Drug Target Strategies
A Quantitative Target Engagement Approach to Profile Compound Affinity and Residence Time Across Enzyme Classes In Live Cells
Tuesday, February 6
4:30 PM - 5:00 PM
Intracellular target selectivity is fundamental to pharmacological mechanism. Although there are currently a number of acellular techniques to quantitatively measure target binding or enzymatic inhibition, no biophysical approach exists that offer quantitative, equilibrium-based analysis of target engagement across enzyme classes in live cells. Here we report the application of an energy transfer technique (NanoBRET) that enables the first quantitative approach to broadly profile target occupancy, compound affinity, and residence time for a variety of target classes including kinases and chromatin modifying enzymes. The NanoBRET method allows for broad kinome profiling of inhibitor selectivity against nearly 200 kinases, and enables a mechanistic interrogation of the potency offsets observed between cellular and acellular analysis. Compared to published biochemical profiling results, we observed an improved intracellular selectivity profile for certain clinically-relevant multi-kinase inhibitors. Due to high levels of intracellular ATP, a number of putative drug targets are unexpectedly disengaged in live cells at a clinically-relevant drug dose. The energy transfer technique can also be performed in real time, allowing for measurements of drug residence time. Broad kinase profiling of compound residence time reveals surprising kinetic selectivity mechanisms.