Abstract A26: PTEN and colonization of the ovary in metastasis of fallopian tube-derived ovarian cancer
Authors: Dean Matthew , Jin Vivian , Russo Angela , Burdette Joanna E. 
Published: August 2018

Abstract
It is now clear that high-grade serous ovarian cancer (HGSOC) can originate in the fallopian tube epithelium (FTE), making the role of the ovary in these tumors unclear. Therefore, the objectives of this research were to 1) determine if colonization of the ovary was an important step in the spread of FTE-derived tumors to the peritoneum and 2) explore the role of ovarian rupture, independent of hormonal changes, in tumor cell colonization of the ovary. First, 50,000 murine oviductal epithelial (MOE) cells stably expressing PTENshRNA+KRASG12V were allografted into the left ovarian bursa (IB) or peritoneal space (PS). IB-grafted mice developed tumors, with all mice requiring sacrifice within 90 days. Tumors were detected throughout the peritoneum and in the contralateral ovary. In contrast, PS-grafted mice failed to develop tumors for the duration of the experiment (150 days). To determine if rupture of the ovary was important in colonization of the ovary, an ex vivo attachment assay was developed in which ovaries was left intact or cut with a scalpel to rupture the ovarian surface (ovulation mimetic) without associated hormonal changes. Ovulation mimetic ovaries had significantly more MOE GFP and OVCAR8 RFP cells attached, particularly in the wounded areas. To confirm that increased attachment was due to exposure of the ovarian stroma, MOE GFP cells were plated on monolayers of murine ovarian surface epithelium (MOSE), monolayers of murine ovarian stroma (MOST), or three-dimensional (3D) type I collagen gels (the primary extracellular matrix protein in the ovarian stroma). MOE GFP cells attached to MOST more than MOSE and to 3D collagen than MOST, supporting the contention that FTE cells adhere to the extracellular matrix (ECM), such as collagen, exposed during ovulation. We next sought to test the ability of cells to survive in an environment rich in 3D collagen. 3D collagen reduced the viability of normal epithelial cells (MOSE, MOE, and IOSE80) but had no effect on HGSOC cells (OVCAR3, OVCAR4, OVCAR5, OVAR8, and OVSAHO), indicating that transformation of FTE is important for survival in the collagen-rich ovarian stroma. RNA-seq showed that 3D collagen decreases genes associated with focal adhesion, regulation of actin cytoskeleton, and ECM-receptor interaction, supporting the idea that normal epithelial cells respond poorly to a microenvironment rich in 3D collagen. In contrast, genes associated with p53 signaling and suppression of MAPK/AKT were increased when cells were grown on 3D collagen. However, plating cells on 3D collagen did not lead to stabilization of p53 and expression of mutant p53 had no effect on viability of MOE cells on 3D collagen. KRASG12V tended to increase viability, but only PTENshRNA completely rescued viability on 3D collagen as compared to 2D. PTENshRNA had no effect on adhesion to 3D collagen, but it did increase invasion through collagen. Cells expressing PTENshRNA showed increased attachment to 3D ovaries that were wounded to mimic ovulation; however, the AKT inhibitor MK2206 did not reduce this adhesion. Interestingly, after 7 days of culture on 3D collagen, MOE PTENshRNA cells formed spheroids while controls did not, leading to the hypothesis that spheroids may mediate metastasis from the fallopian tube to the ovary. In support of this contention, spheroids were observed in the fallopian tube of PAX8+/cre PTENflox/flox mice, which develop fallopian tube-derived ovarian cancer. Furthermore, OVCAR8 RFP spheroids preferentially attached to wounded sites mimicking ovulation relative to intact ovaries. In conclusion, FTE cells attached to exposed ECM of the ovary and colonization of the ovary enhances further spread of FTE-derived tumors beyond the reproductive tract. Inhibition of ovarian colonization may be a new approach to inhibit peritoneal spread of cancer in women at high risk of developing HGSOC.
Citation Format: Matthew Dean, Vivian Jin, Angela Russo, Joanna E. Burdette. PTEN and colonization of the ovary in metastasis of fallopian tube-derived ovarian cancer. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr A26.