Conformationally Constrained CCK8 Analogues Obtained from a Rationally Designed Peptide Library as Ligands for Cholecystokinin Type B Receptor

• Published in: ChemMedChem 2006-09, Vol.1 (9), p.997-1006
• Main Authors: De Luca, Stefania, Saviano, Michele, Della Moglie, Raffaella, Digilio, Giuseppe, Bracco, Chiara, Aloj, Luigi, Tarallo, Laura, Pedone, Carlo, Morelli, Giancarlo
• Format: Article
• Language: English
• Subjects: cholecystokinin; Cholecystokinin - analogs & derivatives; Cholecystokinin - chemical synthesis; Cholecystokinin - chemistry; cyclic constraints; Drug Design; Ligands; Magnetic Resonance Spectroscopy; Models, Molecular; Molecular Conformation; NMR structures; Peptide Library; peptides; Peptides - chemical synthesis; Peptides - chemistry; receptors; Receptors, Cholecystokinin - antagonists & inhibitors
• ISSN: 1860-7179; 1860-7187
• DOI: 10.1002/cmdc.200600054

A library of 14 cyclic peptide analogues derived from the octapeptide C‐terminal sequence of the human cholecystokinin hormone (CCK(26–33), or CCK8) was designed, synthesized, and characterized. The 14 peptide analogues were rationally designed to specifically interact with the CCK type B receptor (CCKB‐R) on the basis of the structure of the bimolecular complex between CCK8 and the third extracellular loop of CCKB‐R, namely CCKB‐R(352–379). The rational design of new ligands for CCKB‐R has relied on stabilization by cyclic constraints of the structural motifs that bring the key residues of the ligand (especially Trp 30, Met 31, and Phe 33) in the proper spatial orientation for optimal interaction with the receptor. The binding affinity of the new ligands for CCKB‐R was assessed by displacement experiments of 111In‐radiolabeled CCK8 in cells that overexpress the CCKB receptor. The new ligands generally showed binding affinities lower than that of parent CCK8, with the best compounds having IC50 values around 10 μM. Structure–activity relationship data show that preservation of the Trp 30–Met 31 motif is essential and that the Phe 33 side chain must be present. NMR conformational studies of the compound with maximal binding affinity (cyclo‐B11, IC50=11 μM) in DPC micelles shows that this compound presents a turn‐like conformation centered at the Trp 30–Met 31 segment, as planned by rational design. Such a conformation is stabilized by its interaction with the micelle rather than by the cyclic constraint. A rationally designed peptide library provided CCK8 analogues as new ligands (in red) for the cholecystokinin type B receptor (blue). In vitro cellular assays were used to determine the binding affinity of the ligands, and IC50≈10 μM for the best compounds. NMR structural data of the lead compound confirm that the structure is stabilized by both the cyclic constraint and by interaction with the micelle.