Nanobodies unlock new mechanisms to target G protein-coupled receptors.

Once considered tools to aid structural studies, single-domain antibodies (nanobodies) have emerged as a new class of G protein-coupled receptor (GPCR) ligands. Aided by camelid immunization and advances in synthetic nanobody library development and computational design platforms, nanobody discovery has expanded to all major GPCR families. Unlike traditional small-molecule ligands, nanobodies can engage extended nonconserved extracellular epitopes to stabilize distinct receptor conformations to achieve high receptor selectivity and enable unique modes of pharmacological modulation. Integration of these properties with classical antibody engineering approaches, such as the generation of bispecific or bivalent constructs, has the potential to enable additional therapeutic strategies, such as cell and tissue-specific targeting. Recent mechanistic studies provide insight into the strategies nanobodies employ to function as agonists, antagonists, inverse agonists, and allosteric modulators. These studies begin to reveal the general principles nanobodies use to selectively engage and regulate GPCRs and uncover strategies to evolve their pharmacological properties. Continued advancements in discovery methods will expand the repertoire of nanobody ligands across additional GPCRs and deepen our mechanistic understanding to accelerate the development of the next generation of GPCR therapeutics. SIGNIFICANCE STATEMENT: G protein-coupled receptors (GPCRs) are one of the most successful targets of small-molecule drugs but are virtually untouched by antibody therapeutics. Aided by advances in discovery strategies, nanobodies have emerged as an ideal antibody scaffold to modulate GPCR signaling. Recent mechanistic studies reveal the molecular approaches nanobodies employ to target GPCRs, providing new insight into their potential as therapeutic tools. These findings provide a path toward developing nanobodies to regulate GPCR function in diverse disease contexts.