A Duplexed High-Throughput Screen to Identify Allosteric Modulators of the Glucagon-Like Peptide 1 and Glucagon Receptors

Injectable, degradation-resistant peptide agonists for the glucagon-like peptide 1 (GLP-1) receptor (GLP-1R), such as exenatide and liraglutide, activate the GLP-1R via a complex orthosteric-binding site and are effective therapeutics for glycemic control in type 2 diabetes. Orally bioavailable orthosteric small-molecule agonists are unlikely to be developed, whereas positive allosteric modulators (PAMs) may offer an improved therapeutic profile. We hypothesize that allosteric modulators of the GLP-1R would increase the potency and efficacy of native GLP-1 in a spatial and temporally preserved manner and/or may improve efficacy or side effects of injectable analogs. We report the design, optimization, and initial results of a duplexed high-throughput screen in which cell lines overexpressing either the GLP-1R or the glucagon receptor were coplated, loaded with a calcium-sensitive dye, and probed in a three-phase assay to identify agonists, antagonists, and potentiators of GLP-1, and potentiators of glucagon. 175,000 compounds were initially screened, and progression through secondary assays yielded 98 compounds with a variety of activities at the GLP-1R. Here, we describe five compounds possessing different patterns of modulation of the GLP-1R. These data uncover PAMs that may offer a drug-development pathway to enhancing in vivo efficacy of both endogenous GLP-1 and peptide analogs.

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Dual-acting peptide with prolonged glucagon-like peptide-1 receptor agonist and glucagon receptor antagonist activity for the treatment of type 2 diabetes

Journal of Endocrinology ◽

10.1677/joe-06-0018 ◽

2007 ◽

Vol 192 (2) ◽

pp. 371-380 ◽

Cited By ~ 35

Author(s):

Thomas H Claus ◽

Clark Q Pan ◽

Joanne M Buxton ◽

Ling Yang ◽

Jennifer C Reynolds ◽

...

Keyword(s):

Type 2 Diabetes ◽

Insulin Secretion ◽

Receptor Agonist ◽

Antagonist Activity ◽

Glucagon Receptor ◽

Glucagon Receptor Antagonist ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Type 2 diabetes is characterized by reduced insulin secretion from the pancreas and overproduction of glucose by the liver. Glucagon-like peptide-1 (GLP-1) promotes glucose-dependent insulin secretion from the pancreas, while glucagon promotes glucose output from the liver. Taking advantage of the homology between GLP-1 and glucagon, a GLP-1/glucagon hybrid peptide, dual-acting peptide for diabetes (DAPD), was identified with combined GLP-1 receptor agonist and glucagon receptor antagonist activity. To overcome its short plasma half-life DAPD was PEGylated, resulting in dramatically prolonged activity in vivo. PEGylated DAPD (PEG-DAPD) increases insulin and decreases glucose in a glucose tolerance test, evidence of GLP-1 receptor agonism. It also reduces blood glucose following a glucagon challenge and elevates fasting glucagon levels in mice, evidence of glucagon receptor antagonism. The PEG-DAPD effects on glucose tolerance are also observed in the presence of the GLP-1 antagonist peptide, exendin(9–39). An antidiabetic effect of PEG-DAPD is observed in db/db mice. Furthermore, PEGylation of DAPD eliminates the inhibition of gastrointestinal motility observed with GLP-1 and its analogues. Thus, PEG-DAPD has the potential to be developed as a novel dual-acting peptide to treat type 2 diabetes, with prolonged in vivo activity, and without the GI side-effects.

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Receptor occupancy of dual glucagon-like peptide 1/glucagon receptor agonist SAR425899 in individuals with type 2 diabetes

Scientific Reports ◽

10.1038/s41598-020-73815-5 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Olof Eriksson ◽

Torsten Haack ◽

Youssef Hijazi ◽

Lenore Teichert ◽

Veronique Tavernier ◽

...

Keyword(s):

Type 2 Diabetes ◽

Receptor Occupancy ◽

Strong Interactions ◽

Glucagon Receptor ◽

Glucose Levels ◽

Positron Emission ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Abstract Unimolecular dual agonists for the glucagon-like peptide 1 receptor (GLP1R) and glucagon receptor (GCGR) are emerging as a potential new class of important therapeutics in type 2 diabetes (T2D). Reliable and quantitative assessments of in vivo occupancy on each receptor would improve the understanding of the efficacy of this class of drugs. In this study we investigated the target occupancy of the dual agonist SAR425899 at the GLP1R in pancreas and GCGR in liver by Positron Emission Tomography/Computed Tomography (PET/CT). Patients with T2D were examined by [68Ga]Ga-DO3A-Tuna-2 and [68Ga]Ga-DO3A-Exendin4 by PET, to assess the GCGR in liver and GLP1R in pancreas, respectively. Follow up PET examinations were performed after 17 (GCGR) and 20 (GLP-1R) days of treatment with SAR425899, to assess the occupancy at each receptor. Six out of 13 included patients prematurely discontinued the study due to adverse events. SAR425899 at a dose of 0.2 mg daily demonstrated an average GCGR occupancy of 11.2 ± 14.4% (SD) in N = 5 patients and a GLP1R occupancy of 49.9 ± 13.3%. Fasting Plasma Glucose levels (− 3.30 ± 1.14 mmol/L) and body weight (− 3.87 ± 0.87%) were lowered under treatment with SAR425899. In conclusion, SAR425899 demonstrated strong interactions at the GLP1R, but no clear occupancy at the GCGR. The study demonstrates that quantitative target engagement of dual agonists can be assessed by PET.

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A synthetic glucagon-like peptide-1 analog with improved plasma stability

Journal of Endocrinology ◽

10.1677/joe.0.1590093 ◽

1998 ◽

Vol 159 (1) ◽

pp. 93-102 ◽

Cited By ~ 44

Author(s):

U Ritzel ◽

U Leonhardt ◽

M Ottleben ◽

A Ruhmann ◽

K Eckart ◽

...

Keyword(s):

Amino Acid ◽

Plasma Insulin ◽

Rat Plasma ◽

Plasma Stability ◽

Terminal Amino Acid ◽

Terminal Amino ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Glucagon-like peptide-1 (GLP-1) is the most potent endogenous insulin-stimulating hormone. In the present study the plasma stability and biological activity of a GLP-1 analog, [Ser]GLP-1(7-36)amide, in which the second N-terminal amino acid alanine was replaced by serine, was evaluated in vitro and in vivo. Incubation of GLP-1 with human or rat plasma resulted in degradation of native GLP-1(7-36)amide to GLP-1(9-36)amide, while [Ser]GLP-1(7-36)amide was not significantly degraded by plasma enzymes. Using glucose-responsive HIT-T15 cells, [Ser]GLP-1(7-36)amide showed strong insulinotropic activity, which was inhibited by the specific GLP-1 receptor antagonist exendin-4(9-39)amide. Simultaneous i.v. injection of [Ser]GLP-1(7-36)amide and glucose in rats induced a twofold higher increase in plasma insulin levels than unmodified GLP-1(7-36)amide with glucose and a fivefold higher increase than glucose alone. [Ser]GLP-1(7-36)amide induced a 1.5-fold higher increase in plasma insulin than GLP-1(7-36)amide when given 1 h before i.v. application of glucose. The insulinotropic effect of [Ser]GLP-1(7-36)amide was suppressed by i.v. application of exendin-4(9-39)amide. The present data demonstrate that replacement of the second N-terminal amino acid alanine by serine improves the plasma stability of GLP-1(7-36)amide. The insulinotropic action in vitro and in vivo was not impaired significantly by this modification.

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Taspoglutide, an Analog of Human Glucagon-Like Peptide-1 with Enhanced Stability and in Vivo Potency

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jcem.95.5.9986 ◽

2010 ◽

Vol 95 (5) ◽

pp. 2516-2516

Author(s):

Elena Sebokova ◽

Andreas D. Christ ◽

Haiyan Wang ◽

Sabine Sewing ◽

Jesse Z. Dong ◽

...

Keyword(s):

Glucagon Like Peptide ◽

Glucagon Like Peptide 1 ◽

Enhanced Stability

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Fibroblast Growth Factor 21 (FGF21) and Glucagon-Like Peptide 1 Contribute to Diabetes Resistance in Glucagon Receptor-Deficient Mice

Diabetes ◽

10.2337/db13-0710 ◽

2013 ◽

Vol 63 (1) ◽

pp. 101-110 ◽

Cited By ~ 50

Author(s):

B. A. Omar ◽

B. Andersen ◽

J. Hald ◽

K. Raun ◽

E. Nishimura ◽

...

Keyword(s):

Growth Factor ◽

Fibroblast Growth Factor ◽

Fibroblast Growth Factor 21 ◽

Fibroblast Growth ◽

Glucagon Receptor ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1 ◽

Deficient Mice

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Enhanced glucagon-like peptide-1 secretion in a patient with glucagonoma: Implications for glucagon-like peptide-1 secretion from pancreatic α cells in vivo

Diabetes Research and Clinical Practice ◽

10.1016/j.diabres.2013.08.005 ◽

2013 ◽

Vol 102 (1) ◽

pp. e1-e4 ◽

Cited By ~ 6

Author(s):

Daisuke Yabe ◽

Mariyo Rokutan ◽

Yoshiyuki Miura ◽

Izumi Komoto ◽

Ryota Usui ◽

...

Keyword(s):

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

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Glucagon-like Peptide 1 Receptor Agonists, Diabetic Retinopathy and Angiogenesis: The AngioSafe Type 2 Diabetes Study

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/clinem/dgz069 ◽

2019 ◽

Vol 105 (4) ◽

pp. e1549-e1560 ◽

Cited By ~ 7

Author(s):

Bénédicte Gaborit ◽

Jean-Baptiste Julla ◽

Samaher Besbes ◽

Matthieu Proust ◽

Clara Vincentelli ◽

...

Keyword(s):

Type 2 Diabetes ◽

Diabetic Retinopathy ◽

Fundus Photography ◽

Endothelial Cell Proliferation ◽

Receptor Agonists ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Abstract Aims Recent trials provide conflicting results on the association between glucagon-like peptide 1 receptor agonists (GLP-1RA) and diabetic retinopathy (DR). The aim of the AngioSafe type 2 diabetes (T2D) study was to determine the role of GLP-1RA in angiogenesis using clinical and preclinical models. Methods We performed two studies in humans. In study 1, we investigated the effect of GLP-1RA exposure from T2D diagnosis on the severity of DR, as diagnosed with retinal imaging (fundus photography). In study 2, a randomized 4-week trial, we assessed the effect of liraglutide on circulating hematopoietic progenitor cells (HPCs), and angio-miRNAs. We then studied the experimental effect of Exendin-4, on key steps of angiogenesis: in vitro on human endothelial cell proliferation, survival and three-dimensional vascular morphogenesis; and in vivo on ischemia-induced neovascularization of the retina in mice. Results In the cohort of 3154 T2D patients, 10% displayed severe DR. In multivariate analysis, sex, disease duration, glycated hemoglobin (HbA1c), micro- and macroangiopathy, insulin therapy and hypertension remained strongly associated with severe DR, while no association was found with GLP-1RA exposure (o 1.139 [0.800–1.622], P = .47). We further showed no effect of liraglutide on HPCs, and angio-miRNAs. In vitro, we demonstrated that exendin-4 had no effect on proliferation and survival of human endothelial cells, no effect on total length and number of capillaries. Finally, in vivo, we showed that exendin-4 did not exert any negative effect on retinal neovascularization. Conclusions The AngioSafe T2D studies provide experimental and clinical data confirming no effect of GLP-1RA on angiogenesis and no association between GLP-1 exposure and severe DR.

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