Abstract 1443: RhoE is underexpressed in hepatocellular carcinoma and suppresses cell motility and invasiveness

Abstract Hepatocellular carcinoma (HCC) is a major malignancy worldwide and is the second commonest fatal cancer in Southeast Asia and Mainland China including Hong Kong, due to the high prevalence of hepatitis B viral infection. Despite definite improvements in the outcome of patients with HCC, the overall prognosis of this cancer is still unsatisfactory because of late presentation and frequent tumor recurrence after surgical resection. In this regard, knowledge of the molecular and cellular targets underlying the development and progression of liver cancer may provide novel opportunities for therapeutic interventions for this cancer. As part of our continual study on the Rho/ROCK pathway in HCC, in this current study, we aimed to find out the role of RhoE (also known as Rnd3), a member of the Rho family of small GTP-binding proteins, in HCC. We have found that RhoE was frequently underexpressed in 43/71 (60.6%) of our human HCCs at mRNA level using qPCR, as compared with their corresponding non-tumorous livers (p <0.001). Moreover, RhoE knockdown in HCC cells by lentiviral infection of shRhoE enhanced cell migration and invasion in vitro. Using transwell migration assay, we showed that stable shRNA knockdown of RhoE in SMMC7721 and BEL7402 HCC cell lines resulted in faster migration of the cells than their corresponding non-target controls. Moreover, stable RhoE knockdown clones also invaded through the matrigel faster in matrigel invasion assay. Previous studies have indicated that phosphorylation of myosin phosphatase target subunit 1 (MYPT1) at Thr850 was essential for phosphorylation of myosin II and crucial for cell contractility. To further delineate the roles of RhoE on actomyosin contractility, the MYPT1 phosphorylation levels in the Rho stable knockdown transfectants in SMMC, BEL and Hep3B HCC cells were assessed by Western blotting and found to be increased as compared with their non-target control. This increased phosphorylation was reversed by ROCK inhibitor Y27632. Furthermore, the ROCK inhibitor Y27632 was able to suppress the accelerated cell migration brought by the knockdown of RhoE in SMMC, suggesting that RhoE may exert its effects by inhibiting the activity of ROCK1. Since MYPT1 is specifically phosphorylated by ROCK at Thr850, the finding suggests that RhoE interacts with and suppresses the activity of ROCK1 in HCC cells. To conclude, our data suggest that RhoE in HCC suppresses both cell motility and invasiveness of HCC cells and that RhoE may exert its effects by inhibiting the activity of ROCK1. (This study was funded by Hong Kong Research Grants Council CRF grants (RGC CRF HKU1/06C and HKU 7/CRG/09) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1443. doi:10.1158/1538-7445.AM2011-1443