(1285-B) First-in-class Selectivity System for Quantitative Kinase Inhibitor Profiling in Live Cells against 192 Kinases
Monday, February 5, 2024
2:00 PM – 3:00 PM EST
Location: Exhibit Halls AB
Abstract: Kinase inhibitor profiling is an invaluable technique during the drug discovery process to understand inhibitor selectivity and determine off-target binding potential. While biochemical assays have been used to assess compound selectivity, the resulting profiles may not reflect activity observed in a complex biological system. We describe the first turnkey system to determine kinase selectivity in live cells and it uses an addition-only multi-well plate format. The NanoBRETâ„¢ Target Engagement K192 Kinase Selectivity System comprises (1) an array of 192 ready-to-transfect vectors, each encoding a full-length human kinase tagged with NanoLuc, and (2) a single NanoBRETâ„¢ TE Intracellular Kinase Assay, K-10. This kinase vector panel broadly represents the kinome and can be easily transfected into HEK293 cells. This system enables a simple, quantitative determination of intracellular compound occupancy for each kinase in the panel. The system has been validated for accuracy and reproducibility using a variety of kinase inhibitors. Using this method, we have also compared the live cell target engagement profile with similar biochemical assay profiles. In cells, the spectrum of activity for multikinase inhibitors may differ dramatically. Furthermore, in cells, a number of novel engagement vulnerabilities can be detected, that are not detectable using competing biochemical methods. These results underscore the value of live cell analysis when evaluating kinase selectivity.