The Dual GLP-1/GIP Receptor Agonist DA4-JC Shows Superior Protective Properties Compared to the GLP-1 Analogue Liraglutide in the APP/PS1 Mouse Model of Alzheimer's Disease

Abstract

Alzheimer's disease (AD) is a neurodegenerative disorder for which there is no cure. Here, we test a dual GLP-1/GIP receptor agonist (DA4-JC) that has a cell penetrating sequence added to enhance blood-brain barrier penetration. We show in a receptor activity study that DA4-JC has balanced activity on both GLP-1 and GIP receptors but not on GLP-2 or Glucagon receptors. A dose-response study in the APP/PS1 mouse model of AD showed both a dose-dependent drug effect on the inflammation response and the reduction of amyloid plaques in the brain. When comparing DA4-JC with the GLP-1 analogue liraglutide at equal doses of 10nmol/kg bw ip. once-daily for 8 weeks, DA4-JC was more effective in reversing memory loss, enhancing synaptic plasticity (LTP) in the hippocampus, reducing amyloid plaques and lowering pro-inflammatory cytokine levels in the brain. The results suggest that DA4-JC may be a novel treatment for AD.

Keywords: TNF-alpha; brain; growth factor; inflammation; insulin; synaptic plasticity.

Figure 1.

Activation of hGLP1-R, hGIP-R, hGLP2-R and hGCG-R by endogenous agonist and DA4-JC and cAMP accumulation in COS-7 cells transfected with (A) hGLP-1 receptor, treated with increasing concentrations of hGLP-1 and DA4-JC (B) hGIP-R treated with increasing concentrations of hGIP and DA4-JC (C) hGLP2-R treated with increasing concentrations of hGLP-2 and DA4-JC (C) hGCG-R treated with increasing concentrations of hGCG and DA4-JC.

Figure 2.

DA4-JC4 dose-response study in the APP/PS1 mouse model of AD. A: Immunohistochemical analysis of amyloid plaque load in the cortex of 9-month old APP/PS1 mice. The 4 different treatments given to the APP/PS1 mice, are shown in the images above, presenting a reduced number of amyloid plaques in the cortex (scale bar 100 µm). Data shown as mean ± S.E.M. (One-way ANOVA, Tukey’s Multiple Comparison post-hoc test, *=p < 0.05, **=p < 0.01, ***=p < 0.001). B: Immunohistochemical analysis of the number of reactive astrocytes in the cortex of 9-month old APP/PS1 mice. DA4-JC treatment reduced the number of activated astrocytes in the brain. The 5 images are illustrating the different treatments given to the APP/PS1 mice, showing the reduction in the number of reactive astrocytes per section within the cortex (scale bar 100 µm). Data shown as mean ± S.E.M. (*=p < 0.05, **=p < 0.01, ***=p < 0.001). C: Immunohistochemical analysis of the number of activated microglia in the cortex of 9-month old APP/PS1 mice. DA4-JC treatment reduced the number of activated microglia in the brain. The 5 images are illustrating the different treatments given to the wild-type and APP/PS1 mice, showing the reduction in activated microglia within the cortex (scale bar 100 µm). Data shown as mean ± S.E.M. (*=p < 0.05, **=p < 0.01, ***=p < 0.001).

Figure 3.

(A): Acquisition times for water maze training. A 2-way ANOVA found an overall difference for drug treatment (p < 0.001) and over time (p < 0.001). Post-hoc tests showed differences between groups. (B): Probe test percentage of target quadrant swim times. A 1-way ANOVA found an overall difference between groups (p < 0.001). Post-hoc tests showed differences between groups. *=p < 0.05; **=p < 0.01; ***=p < 0.001. N = 12 per group.

Figure 4.

Long-term potentiation of synaptic transmission in area CA1 of the hippocampus is much improved by DA4-JC. A 2-way ANOVA found an overall difference between groups (p < 0.001) and over time (P < 0.001). The APP/PS1 DA4-JC group is significantly different from the other 3 groups. The APP/PS1 saline group is significantly different from the wild type (p < 0.001) and the APP/PS1 liraglutide group (p < 0.01). N = 6 per group.

Figure 5.

Quantification of beta-amyloid plaque loads in the neocortex. A 1-way ANOVA found an overall difference between groups (p < 0.001). DA4-JC was more effective in reducing the amyloid load compared to liraglutide in Tukey’s multiple comparison post-hoc tests. *=p < 0.05; **=p < 0.01; **=p < 0.001. N = 6-10 per group. Sample micrographs are shown. A = WT; B = APP/PS1 sal; C = APP/PS1 Lira; D = APP/PS1 DA4-JC. Scale bar = 50 µm.

Figure 6.

Quantification of pro-inflammatory cytokines in the brain. A 1-way ANOVA found an overall difference between groups (p < 0.001). Western blots were repeated 3 times. DA4-JC was more effective in reducing the levels compared to liraglutide in Tukey’s multiple comparison post-hoc tests. *=p < 0.05; **=p < 0.01; **=p < 0.001. Sample bands are shown below.