Exploring the signaling space of a GPCR using bivalent ligands with a rigid oligoproline backbone.

G protein-coupled receptors (GPCRs) are one of the most important drug-target classes in pharmaceutical industry. Their diversity in signaling, which can be modulated with drugs, permits the design of more effective and better-tolerated therapeutics. In this work, we have used rigid oligoproline backbones to generate bivalent ligands for the gastrin-releasing peptide receptor (GRPR) with a fixed distance between their recognition motifs. This allows the stabilization of GPCR dimers irrespective of their physiological occurrence and relevance, thus expanding the space for medicinal chemistry. Specifically, we observed that compounds presenting agonists or antagonists at 20- and 30-Å distance induce GRPR dimerization. Furthermore, we found that 1) compounds with two agonists at 20- and 30-Å distance that induce dimer formation show bias toward Gq efficacy, 2) dimers with 20- and 30-Å distance have different potencies toward β-arrestin-1 and β-arrestin-2, and 3) the divalent agonistic ligand with 10-Å distance specifically reduces Gq potency without affecting β-arrestin recruitment, pointing toward an allosteric effect. In summary, we show that rigid oligoproline backbones represent a tool to develop ligands with biased GPCR signaling.