Targeting a kinetochore-associated motor protein to kill cancer cells.

T umors are characterized by excessive proliferation and, therefore, chemotherapeutics that target proteins involved in tumor cell division can be effective anticancer agents. However, as many of these proteins are involved in core mechanisms in dividing and nondividing cells, these therapeutic agents do not selectively kill tumor cells and their efficacy is limited by general toxicity. As new therapeutics that target proteins involved in these core processes are developed, the likelihood increases of identifying a protein that cancer cells depend on more than normal cells. Wood et al. report the discovery and characterization of an inhibitor of a mitotic motor protein, centromere-associated protein-E (CENP-E, kinesin-7) (1). This inhibitor for a protein whose functions are limited to cell division has the potential to lead to improved cancer chemotherapies. Currently used antimitotic therapeutics target the cytoskeletal protein tubulin, which polymerizes to form microtubules (2). During cell division, segregation of chromosomes requires a microtubulebased bipolar spindle. Only after all chromosome pairs have been attached to the opposite ends of the bipolar spindle through microtubules, the spindle assembly checkpoint is satisfied, chromosomes are separated, and the cell cycle progresses. When normal tubulin polymerization dynamics are disrupted, proper chromosome-spindle attachments are not established, and the cell cycle is blocked by the checkpoint. Through poorly understood mechanisms, this cell-cycle arrest can lead to cell death (3). However, microtubules have essential roles in other cellular processes, such as neuronal transport. Therefore, the use of tubulintargeting antimitotic agents is associated with side effects, including neurotoxicity. A decade ago, a small-molecule inhibitor, monastrol, was reported for kinesin-5 (also called KSP or Eg5) (4). Kinesins are motor proteins that can use ATP hydrolysis to drive transport of cellular cargoes along microtubules. Monastrol was the first chemical inhibitor that targeted a protein, other than tubulin, needed for mitotic spindle assembly. This initial “hit” helped catalyze the development of drugs against kinesin-5, a protein that was not known to have key functions in nondividing cells such as neurons. The kinesin superfamily includes 14 different families and more than 40 individual kinesin genes in humans (5). Differentmitotic kinesins have crucial roles in distinct aspects of spindle assembly and function, including microtubule organization, chromosome movement, and cytokinesis (6). Although more work is needed to determine whether kinesin-5 inhibitors will be more effective than currently used tubulin-targeting chemotherapeutics, the development ofmultiple kinesin-5-targeted chemicals suggests that members of the kinesin superfamily are “druggable” (7). CENP-E is a kinetochore-associated kinesin with an essential role in metaphase chromosome alignment that does not function in nondividing cells (8). Depletion of CENP-E from cultured human cells is characterized by a bipolar mitotic spindle with chromosomes clustered close to either end of the bipolar spindle (i.e., the spindle pole), rather than aligning at the metaphase plate (Fig. 1 A and B) (9). It has been proposed that CENP-E contributes to a cooperative chromosome alignment mechanism in which the motor protein drives the movement of chromosomes from spindle poles to the metaphase plate alongside microtubules that are properly oriented, as these filaments link other aligned chromosomes to spindle poles (10, 11). Recently, Wood et al. conducted an in vitro high-throughput screen to identify inhibitors of CENP-E ATPase activity. Optimization of initial “hits” resulted in GSK923295, a potent inhibitor of CENP-E (1). In dividing cells treated with GSK923295, chromosomes are clustered at the spindle poles as would Fig. 1. GSK923295, an inhibitor of the mitotic kinesin CENP-E. (A) Normal cell division requires alignment of chromosomes at the equator of the bipolar spindle (the metaphase plate). (B) Loss of CENP-E function results in a few chromosomes that remain stuck at spindle poles. (C) Chemical structures of GSK923295, the CENP-E inhibitor, andmonastrol, the kinesin-5 inhibitor. These inhibitors bind at an allosteric site (yellow), which is ∼10 Å from the ADP-binding site (red), in the kinesin ATPase domain, as depicted on the CENP-E motor domain (PDB: 1T5C) (D).