Discovery of CaSR Peptide Agonists via Multistage Screening: In Silico Design, In Vitro Validation, and In Vivo...

Human calcium-sensing receptor (CaSR) is a class C G protein-coupled receptor (GPCR) that directly regulates parathyroid hormone release and maintains calcium homeostasis. The discovery of potent CaSR agonists with computer-aided drug design (CADD) has been appealing. Herein, we report the discovery of a series of new CaSR agonists by enhancing the molecular binding affinity through the replacement of key residues via the iCVETide platform. Ac-_D-Cys(_L-Cys)-_D-Arg-_D-(3-Gu)-Phe-_D-Abu-_D-Arg-_D-Ala-_D-Arg-NH₂ (compound 6g) with an outstanding activity and a satisfactory ADME profile is discovered, and the interaction mode between 6g and CaSR is elucidated through molecular docking and molecular dynamics simulations, showing hydrogen bonds, salt bridges, and π-π stacking. In addition, 6g is capable of activating human CaSR as a calcimimetic positive allosteric modulator. Our results provide a viable alternative to approved calcimimetics and a novel protocol for the discovery of CaSR agonists.