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Research Articles: Therapeutics, Targets, and Development

Induction of apoptosis by monastrol, an inhibitor of the mitotic kinesin Eg5, is independent of the spindle checkpoint

¹ DNAX Research Institute of Molecular and Cellular Biology Research Institute, Palo Alto, California and ² MedImmune, Inc., Gaithersburg, Maryland

Requests for reprints: Ronald Herbst, MedImmune, Inc., Gaithersburg, MD. Phone: 301-398-5253. E-mail: herbstr{at}medimmune.com

Abstract

Spindle poisons such as paclitaxel are widely used as cancer therapeutics. By interfering with microtubule dynamics, paclitaxel induces mitotic arrest and apoptosis. Targeting the kinesin Eg5, which is required for the formation of a bipolar spindle, is a promising therapeutic alternative to drugs that interfere with microtubule dynamics. Recent data suggest that the spindle checkpoint can determine the response of tumor cells to microtubule poisons. The relationship between checkpoint function and Eg5 inhibition, however, has not yet been fully investigated. Here, we used time-lapse video microscopy and biochemical analysis to study the effect of spindle checkpoint abrogation on the response of HeLa cells to monastrol, a selective Eg5 inhibitor. In HeLa cells, monastrol activated the spindle checkpoint, leading to mitotic arrest and apoptosis. Small interfering RNA–mediated depletion of the spindle checkpoint proteins BubR1 or Mad2 significantly shortened drug-induced arrest, causing premature mitotic exit without cell division. Time-lapse microscopy as well as analysis of caspase activation shows that these checkpoint-deficient cells initiate apoptosis after mitotic exit in response to monastrol. Checkpoint-deficient cells treated with paclitaxel, on the other hand, yielded a higher frequency of cells with >4N DNA content and a decreased incidence of apoptotic events, particularly in Mad2-depleted cells. These results indicate that the immediate fate of postmitotic cells is influenced by both the nature of the checkpoint defect and the type of drug used. Furthermore, these results show that inactivation of the kinesin Eg5 can induce apoptosis in tumor cells in the absence of critical spindle checkpoint components. [Mol Cancer Ther 2006;5(10):2580–91]

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

³ Supplementary data for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

⁴ Vijapurkar and Herbst, unpublished data.

⁵ Bookstein and Herbst, unpublished data.

Received 4/11/06; revised 7/ 6/06; accepted 8/16/06.