Achieving signalling selectivity of ligands for the corticotropin‐releasing factor type 1 receptor by modifying the agonist's signalling domain

• Published in: British journal of pharmacology Vol. 151; no. 6; pp. 851 - 859
• Main Authors: Beyermann, M, Heinrich, N, Fechner, K, Furkert, J, Zhang, W, Kraetke, O, Bienert, M, Berger, H
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01-07-2007; Nature Publishing; Nature Publishing Group
• Subjects: Amphibian Proteins; Biological and medical sciences; cAMP; Cell Line; Cell Membrane; Corticotropin-Releasing Hormone - agonists; Corticotropin-Releasing Hormone - pharmacology; CRF receptor; Cyclic AMP - metabolism; functional selectivity; GTP-Binding Protein alpha Subunits, Gi-Go - metabolism; GTP-Binding Protein alpha Subunits, Gs - metabolism; G‐protein coupling; Humans; Ligands; Medical sciences; Peptide Hormones; Peptides; Pharmacology. Drug treatments; Protein Binding; Protein Conformation; receptor binding; Receptors, Corticotropin-Releasing Hormone - metabolism; Research Papers; Signal Transduction; signalling domain; signalling‐selective ligands; Structure-Activity Relationship; Urocortins; Agonist; signalling domain; Ligand; Corticotropin releasing factor; Cyclic AMP; Selectivity; cAMP; G-protein coupling; Hypothalamic hormone; CRF receptor; receptor binding; signalling- selective ligands; Hormone releasing factor; functional selectivity; G protein; Biological receptor
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1038/sj.bjp.0707293

Background and purpose:
Most of the pharmaceuticals target G‐protein‐coupled receptors (GPCRs) which can generally activate different signalling events. The aim of this study was to achieve functional selectivity of corticotropin‐releasing factor receptor type 1 (CRF1) ligands.
Experimental approach:
We systematically substituted urocortin, a natural peptide agonist of CRF1, with bulky amino acids (benzoyl‐phenylalanine, naphthylalanine) and determined the effect of the analogues on coupling of CRF1 to Gs‐ and Gi‐protein in human embryonic kidney cells, using receptor binding, [35S]‐GTPγS binding stimulation, and cAMP accumulation assays.
Key results:
Native ligands stimulated Gs and Gi activation through CRF1, resulting in stimulation and then inhibition of cAMP accumulation. Single replacements in urocortin at positions 6–15 led, dependent on the position and nature of the substituent, to ligands that conserved Gs activity, but were devoid of Gi activity, only stimulating cAMP accumulation, and competitively antagonized the Gi activation by sauvagine. In contrast, analogues with substitutions outside this sequence non‐selectively activated Gs and Gi, as urocortin did.
Conclusions and implications:
Modifications in a specific region, which we have called the signalling domain, in the polypeptide agonist urocortin resulted in analogues that behaved as agonists and, at the same time, antagonists for the activation of different G‐proteins by CRF1. This finding implies significant differences between active conformations of the receptor when coupled to different G‐proteins. A similar structural encoding of signalling information in other polypeptide hormone receptor ligands would result in a general concept for the development of signalling‐selective drug candidates.
British Journal of Pharmacology (2007) 151, 851–859; doi:10.1038/sj.bjp.0707293