B6-hGLP-1R Mice

Figure 1. Schematic representation of the gene editing strategy for generating B6-hGLP-1R mice. Part of the exon 1 sequence and part of the intron 1 sequence in the mouse Glp1r gene was replaced with “hGLP1R Exon 1~2 CDS (without signal peptide), hGLP1R Intron 2 and hGLP1R Exon 3~13 CDS - mGlp1r 3’UTR - hGH pA” while retaining the gene sequence encoding the signal peptide of the mouse Glp1r protein.

● Investigation of the pathogenesis of obesity and type 2 diabetes;

● Development and screening of drugs for obesity and type 2 diabetes;

● Research on other metabolic diseases such as cardiovascular and myocardial diseases ^[5];

● Study of the neuroprotective effect in nervous system diseases.

1. Expression of human GLP-1R gene and protein

Figure 2. Human GLP1R gene and protein expression in different tissues of C57BL6 wild-type (WT) and B6-hGLP-1R mice at 6 weeks of age. (A) qRT-PCR results showed that the human GLP-1R gene was significantly expressed in the lungs, large intestine, and brain of B6-hGLP-1R mice, while no human GLP-1R gene expression was detected in the lungs, large intestine*, and brain of WT mice (*p<0.05, **p<0.01). (B) Western blot results showed that GLP-1R protein was strongly expressed in the pancreas of B6-hGLP-1R mice, and weakly expressed in the lungs*.
*The expression level of this gene in the intestine is relatively low, and the sequence homology between human and mouse genes is high. The amplified product in the WT group may not be a human gene band;
*The GLP-1R antibody used for detection (INVITROGEN: PA5-97789) is a cross-reacting antibody for human and mouse.

 

2. Immunohistochemistry (IHC) of pancreatic tissue

Figure 3. Distribution of human GLP-1R protein in the pancreas of 6-week-old wild-type (C57BL/6N) and homozygous (HO) B6-hGLP-1R mice. Immunohistochemical (IHC) staining of the pancreas revealed strong GLP-1R protein expression in islets of B6-hGLP-1R mice*.
*The GLP-1R antibody used for detection (INVITROGEN: PA5-97789) is a cross-reacting antibody for human and mouse.

 

3. Immunofluorescence (IF) staining of pancreatic tissue

Figure 4. Distribution of human GLP-1R protein in the pancreas of 6-week-old wild-type (C57BL/6N) and homozygous (HO) B6-hGLP-1R mice. Immunofluorescent (IF) staining of the pancreas revealed strong GLP-1R protein expression in islets of B6-hGLP-1R mice*.
*The GLP-1R antibody used for detection (INVITROGEN: PA5-97789) is a cross-reacting antibody for human and mouse.

 

4. Pharmacodynamics (PD) of PF-06882961 in B6-hGLP-1R mice

Figure 5. PF-06882961* significantly reduced blood glucose in B6-hGLP-1R mice under a control diet (CD) and high-fat diet (HFD).
(A~D). Under a control diet (CD), PF-06882961 effectively reduced blood glucose in B6-hGLP-1R mice compared to wild-type mice, as evidenced by the significantly reduced area under the curve (AUC) for the Intraperitoneal glucose tolerance test (IPGTT) (A~B) and food intake (C~D).
(E~H). High-fat diet (HFD) effectively induced obesity in B6-hGLP-1R and wild-type mice (E~F), and PF-06882961 significantly reduced AUC for IPGTT in B6-hGLP-1R mice (G~H), indicating that PF-06882961 can improve glucose tolerance in obese B6-hGLP-1R mice. Since PF-06882961 is a human GLP-1R-specific agonist, the above results suggest that B6-hGLP-1R mice successfully express human GLP-1R protein and interact with PF-06882961 to achieve significant blood glucose lowering (8-week-old male mice, n=6, *p<0.05; **p<0.01; ***p<0.001).
PF-06882961 is a non-peptide GLP-1R agonist (GLP-1RA) that activates the canonical G protein signaling pathway only in GLP-1Rs with the Trp33^ECD structure ^[6].

 

5. Pharmacodynamics (PD) of Semaglutide in B6-hGLP-1R mice

Figure 6. Semaglutide* significantly reduced blood glucose and body weight in B6-hGLP-1R mice under a high-fat diet (HFD).
(A). After 12 weeks of HFD feeding, Semaglutide (30 nmol/kg, twice weekly, subcutaneous injection, male mice, 30-90 min before the start of the dark cycle) was administered to B6-hGLP-1R mice. The body weight, food intake, blood glucose levels before and after administration, and AUC for IPGTT were regularly monitored.
(B~E). Compared to the control group, Semaglutide administration significantly reduced the body weight (B), random blood glucose (C), and food intake of B6-hGLP-1R mice (D~E).
(F~G). Semaglutide significantly decreased the AUC for IPGTT in B6-hGLP-1R mice, suggesting that it can improve the glucose tolerance of B6-hGLP-1R mice and exhibit good blood-glucose-lowering effects.
(H~I). Compared to wild-type and control groups, the body weight of Semaglutide-treated B6-hGLP-1R mice at the endpoint was significantly decreased.
(*p<0.05; **p<0.01; ***p<0.001)
*Semaglutide is a GLP-1R agonist (GLP-1RA) with a longer drug half-life ^[7].

 

6. Pharmacodynamics (PD) of LY3502970 in B6-hGLP-1R mice

Figure 7. The LY3502970* significantly reduces blood glucose levels in 6-week-old male B6-hGLP-1R mice (Normal diet)**. Intraperitoneal glucose tolerance test (IPGTT) was used to detect the hypoglycemic effect of LY3502970, and B6-hGLP-1R mice were treated with different doses of LY3502970 at 5 hours before the start of the detection. The results demonstrate that LY3502970 treatment leads to a significant decrease in blood glucose content in B6-hGLP-1R mice, with a markedly reduced area under the curve (AUC) in the IPGTT. This indicates that the drug improves glucose tolerance in B6-hGLP-1R mice and has a favorable blood glucose-lowering effect (****p<0.0001).
*Orforglipron (LY3502970) is an orally active glucagon-like peptide-1 receptor (GLP-1R) agonist that significantly improves type 2 diabetes ^[8].
**Data provided by Cyagen's client.