2FA-Platform Generates Dual Fatty Acid-Conjugated GLP-1 Receptor Agonist TE-8105 with Enhanced Diabetes, Obesity, and NASH Efficacy Compared to Semaglutide

Abstract

Conjugating two fatty acids (2FAs) to peptide drugs can improve pharmacokinetics and therapeutic effects. However, optimizing FA spacing, chain combination, and attachment site to simultaneously enhance albumin binding and drug efficacy remains challenging. We introduce a multiarm linker technology enabling precise control of 2FA spacing, composition, and attachment. By applying this technology to a modified glucagon-like peptide-1 (GLP-1) and screening various 2FA-GLP-1 conjugates differing in linkage, linker, and FA properties for improved albumin affinity, pharmacokinetics, and pharmacodynamics, TE-8105 emerged as a promising candidate. TE-8105 outperformed semaglutide, showing improved long-term glycemic control, weight loss, and liver health in diabetic mice, and dose-dependent weight loss and favorable body composition changes in obese mice. A distinct advantage of TE-8105 over semaglutide is its low-dose reduction of liver steatosis and improvement of liver health in nonalcoholic steatohepatitis mice. The multiarm linker technology provides a versatile platform for developing improved 2FA-peptide therapeutics.

Figure 1

Schematic of 2FA-GLP-1 conjugates generated using a multiarm linker. (A) General structure of 2FA-bundles created by a multiarm linker. (B) Modified GLP-1 conjugated to a 2FA-bundle at native Lys-26 via a Glu-azide-alkyne reaction. (C) Modified GLP-1 conjugated to a 2FA-bundle at Cys-53 linked by (GGGSG)₃ via thiol-maleimide reaction. n is the number of EG units; L₁ and L₂ denote the number of CH₂ groups, whereas T₁ and T₂ denote the terminal groups in FA-1 and FA-2, respectively; C^ter represents −CH₃ or −H.

Figure 2

Dose–response relationships of semaglutide and TE-8105. Comparison of dose–response relationships between TE-8105 and semaglutide in db/db mice at (A) 3 nmol/kg and (B) 30 nmol/kg. Data are mean ± standard error of the mean (SEM). Comparison of dose–response relationships between semaglutide and TE-8105 in triggering insulin release from isolated mouse pancreatic islets at (C) high glucose concentration (17.5 mM) and (D) low glucose concentration (2.5 mM). Data were shown as mean ± SEM and were analyzed by 2-way ANOVA. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Figure 3

Internalization of GLP-1R under GLP-1, semaglutide, and TE-8105 treatment. Confocal imaging of HEK-GLP-1R-EGFP cells under GLP-1, semaglutide, and TE-8105 induction without FBS (first 3 rows) and with FBS preincubation (last 3 rows).

Figure 4

Chronic treatment of TE-8105 improved glycemic control in db/db mice. (A) Blood glucose levels in db/db mice treated with PBS (gray), semaglutide (blue), or TE-8105 (red) at 10 or 30 nmol/kg dose every 2 days for 8 weeks. (B) HbA1c changes after long-term treatment. (C) Body weight change on day 58 vs day 0. (D) Heart, liver, iWAT, and BAT weights relative to body weights of db/db mice. (E) H&E staining of iWAT and liver sections after long-term treatment; 20× magnification. Arrows indicate macrovesicular steatosis (dotted line); microvascular steatosis (bold); hypertrophy (blue). (F) iWAT cell area and (G) diameter analyzed using StrataQuest Plus Analysis Software. Data shown as mean ± SEM and analyzed using ANOVA with multiple comparisons; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 5

Chronic treatment of TE-8105-induced weight loss in DIO mice. (A) Body weight of DIO mice treated with PBS, and 3–100 nmol/kg of semaglutide or TE-8105 every 4 days for 40 days. (B) Body weight change of day 40 vs day 0. (C) Blood glucose levels and (D) % fat and lean mass post-treatment. (E) Percentage of iWAT cells with diameter ≤60 μM (black) and >60 μM (white) after treatment. (F) Total cholesterol and ALT levels post-treatment. Data shown as mean ± SEM and analyzed using ANOVA with multiple comparisons; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 6

Chronic treatment of TE-8105-induced weight loss in GAN-DIO-NASH mice. (A) Body weight of GAN-DIO-NASH mice treated daily with PBS, semaglutide (5 or 30 nmol/kg), or TE-8105 (5, 10, 30 nmol/kg) for 10 weeks. (B) Body weight change from day 0 to day 70. (C) Fat mass and (D) ALT levels of mice on days 0 and 70. (E) H&E staining of liver; 20× magnification. (F) Liver triglyceride levels post-treatment. Data shown as mean ± SEM. ANOVA with multiple comparisons for (B) and (F), Student’s t test for (C) and (D). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.