N- and C-terminally truncated forms of glucose-dependent insulinotropic polypeptide are high-affinity competitive antagonists of the human GIP receptor

TY - JOUR

T1 - N- and C-terminally truncated forms of glucose-dependent insulinotropic polypeptide are high-affinity competitive antagonists of the human GIP receptor

AU - Hansen, L S

AU - Sparre-Ulrich, A H

AU - Christensen, M.

AU - Knop, F K

AU - Hartmann, B

AU - Holst, J J

AU - Rosenkilde, M M

N1 - This article is protected by copyright. All rights reserved.

PY - 2016/3/1

Y1 - 2016/3/1

N2 - Background and Purpose Glucose-dependent insulinotropic polypeptide (GIP) affects lipid, bone and glucose homeostasis. High-affinity ligands for the GIP receptor are needed to elucidate the physiological functions and pharmacological potential of GIP in vivo. GIP(1-30)NH2 is a naturally occurring truncation of GIP(1-42). Here, we have characterized eight N-terminal truncations of human GIP(1-30)NH2. Experimental Approach COS-7 cells were transiently transfected with human GIP receptors and assessed for cAMP accumulation upon ligand stimulation or competition binding with 125I-labelled GIP(1-42), GIP(1-30)NH2, GIP(2-30)NH2 or GIP(3-30)NH2. Key Results GIP(1-30)NH2 displaced 125I-GIP(1-42) as effectively as GIP(1-42) (Ki 0.75 nM), whereas the eight truncations displayed lower affinities (Ki 2.3-347 nM) with highest affinities for GIP(3-30)NH2 and GIP(5-30)NH2 (5-30)NH2. Only GIP(1-30)NH2 (Emax 100% of GIP(1-42)) and GIP(2-30)NH2 (Emax 20%) were agonists. GIP(2- to 9-30)NH2 displayed antagonism (IC50 12-450 nM) and Schild plot analyses identified GIP(3-30)NH2 and GIP(5-30)NH2 as competitive antagonists (Ki 15 nM). GIP(3-30) NH2 was a 26-fold more potent antagonist than GIP(3-42). Binding studies with agonist (125I-GIP(1-30)NH2), partial agonist (125I-GIP(2-30)NH2) and competitive antagonist (125I-GIP(3-30)NH2) revealed distinct receptor conformations for these three ligand classes. Conclusions and Implications The N-terminus is crucial for GIP agonist activity. Removal of the C-terminus of the endogenous GIP(3-42) creates another naturally occurring, more potent, antagonist GIP(3-30)NH2, which like GIP(5-30)NH2, was a high-affinity competitive antagonist. These peptides may be suitable tools for basic GIP research and future pharmacological interventions.

AB - Background and Purpose Glucose-dependent insulinotropic polypeptide (GIP) affects lipid, bone and glucose homeostasis. High-affinity ligands for the GIP receptor are needed to elucidate the physiological functions and pharmacological potential of GIP in vivo. GIP(1-30)NH2 is a naturally occurring truncation of GIP(1-42). Here, we have characterized eight N-terminal truncations of human GIP(1-30)NH2. Experimental Approach COS-7 cells were transiently transfected with human GIP receptors and assessed for cAMP accumulation upon ligand stimulation or competition binding with 125I-labelled GIP(1-42), GIP(1-30)NH2, GIP(2-30)NH2 or GIP(3-30)NH2. Key Results GIP(1-30)NH2 displaced 125I-GIP(1-42) as effectively as GIP(1-42) (Ki 0.75 nM), whereas the eight truncations displayed lower affinities (Ki 2.3-347 nM) with highest affinities for GIP(3-30)NH2 and GIP(5-30)NH2 (5-30)NH2. Only GIP(1-30)NH2 (Emax 100% of GIP(1-42)) and GIP(2-30)NH2 (Emax 20%) were agonists. GIP(2- to 9-30)NH2 displayed antagonism (IC50 12-450 nM) and Schild plot analyses identified GIP(3-30)NH2 and GIP(5-30)NH2 as competitive antagonists (Ki 15 nM). GIP(3-30) NH2 was a 26-fold more potent antagonist than GIP(3-42). Binding studies with agonist (125I-GIP(1-30)NH2), partial agonist (125I-GIP(2-30)NH2) and competitive antagonist (125I-GIP(3-30)NH2) revealed distinct receptor conformations for these three ligand classes. Conclusions and Implications The N-terminus is crucial for GIP agonist activity. Removal of the C-terminus of the endogenous GIP(3-42) creates another naturally occurring, more potent, antagonist GIP(3-30)NH2, which like GIP(5-30)NH2, was a high-affinity competitive antagonist. These peptides may be suitable tools for basic GIP research and future pharmacological interventions.

U2 - 10.1111/bph.13384

DO - 10.1111/bph.13384

M3 - Journal article

C2 - 26572091

SN - 0007-1188

VL - 173

SP - 826

EP - 838

JO - British Journal of Pharmacology

JF - British Journal of Pharmacology

IS - 5

ER -