Main Research Findings

1) Administration of GLP-3R has been shown to effectively improve body weight and instances of diabetic kidney diseases, while also inhibiting the expression of various inflammatory factors.

2) Long term treatment with GLP-3R over the course of 48 weeks was able to significantly decrease body weight in individuals experiencing obesity.

Selected Data
1) The research team of Ma et al compared the therapeutic effects of peptides Liraglutide, Tirzepatide, and GLP-3R when administered to db/db mice experiencing diabetic kidney disease. For the purpose of this study 7 week old male C57BL/KSJ diabetic db/db and non-diabetic db/m mice were utilized. The mice were housed at the Laboratory Animal Center of Huai’an First People’s Hospital under standardized laboratory conditions, including a temperature of 21 °C, humidity of 45%, and a 12 hour light/dark cycle. Standard sterilized food and water were provided ad libitum. Prior to the initiation of the experiment, the mice underwent a one-week adaptive period to their environment [1].
At 8 weeks of age, the mice were randomly assigned into five groups that were defined as follows: db/m as the healthy control group, db/db as the diabetic control group, db/db + Liraglutide treatment, db/db + Tirzepatide treatment, and db/db + GLP-3R treatment. Each treatment group was housed in two separate cages, with each with three mice per cage. The experimental groups received daily subcutaneous injections of 10 nmol/kg of Liraglutide, Tirzepatide, or GLP-3R for 10 weeks. The db/m and db/db control groups received equivalent volumes of saline delivered via intraperitoneal injection. Doses of the compounds were adjusted weekly based on body weight changes. The mice’s body weight, as well as food and water intake, were also recorded weekly, while (HbA1c) levels in tail vein blood were assessed at the beginning and end of the experiment [1].
After 10 weeks, urine samples were collected over a 24-hour period from individually housed mice in metabolic cages. Urine volume was recorded, and the samples were stored at -80°C for further analysis. The mice were then anesthetized, and blood samples were obtained via the intraorbital venous plexus. Following the experimental period, the mice were euthanized by cervical dislocation. Kidney tissues were harvested, washed with normal saline, and processed for histological and immunohistochemical analysis. The tissues were fixed in 4% paraformaldehyde, dehydrated in ethanol, and embedded in paraffin wax. 3-μm sections were then prepared for hematoxylin and eosin staining as well as periodic acid-Schiff staining. Stained sections were examined under an optical microscope.
Biochemical parameters were measured using an automatic biochemistry analyzer, and the serum levels of alanine aminotransferase, aspartate aminotransferase, total cholesterol, triglycerides, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, blood urea nitrogen, uric acid, creatinine, and urinary albumin-to-creatinine ratio were assessed. Additionally, 24-hour urinary microalbumin excretion rate was quantified using an ELISA kit according to the manufacturer’s protocol. Serum butyrate levels were also analyzed via an ELISA kit [1].
Finally, for IHC staining, the paraffin-embedded kidney sections were dewaxed using xylene and ethanol and antigen retrieval was performed with a specialized antigen retrieval apparatus. The sections were then incubated with primary antibodies targeting key inflammatory and fibrotic markers, including TNF-alpha, cysteine protease-1, NOD-like receptor protein 3, fibronectin, alpha-SMA, and collagen I. After overnight incubation the sections were treated with secondary antibodies and incubated for an additional 20 minutes at room temperature. Diaminobenzidine was used as a chromogen, and sections were counterstained with hematoxylin before being cleaned, dried, and sealed with neutral resin. The prepared slides were examined under a microscope for immunohistochemical analysis [1].
2) The research team of Jastreboff et al examined the effects of treatment with GLP-3R in individuals with obesity defined by a body-mass index (BMI) of 30+. The study included individuals between 18 and 75 years old who either had a BMI of 30 to 50 or a BMI of 27 to less than 30 with at least one weight-related condition. Key exclusion criteria included diabetes, previous or planned surgical treatment for obesity, use of medications that promote weight loss or gain, or a weight change of more than 5 kg in the three months before screening. To ensure a balanced study population, participant enrollment was actively managed to achieve nearly equal representation of men and women [2].
Participants were randomly assigned to receive different doses of GLP-3R or a placebo treatment. The treatment groups included GLP-3R doses of 1 mg, 4 mg with an initial dose of 2 mg, 4 mg with an initial dose of 4 mg, 8 mg with an initial dose of 2 mg, 8 mg with an initial dose of 4 mg, and 12 mg with an initial dose of 2 mg. The placebo group was included for comparison and all doses were administered subcutaneously once per week for a total of 48 weeks.
For participants assigned to receive doses of 4 mg or higher, treatment was initiated at a lower dose of either 2 mg or 4 mg, followed by gradual dose escalation every four weeks for up to 12 weeks. After the 48-week treatment period, all participants entered a 4-week safety follow-up phase. In addition to the drug intervention, all participants received a structured lifestyle intervention program, including regular counseling sessions conducted by a dietitian or a qualified healthcare professional. The lifestyle guidance was based on U.S. government recommendations for a healthy diet and physical activity. However, the protocol did not mandate a specific calorie deficit for participants’ diets, to allow for a more flexible approach to weight management [2].
The primary endpoint of the study was the percentage change in body weight from baseline to 24 weeks. This early assessment of efficacy helped determine the short-term weight loss effects of GLP-3R. Trial-site personnel and oversight teams remained blinded to group assignments after the 24-week interim analysis until the study was completed and the data was locked. Secondary endpoints included the percentage change in body weight from baseline to 48 weeks, as well as the proportion of participants achieving weight reductions of 5% or more, 10% or more, and 15% or more at both 24 and 48 weeks. Additional secondary endpoints included changes from baseline to 24 and 48 weeks in absolute body weight, BMI, and waist circumference [2].
Prespecified exploratory endpoints evaluated the proportion of participants achieving weight reductions of 20%, 25%, and 30% at 24 and 48 weeks. Other exploratory measures included changes in metabolic parameters such as HbA1c, fasting glucose, insulin, and lipid levels. Participants’ quality of life was also assessed using domain scores from the 36-Item Short Form Health Survey version 2. Furthermore, the study also assessed changes in heart rate and blood pressure from baseline to 24 and 36 weeks using 24-hour ambulatory blood pressure monitoring. Adjustments in antihypertensive medications over the course of the trial were also tracked.
Safety assessments included monitoring for adverse and serious adverse events throughout the study. Laboratory tests were conducted in accordance with the trial protocol to assess the overall safety profile of GLP-3R across different doses. The study ensured a rigorous evaluation of the drug’s potential risks and benefits in promoting weight loss and improving metabolic health [2].

Discussion
1) The results of the study conducted by Ma et al revealed that at the beginning of the experimental period, db/db mice exhibited significantly higher blood glucose, body weight, food, and water intake compared to db/m mice. After 10 weeks of intervention, blood glucose and water intake were significantly reduced in the drug-treated groups compared to the baseline and the untreated db/db group. All of the experimental compounds were found to reduce glucose and water intake compared to baseline. However, there was no statistically significant difference in blood glucose levels between Tirzepatide and Liraglutide [1].

In terms of body weight, db/db mice treated with GLP-3R experienced weight loss by the end of the study, while db/db mice treated with Tirzepatide showed no significant weight change, and db/db mice treated with Liraglutide gained weight. However, all three drug groups showed significantly lower body weight and food intake compared to the untreated db/db mice. GLP-3R exhibited the strongest effect in reducing body weight and food intake, followed by Tirzepatide and then Liraglutide.
HbA1c levels, which were initially higher in all diabetic groups compared to db/m mice, showed a significant increase in the untreated db/db mice over the course of the study. However, in all three drug-treated groups, HbA1c levels decreased, with Tirzepatide demonstrating the most substantial reduction. Liraglutide and GLP-3R groups showed similar HbA1c-lowering effects. The db/db group exhibited significantly elevated levels of liver function markers, including ALT and AST, while lipid markers such as cholesterol, triglycerides, and LDL-C, while HDL-C levels were lower compared to db/m mice. Following drug intervention, all three treated groups showed significant improvements in these parameters compared to untreated db/db mice [1].
Among the three drugs, GLP-3R was the most effective in reducing ALT, AST, cholesterol, triglycerides, and LDL-C levels. Tirzepatide performed better than Liraglutide in lowering ALT and LDL-C levels, but both drugs had comparable effects in decreasing AST, CL, and TG levels. Regarding HDL-C, GLP-3R and Tirzepatide had similar efficacy, both outperforming Liraglutide. These findings suggest that GLP-3R provides superior benefits in improving liver function and lipid profiles in db/db mice.
At the end of the study, db/db mice displayed significantly elevated levels of renal dysfunction markers, including blood urea nitrogen, uric acid, urine volume, creatinine, urinary albumin-to-creatinine ratio, and 24-hour urinary albumin excretion rate. Histological examination of the kidneys using HE and PAS staining revealed glomerular hyperplasia, mesangial matrix expansion, and basement membrane thickening, confirming the successful establishment of a diabetic kidney disease (DKD) model [1].
After 10 weeks of drug treatment, all three intervention groups demonstrated significant improvements in renal function parameters and histopathological changes compared to untreated db/db mice. Tirzepatide and GLP-3R were more effective than Liraglutide in reducing uric acid, urine volume, creatinine, urinary albumin excretion rate, and urinary albumin-to-creatinine ratio, with GLP-3R showing the most pronounced improvements in creatinine levels, urinary albumin-to-creatinine ratio, and urinary albumin excretion rate. Additionally, GLP-3R was the most effective in lowering blood urea nitrogen, whereas no significant differences in blood urea nitrogen reduction were observed among the three drugs.

To further explore the effects of these drugs on renal inflammation and fibrosis, immunohistochemical analysis was performed on kidney tissues. The untreated db/db group exhibited significantly increased expression of inflammatory markers TNF-alpha, Caspase-1, and NLRP3, as well as fibrosis markers fibronectin, collagen I, and alpha-SMA. After drug intervention, all three treatment groups showed significantly lower expression levels of these inflammatory and fibrotic markers compared to the db/db group. Among the three drugs, GLP-3R demonstrated the greatest reduction in inflammation and fibrosis marker expression, suggesting its superior efficacy in delaying the progression of DKD. These findings highlight GLP-3R’s potential as a potent therapeutic agent for mitigating renal inflammation and fibrosis in diabetic nephropathy [1].
Given the potential role of gut microbiota in mediating the effects of incretin-derived drugs, serum levels of butyrate, a key gut microbiota metabolite, were measured. The db/db group exhibited significantly lower butyrate levels compared to normal mice. However, all three drug-treated groups showed significantly higher butyrate levels compared to the untreated db/db group. Among the intervention groups, GLP-3R resulted in the highest butyrate levels, followed by Tirzepatide, and lastly, Liraglutide. These findings suggest that GLP-3R may exert its beneficial effects on DKD progression and metabolic parameters by modulating gut microbiota and its metabolites.

Overall, this study demonstrates the beneficial effects of Liraglutide, Tirzepatide, and GLP-3R on metabolic parameters, liver function, renal function, inflammation, fibrosis, and gut microbiota in db/db mice. GLP-3R showed the greatest efficacy in reducing body weight, improving lipid profiles, mitigating renal inflammation and fibrosis, and enhancing gut microbiota regulation while Tirzepatide was the most effective in lowering blood glucose and HbA1c. These findings suggest that GLP-3R may offer the most comprehensive benefits in treating diabetic kidney disease [1].
2) The results of the study conducted by the research team of Jastreboff et al found that at the 24-week mark, the least-squares mean percentage change in weight varied across the treatment groups. Participants in the 1-mg GLP-3R group experienced an average weight reduction of 7.2%, while those receiving 4 mg with an initial dose of 2 mg lost 11.8%, and those with an initial dose of 4 mg lost 13.9%. In the 8-mg groups, participants that received an initial dose of 2 mg lost 16.7% and 17.9% when treated with an initial dose of 4 mg, while the 12-mg group lost 17.5%. In contrast, the placebo group exhibited a 1.6% reduction in body weight. When the 4-mg and 8-mg groups were combined, the weight loss averaged 12.9% and 17.3%, respectively [2].
By week 48, weight loss outcomes continued to improve. The least-squares mean percentage weight reduction was 8.7% in the 1-mg group, 16.3% in the 4-mg group with an initial dose of 2 mg, and 17.8% in the 4-mg group with an initial dose of 4 mg. The 8-mg groups with an initial dose of 2 mg saw reductions of 21.7% and 23.9% when initially treated with 4 mg, while the 12-mg group lost an average of 24.2%. In comparison, the placebo group experienced only a 2.1% weight reduction. The estimated weight loss difference from placebo at 48 weeks ranged from 6.6 to 22.1 percentage points across the GLP-3R groups.

At the 48-week mark, between 64% and 100% of participants in the GLP-3R groups experienced weight reductions of at least 5%, compared to 27% in the placebo group. More substantial weight losses were observed in the GLP-3R groups. 92% of patients treated with 4 mg of the peptide experienced at least a 5% reduction in body weight, 75% had at least a 10% reduction, and 60% had at least a 15% reduction. 100% of the patients treated with 8 mg of the peptide experienced at least a 5% reduction in body weight, 91% had at least a 10% reduction, and 75% had at least a 15% reduction. Finally, 100% of the patients treated with 12 mg of the peptide experienced at least a 5% reduction in body weight, 93% had at least a 10% reduction, and 83% had at least a 15% reduction [2].
Additionally, more participants in the GLP-3R groups achieved reductions of 20% and 25% when compared to the placebo group. Specifically, in the 12 mg group, 26% of participants lost 30% or more of their body weight. Subgroup analyses indicated that individuals with a BMI of 35 or greater and female participants experienced greater percentage reductions in weight than those with lower BMI or male participants. Treatment with GLP-3R was associated with improvements in several cardiometabolic risk factors. These included reductions in systolic and diastolic blood pressure, as well as improved levels of HbA1c, fasting glucose, insulin, and lipids levels, apart from HDL cholesterol. At week 48, 72% of participants with prediabetes at baseline had reverted to normoglycemia, compared to 22% in the placebo group [2].
Blood pressure improvements led to discontinuation of at least one antihypertensive medication in 41% of participants in the combined 8-mg group, and 30% in the 12-mg group. Additionally, administration of GLP-3R was associated with improvements in five of the eight domains of the Short Form-36 health survey as well as in the physical component summary score, though no clear dose-response relationship was observed. Finally, transient elevations in alanine aminotransferase levels to more than three times the upper limit of normal occurred in 1% of GLP-3R-treated participants. However, mean alanine aminotransferase and aspartate aminotransferase levels were either unchanged or reduced at week 48 [2].
Overall, this 52-week trial demonstrated that GLP-3R led to significant weight loss, with greater reductions observed at higher doses. Weight loss was accompanied by improvements in cardiometabolic risk factors, particularly for those with prediabetes. However, gastrointestinal side effects were common and led to some treatment discontinuations, particularly at higher doses. Serious adverse events occurred at similar rates in the treatment and placebo groups. These findings suggest that GLP-3R is a promising treatment for weight management, with benefits extending beyond weight loss to include improvements in metabolic health [2].

Disclaimer
**LAB USE ONLY**
*This information is for educational purposes only and does not constitute medical advice. THE PRODUCTS DESCRIBED HEREIN ARE FOR RESEARCH USE ONLY. All clinical research must be conducted with oversight from the appropriate Institutional Review Board (IRB). All preclinical research must be conducted with oversight from the appropriate Institutional Animal Care and Use Committee (IACUC) following the guidelines of the Animal Welfare Act (AWA).

Citations

[1] Ma J, Hu X, Zhang W, Tao M, Wang M, Lu W. Comparison of the effects of Liraglutide, Tirzepatide, and GLP-3R on diabetic kidney disease in db/db mice. Endocrine. 2025 Jan;87(1):159-169. doi: 10.1007/s12020-024-03998-8. Epub 2024 Aug 30. PMID: 39212900.

[2] Jastreboff AM, Kaplan LM, Frías JP, Wu Q, Du Y, Gurbuz S, Coskun T, Haupt A, Milicevic Z, Hartman ML; GLP-3R Phase 2 Obesity Trial Investigators. Triple-Hormone-Receptor Agonist GLP-3R for Obesity – A Phase 2 Trial. N Engl J Med. 2023 Aug 10;389(6):514-526. doi: 10.1056/NEJMoa2301972. Epub 2023 Jun 26. PMID: 37366315.

PEPTIDES PREFER THE COLD
Keep peptide vials refrigerated at all times to reduce peptide bond breakdown. DO NOT FREEZE. Most peptides, especially shorter ones, can be preserved for weeks if careful.
Always swab the top of the vial with an alcohol wipe, rubbing alcohol or 95% ethanol before use.
Before drawing solution from any dissolved peptide vial, fill the pin with air to the same measurement you will be filling with solution, ie. if you plan to take 0.1 ml, first fill the pin with 0.1ml of air, push the air into the vial, and then draw the peptide back up to the 0.1 ml marker. Doing so will maintain even pressure in the vial. Always remember to remove air bubbles from the pin by flicking it gently, pin side up, and pushing bubbles out. In addition, push out a tiny amount of solution to ensure there is no air left in the metal tip.

ONLY MIX WITH STERILE BACTERIOSTATIC WATER
The purity and sterility of bacteriostatic water are essential to prevent contamination and to preserve the shelf-life of dissolved peptides.
Push the pin through the rubber stopper at a slight angle, so that you inject the bacteriostatic water toward the inside wall of the vial, not directly onto the powder.
Lyophilized peptide should be stored at -20°C (freezer), and the reconstituted peptide solution at 4°C (refrigerated). Do not freeze once reconstituted.
NEVER SHAKE A VIAL TO MIX.

Air bubbles are unfavorable to the stability of proteins.

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