Monitoring PROTAC interactions in biochemical assays using Lumit immunoassays

Abstract

The discovery of new PROTAC molecules is dependent on robust and high-throughput assays to measure PROTAC-protein interactions and ternary complex formation. Here we present the optimization and execution of Lumit Immunoassays to measure PROTAC binding and ternary complex formation in a biochemical format. We demonstrate how Lumit can be used to rank order affinities of small molecules and PROTACs to BRD4(BD1, BD2) and how to measure PROTAC-mediated ternary complex formation of BRD4(BD1, BD2) and E3 Ligase VHL. Results from both biochemical assays correlate with live and lytic cell assays, indicating that Lumit Immunoassays can be used as a high-throughput compatible screening methodology to test new small molecules.

Keywords: BRD4; Immunoassay; Lumit; PROTAC; Proximity assay; Ternary complex; VHL.