Article Figures & Data
Figures
- Fig. 1.
Structural formula of fevipiprant and proposed biotransformation pathways of fevipiprant in humans. *Position of [14C]-radiolabel.
- Fig. 2.
Plasma concentrations of fevipiprant and AG metabolite, and total radioactivity after oral administration of a 200 mg dose of [14C]-fevipiprant to four healthy subjects. Units for fevipiprant and AG metabolite concentrations are ng/ml, for total radioactivity concentrations ng-eq/ml. Concentrations of fevipiprant and AG metabolite were determined by LC-MS/MS, and concentrations of total radioactivity were determined by LSC.
- Fig. 3.
Representative radiochromatograms in plasma (3 hours after dose), urine (time pooled 0–72 hours), and feces extracts (time pooled 0–96 hours) for one subject after oral administration of a 200-mg dose of [14C]-fevipiprant. dpm: disintegrations per minute.
- Fig. 4.
Cumulative excretion of radioactivity in urine and feces after oral administration of a 200-mg dose of [14C]-fevipiprant to four healthy subjects. Radioactivity was determined by LSC.
- Fig. 5.
(A) SDS-PAGE analysis of covalent protein binding after 24 hours of incubation of [14C]-AG metabolite (20 µM) with 1:1 human plasma/PBS at 37°C. Lanes A1–3: untreated incubation sample; lanes B1–3: incubation sample after albumin depletion using a ProteoExtract Albumin Removal Kit (Calbiochem). Lanes A1/B1, A2/B2, and A3/B3 show analyses of 5, 10, and 20 µg of protein, respectively. Protein content was determined using a Protein DC Assay (Bio-Rad Laboratories). Relative intensities of bands calculated using AIDA software: A1, 5878; A2, 9787; A3, 17,839; B1, 2791; B2, 6435; and B3, 10,635. The molecular weight markers on the left side of the gel were drawn with 3H-supplemented ink to mark the location of visible marker proteins (all blue standard; Bio-Rad Laboratories). (B) SEC analysis after 24 hours of incubation of [14C]-AG-metabolite (100 µM) with 1:1 human plasma/PBS at 37°C. (C) Representative SEC analyses of plasma samples from one healthy subject at 2, 6, and 120 hours after oral administration of a 200-mg dose of [14C]-fevipiprant.
Tables
- TABLE 1
PK parameters of total radioactivity in whole blood and plasma and of fevipiprant and its main circulating metabolite (AG metabolite) in plasma
Values are mean ± S.D.
Parameter Total Radioactivity Plasma Whole Blood Plasma Fevipiprant AG Metabolite Tmax (h)a 3.00 (3.00−6.00) 3.00 (3.00−6.00) 2.50 (2.00−6.00) 3.00 (3.00−6.00) Cmax (ng/ml or ng-eq/ml)b 923 ± 216 1750 ± 396 348 ± 139 1020 ± 247 AUC0−240 h (ng*h/ml or ng-eq*h/ml)c 36,400 ± 6960 76,500 ± 9740 2490 ± 312 6890 ± 1240 AUCinf (ng-eq*h/ml) 69,700 ± 10,600 156,000 ± 20,700 — — CL/F (l/h) — — 80.5 ± 9.42 — Vz/F (l) — — 1370 ± 459 — T1/2 (h) 230 ± 54.6 254 ± 24.8 12.3 ± 5.95 11.8 ± 5.19 - TABLE 2
Fevipiprant and metabolites in plasma, based on metabolism profiles
Values are presented as mean ± S.D.
Component [14C]-AUC0−12 ha ng/ml or ng-eq*h/mlb % of Total Fevipiprant 1690 ± 266 15.3 ± 2.33 AG metabolite 6240 ± 922 56.5 ± 5.65 P8.5 125 ± 145 1.18 ± 1.37 Sum of additional components 47.1 ± 52.9 0.405 ± 0.426 Total components detected 8100 ± 852 73.4 ± 2.73 Lost during sample processing 2920 ± 210 26.6 ± 2.73 Total radiolabeled components in sample 11,000 ± 837 100 Component Excretion (% of Dose)a Excretion (% of Dose)b Urine 0−72 h Feces 0−96 h Total Urine 0−240 h Fevipiprant 12.7 ± 2.21 44.5 ± 6.12 57.2 ± 6.29 11.0 ± 1.56 AG metabolite 26.9 ± 2.58 1.24 ± 0.244 28.1 ± 2.40 19.25 ± 2.48 Lactone metabolite 0.133 ± 0.0458 0.177 ± 0.355 0.310 ± 0.389 NA P5.3 0.0956 ± 0.0121 0.00 0.0956 ± 0.0121 NA Total components detected 39.8 ± 3.35 46.0 ± 6.36 85.7 ± 5.16 NA Total excretion in time periodc 41.2 ± 3.78 50.5 ± 6.26 91.7 ± 4.97 NA Incubation Conditions Amount of Covalent Drug Protein Adducts HLM, GSH 1 ± 0.0 pmol/mg protein HLM, NADPH 3 ± 0.4 pmol/mg protein HLM, NADPH, UDPGA 3 ± 0.3 pmol/mg protein HLM, NADPH, UDPGA, GSH 1 ± 0.5 pmol/mg protein HH 8 ± 1.4 pmol/106 cells GSH, glutathione; HLM, human liver microsomes; HH, human hepatocytes; NADPH, nicotine adenine dinucleotide phosphate; UDPGA, uridine diphosphoglucuronic acid.
- TABLE 5
Michaelis-Menten enzyme kinetic parameters for biotransformation of fevipiprant by UGT enzymes
UGT1A3 UGT2B7 UGT2B17 Vmax (mean ± S.D., pmol/min/mg) 5634 ± 1603 35.2 ± 6.8 43.3 ± 0.9 Km (mean ± S.D., µM) 13,352 ± 5854 320 ± 143 53.3 ± 4.5 Derived intrinsic clearance (Vmax/Km, µl/mg/min) 0.422 0.11 0.812
Data Supplement
- Supplemental Data -
SUPPLEMENTAL DATA - Methods
Supplemental Table 1 - Mass spectral data of [14C]-fevipiprant and metabolites
Supplemental Table 2 - Excretion of radioactivity in excreta (0-240 h)
Supplemental Table 3 - Biotransformation of fevipiprant by recombinant uridine diphosphate glucuronosyltransferase (UGTs)
Supplemental Table 4 - Hepatic fevipiprant uptake byhuman hepatocytes
Supplemental Figure 1 - MS/MS spectra and proposed fragmentation of A) fevipiprant B) AG metabolite and C) lactone metabolite
Supplemental Figure 2 - Covalent protein binding after incubation of [14C]-fevipiprant (20 mu-M) or [14C]-AG metabolite (20 mu-M) with 1:1 human plasma/PBS or PBS at 37oC , followed by protein precipitation, filtration and LSC quantification of protein-bound radioactivity
Supplemental Figure 3 - Enzyme kinetics of [14C]-fevipiprant metabolism by recombinant UGT1A3, presented as reaction rate vs substrate concentration, and as reaction rate vs rate/substrate concentration
Supplemental Figure 4 - Enzyme kinetics of [14C]-fevipiprant metabolism by recombinant UGT2B7 presented as reaction rate vs substrate concentration, and as reaction rate vs rate/substrate concentration
Supplemental Figure 5 - Enzyme kinetics of [14C]-fevipiprant metabolism by recombinant UGT2B17 presented as reaction rate vs substrate concentration, and as reaction rate vs rate/substrate concentration
Supplemental Figure 6 - Uptake of fevipiprant by OATP1B3-transporter expressing HEK293 cells presented as CLapp for various substrate concentrations and in the presence and absence of inhibitors of OATP1B3
Supplemental Figure 7 - Uptake of fevipiprant by MDR1-transporter expressing LLCPK1 cells presented as CLapp for two substrate concentrations and in the presence and absence of an inhibitor of MDR1
Supplemental Figure 8 - Uptake of fevipiprant by OAT3 transporter expressing HEK293 cells presented as PSapp for various substrate concentrations
Supplemental Figure 9 - Proposed possible pathways of lactone metabolite formation
Supplemental Figure 10 - Synthetic scheme for [14C]-fevipiprant. i) (14CH2O)n, H2O, Me2NH.HCl; ii) MeI, EtOH, 25°C; iii) NaCN, DMF, 100°C; iv) HCl, 100°C; v) MeOH, H2SO4, 100°C; vi) 5, CsCO3, acetone, reflux; vii 12% NaOH, rt; viii) HCl; ix) crystallization EtOH/H2O
References
- Supplemental Data -
