1. The New Dual Glucagon-GLP-1 Agonist ZP2929 Improves Glycemic Control and Reduces Body Weight in Murine Models of Obesity and Type 2 Diabetes Jens R. Daugaard, Eddi Meier, Ditte Riber, Camilla Æ. Bæk & Kjeld S. Larsen Zealand Pharma A/S, Smedeland 36, DK-2600 Glostrup, Copenhagen, Denmark
Abstract
ZP2929 was created by amino acid substitutions in the glucagon sequence that improved GLP-1R potency, solubility at physiological pH and DPP-IV stability. The in vivo T½ was prolonged.
Oxyntomodulin is released from L cells of the small intestine in response to meal ingestion, and is believed to exert its biological effects by activating both the GLP-1 receptor and the glucagon receptor, i.e. by acting as a dual glucagon-GLP-1 agonist. ZP2929 is a potent dual glucagon-GLP-1 receptor agonist with a pharmacokinetic profile compatible with once daily dosing in humans. The effect of ZP2929 was studied in db/db mice, an animal model of type 2 diabetes and in 8 months high fat fed C57Bl/6J mice, an animal model of obesity. ZP2929 was compared to vehicle and exendin-4. The db/db mice were randomized into groups with similar average glycated haemoglobin (HbA1c). Mice (7-8 weeks of age) were treated twice daily s.c. with exendin-4 (0.2, 0.5, 2, 5, 10, and 20 nmol/kg), ZP2929 (0.6, 1.6, 3.2, 6.4, and 12.7 nmol/kg), or vehicle. Before treatment, and once weekly for six weeks of treatment, blood samples were taken and HbA1c measured. An increase in HbA1c over time was seen in the vehicle treated animals. ZP2929 at 3.2, 6.4 and 12.7 nmol/kg significantly prevented the increase in HbA1c after 14 days. The effect persisted for the full 42 days of treatment. The effect of the highest dose of ZP2929 (12.7 nmol/kg) and of a comparable dose of exendin-4 (10 nmol/kg), were similar over the 42 days. C57Bl/6J male mice were fed a high fat diet for 30 weeks. The mice (36 weeks of age) were treated twice daily s.c. with exendin-4 (10 nmol/kg), ZP2929 (10 nmol/kg) or vehicle for 21 days. ZP2929 induced 24.8 % body weight loss over the 21 days. Exendin-4 only gave an initial small decrease body weight. In conclusion, the new glucagon-GLP-1 agonist ZP2929 improves glycemic control and reduces body weight in murine models of obesity and type 2 diabetes with a possible cardiovascular benefit as a result.
Materials & Methods
GluR and GLP-1R efficacy: HEK293 cells expressing hGluR or hGLP-1-R were incubated with increasing concentrations of test peptides, 100 M IBMX and 6 mM glucose for 15 min at 37 C. cAMP content was measured using the FlashPlate cAMP kit. EC50 and relative efficacies were estimated by computer aided curve fitting.
Pharmacokinetics: Blood samples were collected from mice or monkeys at different time points following administration (s.c. or i.v) and the content of ZP2929 was measured by LC-MS/MS. Pharmacokinetic parameters were estimated by computer aided curve fitting.
Murine model of obesity: 30 weeks high fat fed (HFD) C57BL/6J mice (n = 9/group) were treated (s.c.) twice daily with vehicle (PBS), exendin-4 or ZP2929 for 3 weeks. The body weight was monitored daily and OGTT performed at the end of the experiment. Plasma samples were collected for later analysis of plasma lipids (Cobas cholesterol, HDL, LDL & triglyceride kit).
Murine model of type 2 diabetes: Db/db mice (n = 11/group) were treated (s.c.) twice daily with vehicle (PBS), exendin-4 or ZP2929. The body weight was monitored daily, while HbA1c were measured (Cobas HbA1c kit) on day 0,7, 14, 21, 28, 35 & 42.
Fig. 1. Three weeks treatment of HFD mice with ZP2929 decreases BW with 24.8 % compared to those treated with exendin-4 or vehicle.
Fig. 4. Treatment of db/db mice with ZP2929 diminishes their HbA1c increase by 0.90% compared to controls (1.3% vs. 0.4%).
ZP2929 Attributes
Background
ZP2929 is a full, potent and selective agonist on hGluR and hGLP-1R (tab. 1)
The pharmacokinetics of ZP2929 is well suited for once daily treatment (tab. 2)
ZP2929 improves glycemic parameters in obese and diabetic mice (figs. 2 & 4)
ZP2929 has a superior effect on body weight (fig. 1) and plasma lipid profile (fig. 3) compared to exendin-4 in weight stable obese mice
WHO estimates that the global number of obese diabetics will increase from today’s 220 million to 340 million by The pharmacological treatments of obese diabetics focus primarily on diabetes (decrease of blood glucose) and less so on obesity (decrease of body weight and fat). Dual GluGLP-1 agonists combine the effect of glucagon on fat2 metabolism with GLP-1 on blood glucose levels3.
Results
Table 1. GluR and GLP-1R Efficacy
Fig. 2. Three weeks treatment of HFD mice with ZP2929 improves OGTT.
Peptide
GLP-1R (logavg)
GluR (logavg)
GLP-1R (%)
GluR (%)
GLP-1
0,02
100000
100
Exendin-4
0,03
Liraglutide
0,14 16 99 1
Glucagon
2,3
0,18 7 93
Oxyntomodulin
1,0
0,48 32 68
ZP2929
0,11
0,16 59 41
Rationale
Conclusion
GLP Glucagon
Receptor
Glucagon
ZP2929
The dual GluGLP-1 agonist ZP2929 has clinical potential for once-daily treatment of obese type 2 diabetics with superior weight control and with a possible cardiovascular benefit
Table 2. Pharmacokinetic Parameters of ZP2929
References
Species
Dose (nmol/kg)
T½ I.V. (hours)
T½ S.C (hours)
Tmax (hours)
Fsc
(%)
Mouse 40
3.6
6.2
6.0 30
Monkey 20
5.6 ± 0.3
6.2 ± 0.7
6.0 ± 2.8
36 ± 11
WHO web site
Van Beek et al. 2004; European Journal of Endocrinology 151,
Zander et al. 2002; The Lancet 359,
Wynne et al., 2005; Diabetes 54,
Fig. 3. Three weeks treatment of HFD mice with ZP2929 decreases cholesterol, HDL, LDL and triglycerides and increases HDL/LDL ratio significantly. No significant effects with exendin-4 were observed.