. No significant survival difference was found between patients with levels of staining intensity similar to those of normal ovarian epithelium and those with increased expression (3+). Median survival times were 36 and 27 months, respectively, for the two groups (95% confidence intervals 29 to 45 and 18 to 39 months). Multivariate analysis of possible prognostic factors showed that HER-2/neu overexpression conferred a marginal worsening of survival (p = 0.09) for the subgroup of patients in whom a negative surgical reassessment was not achieved after chemotherapy. HER-2/neu expression does not appear to be an important prognostic factor in patients with advanced epithelial ovarian cancer.

Immunohistochemical localization of c-erbB-2 protein and epidermal growth factor receptor in normal surface epithelium, surface inclusion cysts, and common epithelial tumours of the ovary.

The c-erbB-2 (HER-2/neu) protein is a membrane glycoprotein growth factor receptor showing molecular homology with the epidermal growth factor receptor (EGFR). We examined the immunohistochemical reactivity of monoclonal antibodies against both of these proteins in normal surface epithelium, surface inclusion cysts, and common epithelial tumours of the ovary. The ovarian tumours were classified as benign (16), borderline malignant (2), and malignant (19). Normal surface ovarian epithelium was weakly positive for both c-erbB-2 protein and EGFR. In surface inclusion cysts, however, the epithelial cells lining the lumen exhibited stronger staining for c-erbB-2 protein, but no staining for EGFR. All 16 benign ovarian tumours and the 2 borderline malignant ovarian tumours were positive for c-erbB-2 protein and negative for EGFR. Of the ovarian carcinomas, 13 of the 19 (68.4%) were positive for c-erbB-2 protein and negative for EGFR, while 4 showed positivity for both c-erbB-2 protein and EGFR. Two cases were negative for both proteins. Expression of both c-erbB-2 protein and EGFR was found in endometrioid carcinoma with squamous differentiation and in clinically advanced poorly differentiated serous carcinomas. Expression of c-erbB-2 protein appears to be increased and that of EGFR is reduced in the early stage of epithelial ovarian oncogenesis. The expression of EGFR with c-erbB-2 protein in ovarian carcinoma is related both to histological differentiation and/or advanced clinical stage.

Epidermal growth factor receptor expression in normal ovarian epithelium and ovarian cancer. II. Relationship between receptor expression and response to epidermal growth factor

Previously we have shown that epidermal growth factor acts as a mitogen for some, but not all, ovarian cancer cells in culture. In this study we examined the effect of epidermal growth factor on proliferation of normal human ovarian epithelial cells in monolayer culture. We found that epidermal growth factor stimulated twofold to fourfold increases in proliferation in epithelial cells from each of five normal ovaries (p

Regulation of growth of normal ovarian epithelial cells andovarian cancer cell lines by transforming growth factor-ß

The purpose of this study was to study the role of transforming growth factor-beta in regulation of proliferation of normal and malignant ovarian epithelial cells. We examined production of and responsiveness to transforming growth factor-beta in primary monolayer cultures of epithelial cells from five normal human ovaries and in five ovarian cancer cell lines. In normal ovarian epithelial cells, proliferation always was inhibited by transforming growth factor-beta (greater than 40%) (p less than 0.01). Among the cancer cell lines, proliferation of one was markedly inhibited (greater than 95%) (p less than 0.01), two were only modestly inhibited (15% to 20%) (p less than 0.05), and two were unaffected. In addition, we found that all of the normal ovarian epithelial cells and four of five ovarian cancer cell lines produce transforming growth factor-beta ribonucleic acid and protein. These data suggest that transforming growth factor-beta may act as an autocrine growth inhibitory factor for normal ovarian epithelium in vivo. Because most of the ovarian cancer cell lines are relatively resistant to the growth inhibitory effect of transforming growth factor-beta and because one cell line does not produce transforming growth factor-beta, it is possible that loss of the transforming growth factor-beta pathway may play a role in the development of some ovarian cancers.

Epidermal growth factor receptor expression in normal ovarian epithelium and ovarian cancer: II. Relationship between receptor expression and response to epidermal growth factor

Previously we have shown that epidermal growth factor acts as a mitogen for some, but not all, ovarian cancer cells in culture. In this study we examined the effect of epidermal growth factor on proliferation of normal human ovarian epithelial cells in monolayer culture. We found that epidermal growth factor stimulated twofold to fourfold increases in proliferation in epithelial cells from each of five normal ovaries (p less than 0.01). In addition, Scatchard analysis of binding of epidermal growth factor tagged with iodine 125 indicated the presence of high-affinity receptors in all of the ovarian epithelial cells and ovarian cancer cell lines. The number and affinity of receptors was similar in the normal epithelium and cancer cell lines, and there was no relationship between epidermal growth factor receptor number and responsiveness to epidermal growth factor. We conclude that human ovarian epithelial cells normally express epidermal growth factor receptors and that epidermal growth factor acts as a mitogen for these cells. Although the mitogenic response to epidermal growth factor often is attenuated in ovarian cancer cell lines, loss of responsiveness to epidermal growth factor does not appear to be due to decreased receptor expression.

Epidermal growth factor receptor expression in normal ovarian epithelium and ovarian cancer: I. Correlation of receptor expression with prognostic factors in patients with ovarian cancer

Previous studies in breast and bladder cancer have suggested that epidermal growth factor receptor is expressed by only a proportion of cancers and is associated with poor clinical outcome. We used a monoclonal antibody specifically reactive with the extracellular domain of the epidermal growth factor receptor to localize this receptor immunohistochemically in frozen sections of normal ovary and epithelial ovarian cancer. Normal ovarian epithelium was found to express epidermal growth factor receptor in all cases. Among 87 ovarian cancers, however, 23% did not express immunohistochemically detectable receptor. Epidermal growth factor receptor expression was not related to histologic grade or stage, but was associated with poor survival (p less than 0.05). The median length of survival of patients with tumors that did not express epidermal growth factor receptor was 40 months compared with 26 months in patients with tumors that did express epidermal growth factor receptor. As in breast and bladder cancer, expression of epidermal growth factor receptor in ovarian cancer appears to be a poor prognostic factor.

Regulation of growth of normal ovarian epithelium and ovarian cancer cell lines by transforming growth factor-β

Regulation of growth of normal ovarian epithelium and ovarian cancer cell lines by transforming growth factor-β

Heterogeneity of antigen expression in benign and malignant breast and ovarian epithelial cells

Expression of II distinct antigen families has been studied in normal and malignant epithelial cells from breast and ovary by avidin-biotin immunoperoxidase staining of frozen tissue sections. Each of the antigens was expressed in a fraction of the normal and malignant breast tissues from different individuals. Among breast and ovarian cancers, a similar fraction of tumors expressed each of the determinants. An epitope on a 42-kDa glycoprotein was expressed significantly more often by ovarian carcinomas than by benign ovarian epithelium. Several determinants on 66-, 240-kDa or high-molecular-weight proteins or glycoproteins were expressed significantly more often on benign breast epithelium than on normal ovarian epithelium. Substantial heterogeneity in antigen expression was observed among different tumor specimens. Each of 14 epithelial ovarian cancers expressed a distinctive combination of antigens. Among 18 breast cancers, 16 distinct antigenic phenotypes were observed. Interestingly, comparable heterogeneity was observed in the expression of epitopes on benign breast and ovarian epithelium, compatible with the possibility that the monoclonal reagents recognized polymorphic determinants. Substantial variations in antigen expression were also observed among cells within each neoplasm. However, when antibodies against 5 different determinants were used in combination, a majority of cells could be stained intensely in all 13 breast carcinomas tested and more than 95% of tumor cells could be stained in 10 of the 13.

Expression of MHC products and leucocyte differentiation antigens in gynaecological neoplasms: An immunohistological analysis of the tumour cells and infiltrating leucocytes

Monoclonal antibodies directed against monomorphic determinants of Class I and Class II products of the major histocompatibility complex (MHC) and against leucocyte differentiation antigens were used in an indirect immunoperoxidase technique to compare their expression in normal and malignant disease of the ovary, cervix and endometrium. MHC Class I products, strongly expressed on normal ovarian epithelium, were uniformly absent from 7/8 ovarian carcinomas of varying histology. Lack of Class I expression was also a feature of 6/10 cervical carcinomas and of 4/8 endometrial carcinomas, in comparison with their respective normal tissues. Relative to normal tissue epithelium MHC Class II products, could be either lost or gained, the pattern of expression being either uniform or heterogeneous. Leucocytes were sparse in normal ovary but more numerous in cervix and endometrium. In tumours, with few exceptions, they were abundant, though usually confined to the stroma. T cells, largely of cytotoxic/suppressor (OKT8) phenotype, tended to predominate though in some tumours, particularly cervical carcinoma, large numbers of macrophages and to a lesser extent, B cells were sometimes detected. By contrast, leucocytes of natural killer (NK) phenotype were virtually non-existent in any tumour or normal tissue. The ingress of leucocytes into gynaecological neoplasms does not appear to be a random event and may be evoked by an immune response against tumour-associated antigens. However, the relationship between in situ mononuclear cell infiltration and MHC expression on epithelial tumour cells is complex and remains to be elucidated.

Characterization of the Specificity by Immunohistology of a Monoclonal Antibody to a Novel Epithelial Antigen of Ovarian Carcinomas

An immunohistological study, using the avidin-biotin-peroxidase complex method, was carried out to define the reactivity profile of a murine monoclonal antibody, MOv2, which recognizes a novel glycoprotidic antigen associated with ovarian epithelial tumors. Among the primary ovarian tumors tested, MOv2 immunostained 93% of mucinous and 75% of serous cystadenomas, 100% of mucinous, 81% of serous and 73% of endometrioid carcinomas. Undifferentiated and clear cell tumors revealed more limited reactivity with the antibody, whereas ovarian sex cord-stromal and germinal tumors were immunonegative. Positive reactions were also documented in omental metastases from primary ovarian carcinomas. No immunoreactivity was detected in normal ovarian epithelium, whereas the cells lining Walthard's nests adjacent to the fallopian tubes and a variety of normal epithelia were consistently immunolabeled. These included the lining epithelia of the gastrointestinal tract, bronchi and endocervix, and the epithelium of salivary, biliary and pancreatic ducts and sweat glands. To a lesser extent, positive reactions were detected in other surface epithelia, such as squamous and transitional epithelia. Among tumors other than ovarian, MOv2 consistently reacted with adenocarcinomas and squamous cell carcinomas from different sites, most notably breast, lung and gastrointestinal tract, and with transitional cell carcinomas. In contrast, no staining was demonstrated in non-epithelial malignancies. The antigen defined by MOv2 may be operationally useful as a marker of epithelial lineage in tumor histopathology. Its pattern of immunohistochemical distribution indicates that an antigenic phenotype shared by normal surface epithelia and non-ovarian carcinomas is strongly associated with common epithelial neoplasms of the ovaries.