Abstract 3881: A novel class of structure-based cyclin-dependent kinase inhibitors
Authors: Law Brian K. , Corsino Patrick , Law Mary , Davis Bradley , Parker Nicole , Jahn Stephan 
Published: February 2011

Abstract
Deregulation of the Cdk/Rb/E2F axis is a common feature among cancers. Several promising ATP-competitive Cdk inhibitors are undergoing clinical testing as anticancer therapeutics. However a weakness of ATP-competitive Cdk inhibitors is that all kinases share a highly conserved ATP binding site, making it difficult to selectively inhibit cell cycle Cdks. Further, there is significant functional redundancy among cell cycle Cdks, so the ideal Cdk inhibitor would block the activity of multiple cell cycle Cdks while not affecting other kinases. Nature utilizes several modes of regulation that are specific to the Cdks that involve (1) the inhibitory proteins p21, p27, and p57, (2) inhibitory phosphorylation on a tyrosine residue near the ATP binding pocket, (3) activating phosphorylation on the “T-loop” site, and (4) binding of an activating Cyclin subunit. Usurping one of these natural Cdk-specific control mechanisms may be the best hope for developing potent and selective anticancer Cdk inhibitors.
Comparison of the crystal structures of Cyclin A/Cdk2/p27 and Cyclin A/Cdk2 complexes indicated that p27 binding causes a conformational change, which opens a pocket on the surface of Cdk2. We hypothesized that a small molecule that binds to this pocket would force Cdks to adopt an inactive conformation. In silico molecular docking was performed to identify compounds predicted to bind to this pocket. Multiple compounds identified in this screen inhibited the proliferation of breast cancer cells. Inhibition of proliferation correlated with a decrease in the soluble levels of Cdk1, Cdk2, and Cdk4, and an increase in these proteins in the insoluble fraction. Video microscopy of cells stably expressing a GFP-Cdk4 fusion protein showed aggregate formation over time in the presence of the Cdk inhibitors and that the aggregates dissipated after the drug was removed. Proliferation studies performed in parallel also indicated that the effects of the Cdk inhibitors were largely reversible within the first 24 hours.
One of the compounds, NSC117024, preferentially inhibited the proliferation of BT549 breast cancer cells over nontransformed mammary epithelial cells. Inhibition of BT549 proliferation was observed at NSC117024 concentrations as low as 780 nM. Interestingly, suppression of cancer cell division by the compounds occurred as well or better in cells lacking p21 and p27. The compounds directly inhibited Cdk2 activity in kinase assays. Together the results suggest that the novel family of Cdk inhibitors mimic the actions of p21 and p27 and may preferentially inhibit the proliferation of a subset of breast cancer cells. Current efforts are directed toward optimizing NSC117024 antiproliferative activity, developing a biotinylated NSC117024 affinity probe to verify compound specificity, and identifying the genetic features of cancer cells that render them sensitive to NSC117024.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3881.