N- and C-terminally truncated forms of glucose-dependent insulinotropic polypeptide are high-affinity competitive antagonists of the human GIP receptor
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BACKGROUND AND PURPOSE: Glucose-dependent insulinotropic polypeptide (GIP) impacts lipid, bone, and glucose homeostasis. The GIP receptor belongs to G protein-coupled receptor family B1 and signals through GαS. High affinity ligands for in vivo use are needed to elucidate GIP's physiological functions and pharmacological potential. GIP(1-30)NH2 is a naturally occurring truncation of GIP(1-42). Here we characterize eight N-terminal trrncations of human GIP(1-30)NH2 : GIP(2- to 9-30)NH2 .
EXPERIMENTAL APPROACH: COS-7 cells were transiently transfected with the human GIP receptor and assessed for cAMP accumulation upon ligand stimulation or competition binding with (125) I-GIP(1-42), (125) I-GIP(1-30)NH2 , (125) I-GIP(2-30)NH2 , or (125) I-GIP(3-30)NH2 as radioligands.
KEY RESULTS: GIP(1-30)NH2 displaced (125) I-GIP(1-42) equally to GIP(1-42) (Ki 0.75 nM), whereas the eight variants displayed lower affinities (Ki 2.3-347 nM) with highest affinities of GIP(3-30)NH2 and (5-30)NH2 . Agonism was only observed for GIP(1-30)NH2 with an Emax on 100% of GIP(1-42) and GIP(2-30)NH2 (Emax 20%). GIP(2- to 9-30)NH2 displayed antagonism (IC50 12-450 nM) and right-shifts of the GIP(1-42)-response curve. Schild plot analyses identified GIP(3-30)NH2 and GIP(5-30)NH2 as competitive antagonists (Ki 15 nM). Importantly, GIP(3-30) antagonized with a 26-fold higher potency than GIP(3-42). Binding studies with agonist ((125) I-GIP(1-30)NH2 ), partial agonist ((125) I-GIP(2-30)NH2 ) and competitive antagonist ((125) I-GIP(3-30)NH2 ) revealed distinct receptor conformations for these three ligand classes.
CONCLUSIONS AND IMPLICATIONS: The N-terminus is crucial for GIP agonist functionality. Removal of the C-terminus of the naturally occurring DPP4-product GIP(3-42) creates another naturally occurring, but superior antagonist GIP(3-30)NH2 , that together with GIP(5-30)NH2 were high-affinity competitive antagonist and thus may be suitable tool compounds for basic GIP research and future pharmacological interventions.
|Journal||British Journal of Pharmacology|
|Number of pages||13|
|Publication status||Published - Mar 2016|