Directly Measuring Target Engagement
Measuring target protein-drug occupancy in cells and correlating this target engagement with desired pharmacologic effects is important in drug discovery. NanoBRET™ technology provides a simple method to directly assess target engagement. “The NanoBRET system allowed me to measure engagement of our chemical inhibitors directly to our target in living cells and I am excited to apply the technique to other targets,” said Graeme Walker, Team Leader, Cellular Pharmacology Drug Discovery Unit at the Cancer Research UK Manchester Institute.
Measuring Target Residence in Live Cells
An even greater benefit of NanoBRET technology is the ability to measure target residence time, which is the duration of drug-protein interaction. It allows researchers to optimize drugs for enhanced action despite clearance from the system. Walker added, “[The NanoBRET technology] provided a simple means to measure compound residence time at the target while keeping the cells intact. This capability allowed our project team to elucidate a novel mechanism of action for our chemical probes.”
How NanoBRET Target Engagement Works
To detect drug-target interactions, NanoBRET target engagement uses cell-permeable fluorescent tracers designed to interact with NanoLuc-tagged target proteins. The result of interaction is energy transfer from the NanoLuc protein to the tracer, generating a measurable signal. If a small molecule drug candidate interacts with the target protein, the tracer and drug compete for binding and BRET signal diminishes. Residence time measurement relies on pre-equilibration of compound with cells, removal of excess compound, and addition of tracer. Real-time signal monitoring shows compounds with slow target dissociation impede tracer binding and slow BRET signal production.
An additional publication in Nature Methods—“Application of BRET to monitor ligand binding to GPCRs”—details the application of NanoBRET technology. The article features the study of cell-surface G-protein coupled receptors (GPCRs), an important drug target class.
For more information on the NanoBRET Target Engagement Technology, visit www.promega.com/TargetEngagement.
Promega Corporation is a leader in providing innovative solutions and technical support to the life sciences industry. The company’s 3,500 products enable scientists worldwide to advance their knowledge in genomics, proteomics, cellular analysis, drug discovery and human identification. Founded in 1978, the company is headquartered in Madison, WI, USA with branches in 16 countries and over 50 global distributors. For more information about Promega, visitwww.promega.com.