A novel long-acting glucose-dependent insulinotropic peptide analogue: enhanced efficacy in normal and diabetic rodents

Abstract

Aim: Glucose-dependent insulinotropic peptide (GIP) is an incretin hormone that is released from intestinal K cells in response to nutrient ingestion. We aimed to investigate the therapeutic potential of the novel N- and C-terminally modified GIP analogue AC163794.

Methods: AC163794 was synthesized by solid-phase peptide synthesis. Design involved the substitution of the C-terminus tail region of the dipeptidyl peptidase IV (DPP-IV)-resistant GIP analogue [d-Ala(2) ]GIP(1-42) with the unique nine amino acid tail region of exenatide. The functional activity and binding of AC163794 to the GIP receptor were evaluated in RIN-m5F β-cells. In vitro metabolic stability was tested in human plasma and kidney membrane preparations. Acute insulinotropic effects were investigated in isolated mouse islets and during an intravenous glucose tolerance test in normal and diabetic Zucker fatty diabetic (ZDF) rats. The biological actions of AC163794 were comprehensively assessed in normal, ob/ob and high-fat-fed streptozotocin (STZ)-induced diabetic mice. Acute glucoregulatory effects of AC163794 were tested in diet-induced obese mice treated subchronically with AC3174, the exendatide analogue [Leu(14) ] exenatide. Human GIP or [d-Ala(2) ]GIP(1-42) were used for comparison.

Results: AC163794 exhibited nanomolar functional GIP receptor potency in vitro similar to GIP and [d-Ala(2) ]GIP(1-42). AC163794 was metabolically more stable in vitro and displayed longer duration of insulinotropic action in vivo versus GIP and [d-Ala(2) ]GIP(1-42). In diabetic mice, AC163794 improved HbA1c through enhanced insulinotropic action, partial restoration of pancreatic insulin content and improved insulin sensitivity with no adverse effects on fat storage and metabolism. AC163794 provided additional baseline glucose-lowering when injected to mice treated with AC3174.

Conclusions: These studies support the potential use of a novel GIP analogue AC163794 for the treatment of type 2 diabetes.

Keywords: DPP-IV resistant; GIP; Trp-cage; diabetes; rodents.

Figure 1

In vitro effects of AC163794 on glucose-stimulated insulin secretion in isolated mouse islets. Data are presented as fold increase over non-treated medium control (mean ± SEM). *p < 0.002 versus medium control (n = 3 experiments with three replicates).

Figure 2

Effect of a single bolus injection of AC163794 or glucose-dependent insulinotropic peptide (GIP) on blood glucose in diet-induced obese (DIO) mice treated with the glucagon-like peptide 1 (GLP-1) receptor agonist AC3174 for 4 weeks via continuous subcutaneous (SC) infusion. Data are presented as mean ± SEM of calculated percent change from the glucose values measured immediately prior a single injection of GIP or GIP analogue. Inset represents calculated AUC for this change and dashed line represents theoretical AUC value for the initial glucose concentration remaining unchanged during the study.*p < 0.05 versus GIP, Student’s t test n = 8.

Figure 3

Plasma glucose and insulin excursions in normal rats infused intravenously with glucose-dependent insulinotropic peptide (GIP) analogue peptides at 10, 30 and 100 pmol/kg/min during an intravenous glucose tolerance test (IVGTT). Effects of AC163794 on glucose (A) and insulin (C) excursions. Effects of GIP on glucose (B) and insulin (D) excursions. Insets in graphs A–D represent calculated AUC for specific peptide and specific excursion curves. (E) Calculated slope of pancreatic function curve from experiments presented in A–D. (F) Duration of insulinotropic action of AC163794 and [d-Ala²]GIP(1–42) administered as a single bolus 120 min prior to a glucose challenge (IVGTT). Data are presented as mean ± SEM. *p < 0.05 versus vehicle control (n = 6–9 control group and n = 4–6 peptide-treated groups).

Figure 4

Plasma glucose (A) and insulin (B) excursions in diabetic Zucker fatty diabetic (ZDF) rats infused intravenously with 100 pmol/kg/min AC163794 and glucose-dependent insulinotropic peptide (GIP) during an intravenous glucose tolerance test (IVGTT). Insets represent calculated AUC for glucose (A) and insulin (B) excursion curves. Data are presented as mean ± SEM. *p < 0.05 versus vehicle control (n = 5–7 per group).

Figure 5

Effects of 4-week continuous administration of AC163794 on HbA1c (A), terminal pancreatic insulin content (B) and body weight (C) in diabetic ob/ob mice. Data are presented as mean ± SEM. *p < 0.05 versus vehicle control [n = 9 (A and C), n = 5 (B)].

Figure 6

Effects of 4-week continuous administration of AC163794 on HbA1c (A), terminal pancreatic insulin content (B) and body weight (C) in 16-week old high-fat-fed streptozotocin (HF-STZ) diabetic mice. Mice were fed a HF-diet for 12 weeks prior to the initiation of treatment. Data are presented as mean ± SEM. *p < 0.05 versus vehicle HF-STZ control [n = 16 HF-STZ groups (A and C), n = 6–8 HF-STZ group (B), HF and LF control groups (A and C)].

Figure 7

Plasma glucose (A) and insulin (B) excursions during an oral glucose tolerance test (OGTT) performed in high-fat-fed streptozotocin (HF-STZ) diabetic mice after 4-week continuous treatment with 100 nmol/kg/day AC163794. Insets represent calculated AUC for glucose (A) and insulin (B) excursion curves. *p < 0.05 versus HF-STZ vehicle-treated control (n = 6–8).