Synthesis, Characterization, and Antidiabetic Evaluation of Sequence-Modified Liraglutide Analogs in a Drosophila melanogaster Model.

Liraglutide, a glucagon-like peptide-1 receptor agonist, is widely used as a therapeutic macromolecule for the treatment of type 2 diabetes; however, its large-scale production is limited by high manufacturing costs and the frequent occurrence of closely related deletion impurities during synthesis. In the present study, we describe a novel, impurity-controlled, and industrially feasible synthetic strategy for liraglutide and its sequence-modified analogs. This method utilizes solution-phase incorporation of Pal-γ-Glu-OtBu in combination with a preassembled Boc-His (Boc)-Ala-Glu (OtBu)-OH tripeptide fragment, effectively minimizing the formation of des-His, des-Ala, and des-Glu impurities. The optimized protocol enabled the production of liraglutide and its analogs with consistent isolated yields and high chromatographic purity (> 95% by RP-HPLC), suitable for subsequent biological evaluation. The antidiabetic potential of liraglutide and three sequence-modified analogs, Lira (Trp-O), Lira (desGly), and Lira (Glu), was evaluated using a Drosophila melanogaster model of high-sucrose diet induced diabetes. Among these, Lira (Glu) exhibited the most pronounced metabolic improvements, significantly reducing free glucose (p < 0.001), trehalose (p < 0.001), and triglyceride levels (p < 0.001) when compared to diabetic controls. Furthermore, this analog effectively decreased lipid accumulation and reactive oxygen species in larval gut tissues and enhanced locomotor performance in both larva and adult flies. While Lira (Trp-O) also demonstrated beneficial effects, Lira (desGly) showed comparatively limited efficacy. Collectively, this study presents a cost-effective and impurity-controlled synthetic platform for liraglutide production and identifies Lira (Glu) as a promising analog with enhanced antidiabetic activity, offering valuable insights for peptide manufacturing and GLP-1-based therapeutic development.