The histone H3-lysine 27 (H3K27) methyltransferase EZH2 plays a critical role in regulating gene expression, and its aberrant activity is linked to the onset and progression of cancer. As part of a drug discovery program targeting EZH2, we have identified highly potent, selective, SAM-competitive, and cell-active EZH2 inhibitors, including GSK926 (3) and GSK343 (6). These compounds are small molecule chemical tools that would be useful to further explore the biology of EZH2.

Phosphoinositide 3-kinase R (PI3KR) is a critical regulator of cell growth and transformation, and its signaling pathway is the most commonly mutated pathway in human cancers. The mammalian target of rapamycin (mTOR), a class IV PI3K protein kinase, is also a central regulator of cell growth, and mTOR inhibitors are believed to augment the antiproliferative efficacy of PI3K/AKT pathway inhibition. 2,4-Difluoro-N-{2-(methyloxy)-5-[4-(4-pyridazinyl)-6-quinolinyl]-3-pyridinyl}benzenesulfonamide (GSK2126458, 1) has been identified as a highly potent, orally bioavailable inhibitor of PI3KR and mTOR with in vivo activity in both pharmacodynamic and tumor growth efficacy models. Compound 1 is currently being evaluated in human clinical trials for the treatment of cancer. KEYWORDS GSK2126458, phosphoinositide 3-kinase R, mammalian target of rapamycin, PI3K/AKT pathway

Centromere-associated protein-E (CENP-E) is a kinetochore-associated mitotic kinesin that is thought to function as the key receptor responsible for mitotic checkpoint signal transduction after interaction with spindle microtubules. We have identified GSK923295, an allosteric inhibitor of CENP-E kinesin motor ATPase activity, and mapped the inhibitor binding site to a region similar to that bound by loop-5 inhibitors of the kinesin KSP/Eg5. Unlike these KSP inhibitors, which block release of ADP and destabilize motor-microtubule interaction, GSK923295 inhibited release of inorganic phosphate and stabilized CENP-E motor domain interaction with microtubules. Inhibition of CENP-E motor activity in cultured cells and tumor xenografts caused failure of metaphase chromosome alignment and induced mitotic arrest, indicating that tight binding of CENP-E to microtubules is insufficient to satisfy the mitotic checkpoint. Consistent with genetic studies in mice suggesting that decreased CENP-E function can have a tumor-suppressive effect, inhibition of CENP-E induced tumor cell apoptosis and tumor regression.entromere-associated protein-E (CENP-E; kinesin-7) is a kinetochore-associated kinesin motor protein with an essential and exclusive role in metaphase chromosome alignment and satisfaction of the mitotic checkpoint (1). CENP-E is a likely candidate to integrate the mechanics of kinetochore-microtubule interaction with the mitotic checkpoint signaling machinery responsible for restraining cell-cycle progression into anaphase. CENP-E is a large dimeric protein consisting of an N-terminal kinesin motor domain tethered to a globular C-terminal domain through an extended coiled-coil rod domain (2, 3). The C-terminal, noncatalytic region of CENP-E is not only sufficient to specify localization to kinetochores, but it also mediates interaction of CENP-E with the serine/threonine kinase BubR1, a key effector of mitotic checkpoint signaling that forms complexes with the checkpoint proteins Cdc20, Bub3, and Mad2 to inhibit the ubiquitin ligase activity of the anaphase promoting complex APC/C CDC20 (4-7). The combined interaction of CENP-E with microtubules and a key regulator of APC/C CDC20 has led to the hypothesis that CENP-E functions as the key kinetochore microtubule receptor responsible for silencing mitotic checkpoint signal transduction after capture of spindle microtubules. This hypothesis was further strengthened by the finding that CENP-E could stimulate the kinase activity of BubR1 in a microtubule-sensitive manner (8, 9). In vitro, the addition of CENP-E to BubR1 resulted in a stimulation of BubR1 kinase activity. The addition of microtubules suppressed this stimulatory activity, an effect thought to be mediated by the CENP-E kinesin motor domain. Although the importance of CENP-E interaction with BubR1 and the role of BubR1-mediated phosphorylation in mitotic checkpoint function remain unclear, CENP-E remains a prominent candidate to play a key role in mitotic checkpoint signal transduction.Depletion of CENP-E from ...

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