Abstract
230 Background: TGF-beta functions as a tumor suppressor early in the development of pancreatic ductal adenocarcinoma (PDAC) but during tumor progression after undefined molecular changes TGF-beta switches to a promoter of metastasis, tumor angiogenesis and epithelial to mesenchymal transition (EMT.) A potential strategy is to target TGF-beta-R2, one of the major signaling receptors for TGF-beta. 2G8 is a anti-rat monoclonal antibody that binds specifically to and blocks murine TGF-beta-R2. Methods: Protein expression was analyzed using Western Blot. Cell proliferation was analyzed using Cell Titer Blue Assay and migration was determined using transwell migration assay. EMT protein expression was determined by ICC. PanO2-HY cells were used to establish an orthotopic and splenic metastasis model. In an orthotopic model, mice received saline, gemcitabine (Gem), 2G8 or 2G8 + Gem. For splenic metastasis model,mice received 2G8 on post injection day −1, +1 or +7 days or control IgG. Results: At baseline, PanO2-HY cells express TGF-beta-R2, alk5 and alk1 and express high levels of zeb1 and beta-catenin. 2G8 inhibits phosphorylation of smad2, ERK1/2 and p38 but not smad1/5 in vitro. 2G8 did not inhibit cell proliferation but did inhibit cell migration in vitro. After 7 days of treatment with 2G8, PanO2-HY cells had reduction in nuclear zeb1 staining by ICC. In the orthotopic model, 2G8 significantly inhibited metastasis to liver, peritoneum and gastrointestinal tract(p=0.02). In splenic metastasis assay, 2G8 treated mice livers weighed significantly less than control treated mice (p=0.0005). In the group of mice treated prior to tumor cell injection, none of them developed a primary tumor. Conclusions: Inhibition of TGF-beta-R2 with 2G8, results in inhibition of smad2 dependent pathways as well as non-canonical TGF-beta signaling in vitro. By inhibiting tumor cell migration in vitro and reducing EMT transcription factors, 2G8 may result in delayed tumor progression. In vivo data in both orthotopic and metastatic models suggest that inhibition of TGF-beta signaling via TGF-beta-R2 inhibits tumor cell metastasis. Further analysis are being conducted to understand changes within primary tumor and metastasis within this model.
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