Ureidopeptide GLP-1 analogues with prolonged activity <i>in vivo via</i> signal bias and altered receptor trafficking

Frémaux, Juliette; Venin, Claire; Mauran, Laura; Zimmer, R.; Koensgen, Florian; Rognan, Didier; Bitsi, Stavroula; Lucey, Maria; Jones, Ben; Tomás, Alejandra; Guichard, Gilles; Goudreau, Sébastien R.

doi:10.1039/c9sc02079a

Cited by 32 publications

(51 citation statements)

References 33 publications

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“…This translated to improved insulin secretion in vitro and significantly better anti‐hyperglycaemic effect in vivo. These observations therefore add to the evidence that modifications to GLP‐1 antagonist N‐termini are capable of inducing functionally important signal bias (Zhang et al, 2015; Jones, Buenaventura, et al, 2018; Fremaux et al, 2019).…”

Section: Discussionsupporting

confidence: 57%

Signalling, trafficking and glucoregulatory properties of glucagon‐like peptide‐1 receptor agonists exendin‐4 and lixisenatide

Pickford

Lucey

Fang

et al. 2020

British J Pharmacology

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Background and Purpose Amino acid substitutions at the N‐termini of glucagon‐like peptide‐1 (GLP‐1) receptor agonist peptides result in distinct patterns of intracellular signalling, sub‐cellular trafficking and efficacy in vivo. Here, we to determine whether sequence differences at the ligand C‐termini of clinically approved GLP‐1 receptor agonists exendin‐4 and lixisenatide lead to similar phenomena. Experimental Approach Exendin‐4, lixisenatide and N‐terminally substituted analogues with biased signalling characteristics were compared across a range of in vitro trafficking and signalling assays in different cell types. Fluorescent ligands and new time‐resolved FRET approaches were developed to study agonist behaviours at the cellular and sub‐cellular level. Anti‐hyperglycaemic and anorectic effects of each parent ligand and their biased derivatives were assessed in mice. Key Results Lixisenatide and exendin‐4 showed equal binding affinity, but lixisenatide was fivefold less potent for cAMP signalling. Both peptides induced extensive GLP‐1 receptor clustering in the plasma membrane and were rapidly endocytosed, but the GLP‐1 receptor recycled more slowly to the cell surface after lixisenatide treatment. These combined deficits resulted in reduced maximal sustained insulin secretion and reduced anti‐hyperglycaemic and anorectic effects in mice with lixisenatide. N‐terminal substitution of His1 by Phe1 to both ligands had favourable effects on their pharmacology, resulting in improved insulin release and lowering of blood glucose. Conclusion and Implications Changes to the C‐terminus of exendin‐4 affect signalling potency and GLP‐1 receptor trafficking via mechanisms unrelated to GLP‐1 receptor occupancy. These differences were associated with changes in their ability to control blood glucose and therefore may be therapeutically relevant.

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Section: Discussionsupporting

confidence: 57%

Signalling, trafficking and glucoregulatory properties of glucagon‐like peptide‐1 receptor agonists exendin‐4 and lixisenatide

Pickford

Lucey

Fang

et al. 2020

British J Pharmacology

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“…Non-traditional substitutions can yield GLP-1 analogues that show distinctive behavior and these might also provide insight into agonist conformation. 47–50 To this end, we explored a second set of substitutions at the key Gly residue in GLP-1 and exendin-4. This experimental design was based on previous comparisons of the conformations and biological activities of conventional peptides (comprised entirely of α-amino acid residues) with the properties of analogues in which at least one α residue was replaced with a β-amino acid residue.…”

Section: Resultsmentioning

confidence: 99%

Structural and Functional Diversity among Agonist-Bound States of the GLP-1 Receptor

Zhao

et al. 2021

Preprint

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Recent advances in G protein-coupled receptor (GPCR) structural elucidation have strengthened previous hypotheses that multi-dimensional signal propagation mediated by these receptors is, in part, dependent on their conformational mobility. However, the relationship between receptor function and static structures determined via crystallography or cryo-electron microscopy is not always clear. This study examines the contribution of peptide agonist conformational plasticity to activation of the glucagon-like peptide-1 receptor (GLP-1R), an important clinical target. We employ variants of the peptides GLP-1 and exendin-4 to explore the interplay between helical propensity near the agonist N-terminus and the ability to bind to and activate the receptor. Cryo-EM analysis of a complex involving an exendin-4 analogue, the GLP-1R and Gs protein revealed two receptor conformers with distinct modes of peptide-receptor engagement. Our functional and structural data suggest that receptor conformational dynamics associated with flexibility of the peptide N-terminal activation domain may be a key determinant of agonist efficacy.

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“…Like many other GPCRs [43], entry into the endocytic pathway may be required for a full GLP-1R intracellular signalling response [9,44]. However, with prolonged pharmacological stimulations, mounting evidence from studies using biased agonists with distinctive GLP-1R trafficking characteristics [6,7,11,16,45] suggests that preservation of a minimum level of surface-resident GLP-1R becomes a limiting factor for insulin secretion. A fuller understanding of the range of factors influencing these processes is needed so they can be optimally leveraged for therapeutic benefit.…”

Section: Discussionmentioning

confidence: 99%

Ligand-Specific Factors Influencing GLP-1 Receptor Post-Endocytic Trafficking and Degradation in Pancreatic Beta Cells

Fang

Chen

Manchanda

et al. 2020

IJMS

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The glucagon-like peptide-1 receptor (GLP-1R) is an important regulator of blood glucose homeostasis. Ligand-specific differences in membrane trafficking of the GLP-1R influence its signalling properties and therapeutic potential in type 2 diabetes. Here, we have evaluated how different factors combine to control the post-endocytic trafficking of GLP-1R to recycling versus degradative pathways. Experiments were performed in primary islet cells, INS-1 832/3 clonal beta cells and HEK293 cells, using biorthogonal labelling of GLP-1R to determine its localisation and degradation after treatment with GLP-1, exendin-4 and several further GLP-1R agonist peptides. We also characterised the effect of a rare GLP1R coding variant, T149M, and the role of endosomal peptidase endothelin-converting enzyme-1 (ECE-1), in GLP1R trafficking. Our data reveal how treatment with GLP-1 versus exendin-4 is associated with preferential GLP-1R targeting towards a recycling pathway. GLP-1, but not exendin-4, is a substrate for ECE-1, and the resultant propensity to intra-endosomal degradation, in conjunction with differences in binding affinity, contributes to alterations in GLP-1R trafficking behaviours and degradation. The T149M GLP-1R variant shows reduced signalling and internalisation responses, which is likely to be due to disruption of the cytoplasmic region that couples to intracellular effectors. These observations provide insights into how ligand- and genotype-specific factors can influence GLP-1R trafficking.

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Ureidopeptide GLP-1 analogues with prolonged activity in vivo via signal bias and altered receptor trafficking

Abstract: This study demonstrates the efficacy of single α-amino acids substitution with ureido residues to design long lasting peptides.

Cited by 32 publications

References 33 publications

Signalling, trafficking and glucoregulatory properties of glucagon‐like peptide‐1 receptor agonists exendin‐4 and lixisenatide

Signalling, trafficking and glucoregulatory properties of glucagon‐like peptide‐1 receptor agonists exendin‐4 and lixisenatide

Structural and Functional Diversity among Agonist-Bound States of the GLP-1 Receptor

Ligand-Specific Factors Influencing GLP-1 Receptor Post-Endocytic Trafficking and Degradation in Pancreatic Beta Cells

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