Expression of E-cadherin, alpha-catenin and beta-catenin in normal ovarian surface epithelium and epithelial ovarian cancers.

AIMS: To study the expression of the epithelial adhesion molecule E-cadherin and its associated proteins alpha and beta catenin in paraffin sections of normal ovaries, benign cystadenomas and ovarian carcinomas, and in immortalized normal ovarian surface epithelial cells and ovarian carcinoma cells in culture. METHODS AND RESULTS: Immunocytochemistry was used to study expression of the proteins in paraffin sections and western blotting was used to determine levels of expression of the proteins in cell extracts. E-cadherin expression was found to be absent in ovarian surface epithelial cells in culture and infrequently expressed in normal ovarian surface epithelial cells in vivo, although apical punctate staining was occasionally seen. Seven of nine benign cystadenomas and 29/34 epithelial ovarian carcinomas showed some expression of E-cadherin, but expression was absent in poorly differentiated tumours. Expression of alpha and beta catenin was consistently detected on the lateral membranes of normal ovarian epithelium and benign cystadenomas. alpha and beta catenin expression was lost in 18% and 21% of ovarian carcinomas, respectively: other ovarian carcinomas expressed these proteins at a reduced level. A small number of these tumours showed a diffuse cytoplasmic rather than membranous staining. Reduced staining for alpha and beta catenin appeared to correlate with a more spindly, less adhesive morphology and increased invasive potential in matrigel. CONCLUSIONS: The results suggest that E-cadherin expression is generally induced in well differentiated ovarian cancers. In contrast, alpha and beta catenins are consistently expressed in the normal ovarian surface epithelium and benign tumours, but are sometimes reduced or absent in ovarian carcinomas. It is likely that the catenins associate with membrane proteins other than E-cadherin in ovarian epithelium, and they may possibly function as tumour suppressors in this epithelium.

Transfection of rat ovarian surface epithelium with erb-B2/neu induces transformed phenotypes in vitro and the tumorigenic phenotype in vivo.

The neu/cerb-B2 gene is frequently amplified and/or overexpressed in human epithelial ovarian cancers. We have established an inbred animal model for ovarian cancer that mimics aspects of human ovarian cancer by transducing a spontaneously immortalized rat ovarian surface epithelial cell line in culture with ecotropic retroviruses expressing a mutated rat neu/c-erb-B2 oncogene. Transfectants expressing neu at a high level exhibited altered morphology and behavior in two-dimensional and three-dimensional culture in Matrigel, could be cloned in soft agar, and were more invasive through a Matrigel membrane than control transfectants transduced with a similar retrovirus expressing the beta-galactosidase gene. When injected intraperitoneally, neu-expressing transfectants produced highly invasive, rapidly growing tumors that coated the peritoneal cavity and induced ascites formation. Furthermore, neu transfectants could be grown as solid tumors when injected subepithelially into the ovary. The neu-transfected cells also formed tumors when injected subcutaneously into the mammary fat pad, although they grew relatively poorly and often regressed. Transfectants expressing beta-galactosidase failed to produce tumors at any of the sites injected. A second rat ovarian surface epithelial cell line was similarly transduced with the neu/c-erb-B2-expressing retrovirus. However, transformed phenotypes and tumorigenicity were not induced in this cell line. These experiments show directly that overexpression of neu in an established line of rat ovarian epithelium is extremely oncogenic. This animal model system may prove useful for the study of ovarian cancer biology in immunocompetent animals.

Overexpression of cyclin D1 in epithelial ovarian cancers.

Amplification and overexpression of the cell cycle-related gene cyclin D1 have been demonstrated in several human malignancies and have been shown to be directly oncogenic in breast epithelium and lymphocytes. Overexpression of the gene can occur in the absence of gene amplification. We have investigated whether cyclin D1 is overexpressed in a panel of 43 sporadic epithelial ovarian cancers using immunohistochemistry. Cyclin D1 was overexpressed in 26% of these tumors. Overexpression of cyclin D1 is associated with borderline or well-differentiated, grade 1 tumors but does not correlate with a particular histological type, overexpression of the c-erb-B2 oncogene, or presence of estrogen receptors. It is suggested that overexpression of cyclin D1 may contribute to the pathogenesis of epithelial ovarian cancers, including a subset of tumors different from those overexpressing the c-erb-B2 oncogene.

A differential PCR assay for the detection of c-erbB 2 amplification used in a prospective study of breast cancer.

AIMS: To establish a robust differential polymerase chain reaction (PCR) assay for the detection of c-erbB 2 amplification in breast cancer that can be used in a routine pathology laboratory. Once established, the assay was used in a prospective study of breast tumours to investigate the relation between c-erbB 2 amplification and both recognised prognostic features and short term clinical outcome. METHODS: The differential PCR was used for the co-amplification of c-erbB 2 and a reference gene from 48 tumour DNA samples and control DNA samples. The ratio of the two genes was determined by image analysis of the PCR products electrophoresed on a highly resolving agarose gel. RESULTS: The differential PCR assay was shown to be accurate and reproducible using the conditions outlined. Twenty six per cent of the breast cancer patients were shown to have c-erbB 2 amplification in their tumour biopsies. Twenty eight per cent of the patients died of their disease or had disease recurrence during the follow up period and 73% of these patients had amplification of c-erbB 2. CONCLUSIONS: A significant association was found between c-erbB 2 amplification and early disease recurrence. This assay could be used to provide a marker for poor prognosis in breast cancer.

Cyclin D1 amplification and expression in human breast carcinoma: correlation with histological prognostic markers and oestrogen receptor expression.

Aims-To study the amplification of the Cyclin D1 gene (CCND1) in human breast carcinoma; to relate this to Cyclin D1 protein expression; to relate these parameters to recognised pathological prognostic factors, including oestrogen receptor (ER) status.Methods-DNA extracted from frozen sections of breast tumours (n = 36) was used for Southern blotting. Probes for CCND1, c-myc and the immunoglobulin heavy chain locus (IgH) were hybridised to tumour DNA. Immunocytochemical expression of Cyclin D1 protein and ER was studied in paraffin wax sections from the same tumours.Results-Amplification of CCND1 was observed in 11% (four of 36) of tumours studied. Over expression of Cyclin D1 protein was observed in 73% (30/41) of tumours. There was no correlation between recognised histological prognostic markers and either gene amplification or expression. However, a weak association was seen between Cyclin D1 expression and ER status.Conclusions-A disparity exists between locus amplification and over expression of Cyclin D1, suggesting the existence of another mechanism for raised protein expression. No significant correlation was detected between either Cyclin D1 amplification or over expression and established prognostic markers.