Simeprevir is a potent HCV NS3/4A protease inhibitor, and inhibits HCV replication with EC₅₀ of 8 nM.

EC50: 8 nM

In Huh7-Luc cells, antiviral activity of simeprevir (TMC435350) is dose dependent, and the EC₅₀ and EC₉₀ values determined for TMC435350 are 8 nM and 24 nM, respectively. Inhibition of TMC435350 on NS3/4A protease is time dependent, and the overall K_is are estimated to be 0.5 nM for genotype 1a and 0.4 nM for genotype 1b, respectively^[1]. TMC435350 is a potent inhibitor of HCV NS3/4A protease (K_i=0.36 nM) and viral replication (replicon EC₅₀=7.8 nM)^[2].

In rats, TMC435350 (40 mg/kg, p.o.) is extensively distributed to the liver and intestinal tract (tissue/plasma area under the concentration-time curve ratios of >35), and the absolute bioavailability is 44%^[1].

• [1]. Lin TI, et al. In vitro activity and preclinical profile of TMC435350, a potent hepatitis C virus protease inhibitor.Antimicrob Agents Chemother. 2009 Apr;53(4):1377-85. Epub 2009 Jan 26.

  [2]. Raboisson P, et al. Structure-activity relationship study on a novel series of cyclopentane-containing macrocyclic inhibitors of the hepatitis C virus NS3/4A protease leading to the discovery of TMC435350. Bioorg Med Chem Lett. 2008 Sep 1;18(17):4853-8.

In vitro inhibition of NS3/4A activity is determined using a fluorescence resonance energy transfer cleavage assay with the RetS1 peptide substrate, derived from the genotype 1a NS4A-4B junction, and bacterially expressed full-length NS3 protease domain, supplemented with an NS4A peptide. Briefly, NS3/4A is preincubated in the presence of TMC435350 for 10 min, and then the RetS1 substrate is added and fluorescence is continuously measured for 20 min (excitation, 355 nm; emission, 500 nm). Cleavage of the substrate is expressed as a percentage of the cleavage seen with the vehicle control. MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Simeprevir is diluted in in a final DMSO concentration of 0.5% in the absence of G418.

Huh7-Luc cells are seeded at a density of 2,500 cells/well in a 384-well plate in Dulbecco"s modified Eagle"s medium plus 10% fetal calf serum and incubated with a range of concentrations of serially diluted simeprevir (TMC435350), in a final DMSO concentration of 0.5% in the absence of G418. After 72 h of incubation, Steady Lite reagent is added in a 1:1 ratio to the medium, and luciferase signal is measured using a ViewLux reader.  MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Twenty-four male specific-pathogen-free Sprague-Dawley rats, weighing between 200 and 300 g at the time of dosing, are divided into eight groups of three rats each. Seven groups are dosed orally (p.o.) by gastric intubation of a vitamin E acetate-d-α-tocopheryl polyethylene glycol 1000 succinate-polyethylene glycol 400 solution of Simeprevir (TMC435350) at 2 mL/kg body weight to provide a dose of 40 mg/kg. One group is dosed intravenously (i.v.) by slow bolus injection in a tail vein of a 20% 2-hydroxypropyl-β-cyclodextrin formulation of TMC435350 (containing TMC435350, 100 mg/mL 2-hydroxypropyl-β-cyclodextrin, 0.1 N NaOH to pH 8.0±0.1, and mannitol-and pyrogen-free water) at 2 mL/kg body weight to provide a dose of 4 mg/kg. Water and food are available ad libitum during the study. MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Lin TI, et al. In vitro activity and preclinical profile of TMC435350, a potent hepatitis C virus protease inhibitor.Antimicrob Agents Chemother. 2009 Apr;53(4):1377-85. Epub 2009 Jan 26.

  [2]. Raboisson P, et al. Structure-activity relationship study on a novel series of cyclopentane-containing macrocyclic inhibitors of the hepatitis C virus NS3/4A protease leading to the discovery of TMC435350. Bioorg Med Chem Lett. 2008 Sep 1;18(17):4853-8.

749.94

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Room temperature in continental US; may vary elsewhere

DMSO

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