Inhibition of Invasion and Metastasis of Hepatocellular Carcinoma Cells via Targeting RhoC In vitro and In vivo

Abstract

Purpose: Hepatocellular carcinoma (HCC) is one of the most deadly human cancers because of its high incidence of metastasis. Our previous work identified a strong correlation between increased expression of RhoC and HCC metastasis. Here, we investigate to define the role of RhoC in HCC metastasis. Furthermore, we sought to determine whether inhibition of the expression of RhoC might block the metastasis of HCC in vivo. Experimental Design: A stable retroviral small interfering RNA approach was employed to selectively knockdown the expression of RhoC in vitro and in vivo. Invasion and migration assay, MTT and fluorescence-activated cell sorting analysis, Rho activity assay, and immunofluorescence staining were carried out to characterize RhoC in vitro. An anti-RhoC retroviral gene delivery BALB/c nude mice model was established to investigate whether knockdown of the expression of RhoC might inhibit the metastasis of HCC in vivo. Results: We confirmed the correlation of RhoC expression and metastatic potentials of HCC cell lines.We also showed that suppression of RhoC expression resulted in inhibition of invasion and migration without an apparent effect on cell survival and proliferation in HCCLM3 cells. Furthermore, a similar effect of RhoC on autotaxin-induced invasion of HCCLM3 cells was also observed. Significantly, we successfully adopted an HCC metastatic mouse model that allowed us to show that knockdown of the RhoC expression resulted in inhibition of metastasis of HCC in vivo for the first time. Conclusions: Our results show a critical role of RhoC in metastasis of HCC, implicating RhoC as a potential therapeutic target to block HCC metastasis.Hepatocellular carcinoma (HCC) is one of the most common malignancies in Asia and Africa, especially in China (1, 2). During the past decade, the long-term survival remains unsatisfactory because of a high incidence of recurrence and metastasis after the hepatic resection, with a 5-year actuarial recurrence rate of 75% to 100% reported in the literature (3). Thus, the inhibition of invasion and metastasis is of great importance in the HCC therapies.Available information indicates that invasion and metastasis is to a large extent attributable to the ability of cell migration (4, 5). Rho-GTPases, members of Ras superfamily of small GTPases, shuttle between inactive GDP-bound and active GTPbound form and exhibit intrinsic GTPase activities. Activation of the Rho protein leads to the assembly of the actin-myosin contractile filaments into focal adhesion complexes that lead to cell polarity and facilitate motility (6). Recently, RhoC has attracted interest because its increased expression has been linked to increased invasion in breast, melanoma, pancreatic, colon, bladder, hepatocellular, non-small cell lung carcinoma, and primary gastric tumor or cell lines (7 -13). Our previous study revealed that RhoC was a prognostic marker with HCC and its expression was correlated with invasion and metastasis of HCC (14 -16). Interestingly, a recent study using RhoCdefic...

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YMO1 suppresses invasion and metastasis by inhibiting RhoC signaling and predicts favorable prognosis in hepatocellular carcinoma

“…However, YMO1 didn't increased ubiquitination of RhoC (Supplementary Figure S8). Our previous study has indicated that RhoC expression was remarkably increased in HCC tissues and was associated with metastasis of HCC [21]. Rho-GTPases, members of Ras superfamily of small GTPases, shuttle between inactive GDP-bound and active GTP-bound form and exhibit intrinsic GTPase activities [22].…”

The MLK-related Kinase (MRK) Is a Novel RhoC Effector That Mediates Lysophosphatidic Acid (LPA)-stimulated Tumor Cell Invasion*

“…In particular, RhoC has emerged as a key factor in tumor progression in many types of cancer that include invasive breast carcinoma (5,15), gastric cancer (13), non small cell lung carcinoma (14), ovarian carcinoma (8), and esophageal squamous cell carcinoma (70). In addition, RhoC has been shown to be essential for metastasis in animal models (10,11,12,72).…”