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Development of a Mechanism of Action-Reflective Cell-Based Reporter Gene Assay for Measuring Bioactivities of
Xiaoming Zhang1,2, Chunyan Li3, Zhe Deng4
1NHC Key Laboratory of Research on Quality and Standardization of Biotech Products, NMPA Key Laboratory for Quality Research and Evaluation of Biological Products, National Institutes for Food and Drug Control, Beijing 102629, China.
Abstract:
Glucagon-like peptide-2 (GLP-2) is a gut hormone that plays a pivotal role in regulating intestinal epithelial cell growth and function, making it a promising therapeutic agent for intestinal damage and bone-related diseases. Nonetheless, the therapeutic potential of GLP-2 is substantially diminished due to its inactivation by dipeptidyl peptidase 4 (DPP-4). In recent years, advancements have been made in developing dipeptidyl peptidase 4 (DPP-4) resistant GLP-2 analogues with an extended half-life. The murine model with extensive experimental bowel resection maintained on parenteral nutrition has been used for assessing the physiology and pharmacology of GLP-2, and for the preclinical validation of GLP-2 analogues. However, it possesses certain limitations, such as complex procedure, considerable variability, and time-consuming nature. Consequently, there is a pressing need for the development of a cell-based bioassay to assess GLP-2 analogues. Here, we successfully developed a mechanism-of-action (MOA)-reflective cell-based reporter gene assay (RGA), utilizing a stable HEK293 cell line expressing the GLP-2 receptor and a luciferase reporter gene. This innovative approach allows for precise quantification of the potency of GLP-2 analogues. The RGA demonstrated good accuracy, linearity, precision, and specificity, with potential applications in stability testing, drug screening, and therapeutic monitoring of GLP-2 analogues. Moreover, RNA sequencing reveals the multi-target regulatory effect of GLP-2 analogues. The establishment of this RGA provides a valuable tool for evaluating the potency of GLP-2 analogues and the screening of potential therapeutic drugs targeting to GLP-2 receptor.
Insights
A new cell-based reporter gene assay accurately measures glucagon-like peptide-2 (GLP-2) analogue potency. This assay offers a valuable tool for evaluating GLP-2 drugs, overcoming limitations of traditional animal models.
Area of Science:
- Biochemistry
- Pharmacology
- Cell Biology
Background:
- Glucagon-like peptide-2 (GLP-2) is crucial for intestinal health and bone metabolism.
- Its therapeutic use is limited by rapid inactivation by dipeptidyl peptidase 4 (DPP-4).
- Existing preclinical models for GLP-2 analogues have significant drawbacks.
Purpose of the Study:
- To develop a novel, mechanism-of-action (MOA)-reflective cell-based assay for assessing GLP-2 analogues.
- To provide a more efficient and reliable method for evaluating GLP-2 analogue potency compared to animal models.
Main Methods:
- Development of a stable HEK293 cell line expressing the GLP-2 receptor and a luciferase reporter gene.
- Utilized a reporter gene assay (RGA) for precise quantification of GLP-2 analogue potency.
- RNA sequencing was employed to investigate the multi-target regulatory effects of GLP-2 analogues.
Main Results:
- The developed RGA demonstrated high accuracy, linearity, precision, and specificity.
- The assay successfully quantified the potency of GLP-2 analogues.
- RNA sequencing revealed the complex regulatory actions of GLP-2 analogues.
Conclusions:
- The established RGA is a robust and valuable tool for evaluating GLP-2 analogue potency.
- This assay facilitates drug screening, stability testing, and therapeutic monitoring of GLP-2 receptor-targeting drugs.
- The RGA offers an improvement over traditional animal models for GLP-2 analogue assessment.
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