Quick evaluation of kinase inhibitors by surface plasmon resonance using single-site specifically biotinylated kinases.
In evaluating kinase inhibitors, kinetic parameters such as association/dissociation rate constants are valuable information, as are equilibrium parameters KD and IC50 values. Surface plasmon resonance (SPR) is a powerful technique to investigate these parameters. However, results are often complicated because of impaired conformations by inappropriate conditions required for protein immobilization and/or heterogeneity of the orientation of immobilization. In addition, conventional SPR experiments are generally time-consuming. Here we introduce the use of single-site specifically biotinylated kinases combined with a multichannel SPR device to improve such problems. Kinetic parameters of four compounds-staurosporine, dasatinib, sunitinib, and lapatinib-against six kinases were determined by the ProteOn XPR36 system. The very slow off-rate of lapatinib from the epidermal growth factor receptor and dasatinib from Bruton's tyrosine kinase and colony stimulating factor 1 receptor (CSF1R) were confirmed. Furthermore, IC50 values were determined by an activity-based assay. Evaluating both physicochemical and biochemical properties would help to understand the detailed character of the compound.
1Carna Biosciences, Inc., Kobe, Japan.
Daisuke
D
Kitagawa
Masaki
M
Gouda
Yasuyuki
Y
Kirii
D000818:Animals; D019921:Biotinylation; D002460:Cell Line; D055808:Drug Discovery; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015870:Gene Expression; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D010770:Phosphotransferases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020349:Surface Plasmon Resonance / Q000379:methods*
D004791:Enzyme Inhibitors; D010770:Phosphotransferases
biotinylation; drug discovery; kinase inhibitor; surface plasmon resonance; tyrosine kinase