Validating GLP-1RA Therapy in Preclinical Mouse Models of Metabolic Dysfunction-Associated Steatohepatitis

Introduction: GLP-1RA Therapeutics for Metabolic Diseases

As the global metabolic disease crisis continues to grow, GLP-1 receptor agonists (GLP-1RAs) have emerged as groundbreaking therapeutics with applications extending far beyond their initial diabetes indications. Additionally, this drug class has become one of the most profitable investment areas for biopharmaceutical companies. With the GLP-1 receptor market projected to reach nearly $56 billion by 2031,^[2] pharmaceutical researchers are intensifying efforts to validate these compounds across metabolic conditions—particularly for metabolic dysfunction-associated steatohepatitis (MASH), a condition affecting millions worldwide with limited treatment options.

Market Growth and Clinical Relevance

This article explores how Cyagen's advanced preclinical MASH mouse models are providing critical validation for GLP-1RA therapeutic applications. Through rigorous pharmacodynamic testing of semaglutide in clinically relevant animal models, we demonstrate how these platforms are supporting translational research that mirrors recent breakthrough clinical findings in human MASH patients.

Mechanism of Action: How GLP-1RAs Treat Metabolic Disorders

Glucagon-like peptide-1 (GLP-1) is an incretin hormone that plays a central role in glucose regulation, lipid metabolism, and numerous vital physiological functions. GLP-1 receptor (GLP-1R) is an important member of the G protein-coupled receptor (GPCR) family, widely distributed across various human cell surfaces and is the biological target for both endogenous GLP-1 and therapeutic interventions.

GLP-1 receptor agonists (GLP-1RAs) are medications specifically designed to mimic the action of endogenous GLP-1 by binding to and activating the GLP-1 receptor. In recent years, glucagon-like peptide-1 (GLP-1) drugs have become a popular area of medical research and drug development due to their unique mechanism of action, excellent therapeutic effects, and broad clinical application prospects.^[1] The FDA has approved several GLP-1 receptor agonists as key medications for the treatment of type 2 diabetes (T2D) and obesity, establishing this drug class as one of the most lucrative investment areas for biopharmaceutical companies.  

Clinical Evidence Supporting GLP-1RA Use for MASH

Diabetes and obesity are associated with metabolic dysfunction-associated steatohepatitis (MASH/NASH),  positioning GLP-1RAs as promising treatment candidates. In the Q4 2024, Novo Nordisk announced the primary results from their initial trial evaluating semaglutide for MASH patients. This Phase 3, double-blind trial enrolled 1,200 patients with MASH and moderate to severe liver fibrosis (stage 2 or 3), assessing the efficacy of weekly 2.4 mg semaglutide in the first 800 randomized patients.

Phase 3 Semaglutide Trial Overview

The trial successfully achieved its primary endpoint: compared to placebo, semaglutide significantly improved liver fibrosis without worsening steatohepatitis, and actually promoted the remission of steatohepatitis without worsening liver fibrosis. After 72 weeks, 37.0% of patients showed improvement in liver fibrosis (placebo group: 22.5%), and 62.9% of patients experienced remission of steatohepatitis (placebo group: 34.1%).^[3]

Cyagen’s Preclinical Mouse Model of MASH

To support research targeting this disease, Cyagen's metabolic platform has developed a high-fat diet plus carbon tetrachloride (HFD+CCl4)-induced MASH mouse model, which closely mimics the clinical fibrosis characteristics of human MASH. The effectiveness of therapeutic targets and the clinical translational potential of this model were validated using two drugs, Resmetirom and Semaglutide (a GLP-1RA). Today, we will introduce the pharmacodynamic testing of Semaglutide in the mouse MASH model.

Model Design: HFD+CCl4-Induced Liver Fibrosis

C57BL/6 mice were fed a 60% high-fat diet (HFD) for 12-16 weeks to successfully establish a diet-induced obesity (DIO) model, exhibiting symptoms such as obesity and hyperlipidemia. Liver fibrosis was induced through 6 weeks of CCl4 administration to yield the HFD+CCl4-induced MASH mouse model.

Validation with Semaglutide and Resmetirom

Therapeutic evaluation involved administration of semaglutide subcutaneously three times per week. The study endpoints included analysis of body composition, blood lipid levels, and pathology, revealing that Semaglutide significantly reduced hyperlipidemia, liver lipid accumulation, and fibrosis phenotypes.

Figure 1. Overview of the HFD+CCl4-induced MASH (NASH) Mouse Study

Pharmacodynamic Validation of Semaglutide Efficacy in HFD+CCl4 MASH Mouse Model

Mice underwent high-fat diet (HFD) induction for 12-16 weeks, reaching body weights exceeding 45g. They were randomly divided into groups based on blood lipid levels and body weight. Semaglutide (30 nmol/kg) was administered subcutaneously three times per week for 6 weeks, while CCl4 was administered intraperitoneally twice a week during this period.

Key Findings from Body Composition Analysis

Figure 2. Weight and Body Composition Analysis

Body weight was measured twice weekly. Semaglutide significantly reduced mouse body weight and fat content. Data are expressed as mean ± standard error, n=5. *P<0.05, **P<0.01, and ***P<0.001 compared to the normal diet control group; #P<0.05, ##P<0.01, and ###P<0.001 compared to the high-fat diet + CCl4 group, analyzed using one-way ANOVA.

Lipid and Liver Weight Results

Figure 3. Liver Weight and Blood Lipid Analysis

At the study endpoint, liver weight was measured. The Semaglutide group significantly reduced liver weight. Before and after administration, the Semaglutide-treated group significantly reduced the hyperlipidemia phenotype induced by HFD. Data are expressed as mean ± standard error, n=5. *P<0.05, **P<0.01, and ***P<0.001 compared to the normal diet control group; #P<0.05, ##P<0.01, and ###P<0.001 compared to the high-fat diet + CCl4 group, analyzed using one-way ANOVA.

Histological Findings (H&E, Sirius Red)

Figure 4. H&E Staining Pathological Analysis

At the study conclusion, liver tissue samples were collected for histopathological analysis. Compared to the DIO model group, Semaglutide reduced ballooning and steatosis. After composite scoring, the Semaglutide-treated group significantly decreased the Non-alcoholic Fatty Liver Disease (NAFLD) activity score (NAS) compared to both the DIO and DIO+CCl4 groups.

Yellow arrows indicate steatosis, blue arrows indicate hepatocyte ballooning, red arrows show lymphocyte infiltration in different regions of the liver tissue, green arrows represent congestion in the central vein and portal vein, and brown arrows indicate occasional fibroblast proliferation. Data are expressed as mean ± standard error, n=5. *P<0.05, **P<0.01, and ***P<0.001 compared to the normal diet control group; #P<0.05, ##P<0.01, and ###P<0.001 compared to the high-fat diet + CCl4 group, analyzed using one-way ANOVA.

Figure 5. Sirius Red Staining and Pathological Analysis

Sirius Red staining results showed that the CCl4 model group had a significantly increased degree of fibrosis compared to the DIO model group, while Semaglutide significantly reduced CCl4-induced liver fibrosis. Black arrows indicate the proliferation of collagen fibers around the central vein in the liver tissue. Data are expressed as mean ± standard error, n=5. *P<0.05, **P<0.01, and ***P<0.001 compared to the normal diet control group; #P<0.05, ##P<0.01, and ###P<0.001 compared to the high-fat diet + CCl4 group, analyzed using one-way ANOVA.

Summary of Therapeutic Benefits in Preclinical MASH Models

DIO mice induced by a high-fat diet (HFD) for 12-16 weeks demonstrated increases in both body weight and liver weight compared to mice maintained on regular diets. The levels of TC, HDL-C, and LDL-C in the serum of DIO (HFD) mice were significantly increased, effectively mimicking the symptoms of MASH. Notably, the combined HFD+CCl4 model accelerated and aggravated the pathological processes of the MASH mouse model, with CCl4 reducing the induction period while exacerbating liver fibrosis in DIO/HFD mice. Semaglutide has protective and therapeutic effects on the HFD+CCl4-induced MASH model, effectively reducing steatosis and fibrosis.

This validated model is widely utilized in preclinical efficacy studies to evaluate the therapeutic and protective effects of novel drug candidates for MASH. Our results confirm that Semaglutide, a GLP-1 receptor agonist, delivers significant protective and therapeutic benefits in the HFD+CCl4-induced MASH model, effectively reducing both steatosis and fibrosis—mirroring the clinical benefits observed in human trials.

Cyagen’s Solutions for Metabolic Disease Research

Based on the successful application of this model in preclinical efficacy studies, Cyagen's metabolic platform leverages advanced animal model development technology and comprehensive pharmacodynamic evaluation systems to support in-depth metabolic disease research. We offer numerous mouse models, custom modeling services, and pharmacodynamic evaluation platforms for research in obesity, liver disease, diabetes, atherosclerosis, and other metabolic and cardiovascular diseases.

Our team is committed to delivering premium research services and generating reliable, reproducible data to support both foundational disease research and preclinical drug development, ultimately accelerating therapeutic development timelines. For information about our related services and customized research solutions, please contact our scientific team.

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References:

[1] Zheng, Z., Zong, Y., Ma, Y. et al. Glucagon-like peptide-1 receptor: mechanisms and advances in therapy. Sig Transduct Target Ther 9, 234 (2024).

[2] https://www.biospace.com/drug-development/7-indications-for-glp-1s-beyond-weight-loss

[3] https://www.novonordisk.com/news-and-media/news-and-ir-materials/news-details.html?id=171971