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Development of high-affinity agonist- and antagonist radioligands for the GLP-2 receptor - powerful tools for the study of GLP-2 pharmacology
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  • Sarina Gadgaard,
  • Wijnand van der Velden,
  • Sine Schiellerup ,
  • Jenna Hunt ,
  • Maria Gabe ,
  • Johanne Windeløv,
  • Geke Boer,
  • Hannelouise Kissow,
  • Cathrine Ørskov,
  • Jens Holst,
  • Bolette Hartmann,
  • Mette Rosenkilde
Sarina Gadgaard
University of Copenhagen Faculty of Health and Medical Sciences
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Wijnand van der Velden
University of Copenhagen
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Sine Schiellerup
University of Copenhagen
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Jenna Hunt
University of Copenhagen
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Maria Gabe
University of Copenhagen
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Johanne Windeløv
University of Copenhagen
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Geke Boer
University of Copenhagen
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Hannelouise Kissow
University of Copenhagen
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Cathrine Ørskov
University of Copenhagen
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Jens Holst
University of Copenhagen
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Bolette Hartmann
University of Copenhagen
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Mette Rosenkilde
University of Copenhagen
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Abstract

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.

Peer review status:IN REVISION

18 Aug 2020Submitted to British Journal of Pharmacology
20 Aug 2020Assigned to Editor
20 Aug 2020Submission Checks Completed
09 Sep 2020Reviewer(s) Assigned
07 Dec 2020Review(s) Completed, Editorial Evaluation Pending
11 Dec 2020Editorial Decision: Revise Minor
22 Jun 20211st Revision Received