Precision Aptamers Against a Native GPCR through Ligand-Guided Selection

Abstract G protein–coupled receptors (GPCRs) constitute the largest and most diverse class of membrane receptors encoded in the human genome. They detect a wide range of chemical and physical stimuli and transduce these signals into intracellular responses through highly regulated pathways. Reflecting their central role in physiology, GPCRs are among the most prominent targets in drug discovery. However, identifying ligands that recognize GPCRs in their native conformational and membrane context remains a significant challenge. Here, we report an expanded aptamer discovery platform based on ligand-guided selection (LIGS) to isolate aptamers against GPCRs in their native cellular state. Using the β2-adrenergic receptor (β2AR) as a model system and employing agonists and antagonists as competing ligands, we identified three aptamers with high specificity for β2AR. These aptamers exhibit selective binding to cell-surface β2AR, showing higher apparent affinity towards cell-membrane bound β2AR than toward the purified receptor, which is consistent with recognition of native receptor context. Beyond target recognition, we show that the selected aptamers induce rapid internalization, indicating functional engagement. Together, these findings establish ligand-guided selection as a generalizable strategy for the discovery of conformationally sensitive aptamers targeting GPCRs in their native membrane environments. Significance The ability to discover ligands for receptors that undergo dynamic conformational changes is essential for advancing targeted therapeutics. G protein–coupled receptors (GPCRs), among the most sought-after drug targets, exist in transient and heterogeneous conformational states that are difficult to replicate in purified or artificial systems. Here, we introduce a ligand discovery platform that leverages native receptor interactions with agonists and antagonists to enable the selection of nucleic acid ligands (aptamers) directly against GPCRs in their cellular context, eliminating the need for purified receptors. The resulting aptamers exhibit selective binding to membrane-bound receptors and display intracellular functionality, highlighting a broadly applicable strategy for discovering ligands that recognize and modulate GPCRs in their native environments. Competing Interest Statement The authors have declared no competing interest.