Abstract 2880: Targeting Rho GTPases in ovarian clear cell cancer
Authors: Arildsen Nicolai S. , Hedenfalk Ingrid 
Published: August 2018

Background: Ovarian clear cell carcinomas (OCCC) constitute a rare subtype of epithelial ovarian cancer with distinct clinical features. Resistance to first-line platinum-based treatment is common. We recently reported deregulated Rho GTPase signaling in OCCC. Rho GTPases are small signaling G proteins, and a subfamily of the RAS superfamily. Rho GTPases are reported to regulate actin organization and cytoskeletal organization. We hypothesized that inhibiting Rho GTPases would have a potent cytotoxic effect on OCCC cell lines.

Materials and Methods: Sulphorhodamine B (SRB) assays were used to assess the antiproliferative effects of 72h of treatment with the cholesterol-lowering agent simvastatin, the pan GTPase inhibitor CID1067700 (CID) and carboplatin (as control) in the two OCCC cell lines JHOC5 and OVMANA. Single effects (IC50) and combination effects were calculated using the Chou-Talalay method. We assessed cytoskeletal integrity using phalloidin staining of actin and DAPI for cell death. Western blotting was used to assess the expression of cleaved PARP (cPARP) after 4h and 24h of treatment.

Results: All three drugs had similar IC50 values in both OCCC cell lines; however, they were significantly different from the high-grade serous ovarian cancer control cell line CAOV3. JHOC5 and OVMANA were more resistant to carboplatin (IC50: 65.3 µM and 76.2 µM, respectively) compared to CAOV3 (IC50: 16.4 µM). JHOC5 and OVAMANA were more sensitive to simvastatin and CID (JHOC5: 6.7 µM/69.2 µM, OVMANA: 10.3 µM/84.9 µM, simvastatin/CID1067700) compared to CAOV3 (27.7 µM/122.5 µM). The combination index (CI) for either CID or simvastatin in combination with carboplatin was moderately antagonistic for all cell lines (CI: 1.6-2.4). Combinations of CID and simvastatin were mildly to moderately synergistic (CI: 0.6-0.8) for all cell lines. Simvastatin and CID were found to interfere with actin organization, compared to carboplatin, with simvastatin also interfering with cell membrane integrity. A small increase in cPARP was observed after 24h of treatment with all drugs in single regimens.

Conclusion: Cytotoxic effects of simvastatin and CID both in single regimens or in combination were achieved with lower doses in OCCC cell lines than in the high-grade serous cells; however, combinations of simvastatin and CID were found to be mildly synergistic in all cell lines. Simvastatin was found to be more potent than CID. Interestingly, all combinations with carboplatin were found to be moderately antagonistic. Actin organization was more extensively disrupted with simvastatin than CID. An initial assessment of cell death mechanisms suggested a mechanism working through cleaved PARP. However, these findings provide a basis for further studies into the molecular mechanisms of simvastatin and CID in OCCC. These studies are currently under way, including an in-depth assessment of the underlying signaling and cell death mechanisms.

Citation Format: Nicolai S. Arildsen, Ingrid Hedenfalk. Targeting Rho GTPases in ovarian clear cell cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2880.