Involvement of Cdc25A phosphatase in Hep3B hepatoma cell growth inhibitioninduced by novel K vitamin analogs

We previously found that K vitamin analogues caused cell growth inhibition in Hep3B hepatoma cells in vitro, which was associated with their inhibitor y effects on protein tyrosine-phosphatases. In this study, we show that Cdc 25A, a protein phosphatase, was inactivated by novel arylating K vitamin an alogues. The inactivation of Cdc25A correlated with their effects on cell g rowth inhibition. Cyclin-dependent kinase (Cdk) 4, an important regulator f or G(1) progression, was found to be tyrosine-phosphorylated by the arylati ng analogues, and this phosphorylation was correlated with the inhibitory e ffects of the analogues on Cdc25A activity. Furthermore, Cdk4 dephosphoryla tion experiments showed that Compound (Cpd) 5, a prototype arylating analog ue, inhibited Cdc25A-mediated Cdk4 dephosphorylation, whereas Cpd 26, a non arylating vitamin K analogue, had no effect on this event. We also examined Cdk4 kinase activity using retinoblastoma protein as a substrate and found that Cpd 5 inhibited retinoblastoma protein phosphorylation in a concentra tion-dependent manner, indicating that Cdk4 activity was inhibited by Cpd 5 treatment Moreover, the thiol-antioxidants glutathione and N-acetyl-L-cyst eine antagonized the Cpd 5-induced Cdk4 tyrosine phosphorylation, whereas t he nonthiol-antioxidants catalase and superoxide dismutase did not. These r esults suggest that Hep3B cell growth inhibition by these K vitamin analogu es may be related in part to inactivation of Cdc25A activity and support th e hypothesis that Cdc25A is an attractive target for drugs designed to inhi bit cancer cell growth.