Blocking transforming growth factor-beta up-regulates E-cadherin and reduces migration and invasion of hepatocellular carcinoma cells

Abstract

Hepatocellular carcinoma (HCC) treatment is challenging because the mechanisms underlying tumor progression are still largely unknown. Transforming growth factor (TGF)-␤1 is considered a crucial molecule in HCC tumorigenesis because increased levels of patients' serum and urine are associated with disease progression. The aim of the present study was to investigate the inhibition of TGF-␤ signaling and its impact on HCC progression. Human HCC cell lines were treated with a TGF-␤ receptor kinase inhibitor (LY2109761) whose selectivity was determined in a kinase assay. Exogenous TGF-␤1 phosphorylates the TGF-␤ receptor, consequently activating Smad-2, whereas the drug selectively blocks this effect and dephosphorylates autocrine p-Smad-2 at concentrations ranging from 0.001 to 0.1 M. A cytotoxic effect documented by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), trypan blue, and propidium iodide staining assays was observed at 10 M, whereas the drug inhibits (P < 0.001) the migration of HCC cells on fibronectin, laminin-5, and vitronectin and invasion through Matrigel (P < 0.001) at concentrations up to 0.1 M. LY2109761 up-regulates (P < 0.001) E-cadherin mRNA and protein levels. This increase was localized at the cellular membrane where E-cadherin mediates anchorage that is cell-cell dependent. Consistently, a functional monoclonal antibody that inhibits E-cadherin-dependent cell-cell contact restores the migratory and invasive activity. Finally, nonmetastatic HCC tissues from 7 patients were cultured with TGF-␤1 in the presence or absence of LY2109761. E-cadherin expression was reduced by TGF-␤1 and was significantly (P < 0.0001) increased by LY2109761 treatment, measured by quantitative real-time PCR on microdissected tissues and by immunohistochemistry on serial sections. In 72 patients, E-cadherin tissue expression was more weakly expressed in metastatic than in nonmetastatic HCC (P < 0.0001). Conclusion: LY2109761 blocks migration and invasion of HCC cells by upregulating E-cadherin, suggesting that there could be a mechanistic use for this molecule in clinical trials. (HEPATOLOGY 2008;47:1557-1566 H epatocellular carcinoma (HCC) is a highly malignant cancer that is the third most frequent cause of tumor-related death in the United States and Europe. 1 Current therapeutic options are invasive and aim to physically remove or destroy the tumor mass. However, later recurrence and/or metastatic spread are common and negatively affect survival. The overall prognosis is still unsatisfactory, and little progress has been made in finding new treatment options. However, a recent study with sorafenib suggests that targeting the vasculature may provide additional insights into how to develop future treatment options for patients with HCC. 2 Based on the recent clinical observations with sorafenib, the focus for developing new treatments in HCC has shifted from targeting the cancer to targeting the tissue microenvironment and its role in modulating the biological behavior of HCC. 3 Transforming grow...

Targeting TGF-β Signaling for Therapeutic Gain

“…Expression of E-cadherin, which can suppress tumor progression, is commonly down-regulated within many carcinomas during EMT (Caulin et al 1995;Portella et al 1998;Lacher et al 2006;Fransvea et al 2008;Hoot et al 2008). The TGF-b-Smad pathway mediates expression of high-mobility group A2 (HMGA2), which contributes to the induction of the expression of Snail and Slug, two zinc-finger transcription factors that repress the E-cadherin gene (Padua and Massagué 2009).…”

HAb18G/CD147 promotes epithelial–mesenchymal transition through TGF-β signaling and is transcriptionally regulated by Slug

“…Increased levels of TGF-b were detected in patients' serum, urine and liver neoplastic nodules, indicating that TGF-b is associated with HCC progression (Bedossa et al, 1995;Tsai et al, 1997;Song et al, 2002). It is believed that TGF-b-induced EMT has a pivotal role in the dissemination of malignant hepatocytes during HCC progression (Giannelli et al, 2005) because blocking TGF-b upregulates E-cadherin and reduces the migration and invasion of HCC cells (Fransvea et al, 2008). In this study, we showed that CD147 is upregulated by TGF-b and mediates EMT in tumor formation, and that this change is coupled with the induction of MMP-2 secretion, the inhibition of apoptosis and the acceleration of the cell cycle.…”

Inhibition of transforming growth factor β receptor I kinase blocks hepatocellular carcinoma growth through neo-angiogenesis regulation

“…10,11 Recently, we have shown that LY2109761 displays an antimetastatic activity in HCC in vivo models by increasing the expression of E-cadherin on the HCC cellular surface, thus blocking the invasion into the surrounding stroma. 12 Furthermore, it also inhibits the spread of HCC through blood vessels, dephosphorylating the intracytoplasmic tail of ␤1 integrin, the main receptor responsible for fibrinogen and fibronectin-dependent migration. Because these two extracellular matrix proteins are the major internal and external constituents of the blood vessel wall, LY2109761 blocks HCC cells intravasation by way of a a selective inhibition of the transforming growth factor ␤1 (TGF-␤1) SMAD-dependent pathway.…”