ABSTRACT
Human eyes absent (EYA) proteins possess Tyr phosphatase activity, which is critical for numerous cancer and metastasis promoting activities, making it an attractive target for cancer therapy. In this work, we demonstrate that the inhibitor-bound form of EYA2 does not favour binding to Mg2+ , which is indispensable for the Tyr phosphatase activity. We further describe characterization and optimization of this class of allosteric inhibitors. A series of analogues were synthesized to improve potency of the inhibitors and to elucidate structure-activity relationships. Two co-crystal structures confirm the binding modes of this class of inhibitors. Our medicinal chemical, structural, biochemical, and biophysical studies provide insight into the molecular interactions of EYA2 with these allosteric inhibitors. The compounds derived from this study are useful for exploring the function of the Tyr phosphatase activity of EYA2 in normal and cancerous cells and serve as reference compounds for screening or developing allosteric phosphatase inhibitors. Finally, the co-crystal structures reported in this study will aid in structure-based drug discovery against EYA2.
Subject(s)
Intracellular Signaling Peptides and Proteins , Nuclear Proteins , Protein Tyrosine Phosphatases , Enzyme Inhibitors/chemistry , Humans , Intracellular Signaling Peptides and Proteins/antagonists & inhibitors , Intracellular Signaling Peptides and Proteins/chemistry , Nuclear Proteins/antagonists & inhibitors , Nuclear Proteins/chemistry , Protein Tyrosine Phosphatases/antagonists & inhibitors , Protein Tyrosine Phosphatases/chemistry , Structure-Activity RelationshipABSTRACT
AXL is a member of the TAM (TYRO3, AXL, MER) subfamily of receptor tyrosine kinases. It is upregulated in a variety of cancers and its overexpression is associated with poor disease prognosis and acquired drug resistance. Utilizing a fragment-based lead discovery approach, a new indazole-based AXL inhibitor was obtained. The indazole fragment hit 11, identified through a high concentration biochemical screen, was expeditiously improved to fragment 24 by screening our in-house expanded library of fragments (ELF) collection. Subsequent fragment optimization guided by docking studies provided potent inhibitor 54 with moderate exposure levels in mice. X-ray crystal structure of analog 50 complexed with the I650M mutated kinase domain of Mer revealed the key binding interactions for the scaffold. The good potency coupled with reasonable kinase selectivity, moderate in vivo exposure levels, and availability of structural information for the series makes it a suitable starting point for further optimization efforts.
Subject(s)
Drug Discovery , Indazoles/pharmacology , Protein Kinase Inhibitors/pharmacology , Proto-Oncogene Proteins/antagonists & inhibitors , Receptor Protein-Tyrosine Kinases/antagonists & inhibitors , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Humans , Indazoles/chemical synthesis , Indazoles/chemistry , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Proto-Oncogene Proteins/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , Structure-Activity Relationship , Axl Receptor Tyrosine KinaseABSTRACT
Dysregulation of translation initiation factor 4E (eIF4E) activity occurs in various cancers. Mitogen-activated protein kinase (MAPK) interacting kinases 1 and 2 (MNK1 and MNK2) play a fundamental role in activation of eIF4E. Structure-activity relationship-driven expansion of a fragment hit led to discovery of dual MNK1 and MNK2 inhibitors based on a novel pyridine-benzamide scaffold. The compounds possess promising in vitro and in vivo pharmacokinetic profiles and show potent on target inhibition of eIF4E phosphorylation in cells.
Subject(s)
Intracellular Signaling Peptides and Proteins/antagonists & inhibitors , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Cell Line, Tumor , Crystallography, X-Ray , Drug Discovery , Eukaryotic Initiation Factor-4E/antagonists & inhibitors , Humans , Models, Molecular , Phosphorylation , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/pharmacokinetics , Structure-Activity RelationshipABSTRACT
Protein kinase C iota (PKC-ι) is an atypical kinase implicated in the promotion of different cancer types. A biochemical screen of a fragment library has identified several hits from which an azaindole-based scaffold was chosen for optimization. Driven by a structure-activity relationship and supported by molecular modeling, a weakly bound fragment was systematically grown into a potent and selective inhibitor against PKC-ι.
