Abstract POSTER-TECH-1108: A novel spontaneously transformed model of early-stage serous cancer from murine oviductal epithelial cells

Abstract

Abstract High-grade serous carcinoma (HGSC) is the most lethal gynecological malignancy. Many studies have implicated the ovarian surface epithelia as the origin of HGSC origin. Newer studies suggest other sources of HGSC, such as the distal fallopian tube (oviduct in mice). Current models have to use multiple genetic manipulations of either isolated human fallopian tube secretory cells (FTSECs) or in transgenic mice to form HGSC-like tumors. While these models are vital for identifying and understanding the key players in HGSC, they do not provide information regarding the step-wise development of HGSC. This information is critical for earlier diagnosis and treatments, since most women develop HGSC spontaneously (85%-90% of cases). Thus, development of a spontaneous transformation model offers the potential to interrogate, in a step-wise manner, premalignant changes that initially drive HGSC progression. A murine oviductal epithelial (MOE) cell line was developed from oviducts of CD-1 mice that were validated using known markers: oviductal glycoprotein 1, Pax8, and acetylated tubulin. The low passage cells, designated MOELOW, (<25 passages) contain both epithelial cell subtypes, ciliated and secretory cells. A subpopulation of the MOELOW cells was then continuously passaged in culture to simulate the aging process, designated MOEHIGH (<85 passages). Molecular comparisons were done by Western blot and next-generation RNA sequencing (RNA-Seq). These cell lines were compared using cellular assays, like proliferation, cell death, 2D foci formation, soft agar growth, and xenografted into nude mice to evaluate tumorgenicity. As compared to MOELOW cells, MOEHIGH cells were significantly more proliferative, resistant to damage by bulky DNA adducts, displayed increased clonogenicity, and anchorage-independent colony formation. RNA-Seq analysis elicited 7,425 differentially expressed transcripts. Database comparisons uncovered several well-known pathways altered in HGSC and a few unique pathways altered upon MOE transformation. Consistent with the results from The Cancer Genome Atlas (TCGA) both the Rb and FoxM1 pathways were altered with increased expression of cyclin E1, c-Myc, PCNA; and reduced expression of p16INK4A and hyperphosphorylation of Rb. Remarkably, the MOEHIGH cells do not contain a mutation in Trp53, the HGSC hallmark expressed in 96% of all cases, but rather an alternatively spliced transcript. This p53 transcript, termed p53AS, has both a frameshift and truncation within the negative-regulation domain at the C-terminus. This region loses the lysine residues for MDM2-mediated degradation thereby stabilizing the protein. When wild-type p53 is transiently transfected back into MOEHIGH cells, the cells still cannot induce p21 expression, functioning similar to a p53 DNA-binding mutation. Nude mice injected subcutaneously with MOEHIGH cells all formed tumors but MOELOW mice did not. MOEHIGH forms densely packed and necrotic tumors, stained for Ki67, Pax8, and OVGP1. Intraperitoneal injections of MOEHIGH cells did not metastasize in any mice. This spontaneous transformation model allows for the evaluation of the stepwise progression of HGSC and since tumors are not metastatic, this will be one of the oviductal-derived first early stage serous carcinoma cell lines. Citation Format: Michael P. Endsley, Georgette Moyle-Heyrman, Suzanne M. Quartuccio, Daniel D. Lantvit, Joanna E. Burdette. A novel spontaneously transformed model of early-stage serous cancer from murine oviductal epithelial cells [abstract]. In: Proceedings of the 10th Biennial Ovarian Cancer Research Symposium; Sep 8-9, 2014; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(16 Suppl):Abstract nr POSTER-TECH-1108.