Article Figures & Data
Figures
- Fig. 1.
Structure of E7766, a novel agonist of STING pathway.
- Fig. 2.
Phenotyping of hepatobiliary transporters involved in the disposition of E7766. (A) Uptake of E7766 was evaluated in SLC transporter–expressing HEK293 cells; (B) transport of E7766 was evaluated on ATP-binding cassette transporter–expressing membrane vesicles. n.s., not significant.
- Fig. 3.
In vitro transporter kinetics of E7766 were measured in HEK-293 cells expression OATP1B1 or OATP1B3. (A) Kinetics and Michaelis-Menten parameters of OATP1B1-mediated uptake of E7766. (B) Kinetics and Michaelis-Menten parameters of OATP1B3-mediated uptake of E7766.
- Fig. 4.
Pharmacokinetics and disposition of E7766 after intravenous administration in wild-type and OATP1B1/1B3-humanized mice. (A) Plasma concentrations of E7766 were measured with or without coadministration with Rifampicin after intravenous administration of E7766 and Rifampicin. (B) Blood concentrations of E7766 were measured with or without coadministration with Rifampicin after intravenous administration of E7766 and Rifampicin. (C) Excretion with or without coadministration with Rifampicin of E7766 in urine, bile, and feces was determined after intravenous administration of E7766 and Rifampicin. Hu, humanized mouse.
- Fig. 5.
Summary of PK parameters and DDI profile of E7766 from PBPK model. (A) Contribution of passive diffusion and OATP1B1- and OATP1B3-mediated uptake to overall hepatic uptake clearance of E7766. Simulated plasma (B) and liver (C) concentration-time profiles of E7766 after intravenous administration of 1-mg dose with and without 600-mg oral dose of Rifampicin. Sensitivity analysis of changes in E7766 area under the curve ratio as a function of REF (D) and kinetic parameters for OATP1B1 (E) and OATP1B3 (F).
Tables
- TABLE 1
Summary of in vitro parameters for E7766 estimated from the sandwich-cultured human hepatocytes
Test Article Target Conc. (µmol/l) Temperature Time (min) Uptake CLint,T (µl/min per 106 cells) Efflux CLint,T (µl/min per 106 cells) BEI % E7766 0.3 37°C 1 7.69 ± 0.47 ― ― 5 3.83 ± 0.26 ― ― 10 3.12 ± 0.26 ― ― 20 Not determined 2.05 ± 0.07 85.2 ± 1.7 4°C 10 0.13 ± 0.00 ― ― 1.0 37°C 1 5.46 ± 0.10 ― ― 5 2.70 ± 0.14 ― ― 10 1.85 ± 0.13 ― ― 20 Not determined 1.42 ± 0.08 86.2 ± 0.40 4°C 10 0.14 ± 0.02 ― ― 10.0 37°C 1 1.22 ± 0.11 ― ― 5 0.47 ± 0.02 ― ― 10 0.33 ± 0.01 ― ― 20 Not determined 0.15 ± 0.01 70.9 ± 1.3 4°C 10 0.58 ± 0.73 ― ― d8-TCA 5 37°C 10 14.2 ± 0.82 11.0 ± 1.74 72.6 ± 5.8 4°C 0.27 ± 0.02 ― ― Rosuvastatin 10 37°C 10 6.22 ± 0.31 2.23 ± 0.38 42.7 ± 5.8 4°C 0.21 ± 0.04 ― ― Efflux CLint,T, intrinsic biliary efflux clearance; uptake CLint,T, intrinsic hepatic uptake clearance. Values represent the mean ± S.D. (n = 3).
Dose BDC Rat Intravenous Bolus, 1 mg/kg Dog Intravenous Bolus, 0.075 mg/kg CLtot,p (l/h per kilogram) 6.50 ± 0.429 1.29 ± 0.369 Vss (l/kg) 2.47 ± 0.849 0.553 ± 0.345 Ae renal (%) 13.7 ± 2.80 4.96 ± 4.37 CLrenal (l/h per kilogram) 0.895 ± 0.189 0.0528 ± 0.0293 Ae biliary (%) 92.1 ± 7.26 87.9 ± 30.5 CLbiliary (l/h per kilogram) 6.04 ± 0.798 1.19 ± 0.637 Ae fecal (%) 0.366 ± 0.208 ― CLfecal (l/h per kilogram) 0.0227 ± 0.0138 ― Values represent the mean ± S.D. (n = 4 for BDC rats and n = 3 for dogs).
- TABLE 3
Liver and systemic exposure of E7766 in wild-type and humanized mice in the presence or the absence of Rifampicin
Parameters WT Mice 0.5 mg/kg E7766 WT Mice 0.5 mg/kg E7766 +Rifampicin Hu Mice 1.0 mg/kg E7766 Hu Mice 0.5 mg/kg E7766 +Rifampicin AUCtotal (ng∙h/ml) 56.0 300 126 ± 48.7 302 ± 92.2 AUCtotal/dose (ng∙h/ml/[mg/kg]) 112 600 126 ± 48.7 604 ± 184 CLtot,p (l/h per kilogram) 8.93 1.72 9.20 ± 4.65 1.80 ± 0.619 Vss (l/kg) 1.66 0.520 5.93 ± 5.96 1.29 ± 0.243 Liver AUCtotal (ng·h/g) 4460 4250 ― ― Liver Kp,total 79.6 14.2 ― ― AUCtotal/dose, area under the total plasma conc.-time curve normalized by dose; Hu, OATP1B1/OATP1B3-humanized mouse; liver AUCtotal, area under the total liver conc.-time curve. Values represent the mean ± S.D. (n = 3).
Parameter Scenario 1 Scenario 2 Source PhysChem and blood binding Mol. wt. (g/mol) 746 746 Calculated Log P 1.31 1.31 Measured Compound type Monoprotic acid Monoprotic acid ― pKa 3.41 3.41 Calculated B/P 0.55 0.55 Measured fu 0.50 0.50 Measured Distribution model Full PBPK model Full PBPK model ― Vss (l/kg) 0.637 0.637 SimCYP predicted (method 2, the Rodgers-Rowland method) Kp scalar 4.0 4.0 Fitted based on preclinical data, see Materials and Methods for details CLrenal (l/h) 3.0 3.0 Estimate as fu × glomerular filtration rate Hepatic transport (permeability-limited liver module) Passive diffusion CLPD (ml/min per 106 cells) 0.00013 0.00013 Obtained from E7766 uptake measured at 0.3 µmol/l at 4 °C with SCHH model (Table 1) fuIW 1.00 1.00 SimCYP predicted fuEW 0.657 0.657 SimCYP predicted Jmax (pmol/min per 106 cells) for OATP1B1 8.34 ― Vmax in the unit of pmol/min/mg protein (Fig. 3) was converted to Jmax in the unit picomoles per minute per 106 cells by incorporating measured protein abundance data of HEK293 cells (0.3 mg protein per 106 HEK293 cells) Km (µmol/l) for OATP1B1 2.20 ― Obtained from transporter kinetic assays (Fig. 3) fuinc for OATP1B1 1.00 ― SimCYP predicted REF for OATP1B1 0.10 ― Calculated by eq. 4, see Supplemental Table 2 for details of transporter protein expression. Jmax (pmol/min per 106 cells) for OATP1B3 24.39 ― Fig. 3. Units were converted as shown above for OATP1B1 Km (µmol/l) for OATP1B3 3.97 ― Obtained from transporter kinetic assays (Fig. 3) fuinc for OATP1B3 1.00 1.00 SimCYP predicted REF for OATP1B3 2.80 ― Calculated by eq. 5, see Supplemental Table 2 for details of transporter protein expression. Uptake CLint,T (µl/min per 106 cells) ― 7.7 For scenario 2, the uptake CLint,T for E7766 measured at 0.3 µmol/l after 1 min incubation at 37 °C (Table 1) was assigned as input value for uptake CLint,T in SCHH, as early time point and lower conc. can better represent the initial linear uptake phase. REFSCHH ― 1 The REFSCHH is assumed to be one based on literature reported data shown that OATP1B1 and OATP1B3 expression levels are comparable between SCHH and primary hepatocyte if from the same lot (Kimoto, 2012) Efflux CLint,T (µl/min per 106 cells) 2.1 2.1 For both scenarios 1 and 2, SCHH efflux CLint,T for E7766 measured at 0.3 µmol/l after 20-min incubation at 37 °C (Table 1) was used as input value for efflux CLint,T. B/P, blood-to-plasma partition ratio; CLPD, passive diffusion clearance; fu, unbound drug fraction in plasma; fuIW, unbound drug fraction in intracellular water; fuEW, unbound drug fraction in extracellular water; fuinc, unbound drug fraction in in vitro incubation system; Kp, tissue-to-plasma partition coefficient; uptake CLint,T, intrinsic uptake clearance obtained from SCHH assay; efflux CLint,T, intrinsic biliary efflux clearance obtained from SCHH assay. For scenario 1, hepatic uptake clearance was assigned from the transporter kinetics measured in HEK293 cells and for scenario 2, intrinsic active uptake clearance measured in from SCHH was assigned to OATP1B3.
- TABLE 5
Summary and comparison of simulated PK and DDI parameters of E7766 from PBPK models using two scenarios
SimCYP default compound for Rifampicin was used for simulations. For scenario 1, hepatic uptake clearance was assigned from the transporter kinetics measured in HEK293 cells, and for scenario 2, intrinsic active uptake clearance measured in from SCHH was assigned to OATP1B3.
PK Profile Parameters of E7766 Scenario 1 Scenario 2 (−) Rifampicin (+) Rifampicin (−) Rifampicin (+) Rifampicin AUCtotal (nmol·h/l) 47.38 131.06 67.26 174.07 AUCtotal ratio 2.77 2.59 Cmax,tot (nmol/l) 474.01 479.92 476.24 480.12 Cmax,tot ratio 1.01 1.01 CLtot,p (l/h) 29.62 11.88 21.55 8.66 CLtot,p ratio 0.40 0.40 Liver intracellular AUCfree (nmol·h/l) 76.19 69.06 71.35 61.28 Liver intracellular AUCfree ratio 0.91 0.86 AUCtotal ratio, ratio of AUCtotal in the presence and absence of the inhibitor; CLtot,p ratio, ratio of CLtot,p in the presence and absence of the inhibitor; Cmax,tot, maximum total plasma conc.; Cmax,tot ratio, ratio of Cmax,tot in the presence and absence of the inhibitor; liver intracellular AUCfree ratio, ratio of area under free intrahepatocellular conc.-time curve in the presence and absence of the inhibitor.
Data Supplement
- Supplemental Figure 1 -
Supplemental Figure 1 - Uptake activity of positive controls in the presence and absence of inhibitors in uptake transporter-expressing cells (A) or efflux transporter-expressing vesicles (B).
- Supplemental Figure 2 -
Supplemental Figure 2 - Time-dependent uptake of E7766 with HEK293-control (empty circle, A, B), HEK293-OATP1B1 (solid circle, A) and HEK293-OAPT1B3 (solid circle, B) cells. All experiments were run in triplicates.
- Supplemental Figure 3 -
Supplemental Figure 3 - Systemic unbound concentration versus time profile of Rifampicin in mice after i.v. bolus coadministration of E7766 and Rifampicin. Data are presented as mean ± SD (N=3).
- Supplemental Table 1 -
Supplemental Table 1 - Permeability assessment of E7766 in LLC-PK1 cells.
- Supplemental Table 2 -
Supplemental Table 2 - OATP1B1 and OATP1B3 protein expression levels in human hepatocytes and HEK293FT overexpressing cell lines.
- Supplemental Table 3 -
Supplemental Table 3 - Demographic information of hepatocyte donor used in SCHH study.
- Supplemental Table 4 -
Supplemental Table 4 - Summary of input parameters of Rifampicin used to conduct drug-drug interaction simulation of E7766.
- Supplemental Figure 1 -
In this issue
Transporter-Mediated Drug-Drug Interactions for E7766
