RHOV is a Detachment-Responsive Rho GTPase Necessary for Early Metastatic Reprogramming in Ovarian Cancer

Transcoelomic dissemination is the predominant route of ovarian cancer metastasis and a major driver of patient mortality. In our pursuit to identify novel tumor cell adaptations necessary for metastatic spread, we identified a conserved transcriptomic signature activated shortly after matrix-detachment across multiple ascites-derived cell lines and patient specimens. Within this signature, RHOV, an atypical and constitutively active Rho GTPase, emerged as the top upregulated gene. We show that RHOV is essential for anoikis resistance, cellular aggregation in anchorage-independence, migration, invasion, in vivo metastatic colonization, and is elevated in patient omental metastases. Mechanistically, RHOV integrates c-Jun dependent cytoskeletal remodeling with early endosomal trafficking to support pro-metastatic signaling, which is dependent on both RHOV's GTPase activity and membrane localization. For the first time this study identifies RHOV as a necessary mediator of ovarian cancer metastasis, and highlights the potential of immediate early detachment-responsive transcriptional changes as promising therapeutic targets in metastatic disease. Overall design: To investigate the role of RHOV in metastasizing ovarian cancer cells, we performed bulk RNA sequencing on OVCA433 cells following CRISPR/Cas9-mediated RHOV knockout in a validated RHOV-KO clone (#D4). Wild-type and RHOV-KO (D4) cells were cultured under three conditions: adherent (2D), early anchorage independence (3D-2Hrs post-detachment), and late anchorage independence (3D-24Hrs post-detachment); RNA from each condition was extracted and sequenced.