Glucagon-like peptide-1 (GLP-1) receptor agonism or DPP-4 inhibition does not accelerate neoplasia in carcinogen treated mice

doi:10.1016/j.regpep.2012.08.016

Regulatory Peptides

Volume 179, Issues 1–3, 10 November 2012, Pages 91-100

https://doi.org/10.1016/j.regpep.2012.08.016 Get rights and content

Abstract

Introduction

Glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2) are secreted in parallel from the intestinal endocrine cells after nutrient intake. GLP-1 is an incretin hormone and analogues are available for the treatment of type 2 diabetes mellitus (T2DM). GLP-2 is an intestinal growth hormone and is shown to promote growth of colonic adenomas in carcinogen treated mice. Both peptides are degraded by dipeptidyl peptidase-4 (DPP-4) into inactive metabolites. DPP-4 inhibitors are therefore also in use for treatment of T2DM. It is possible that DPP-4 inhibition by enhancing the exposure of endogenous GLP-2 to the intestinal epithelia also might mediate growth and promote neoplasia. We investigated the intestinal growth effect of the GLP-1 receptor agonists (GLP-1 RAs) (liraglutide and exenatide) and DPP-4 inhibition (sitagliptin) in healthy mice. We also investigated the potential tumour promoting effect of liraglutide and sitaglitin in the colon of carcinogen treated mice. We used GLP-2 as a positive control.

Methods

For the growth study we treated healthy CD1 mice with liraglutide (300 μg × 2), exenatide (12.5 μg × 2) or vehicle subcutaneously and sitagliptin (8 mg × 2) or water by oral gavage for 10 or 30 days. We measured intestinal weight, cross sectional area, villus height and crypt depth. For the tumour study we treated carcinogen treated mice (1,2 dimethylhydrazine 21 mg/kg/week for 12 weeks) with liraglutide (300 μg × 2), Gly2-GLP-2 (25 μg × 2) or vehicle subcutaneously and sitagliptin (8 mg × 2) or water by oral gavage for 45 days. We counted aberrant crypt foci (ACF), mucin depleted foci (MDF) and adenomas in the colon. Using COS-7 cells transfected with a GLP-2 receptor, we tested if liraglutide or exenatide could activate the receptor.

Results

In the 10 days experiment the relative small intestinal weight was increased with 56% in the liraglutide group (p < 0.001) and 26% in the exenatide group (p < 01) compared with vehicle treated mice.

After 30 days of treatment, liraglutide did also increase the colonic weight (p < 0.01). By morphometry the growth pattern mimicked that of GLP-2. Sitagliptin treatment had only a minor effect.

In the carcinogen treated mice we found no increase of ACF in any of the groups, the numbers of MDF and adenomas after liraglutide and sitagliptin treatments were similar to their respective control groups. Neither liraglutide nor exenatide stimulated cAMP release from GLP-2 receptor transfected cells.

Conclusion

Both GLP-1 analogues were potent growth stimulators of the healthy mouse intestine. No agonism was found for GLP-1 RAs at the GLP-2 receptor. Despite of the growth effect, liraglutide did not promote dysplasia in the colon. Sitagliptin did not show any tumour promoting effects, and non considerable growth effects.

Highlights

► GLP-1 receptor agonists and DPP-4 inhibitors are extensively used as anti-diabetic drugs. ► We investigated if GLP-RAs and a DPP-4 inhibitor had trophic effects in the mouse intestine. ► We also investigated if a GLP-RA and a DPP-4 inhibitor had tumour promoting effects in the colon of carcinogen treated mice. ► GLP-1 RAs significantly increased intestinal growth, but we found no tumour promoting effect in the colon.

Introduction

Glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2) are encoded by the proglucagon gene and, after nutrient stimulation, are secreted in parallel from the enteroendocrine L-cells of the intestinal epithelium. GLP-1 is an incretin hormone, which stimulates the pancreatic beta (ß)-cells to secrete insulin in a glucose-dependent manner [1]. GLP-2 plays a role in the adaptive response of the intestine by regulating the proliferation and inhibition of apoptosis in the intestinal mucosa [2]. Both peptides are rapidly degraded by the enzyme dipeptidyl peptidase-4 (DPP-4), which cleaves the peptides after the first two amino acids. GLP-1 has an in vivo half-life of approximately 1–2 min [1], whereas GLP-2 has a half-life of approximately 7 min [3]. Because of the anti-diabetic properties of GLP-1, DPP-4-resistant GLP-1 receptor agonists (GLP-1 RAs) have been developed. Exenatide (Byetta) is a synthetic version of exendin-4, a hormone found in the saliva of the Gila monster, which has a 50% amino-acid homology to GLP-1 and a longer in vivo half-life [4]. Liraglutide (Victoza) is an acylated GLP-1 analogue that binds to albumin and thereby avoids DPP-4 degradation [5]. Both drugs are administered by subcutaneous injection; exenatide is administered twice daily, and liraglutide is administered once daily.

By inhibiting the enzyme DPP-4, it is possible to increase the level of endogenous GLP-1 and thereby lower blood glucose. Inhibitors of DPP-4 are now available for the treatment of type 2 diabetes mellitus; the first to enter the market was sitagliptin (Januvia). In addition to their anti-diabetic properties, GLP-1 RAs have the potential to treat obesity as well [6]. GLP-1 has been shown to increase ß-cell mass by stimulating ß-cell proliferation [7], [8] and inhibiting ß-cell apoptosis [9]. In 2007, Simonsen et al. described that like GLP-2, prolonged treatment with exendin-4 increased the weight of the small intestine in rats [10]. We have previously demonstrated that the intestinal growth-promoting effect of GLP-2 is associated with accelerated rates of colonic neoplasia in carcinogen-treated mice [11]. Because GLP-1 also appears to stimulate intestinal growth it is necessary to investigate whether GLP-1 RAs have a similar effect. Moreover, DPP-4 inhibition might result in higher levels of both endogenous GLP-1 and GLP-2 because GLP-2 degradation is also inhibited. The aims of our study were to determine the growth effects of exenatide, liraglutide and sitagliptin on the small intestine and colon of mice and to determine whether liraglutide or sitagliptin, like exogenous GLP-2, promotes colonic neoplasia in carcinogen-treated mice.

Section snippets

Animals

The animal studies were approved by the Danish National Committee for Animal Studies (2008/561-1579), and standards equivalent to the EU Directive 2010/63/EU for animal experiments were followed. A total of 128 female CD-1 mice (Taconic, Ejby, Denmark) weighing approximately 25 g were used in the intestinal growth study, and 120 female C57 black 6 mice were used in the tumour study. The mice were housed in air-conditioned (21 °C), humidity-controlled (55%) rooms with a 12-hour light/12-hour dark

Body weight changes and intestinal weight

No differences with regard to body weight changes were observed between any of the groups, except for a small increase in body weight gain in mice treated with Gly2-GLP-2 for 30 days (Table 1).

When the intestinal weights were measured after 10 days, liraglutide, exenatide and Gly2-GLP-2, had a clear and significant trophic effect in the small intestine. The weight of the small intestine increased to 6.1% (± 0.22%) in the liraglutide-treated mice, 4.9% (± 0.23%) in the exenatide-treated mice and

Discussion and conclusion

A primary finding in the present study is that liraglutide significantly increases the weight of the small intestine; this was observed both after 10 and 30 days of treatment. Treatment with exenatide for 10 or 30 days did also increase small intestinal weight significantly. There was also a significant increase in the colon weight following liraglutide treatment (Fig. 1).

A second primary finding was that neither liraglutide nor sitagliptin accelerated colonic dysplasia, though we confirmed our

Funding

This study was funded by the Danielsen Foundation of Denmark. The foundation had no role in the design and conduct of the study, the collection, management, analysis and interpretation of the data, or the preparation, review or approval of the manuscript.

Author contributions

HK designed and performed the majority of the experiments and wrote the manuscript. BH, KH and NV assisted in designing and performing the experiments and revised the manuscript. BH analysed the plasma samples. LH and MR designed and performed the receptor-signalling study. JJH and SSP supervised the experiments and wrote and revised the manuscript. All authors have approved the final article.

Disclosure statement

HK, KH, SSP, LH, BH and MR declare neither financial relationships with any organisations that might have an interest in the submitted work during the previous three years nor any other relationships or activities that could appear to have influenced the submitted work. JJH declares serving as a consultant or advisor to Novartis Pharmaceuticals, Novo Nordisk, Merck, Sharp and Dome and Roche and to have received a fee for speaking from Novo Nordisk, Merck, Sharp and Dome and Glaxo SmithKline

Acknowledgements

The authors thank Heidi Marie Paulsen and Lise Strange for their technical support, Lars Thim for the synthesis of Gly2-GLP-2 and Grazyna Hahn for artwork support.

References (37)

J. Eng et al.
Isolation and characterization of exendin-4, an exendin-3 analogue, from Heloderma suspectum venom. Further evidence for an exendin receptor on dispersed acini from guinea pig pancreas
J Biol Chem
(1992)
A. Astrup et al.
Effects of liraglutide in the treatment of obesity: a randomised, double-blind, placebo-controlled study
Lancet
(2009)
B. Hartmann et al.
Structure, measurement, and secretion of human glucagon-like peptide-2
Peptides
(2000)
R.P. Bird
Observation and quantification of aberrant crypts in the murine colon treated with a colon carcinogen: preliminary findings
Cancer Lett
(1987)
F.L. Graham et al.
A new technique for the assay of infectivity of human adenovirus 5 DNA
Virology
(1973)
Y. Wei et al.
Tissue-specific expression of the human receptor for glucagon-like peptide-I: brain, heart and pancreatic forms have the same deduced amino acid sequences
FEBS Lett
(1995)
J.L. Dunphy et al.
Tissue distribution of rat glucagon receptor and GLP-1 receptor gene expression
Mol Cell Endocrinol
(1998)
E.J. Gallagher et al.
The proliferating role of insulin and insulin-like growth factors in cancer
Trends Endocrinol Metab
(2010)
K. Masur et al.
DPPIV inhibitors extend GLP-2 mediated tumour promoting effects on intestinal cancer cells
Regul Pept
(2006)
J. Thulesen et al.
The truncated metabolite GLP-2 (3–33) interacts with the GLP-2 receptor as a partial agonist
Regul Pept
(2002)

J.J. Holst

The physiology of glucagon-like peptide 1

Physiol Rev

(2007)

M.A. Ghatei et al.

Proglucagon-derived peptides in intestinal epithelial proliferation: glucagon-like peptide-2 is a major mediator of intestinal epithelial proliferation in rats

Dig Dis Sci

(2001)

B. Hartmann et al.

In vivo and in vitro degradation of glucagon-like peptide-2 in humans

J Clin Endocrinol Metab

(2000)

A. Lund et al.

Emerging GLP-1 receptor agonists

Expert Opin Emerg Drugs

(2011)

D.J. Drucker

Glucagon-like peptides: regulators of cell proliferation, differentiation, and apoptosis

Mol Endocrinol

(2003)

G. Xu et al.

Exendin-4 stimulates both beta-cell replication and neogenesis, resulting in increased beta-cell mass and improved glucose tolerance in diabetic rats

Diabetes

(1999)

J. Buteau et al.

Glucagon-like peptide-1 prevents beta cell glucolipotoxicity

Diabetologia

(2004)

L. Simonsen et al.

Exendin-4, but not dipeptidyl peptidase IV inhibition, increases small intestinal mass in GK rats

Am J Physiol Gastrointest Liver Physiol

(2007)

Cited by (81)

Immunomodulation and inflammation: Role of GLP-1R and GIPR expressing cells within the gut
2024, Peptides
Glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are peptide hormones produced by enteroendocrine cells in the small intestine. Despite being produced in the gut, the leveraging of their role in potentiating glucose-stimulated insulin secretion, also known as the incretin effect, has distracted from discernment of direct intestinal signaling circuits. Both preclinical and clinical evidence have highlighted a role for the incretins in inflammation. In this review, we highlight the discoveries of GLP-1 receptor (GLP-1R)+ natural (TCRαβ and TCRγδ) and induced (TCRαβ+CD4+ cells and TCRαβ+CD8αβ+) intraepithelial lymphocytes. Both endogenous signaling and pharmacological activation of GLP-1R impact local and systemic inflammation, the gut microbiota, whole-body metabolism, as well as the control of GLP-1 bioavailability. While GIPR signaling has been documented to impact hematopoiesis, the impact of these bone marrow-derived cells in gut immunology is not well understood. We uncover gaps in the literature of the evaluation of the impact of sex in these GLP-1R and GIP receptor (GIPR) signaling circuits and provide speculations of the maintenance roles these hormones play within the gut in the fasting-refeeding cycles. GLP-1R agonists and GLP-1R/GIPR agonists are widely used as treatments for diabetes and weight loss, respectively; however, their impact on gut homeostasis has not been fully explored. Advancing our understanding of the roles of GLP-1R and GIPR signaling within the gut at homeostasis as well as metabolic and inflammatory diseases may provide targets to improve disease management.
Intestinal adaptation and rehabilitation
2023, Seminars in Pediatric Surgery
Massive intestinal resection is a regrettably necessary but life-saving intervention for progressive or fulminant necrotizing enterocolitis (NEC). However, the resultant short bowel syndrome (SBS) poses its own array of challenges and complications. Within hours of such an abrupt loss of intestinal length, the intestine begins to adapt. Our ability to understand this process of intestinal adaptation has proven critical in our ability to clinically treat the challenging problem of short bowel syndrome. This review first highlights key data relating to intestinal adaptation including structural and functional changes, biochemical regulation, and other factors affecting the magnitude of intestinal adaptation responses. We then focus on intestinal rehabilitation as it relates to strategies to enhance intestinal adaptation while meeting nutritional needs and preventing complications of parenteral nutrition.
The beneficial mechanism of chitosan and chitooligosaccharides in the intestine on different health status
2022, Journal of Functional Foods
Citation Excerpt :
In the study related to intestinal Caco-2 cells and 3 T3-L1 cells, chitosan at a low molecular weight was found to inhibit the activities of intestinal α-glucosidase, Sodium/Glucose Cotransporter 1 (SGLT1) and glucose transporter-2 (GLUT2) (Yu, Kwon, Lee, Apostolidis, & Kim, 2017). Furthermore, diabetes is generally accompanied by an imperfection of the intestinal barrier and inflammation (Kissow et al., 2012). It was demonstrated that COS treatment could inhibit the phosphorylation of P38, so as to reverse the reduction of AMPK protein in proximal colon of diabetic mice, which can enhance the tight junction of intestine by up-regulating the expressions of ZO-1 and occluding (Zheng et al., 2018).
Chitosan and Chitooligosaccharides can serve as prominent food materials with high biological activities, of which the intestinal flora regulation is critical to probiotic regulation as reported by increasing studies. However, no such review has summarized their regulation about intestinal flora and the molecular mechanisms on the intestinal part. In this review, their intestinal manipulating mechanisms on healthy, obese, diabetic individuals and those with other diseases are mainly summarized. In sum, they improve the intestinal villi on healthy individuals by up-regulating Bifidobacteri and Lactobacill and reducing the lipids absorption in the obese intestine by up-regulating weight reducing bacteria (e.g., Coprobacillus cateniformis and Clostridium leptum). Moreover, they inhibit the intestinal absorption of glucose and regulate certain diabetes-related bacteria, reducing intestinal inflammation and up-regulating the tight junction protein expression in epithelial cells of their anti-diabetic effects. This review is aiming to provide some mechanism study of chitosan and chitooligosaccharides as food materials.
Crosstalk between incretin hormones, Th17 and Treg cells in inflammatory diseases
2022, Peptides
Citation Excerpt :
Studies have shown that GLP-1 induces activation of B-RAF/ extracellular signal-regulated kinase (ERK) signaling in human islet cells [144]. The action of GLP-1 analogs and agonists on GLP-1R in a colorectal cancer model has been investigated, although these drugs did not affect tumor mass growth [131]. Nevertheless, in an experimental model of spontaneous colorectal cancer development (APCmin/+model), animals without GLP-1/GLP-1R signaling showed decreased tumorigenesis [145].
Intestinal epithelial cells constantly crosstalk with the gut microbiota and immune cells of the gut lamina propria. Enteroendocrine cells, secrete hormones, such as incretin hormones, which participate in host physiological events, such as stimulating insulin secretion, satiety, and glucose homeostasis. Interestingly, evidence suggests that the incretin pathway may influence immune cell activation. Consequently, drugs targeting the incretin hormone signaling pathway may ameliorate inflammatory diseases such as inflammatory bowel diseases, cancer, and autoimmune diseases. In this review, we discuss how these hormones may modulate two subsets of CD4 + T cells, the regulatory T cells (Treg)/Th17 axis important for gut homeostasis: thus, preventing the development and progression of inflammatory diseases. We also summarize the main experimental and clinical findings using drugs targeting the glucose‐dependent insulinotropic polypeptide (GIP) and glucagon‐like peptide (GLP-1) signaling pathways and their great impact on conditions in which the Treg/Th17 axis is disturbed such as inflammatory diseases and cancer. Understanding the role of incretin stimulation in immune cell activation and function, might contribute to new therapeutic designs for the treatment of inflammatory diseases, autoimmunity, and tumors.
GIP and GLP-2 together improve bone turnover in humans supporting GIPR-GLP-2R co-agonists as future osteoporosis treatment
2022, Pharmacological Research
Citation Excerpt :
The cells were used within the passage number of 15–40. Transient transfection of COS-7 cells was performed using the calcium phosphate precipitation method [31]. In brief, receptor DNA was added to a Tris EDTA buffer and a 2 M CaCl2 solution was added.
The intestinal hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-2 (GLP-2) are key regulators of postprandial bone turnover in humans. We hypothesized that GIP and GLP-2 co-administration would provide stronger effect on bone turnover than administration of the hormones separately, and tested this using subcutaneous injections of GIP and GLP-2 alone or in combination in humans. Guided by these findings, we designed series of GIPR-GLP-2R co-agonists as template for new osteoporosis treatment. The clinical experiment was a randomized cross-over design including 10 healthy men administered subcutaneous injections of GIP and GLP-2 alone or in combination. The GIPR-GLP-2R co-agonists were characterized in terms of binding and activation profiles on human and rodent GIP and GLP-2 receptors, and their pharmacokinetic (PK) profiles were improved by dipeptidyl peptidase-4 protection and site-directed lipidation. Co-administration of GIP and GLP-2 in humans resulted in an additive reduction in bone resorption superior to each hormone individually. The GIPR-GLP-2R co-agonists, designed by combining regions of importance for cognate receptor activation, obtained similar efficacies as the two native hormones and nanomolar potencies on both human receptors. The PK-improved co-agonists maintained receptor activity along with their prolonged half-lives. Finally, we found that the GIPR-GLP-2R co-agonists optimized toward the human receptors for bone remodeling are not feasible for use in rodent models. The successful development of potent and efficacious GIPR-GLP-2R co-agonists, combined with the improved effect on bone metabolism in humans by co-administration, support these co-agonists as a future osteoporosis treatment.
The GLP-1R agonist liraglutide limits hepatic lipotoxicity and inflammatory response in mice fed a methionine-choline deficient diet
2021, Translational Research
Nonalcoholic fatty liver disease (NAFLD) is the most common hepatic disorder related to type 2 diabetes (T2D). The disease can evolve toward nonalcoholic steatohepatitis (NASH), a state of hepatic inflammation and fibrosis. There is presently no drug that effectively improves and/or prevents NAFLD/NASH/fibrosis. GLP-1 receptor agonists (GLP-1Ra) are effective in treating T2D. As with the endogenous gut incretins, GLP-1Ra potentiate glucose-induced insulin secretion. In addition, GLP-1Ra limit food intake and weight gain, additional beneficial properties in the context of obesity/insulin-resistance. Nevertheless, these pleiotropic effects of GLP-1Ra complicate the elucidation of their direct action on the liver. In the present study, we used the classical methionine-choline deficient (MCD) dietary model to investigate the potential direct hepatic actions of the GLP-1Ra liraglutide. A 4-week infusion of liraglutide (570 µg/kg/day) did not impact body weight, fat accretion or glycemic control in MCD-diet fed mice, confirming the suitability of this model for avoiding confounding factors. Liraglutide treatment did not prevent lipid deposition in the liver of MCD-fed mice but limited the accumulation of C16 and C24-ceramide/sphingomyelin species. In addition, liraglutide treatment alleviated hepatic inflammation (in particular accumulation of M1 pro-inflammatory macrophages) and initiation of fibrosis. Liraglutide also influenced the composition of gut microbiota induced by the MCD-diet. This included recovery of a normal Bacteroides proportion and, among the Erysipelotrichaceae family, a shift between Allobaculum and Turicibacter genera. In conclusion, liraglutide prevents accumulation of C16 and C24-ceramides/sphingomyelins species, inflammation and initiation of fibrosis in MCD-diet-fed mice liver, suggesting beneficial hepatic actions independent of weight loss and global hepatic steatosis.

View all citing articles on Scopus

¹: Present address: Department of Cardiothoracic Surgery, Copenhagen University Hospital, Blegdamsvej 9, DK-2100 Copenhagen, Denmark.

²: Present address: Department of Gynecology and Obstetrics, Hvidovre Hospital, Kettegaard Alle 30, DK-2650 Hvidovre, Denmark.

View full text

Glucagon-like peptide-1 (GLP-1) receptor agonism or DPP-4 inhibition does not accelerate neoplasia in carcinogen treated mice

Abstract

Introduction

Methods

Results

Conclusion

Highlights

Introduction

Section snippets

Animals

Body weight changes and intestinal weight

Discussion and conclusion

Funding

Author contributions

Disclosure statement

Acknowledgements

J Biol Chem

Lancet

Peptides

Cancer Lett

Virology

FEBS Lett

Mol Cell Endocrinol

Trends Endocrinol Metab

Regul Pept

Regul Pept

The physiology of glucagon-like peptide 1

Physiol Rev

Proglucagon-derived peptides in intestinal epithelial proliferation: glucagon-like peptide-2 is a major mediator of intestinal epithelial proliferation in rats

Dig Dis Sci

In vivo and in vitro degradation of glucagon-like peptide-2 in humans

J Clin Endocrinol Metab

Emerging GLP-1 receptor agonists

Expert Opin Emerg Drugs

Glucagon-like peptides: regulators of cell proliferation, differentiation, and apoptosis

Mol Endocrinol

Exendin-4 stimulates both beta-cell replication and neogenesis, resulting in increased beta-cell mass and improved glucose tolerance in diabetic rats

Diabetes

Glucagon-like peptide-1 prevents beta cell glucolipotoxicity

Diabetologia

Exendin-4, but not dipeptidyl peptidase IV inhibition, increases small intestinal mass in GK rats

Am J Physiol Gastrointest Liver Physiol