Abstract
Abstract Background: More than 80% of Hepatocellular carcinoma (HCC) patients present at advanced stages not amenable to curative therapies. Currently existing palliative therapies achieve a minimal survival benefit of 3 months. Hence, there is an urgent need for novel therapeutic strategies. Tumor Growth Factor-β (TGF-β) is a potent tumor suppressor via its Smad2/3 signaling axis. Inactivation of this axis has been implicated in hepatocarcinogenesis. In patients with advanced HCC, TGF- β serum concentrations are elevated and correlated to disease stage, metastases and worse prognosis. Our aims were to evaluate the status and function of the TGF-β/Smad2 signaling axis in advanced HCC. Material and Methods: Immunohistochemical analysis for pSer465/467-Smad2 in human HCC samples was performed using Tissue Micro Arrays. In vitro studies were performed using established human HCC cell lines (HepG2, Hep3B, PLC/PRF/5, HUH-7, SNU-398, SNU-449). Pathway activation following TGF-β stimulation was evaluated by immunoblot analysis and immunofluorescence microscopy. Cell proliferation was assessed by cell quantification and MTS-assay. Apoptosis was quantified morphologically by DAPI stain and fluorescence microscopy, and biochemically by caspase-3/7 assay. Migration was assessed by gap closure assay, and invasion by Matrigel invasion assay. Results: Intranuclear pSer465/467-Smad2 was identified in up to 51% of advanced HCC (n=73) and in 89% of HCC metastases (n=9). In 50% of human HCC cell lines, TGF-β treatment (10 ng/mL) resulted in Ser465/467-phosphorylation and nuclear translocation of the transcription factor Smad2. Exclusively in HCC cells with an intact TGF-β/Smad2-signaling cascade, TGF-β stimulation induced Tyr705-phosporylation of the transcription factor STAT3. TGF-β induced STAT3 Tyr705-phosporylation is inhibited by Smad2-directed gene silencing. Further, pyridine 6 mediated Janus Kinase-1/2 (JAK1/2) inhibition prevented TGF-β induced Tyr705-phosporylation of STAT3. Cell proliferation of HCC cells was inhibited by TGF-β. However, TGF-β treatment significantly induced invasion and migration in HCC cell lines with intact TGF-β/Smad2 signaling. Further, TGF-β induced resistance to TRAIL-induced apoptosis in these cells. However, co-treatment with the STAT3-inhibitor Stattic (10 nM) completely abrogates TGF-β mediated HCC cell migration and invasion, and sensitizes these HCC cell lines to TRAIL-mediated apoptosis. Conclusion: In advanced HCC, the TGF-β/Smad signaling axis is constitutively activated in the majority of primary HCC tumors and HCC metastases. The function of TGF-β/Smad2 in human HCC cell lines has switched from tumor inhibitor to a driver of tumor progression via JAK/STAT3 signaling. STAT3-inhibition reverses the TGF-β tumor-promoting effects. Hence, JAK/STAT3 targeting might be a promising new strategy to restore TGF-β tumor suppressor function in advanced HCC. Citation Format: Andres Rojas, Jessica Cromheecke, Mong-Hong Lee, Mien-Chie Hung, Boris Blechacz. TGF-β drives tumor-progression via JAK/STAT3-signaling in advanced HCC. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2046. doi:10.1158/1538-7445.AM2013-2046
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