A high throughput kinase inhibitor binding kinetics assay provides indicators of clinical success

Abstract: Drug development is a lengthy and costly process with high attrition rates. More than half of the drug candidates going into phase 2 clinical trials fail mainly due to lack of effectiveness. A feature associated to the efficacy and safety of drug candidates is how fast they bind (k_on), and dissociate (k_off) from their targets, and accordingly for how long they can occupy them (residence time). Ideally these kinetic parameters are determined in the preclinical stages, creating a need for high-throughput methods to profile hundreds and thousands of compounds. Moreover, comprehensive retrospective profiling of existing molecules from different clinical development stages can provide insights into the impact of target binding kinetics on the pharmacological profiles of drugs. Here, we present the determination of k_on and k_off values in a microtiter plate reader using time-resolved FRET (TR-FRET) for a panel of 40 kinase drug targets in cancer therapies against 270 inhibitors. This method features the rapid data acquisition needed to determine binding parameters and should be a useful tool in making informed choices about which drug candidates to promote into clinical trials.