Background and Purpose The gut hormone glucose-dependent insulinotropic polypeptide (GIP) signals via the GIP receptor (GIPR) resulting in postprandial processes such as potentiation of glucose-stimulated insulin secretion. Translation of results from rodent to human studies has, however, been challenged by contradictive therapeutic effects of GIPR-targeting compounds. We, therefore, investigated the variation between species focusing on GIPR desensitization and the role of the C-terminus. Experimental Approach Species variants of the GIPR were studied in vitro for endogenous ligand affinity, G protein activation (cAMP accumulation), recruitment of beta-arrestin, and internalization. Variants of the mouse, rat, and human GIPRs with swapped C-terminal tails were studied in parallel. Key Results The human GIPR is more prone to internalization than rodent GIPRs. Despite similar agonist affinities and potencies for Gαs-activation especially the mouse GIPR has a reduced receptor desensitization, internalization, and beta-arrestin recruitment. Using an enzyme-stable, long-acting GIP analogue, the species differences were even more pronounced. “Tail swapped” human, rat, and mouse GIPRs were all fully functional in their Gαs-coupling and the mouse GIPR regained internalization and beta-arrestin 2-recruitment properties with the human tail while the human GIPR lost the ability to recruit beta-arrestin 2 when its own C-terminus was replaced by the rat or mouse tail. Conclusion and Implications Desensitization of the human GIPR is dependent on the C-terminal tail. The diverse functionality of the C-terminal tail as well as receptor internalization patterns between species, especially human and mouse GIPRs, are important factors that could influence the preclinical therapeutic evaluation of GIPR targeting compounds.

Background and purpose The incretin hormone, glucose-dependent insulinotropic polypeptide (GIP), secreted by the enteroendocrine K-cells in the proximal intestine, may regulate lipid metabolism and adiposity but its exact role in these processes is unclear. Experimental approach We characterized in vitro and in vivo antagonistic properties of a novel GIP analogue, mGIPAnt-1. We further assessed the in vivo pharmacokinetic profile of this antagonist, as well as its ability to affect high-fat diet (HFD)-induced body weight gain in ovariectomized mice during an 8-week treatment period. Key results mGIPAnt-1 showed competitive antagonistic properties to the GIP receptor (GIPR) in vitro as it inhibited GIP-induced cAMP accumulation in COS-7 cells. Furthermore, mGIPAnt-1 was capable of inhibiting GIP-induced glucoregulatory and insulinotropic effects in vivo and has a favourable pharmacokinetic profile with a half-life of 7.2 hours in C57Bl6 female mice. Finally, sub-chronic treatment with mGIPAnt-1 in ovariectomized HFD mice resulted in a reduction of body weight and fat mass. Conclusion and Implications mGIPAnt-1 successfully inhibited acute GIP-induced effects in vitro and in vivo and sub-chronically induces resistance to HFD-induced weight gain in ovariectomized mice. Our results support the development of GIP antagonists for the therapy of obesity.

Background: Glucagon-like peptide-2 (GLP-2) is a 33 amino acid pro-glucagon-derived hormone produced in the intestinal enteroendocrine L-cells with trophic actions on both the gut and bones. GLP-2(1-33) is cleaved by the ubiquitous protease dipeptidyl peptidase-4 (DPP-4), resulting in GLP-2(3-33) with competitive antagonistic properties on the GLP-2 receptor (GLP-2R). Here we present two new hGLP-2 radioligands with different pharmacodynamic profiles. Experimental Approach: The methionine in position 10 of GLP-2(1-33) was substituted with tyrosine to enable oxidative iodination with incorporation of the iodine isotope [125I]. Similar substitution was done in GLP-2(3-33), thereby creating two new radioligands; an agonist [125]-hGLP-2(1-33,M10Y) and an antagonist [125]-hGLP-2(3-33,M10Y). Both were characterized regarding competition binding, binding kinetics and target tissue autoradiography. Key results: High and similar binding affinities for the human GLP-2R were observed for [125I]-hGLP-2(1-33,M10Y) and [125I]-hGLP-2(3-33,M10Y) with KD values of 59.3 nM and 40.6 nM, respectively. The M10Y substitution did not change the functional properties of GLP-2(1-33) or GLP-2(3-33). The antagonist [125I]-hGLP-2(3-33,M10Y) had higher Bmax and faster on-rate for the hGLP-2R compared to the agonist [125I]-hGLP-2(1-33,M10Y). Using autoradiography in mice strong labeling was observed in subepithelial myofibroblasts (SEMF) and pancreas islet cells. Both radioligands were selective for the GLP-2R, except for a low affinity binding to the GLP-1R (IC50 of 130 and 330 nM, respectively) Conclusion and implications: We successfully developed two new high affinity radioligands for GLP-2R studies and identified SEMF and pancreatic islets as target for GLP-2. It is uncertain whether binding in the pancreas islets results from GLP-2R or GLP-1R binding.