SKI-606 decreases growth and motility of colorectal cancer cells by preventing pp60(c-Src)-dependent tyrosine phosphorylation of β-catenin and its nuclear signaling

Inhibition of deregulated protein tyrosine kinases represents an attractive strategy for controlling cancer growth. However, target specificity is an essential aim of this strategy. In this report, pp60(c-Src) kinase and β-catenin were found physically associated and constitutively activated on tyrosine residues in human colorectal cancer cells. The use of specific small-interfering RNAs (siRNA) validated pp60(c-Src) as the major kinase responsible for β-catenin tyrosine phosphorylation in colorectal cancer. Src-dependent activation of β-catenin was prevented by SKI-606, a novel Src family kinase inhibitor, which also abrogated β-catenin nuclear function by impairing its binding to the TCF4 transcription factor and its trans-activating ability in colorectal cancer cells. These effects were seemingly specific, as cyclin D1, a crucial β-catenin/TCF4 target gene, was also down-regulated by SKI-606 in a dose-dependent manner accounting, at least in part, for the reduced growth (IC50, 1.5-2.4 μmol/L) and clonogenic potential of colorectal cancer cells. Protein levels of β-catenin remained substantially unchanged by SKI-606, which promoted instead a cytosolic/membranous retention of β-catenin as judged by immunoblotting analysis of cytosolic/nuclear extracts and cell immunofluorescence staining. The SKI-606-mediated relocalization of β-catenin increased its binding affinity to E-cadherin and adhesion of colorectal cancer cells, with ensuing reduced motility in a wound healing assay. Interestingly, the siRNA-driven knockdown of β-catenin removed the effect of SKI-606 on cell-to-cell adhesion, which was associated with prolonged stability of E-cadherin protein in a pulse-chase experiment. Thus, our results show that SKI-606 operates a switch between the transcriptional and adhesive function of β-catenin by inhibiting its pp60(c-Src)-dependent tyrosine phosphorylation; this could constitute a new therapeutic target in colorectal cancer. ©2006 American Association for Cancer Research.