Fostriecin-mediated G2-M-phase growth arrest correlates with abnormal centrosome replication, the formation of aberrant mitotic spindles, and the inhibition of serine/threonine protein phosphatase activity.
Fostriecin, a structurally unique phosphate ester, is presently under evaluation in clinical trials to determine its potential use as an antitumor drug in humans. Fostriecin has been reported as having inhibitory activity against DNA topoisomerase type II and protein phosphatases implicated in cell-cycle control. However, the relative contribution of these mechanisms to the antitumor activity of fostriecin has not yet been elucidated. In this study, after confirming that fostriecin is a potent inhibitor of serine/threonine protein phosphatase type 2A and a weak inhibitor of serine/threonine protein phosphatase type 1, we show that fostriecin inhibits approximately 50% of the divalent cation independent serine/threonine protein phosphatase (PPase) activity contained in whole cell homogenates of Chinese hamster ovary cells at concentrations associated with antitumor activity (1-20 microM). Investigations into the cellular effects produced by fostriecin treatment reveal that 1-20 microM fostriecin induces a dose-dependent arrest of cell growth during the G2-M phase of the cell cycle. Immunostaining of treated cells indicates that growth arrest occurs before the completion of mitosis and that fostriecin-induced growth arrest is associated with the aberrant amplification of centrosomes, which results in the formation of abnormal mitotic spindles. The "mitotic block" induced by fostriecin is reversible if treatment is discontinued in <24 h. However, after approximately 24-30 h of continuous treatment, growth arrest is not reversible, and treated cells die even when placed in fostriecin-free media. Correlative studies conducted with established PPase inhibitors reveal that, when applied at concentrations that inhibit PPase activity to a comparable extent, both okadaic acid and cantharidin also induce aberrant centrosome replication, the appearance of multiple aberrant mitotic spindles, and G2-M-phase growth arrest. These studies add additional support to the concept that PPase inhibition underlies the antitumor activity of fostriecin and suggest that other type-selective PPase inhibitors should be evaluated for potential antitumor activity.
Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile 36688, USA.
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A
Cheng
R
R
Balczon
Z
Z
Zuo
J S
JS
Koons
A H
AH
Walsh
R E
RE
Honkanen
D000475:Alkenes / Q000494:pharmacology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D016466:CHO Cells / Q000187:drug effects / Q000201:enzymology; D002193:Cantharidin / Q000494:pharmacology; D018385:Centrosome / Q000187:drug effects*; D006224:Cricetinae; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D016195:G2 Phase / Q000187:drug effects*; D008938:Mitosis / Q000187:drug effects*; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011090:Polyenes; D011753:Pyrones; D012333:RNA, Messenger / Q000378:metabolism; D008941:Spindle Apparatus / Q000187:drug effects*; D016159:Tumor Suppressor Protein p53 / Q000187:drug effects / Q000378:metabolism
D000475:Alkenes; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D011090:Polyenes; D011753:Pyrones; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases; D002193:Cantharidin; C040313:fostriecin