476637-05-5Relevant articles and documents
Hybrid compound design to overcome the gatekeeper T338M mutation in cSrc
Getlik, Matth?us,Grütter, Christian,Simard, Jeffrey R.,Klüter, Sabine,Rabiller, Matthias,Rode, Haridas B.,Robubi, Armin,Rauh, Daniel
, p. 3915 - 3926 (2009)
The emergence of drug resistance remains a fundamental challenge in the development of kinase inhibitors that are effective over long-term treatments. Allosteric inhibitors that bind to sites lying outside the highly conserved ATP pocket are thought to be more selective than ATP-competitive inhibitors and may circumvent some mechanisms of drug resistance. Crystal structures of type I and allosteric type III inhibitors in complex with the tyrosine kinase cSrc allowed us to employ principles of structure-based design to develop these scaffolds into potent type II kinase inhibitors. One of these compounds, 3c (RL46), disrupts FAK-mediated focal adhesions in cancer cells via direct inhibition of cSrc. Details gleaned from crystal structures revealed a key feature of a subset of these compounds, a surprising flexibility in the vicinity of the gatekeeper residue that allows these compounds to overcome a dasatinib-resistant gatekeeper mutation emerging in cSrc.
Development of a fluorescent-tagged kinase assay system for the detection and characterization of allosteric kinase inhibitors
Simard, Jeffrey R.,Getlik, Matthaeus,Gruetter, Christian,Pawar, Vijaykumar,Wulfert, Sabine,Rabiller, Matthias,Rauh, Daniel
supporting information; experimental part, p. 13286 - 13296 (2010/01/30)
Kinase disregulation disrupts the intricate network of intracellular signaling pathways and contributes to the onset of diseases such as cancer. Although several kinase inhibitors are on the market, inhibitor selectivity and drug resistance mutations persist as fundamental challenges in the development of effective long-term treatments. Chemical entities binding to less conserved allosteric sites would be expected to offer new opportunities for scaffold development. Because no high-throughput method was previously available, we developed a fluorescence-based kinase binding assay for identifying and characterizing ligands which stabilize the inactive kinase conformation. Here, we present a description of the development and validation of this assay using the serine/threonine kinase p38R. By covalently attaching fluorophores to the activation loop of the kinase, we were able to detect conformational changes and measure the Kd, kon, and koff associated with the binding and dissociation of ligands to the allosteric pocket. We report the SAR of a synthesized focused library of pyrazolourea derivatives, a scaffold known to bind with high affinity to the allosteric pocket of p38R. Additionally, we used protein X-ray crystallography together with our assay to examine the binding and dissociation kinetics to characterize potent quinazoline- and quinoline-based type II inhibitors, which also utilize this binding pocket in p38α. Last, we identified the b-Raf inhibitor sorafenib as a potent low nanomolar inhibitor of p38α and used protein X-ray crystallography to confirm a unique binding mode to the inactive kinase conformation.
