Predicting Dopamine D₂ Receptor Occupancy Of Ropinirole In Rats Using PET And Pharmacokinetic-Pharmacodynamic Modeling

Objective: To investigate dopamine D2 receptor occupancy of anti-Parkinson drug ropinirole in striatum of rats using a 18F-Fallypride PET scan method and measure plasma and striatum pharmacokinetic properties, then develop a combined pharmacokinetic-pharmacodynamic model for the concentration-occupancy relationship.Methods: Male Sprague Dawley rats were selected for the following experiments. Ropinirole solution was administrated by intravenous injection via the tail vein. Dose levels were set as 5, 15, 30 and 60 mg/kg, respectively.(1)18F-Fallypride PET scan of dopamine D2 receptor binding. 4 rats at each scanning batch underwent a pre-treatment scan and 4 rats of each scanning time point at each dose level were then selected for post-treatment PET scan. Radiotracer was bolus infused intravenously followed by static, three-dimensional mode acquisition scans of 10 minutes. Regions of interests were drawn for striatum and cerebellum and transferred to standard uptake values. Binding potential and receptor occupancy were calculated according to simplified reference tissue model.(2) Study on pharmacokinetics of ropinirole in rats. Blood samples were withdrawn before and after dosing at different time points of 6 rats per dose level. Another 4 rats at each selected time point of each dose level underwent blood collection and striatum dissection. Concentration of plasma and striatum homogenate samples were determined via a UPLC-MS/MS method and their correlations were evaluated. Pharmacokinetic parameters were estimated by non-compartmental analysis using Win Nonlin software.(3) Pharmacokinetic-pharmacodynamic modeling based on plasma concentration and dopamine D2 receptor occupancy of ropinirole. Win Nonlin software was applied to combine plasma pharmacokinetic parameters and occupancy data to fit the best PK-PD model and describe the time course of plasma concentration and receptor occupancy by ropinirole. Bootstrap was applied to validate the model and Monte-Carlo method was used to simulate individual data.Results:(1) Dopamine D2 receptor binding profile of ropinirole in rats. Binding potential from 4 pre-treatment scans showed no significant difference(p> 0.05) through one-way ANOVA analysis, indicating a reliable and repeatable scanning method. Scan results of 15 mg/kg dose level showed that receptor occupancy decline from 56.23 ± 6.39% to 15.19 ± 13.60%(p < 0.05) with the extension of time after administration.Binding between ropinirole and dopamine D2 receptor at 0.17 h after dosing revealed dose-dependent manner that receptor occupancy increased from 6.09±15.32% to 92.89±2.83%(p < 0.05).(2) Validation of UPLC-MS/MS method for the determination of ropinirole in plasma and striatum homogenate. UPLC-MS/MS method was used for simultaneous determination of ropinirole in rat plasma and striatum homogenate. The calibration curve was linear during the range of 1 ~ 5000 ng/m L. Good precision and accuracy were found, and the matrix effect and stability met the requirements of biological sample analysis as well.(3) Plasma pharmacokinetic profile of ropinirole in rats. Ropinirole eliminated rapidly in plasma after administration. Pharmacokinetic parameters of four dose levels analyzed by non-compartmental model were as follows respectively: elimination half-life(t1/2): 0.84 ± 0.12, 0.75 ± 0.25, 0.98 ± 0.17 and 1.17 ± 0.13 h; volume of distribution(Vz): 8593.79±1363.21, 3964.02±1345.75, 6897.94±974.83 and 5284.12±543.23 m L/kg; clearance(Cl): 7044.32±316.91, 3685.73±488.32, 4912.63±520.31 and 3135.86±96.61 m L/h/kg.(4) Striatum pharmacokinetic profile of ropinirole in rats. Distribution trend of ropinirole were similar in striatum and plasma with significant correlation(r2= 0.9499, p < 0.0001) and also consistent with the changes of receptor occupancy. Elimination half-life(t1/2) in striatum of 15 mg/kg dose level was 1.31±0.25 h.(5) Pharmacokinetic-pharmacodynamic modeling of ropinirole in rats. Two-compartment model with first-order elimination assuming a maximum occupancy model was applied to the PK-PD analysis. The estimated parameters for the final model were presented below: apparent clearance from central compartment(Cl): 3309.09±565.13 m L/h/kg; central volume of distribution(V1): 523.54±96.03 m L/kg; intercompartmental clearance(Q): 7511.72±1114.50 m L/h/kg; peripheral volume of distribution(V2): 1526.57±251.00 m L/kg; half maximal effective concentration(EC50): 1390.70±96.49 ng/m L; equilibrium rate constant(ke0): 1.09±0.08 /h. Bootstrap resampling proved that model was stable and accurate. Concentration-occupancy profile under multiple doses and time after dose was obtained from Monte- Carlo Simulation.Conclusion: Binding with dopamine D2 receptor of ropinirole in rats was dose- and time-dependent and similar pharmacokinetic properties were found in plasma and striatum. The concentration-occupancy relationship can be dynamically and quantitatively descripted by the PK-PD model established in our paper, which can provide a brand new technology platform for objective and direct evaluation of anti-Parkinson drug treatment for clinics.