Designing allosteric modulators to change GPCR G protein subtype selectivity

G-protein-coupled receptors (GPCRs) convert extracellular signals into intracellular responses by signalling through 16 subtypes of Gα proteins and two β-arrestin proteins. Biased compounds-molecules that preferentially activate a subset of these proteins-engage therapy-relevant pathways more selectively¹ and promise to be safer, more effective medications than compounds that uniformly activate all pathways². However, the determinants of bias are poorly understood, and we lack rationally designed molecules that select for specific G proteins. Here, using the prototypical class A GPCR neurotensin receptor 1 (NTSR1), we show that small molecules that bind to the intracellular GPCR-transducer interface change G protein coupling by subtype-specific and predictable mechanisms, enabling structure-guided drug design. We find that the intracellular, core-binding compound SBI-553 switches the G protein preference of NTSR1 through direct intermolecular interactions^3-5, promoting or preventing association with specific G protein subtypes. Modifications to the SBI-553 scaffold produce allosteric modulators with distinct G protein selectivity profiles. Selectivity profiles are probe independent, conserved across species and translate to differences in activity in vivo. Our studies show that G protein selectivity can be tailored with small changes to a single chemical scaffold targeting the receptor-transducer interface. Moreover, given that this pocket is broadly conserved, our findings could provide a strategy for pathway-selective drug discovery that is applicable to the diverse GPCR superfamily.