Bias from within: intracellular allosterism and the ICL2 hub.

Abstract

Targeting the orthosteric site of G protein-coupled receptors (GPCRs) has long dominated drug discovery, but intracellular allosteric modulators (AMs) provide new avenues to enhance selectivity and fine-tune signaling. Recent structural advances have illuminated intracellular allosteric pockets that act as conformational control hubs for transducer-selective engagement, biased signaling, and noncanonical receptor activation. Among these, the transmembrane helix 3 (TM3)-intracellular loop 2 (ICL2)-TM4 intracellular pocket (hereafter, the ICL2 site) has emerged as a key determinant of GPCR signaling bias and a strategic target for intracellular AM development. We review emerging structure-activity relationships, mechanistic and evolutionary determinants of ICL2-targeted intracellular allostery, and key translational challenges including membrane accessibility, pharmacokinetics, and species divergence. Finally, we highlight structure-guided and affinity-based discovery strategies for engineering pathway-selective intracellular AMs with greater transducer control than conventional orthosteric ligands.