Rho-associated coiled-coil containing kinases (ROCK1/ROCK2) are critical regulatory enzymes implicated in controlling cytoskeletal functions, cell morphology, migration, metastasis and epithelial-mesenchymal transition (EMT) in cancer cells. Inhibition of ROCK enzyme activity therefore represents a potential novel therapeutic approach for the treatment of metastatic cancer. Multiple potential physiological substrates of ROCK have been identified as possible downstream signaling effectors, but it is currently unclear which of these represent essential elements for specific functions of ROCK in cancer cells. Furthermore, there are several additional kinases that have been reported to regulate one or more of the established targets of ROCK under certain conditions (including MRCK, MLCK, DMPK, CRIK), but the relative role of each enzyme remains to be established. We have used siRNA to evaluate the role of ROCK isoforms in signaling events in pancreatic cancer cells with mesenchymal characteristics, and established the relative contributions of ROCK1 and ROCK2 in signaling to the cytoskeleton. SiRNA knockdown of ROCK protein levels reduced phosphorylation of the regulatory light chain subunit of myosin and the myosin binding subunit of protein phosphatase 1, but had limited effects on LIMK/cofilin signaling. Similar effects were observed with selective inhibitors of the kinase activity of ROCK, and these effects were associated with inhibitory activity in cell migration assays performed in vitro sug gesting that regulation of certain key cytoskeletal signaling events by ROCK is critical for the migratory phenotype of pancreatic cancer cells with mesenchymal characteristics.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]