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The activation of sonic hedgehog (Shh) signaling has been implicated in the progression of various tumor types, including gastric carcinoma. Transforming growth factor (TGF- β) is a potent tumor suppressor but, paradoxically, TGF-β enhances tumor cell motility and invasiveness in different kinds of cells. In the present study, the effect of Shh on the TGF-β-mediated invasion potential of gastric cancer cells was investigated. In vitro migration and cell invasion assays with the gastric cancer cell lines AGS and MKN-28 revealed that the increased concentration of Shh N-terminal peptide enhanced TGF-β-mediated cell motility and invasiveness in these cells. However, TGF-β-mediated migration and invasiveness was blocked by treatment with KAAD-cyclopamine (a Shh signaling inhibitor), neutralizing anti-Shh or TGF-β antibodies. To analyze the effects of Shh on TGF-β regulated transcriptional activity, we conducted luciferase reporter assay using TGF-β response element reporter plasmid. The results showed that TGF-β-mediated transcriptional response was enhanced by treatment with Shh N-terminal peptide, but not with KAAD-cyclopamine or neutralizing anti-Shh antibodies. Western blot showed that the phosphorylation of Akt or Smad 3 and expression of ALK 5 protein was up-regulated by treatment with Shh N-terminal peptide, whereas was blocked by treatment with KAAD-cyclopamine (a Shh signaling inhibitor), anti-shh or TGF-β antibodies. Knockdown of the Akt, Smad 3 or ALK 5 (activin receptor-like kinase 5) using specific inhibitor or small interfering RNA significantly inhibited the Shh-induced cell motility and invasiveness. Taken together, these data indicate that Shh promotes motility and invasiveness of gastric cancer cells through TGF-β-mediated activation of Akt and/or Smad 3 pathway.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA