Orchestrated expression of the atypical RhoGTPase, RHOV, in response to matrix detachment of ovarian cancer cells

Abstract Ovarian cancer (OVCA) metastasis occurs through the process of transcoelomic spread, where cells disseminate into the peritoneal fluid and form multi-cellular aggregates (MCA) that mediate Anchorage-Independent (A-I) survival and invasion of the peritoneal organs. Isolation of MCAs from patient ascites has been extensively reported in the literature and found to correlate with poor patient survival and resistance to therapy. However, the mechanisms promoting OVCA MCA formation and survival in A-I are not fully understood. We previously demonstrated that MCAs upregulate their mitochondrial antioxidant defenses to maintain optimal survival in A-I, and that this is an early event following cellular detachment. To identify additional key molecular players promoting MCA formation and fitness in A-I, we employed sequential RNA seq analysis to compare the transcriptome of cells in attached, early A-I, and late A-I conditions using the serous cell line OVCA433. Analysis of early transcriptomic changes observed within 2hrs of A-I revealed that RHOV, a recently identified atypical member of the Rho GTPase family, is the top gene significantly upregulated in early A-I. Moreover, we found that RHOV expression rapidly declines in later A-I timepoints indicative of a tight temporal regulation of RHOV expression following cellular detachment. Next, we compared the expression of RHOV in primary ovarian tumor cells and matching malignant ascites cells from the publicly available dataset (GEO: GSE85296) and found increased RHOV expression specifically in the metastatic ascites of ovarian cancer patients when compared to the attached tumor. Previously, RHOV expression was found to regulate intercellular adhesion dynamics during specific stages of neural crest development, a physiological process mimicking epithelial to mesenchymal transition (EMT). RHOV has also been reported to be overexpressed in lung cancer where it was found to correlate with poor patient outcome and resistance to therapy. However, the role of RHOV in ovarian cancer metastasis remains unexplored. To test the consequences of increased RHOV transcription in A-I: its role in migration, aggregate formation, and anoikis resistance is being tested following silencing of RHOV expression using both CRISPR and siRNA. Current work is exploring how RHOV is rapidly transcribed following matrix detachment and how increased expression alters cellular signaling events in early A-I stages to promote optimal MCA fitness. This work aims to contribute new knowledge on the novel oncogenic role of an understudied member of the Rho family of GTPases, RHOV. Moreover, given that increased MCA fitness has been correlated with increases metastatic potential and resistance to therapy, we provide a unique proof-of-concept study to target key components of early MCA adaptations as a novel therapeutic strategy for prolonging survival rates in OVCA patients diagnosed in metastatic stages. Citation Format: Amal T. Elhaw, Yeon Soo Kim, Zaineb Javed, Priscilla Tang, Weihua Pan, Nadine Hempel. Orchestrated expression of the atypical Rho-GTPase, RHOV, in response to matrix detachment of ovarian cancer cells [abstract]. In: Proceedings of the AACR Special Conference: Cancer Metastasis; 2022 Nov 14-17; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_2):Abstract nr A034.