Heterochiral Glucagon and PTH(1-34) Analogues Achieve Potent, Biased Class B1 GPCR Activation

Current treatments for conditions like severe hypoglycemia (using Glucagon) and osteoporosis (using PTH(1-34)) rely on peptides that activate specific Class B1 G-protein-coupled receptors (GPCRs). While effective, these peptides can induce broad signaling pathways, including those mediated by β-arrestin, which may contribute to side effects or limit therapeutic windows. Understanding and manipulating the conformational states of these peptides in their bound state, particularly their α-helical structure, offers a route to developing biased agonists that selectively activate desired G-protein pathways while minimizing β-arrestin recruitment, potentially leading to safer and more effective therapies.

Researchers developed a novel design strategy involving the replacement of multiple L-α-amino-acid residues with D-α-amino-acid residues at selected positions within Glucagon and PTH(1-34) sequences. This heterochiral modification was intended to destabilize the native α-helical conformation of the peptides. The team then synthesized these heterochiral glucagon analogues and heterochiral PTH(1-34) analogues and evaluated their activity in vitro. Primary endpoints included assessing receptor activation potency via cAMP production assays and evaluating signaling bias by measuring β-arrestin recruitment and receptor internalization relative to the all-L peptide counterparts.

The heterochiral design strategy successfully yielded potent agonists for both Glucagon and PTH(1-34) receptors. Despite the intended destabilization of the helical conformation, the synthesized heterochiral analogues demonstrated receptor activation potency (measured by cAMP production) that approached that of the all-L peptides. A significant finding was the observed signaling bias: the heterochiral agonists were consistently biased away from β-arrestin recruitment relative to their all-L peptide counterparts. This suggests a preferential activation of G-protein signaling over β-arrestin-mediated pathways. Furthermore, for the glucagon analogues, an additional bias was observed: they were also biased away from receptor internalization when compared to native glucagon. This dual bias (reduced β-arrestin and reduced internalization for glucagon) represents an unanticipated and potentially beneficial pharmacological profile. These findings were consistent across two distinct Class B1 GPCRs, highlighting the broad applicability of this heterochiral design approach.

The heterochiral agonists approached the all-L peptide in receptor activation potency (cAMP production) and were biased away from β-arrestin recruitment.