| 細胞實驗 | Cytotoxicity is determined by incubating cells (3,000 to 10,000 cells/well) with serially diluted test compounds or DMSO for 5 days (MT-2 cells) or 4 days (all other cell types). Cell viability is quantitated using an MTS assay for MT-2 or a Cell-Titer Blue reagent assay for HEK-293, HuH-7, HepG2, and MRC5 cells, and 50% cytotoxic concentrations (CC50s) are calculated. |
| 動物實驗 | Mice (n=9 per group; overnight fast) receive Asunaprevir (ASV) by oral gavage (5 mg/kg; a vehicle of PEG-400-ethanol, 9:1). Blood samples (-0.2 mL) are obtained by retro-orbital bleeding at 0.25, 0.5, 1, 3, 6, 8, and 24 h after dosing. Within each group, three animals are bled at 0.25, 3, and 24 h, three at 0.5 and 6 h, and three at 1 and 8 h, resulting in a composite pharmacokinetic profile. Livers and brains are also removed from mice at the terminal sampling points. Rats (n=3 per group; overnight fast) receive ASV (amorphous free acid) by oral gavage (3, 5, 10, and 15 mg/kg) in PEG-400-ethanol (9:1). Serial blood samples (-0.3 mL) are obtained from the jugular vein predosing (0 h) and at 0.25, 0.5, 0.75, 1, 2, 4, 6, 8, 24, and 48 h post dosing. To assess tissue exposure, rats are orally administered ASV (5 or 15 mg/kg, same vehicle as above), and blood, liver, and heart samples from two rats/group are obtained at 0.17, 0.5, 1, 2, 4, 6, 8, 24, 48, and 72 h after dosing. |