Abstract GMM-058: CBX2 IDENTIFIED AS DRIVER OF ANOIKIS ESCAPE AND DISSEMINATION IN HIGH GRADE SEROUS OVARIAN CANCER
Authors: Lindsay J. Wheeler, Zachary L. Watson, Lubna Qamar, Alexandra McMellen, Tomomi Yamamoto, Kian Behbakht, Benjamin G. Bitler
Journal: Clinical Cancer Research (2019)

Abstract The vast majority of high grade serous ovarian carcinomas (HGSOC) are diagnosed at an advanced stage. Chromobox 2 (CBX2), a polycomb repressor complex subunit, plays an oncogenic role in a variety of cancers. In prostate cancer, CBX2 is a driver of metastatic progression. Little is known about the role of CBX2 in HGSOC. Our hypothesis is that CBX2 upregulation promotes advanced HGSOC by promoting a stem-like transcriptional profile and inhibiting anoikis. Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) were queried to establish the role of CBX2 in HGSOC. In vitro evaluation of CBX2 occurred in OVCAR4, PEO1, and OVCAR8 HGSOC cell lines. PolyHEMA-coated plates forced cells to grow in suspension and simulated anoikis-escape. Quantitative polymerase chain reaction and immunoblots evaluated CBX2 expression. Small hairpin RNAs (shRNAs) knocked CBX2 down for loss of function studies. To mimic HGSOC progression several culture conditions, including 2D, colony formation, and 3D, spheroid, were examined. Secreted Gaussia luciferase (gLuc) activity was utilized as an indicator of proliferation. Stemness was tested with the Aldefluor assay, measuring aldehyde dehydrogenase activity (ALDH). Using patient tumors derived from the Gynecology Tissue and Fluid Bank (GTFB) and a HGSOC tissue microarray (TMA) with matched primary, metastatic, and lymph nodes, a CBX2 expression profile was established. Student's t-test was used to define statistical significance, with a p value of &amp;lt; 0.05. Analysis of GEO databases established CBX2 is upregulated in HGSOC tumors compared to benign tissues. Analysis of TCGA found CBX2 expression conveyed worse disease-free survival (11.7 vs 17.6 months, Log-rank test p-value &amp;lt; 0.005) and overall survival (34 vs. 44.8 months, Log-rank test p-value &amp;lt;0.005). Examination of primary HGSOC tumors confirmed CBX2 was upregulated at the protein level in HGSOC compared to benign tissue. In vitro, OVCAR4, PEO1, and OVCAR8 cells upregulate CBX2 when grown in suspension compared to adherent conditions. CBX2 knockdown led to a significant inhibition of proliferation in 2D, 3D, and in suspension. Forced suspension promoted increased ALDH3A1 expression and ALDH activity. CBX2 knockdown led to a decrease in both ALDH3A1 expression and ALDH activity. HGSOC cells grown in suspension were found to be more chemoresistant compared cells grown under adherent conditions. Similarly, knockdown of CBX2 sensitized HGSOC cells to cisplatin. Examination of primary tissue in a TMA of matched patient samples revealed CBX2 is expressed in primary and metastatic disease. We conclude that CBX2 directly impacts proliferation and is overexpressed in HGSOC. Our work indicates that CBX2 is an important regulator of stem-ness, which could play a role in anoikis escape, HGSOC dissemination, and chemoresistance, suggesting that CBX2 may be associated with more advanced disease. This exploration expands our understanding of molecular drivers of HGSOC progression and potentially identifies a novel therapeutic target. Citation Format: Lindsay J. Wheeler, Zachary L. Watson, Lubna Qamar, Alexandra McMellen, Tomomi Yamamoto, Kian Behbakht, and Benjamin G. Bitler. CBX2 IDENTIFIED AS DRIVER OF ANOIKIS ESCAPE AND DISSEMINATION IN HIGH GRADE SEROUS OVARIAN CANCER [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr GMM-058.