Abstract 5021: A 3D model of genetic transformation of normal ovarian epithelial cells identifies candidate genes associated with the initiation and development of epithelial ovarian cancer
Authors: Simon A. Gayther, Kate Lawrenson, Maria Notaridou, Elizabeth Benjamin, Ian J. Jacobs, Dimitra Dafou
Journal: Cancer Research (2010)

Abstract Novel biomarkers associated with early-stage ovarian cancer (OC) development may represent susceptibility makers that initiate ovarian cancers or screening markers for early disease detection. Little is known about the biology underlying OC development and heterogeneity. It has been hyporthesis that OCs can arise from the ovarian surface epithelium (OSE), a monolayer of epithelial cells covering the surface of the ovary. We have shown that immortalised OSE cells (IOSE) bypass replicative senescence but show no other features of neoplastic transformation. Here, we established an in vitro three-dimensional (3D) genetic model of early-stage ovarian carcinogenesis. By first overexpressing the CMYC oncogene, and then expressing mutant alleles of the KRAS or BRAF oncogenes in IOSE cells, we were able to create a stepwise model of neoplastic transformation of OSE. Overexpression of CMYC in IOSE cells (IOSE-CMYC) induces a significant increase in anchorage-dependent and -independent growth, enhanced progression through the cell cycle and decreased apoptosis. Subsequent expression of mutant KRAS-G12V or BRAF-V600E alleles in IOSE-CMYC cells caused differential rates of anchorage-independent growth, invasiveness and proliferation showing that the different genetic elements mimics the phenotypic heterogeneity seen in primary ovarian tumors in vivo. 3D models of oncogene-expressing clones revealed many characteristics of malignant cells in vivo that could not be detected in 2D monolayer cultures. Gene expression microarray profiling of the different stages of this model identified several candidate tumour suppressor genes that are associated with neoplastic transformation. These included: THBS1, an inhibitor of angiogenesis in several cancer types; FEZ1, which is involved in cell growth and shows loss of expression in ∼40% of OCs; and RGS4, (a regulator of G-protein signaling) shown to promote breast cancer migration and invasion. Novel activated genes were also identified including PITX1, previously shown to be overexpressed in a murine model of ovarian granulosa cell tumours; and HIST1H4C, which is a overexpressed in therapy related myloid leukemias. We used the model to evaluate the functional role of genes that have recently been shown to increase susceptibility to ovarian cancer using genome wide association studies. One locus is 8q24, near the CMYC oncogene, highlighting its significance in OC development. Two other loci are BNC2 at 9q21 and TiPARP at 3q25. Both genes were down-regulated in response to CMYC over-expression and showed significant loss of function with further transformation after KRAS-G12V activation (p=7.8 × 10-7 and p=6.3 × 10-4, respectively). This suggests both behave as tumor suppressor genes. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. 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 5021.