Fig. 5: Evaluation of serum stability, BBB penetration, and intravenous/subcutaneous pharmacokinetics of AG10-L2-Nal, naloxone, and naloxegol.

a Serum stability of AG10-L2-Nal was evaluated in mouse, rat, and human sera. AG10-L2-Nal (100 μM) was incubated in sera, and the concentration remaining in respective media was determined at 0 and 48 h using HPLC. Bar graphs represent the mean of % compound remaining ± s.d. (n = 3). b Brain to plasma ratios and c cerebrospinal fluid (CSF) to plasma ratios of AG10-L2-Nal, naloxegol, and naloxone. Male Sprague Dawley rats were dosed intravenously with 4.84 µmol/kg of test compounds (equivalent to 1.6 mg/kg, 3.2 mg/kg, and 4 mg/kg for naloxone, naloxegol, and AG10-L2-Nal, respectively). The plasma, brain tissue, and CSF were collected at 30 min after dosing. The ratio of the brain (ng/g) versus plasma concentration (ng/mL) is expressed as the percentage brain to plasma ratio. The ratio of the CSF (ng/mL) versus plasma concentration (ng/mL) is expressed as the percentage CSF to plasma ratio. Bar graphs show the respective mean (±s.d.) (n = 3 for each group). Statistical differences were determined using one-way ANOVA followed by Tukey’s Post Hoc test (*P < 0.05 compared to naloxone). For the brain to plasma ratio experiment F(2,6) = 678.0, P < 0.0001 and for the CSF to plasma ratio experiment F(2,6) = 174.6, P < 0.0001. d a single intravenous bolus dose of 4.84 µmol/kg and e a single subcutaneous injection of 16 µmol/kg dose of antagonists (equivalent to 5.2 mg/kg, 10.4 mg/kg, and 13.2 mg/kg for naloxone, naloxegol, and AG10-L2-Nal, respectively) were administered to rats (n = 3 for each group). The concentration of the test compounds in plasma was determined at different time points and expressed as means ± s.d. of three biological replicates. Source data are provided as a Source Data file.