ABSTRACT
The identification of a novel series of PKCθ inhibitors and subsequent optimization using docking based on a crystal structure of PKCθ is described. SAR was rapidly generated around an amino pyridine-ketone hit; (6-aminopyridin-2-yl)(2-aminopyridin-3-yl)methanone 2 leading to compound 21 which significantly inhibits production of IL-2 in a mouse SEB-IL2 model.
Subject(s)
Aminopyridines/chemistry , Protein Kinase C/antagonists & inhibitors , Protein Kinase Inhibitors/chemistry , Aminopyridines/pharmacology , Animals , Mice , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/pharmacology , Structure-Activity RelationshipABSTRACT
The conjugation of ubiquitin to proteins involves a cascade of activating (E1), conjugating (E2), and ubiquitin-ligating (E3) type enzymes that commonly signal protein destruction. In TGFbeta signaling the inhibitory protein Smad7 recruits Smurf2, an E3 of the C2-WW-HECT domain class, to the TGFbeta receptor complex to facilitate receptor degradation. Here, we demonstrate that the amino-terminal domain (NTD) of Smad7 stimulates Smurf activity by recruiting the E2, UbcH7, to the HECT domain. A 2.1 A resolution X-ray crystal structure of the Smurf2 HECT domain reveals that it has a suboptimal E2 binding pocket that could be optimized by mutagenesis to generate a HECT domain that functions independently of Smad7 and potently inhibits TGFbeta signaling. Thus, E2 enzyme recognition by an E3 HECT enzyme is not constitutively competent and provides a point of control for regulating the ubiquitin ligase activity through the action of auxiliary proteins.
Subject(s)
DNA-Binding Proteins/metabolism , Trans-Activators/metabolism , Ubiquitin-Conjugating Enzymes/metabolism , Ubiquitin-Protein Ligases/metabolism , Amino Acid Motifs/genetics , Amino Acid Sequence , Binding Sites/genetics , Catalysis , Cell Line , Crystallography, X-Ray , DNA-Binding Proteins/genetics , DNA-Binding Proteins/physiology , Enzyme Activation , Humans , Models, Molecular , Molecular Sequence Data , Mutagenesis, Site-Directed , Mutation , Peptide Fragments/genetics , Protein Binding , Protein Structure, Tertiary , Receptors, Transforming Growth Factor beta/genetics , Receptors, Transforming Growth Factor beta/metabolism , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Signal Transduction/physiology , Smad7 Protein , Trans-Activators/genetics , Trans-Activators/physiology , Transfection , Ubiquitin/metabolism , Ubiquitin-Activating Enzymes/metabolism , Ubiquitin-Conjugating Enzymes/chemistry , Ubiquitin-Conjugating Enzymes/genetics , Ubiquitin-Protein Ligases/chemistry , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/physiologyABSTRACT
The Aurora kinases are essential for the regulation of chromosome segregation and cytokinesis during mitosis. Aberrant expression and activity of these kinases occur in a wide range of human tumors, and lead to aneuploidy and tumorigenesis. Here we report the discovery of a highly potent and selective small-molecule inhibitor of Aurora kinases, VX-680, that blocks cell-cycle progression and induces apoptosis in a diverse range of human tumor types. This compound causes profound inhibition of tumor growth in a variety of in vivo xenograft models, leading to regression of leukemia, colon and pancreatic tumors at well-tolerated doses. Our data indicate that Aurora kinase inhibition provides a new approach for the treatment of multiple human malignancies.
