Back-scattering interferometry (BSI) is an emerging label-free, conformation-sensitive detection technology for quantitative mass- and matrix-independent biophysical characterization of small molecule interaction with complex drug target proteins under native-like conditions (1). Integral membrane proteins such as GPCRs are critical targets for drug discovery but present a host of challenges to the investigation of their biophysical properties. Of paramount interest to drug discovery efforts is the characterization of the interaction of GPCRs with small molecule compounds as a component of library screening, mechanism of action (MOA) determination, drug candidate profiling, and other aspects of intermolecular binding that inform pharmacology and medicinal chemistry. The difficulty associated with obtaining small molecule affinity data for functionally intact GPCRs effectively restricts the range of assay techniques suited to quantifying these interactions in vitro.

Herein, we describe the application of BSI to the characterization of small molecule ligand binding to human GPR39 overexpressed in crude membrane fractions in free solution. GPR39 is a Zn2+-responsive GPCR under investigation as a therapeutic target for type-2 diabetes (2). The ability to measure the affinity of small molecule agonists such as Zn2+is especially novel, given the unfavorable mass ratio and fast off rate that complicates the use of more established binding assays. Results from screening representatives from multiple novel GPR39 agonist series is presented, including how BSI-derived affinity and functional assay-derived potency correlate for compounds of varying scaffolds.