Identification and characterization of a selective inhibitor of the PIM-1 kinase

The PIM-1 kinase is a serine/threonine kinase that has been implicated in the development of many human cancers, including leukemias, lymphomas, and prostate cancer. We have endeavored to identify and characterize a selective inhibitor of the PIM-1 kinase. Such an inhibitor would have utility as a laboratory tool for the study of PIM-1 kinase function and as a template for the design of molecular therapeutics for diseases in which PIM-1 kinase activity is dysregulated.; Using phage display techniques we identified the peptide, HGVKKRPHNPYG, as a probable pseudosubstrate of the PIM-1 kinase with inhibitory activity in the low micromolar range in in vitro kinase assays. However, the peptide lacked sufficient potency for our objectives; hence we employed a second approach---the computational design of a novel PIM-1 kinase antagonist.; We created a homology model of the PIM-1 kinase that was subsequently determined to be structurally similar to a recently published crystal structure of the PIM-1 kinase. Docking studies using the PIM-1 kinase model demonstrated that the AFFINITY module of InsightII, under the parameters employed, was unable to accurately predict the potency of known PIM-1 kinase inhibitors. Consequently, we utilized comparative molecular field analysis, which resulted in a strong correlation of predicted IC50 vs. experimental IC 50 values for known flavonoid inhibitors of the PIM-1 kinase. Thus we have created the first predictive model that may be used for the rational design of small molecule inhibitors of the PIM-1 kinase.; We have also performed an extensive screen of small molecules with structures similar to known kinase inhibitors and identified quercetagetin as a potent inhibitor of the PIM-1 kinase, with an in vitro IC50 of 0.34 muM. Lineweaver-burk analyses of the reaction kinetics demonstrated that quercetagetin is an ATP competitive inhibitor the PIM-1 kinase. We have also determined that quercetagetin is selective for the PIM-1 kinase over many other kinases and that quercetagetin successfully inhibits PIM-1 kinase activity in mammalian cells. Therefore quercetagetin may be an effective tool for the laboratory study of the PIM-1 kinase, and may serve as a template for the design of molecular therapeutics directed against the PIM-1 kinase.