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The biochemical and cell cycle effects of the antitumour drug postriecin

University of British Columbia

Fostriecin is an antitumour drug in phase I clinical trials. It has been recently shown that it is a potent inhibitor of protein phosphatases 1 and 2A in vitro, a property not previously described for an antitumour drug. We have investigated its effects on protein phosphorylation in Baby Hamster Kidney cells. Fostriecin strongly stimulated the phosphorylation of a single protein, which we identified as the intermediate filament vimentin. Fostriecin also caused rounding of the cells and a reorganization of the vimentin filaments. These effects are similar to those of the known inhibitors of protein phosphatase 1 and 2A, okadaic acid and calyculin A, which are actually tumour promoters. Fostriecin induced hyperphosphorylation of vimentin mostly at two sites which were sensitive to staurosporine and could be phosphorylated by protein kinase C in vitro. Fostriecin-induced hyperphosphorylation of vimentin also occurred in cells that lack cdc2 activity. These results suggest that protein kinase C plays a role in the hyperphosphorylation of vimentin during exposure to fostriecin. Furthermore, they provide strong evidence that fostriecin inhibits protein phosphatases 1 and 2A in vivo and raise the possibility that it may have tumour promoting activity. We further studied the effects of fostriecin on the cell cycle. When two human cancer cell lines, Hela and Jurkat, were treated with fostriecin, premature degradation of cyclin A and cyclin B was induced independently of mitosis. This degradation pathway required the activity of staurosporinesensitive kinase(s) and ubiquitin-proteasome activity. These results suggest that protein phosphatase activity is required for the proper timing of cyclin degradation. Upon fostriecin treatment and phosphatase inhibition, the degradation pathway is activated and cyclins are degraded. Since cyclins are essential for the progression of the cell cycle, we determined whether this premature cyclin degradation caused the cells to die. Instead we found that general cysteine neutral protease activity was required in part for the cytotoxicity of fostriecin and the degradation of other proteins besides cyclins is required for cell death.

Thesis/Dissertation

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2009-01-28

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http://hdl.handle.net/2429/3970

Biochemistry and Molecular Biology

University of British Columbia

UBCV

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