A Novel Assay Platform for the Detection of Kinase-Inhibitor Binding in Intact Mammalian Cells

Version:: V1
File Name/Number:: 2013 SLAS
Year:: 2013

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Assays that directly detect compound binding inside cells can be of great utility in translating in vitro data to more biologically relevant models. However, these types of intracellular binding assays have not been possible so far. Cellular assays for serveral targets are limited to downstream activity-based readouts which can be difficult to develop, may not be proximal to the target, or may require reagents or knowledge that does not currently exist. Here, we describe the development of a novel cell-based assay platform for the detection of ompound binding to intracellular proteins termed InCELL Hunter™. This assay platform detects the binding of a small molecule to a protein target in intact mammalian cells by monitoring changes in protein structure and stability. Using this assay, we have profiled a number of kinase targets and show that the system can detect allosteric as well as ATP-competitive inhibitors and can appropriately rank-order compounds for cellular potency. Since the system is activity-independent, HTS-friendly cell-based assays can be developed for kinases where the substrates are unknown or antibodies have not been developed, greatly expanding the number of immediately tractable kinase targets.