Mechanism of Action

Vaniprevir is a reversible, macrocyclic inhibitor of HCV NS3/4A protease which is essential for HCV replication.   HCV NS3/4A protease is involved in proteolytic proce- ssing of the HCV nonstructural polyprotein by cleaving four known sites along the virally encoded polyprotein.

In a biochemical assay, vaniprevir inhibited the protelytic activity of recombinant HCV NS3/4A proteases from all 6 HCV genotypes.

●    Genotype 1a/b, 2a, 4a, 5a, 6a: IC₅₀ = 0.016-0.54 nM.

●    Genotype 2b: IC₅₀ = 1.4 nM.

●    Genotype 3a: IC₅₀ = 53 nM.

●    Mutant genotype 1b: IC₅₀ = 6-101 nM.

Vaniprevir showed high selectivities (>10000 fold) for a panel of 169 receptors, ion channels and enzymes.

In Vitro Efficacy

Anti-HCV replicons in HCV replicon cells:

●    GT 1a/1b: EC₅₀ = 1-3 nM, EC₉₀ = 3.9-4 nM.

●    GT 2a: EC₅₀ = 2.7-7 nM, EC₉₀ = 35.5 nM.

●    GT 2b/5a: EC₅₀ = 1.8-61 nM, EC₉₀ = 5.9-157 nM.

●    GT 3a: EC₅₀ = 70 nM, EC₉₀ = 337 nM.

Inhibitions of vaniprevir in the presence of human serum proteins in Con1 cell line:

●    GT 1b 50% NHS: 5.7 fold increase in EC₅₀.

●    GT 1b 40% NHS: 3.1 fold increase in EC₅₀.

Substitutions changes during vaniprevir selection:

●    GT 1a/1b mutation in NS3/4A: A156 and D168.

Anti-HCV activities against HCV containing NS3/4A substitutions in cell culture:

●    GT 1a in replicon assays:

v    V36, Q41, T54, V55, Q80, V107: EC₅₀ = 0.5-2.1 nM (0.6-2.3 fold change).

v    R155, D168A/E: EC₅₀ = 39.9- >1000 nM (44.3- >1000 fold change).

v    A156S, V158I, D168N: EC₅₀ = 1.0-6.3 nM (1.1-7 fold change).

●    GT 1b in replicon assays:

v    V36, Q41, T54, V55, Q80: EC₅₀ = 0.7-14.6 nM (0.4-7.7 folds).

v    R155, A156, D168: EC₅₀ = 29- >2000 nM (15.3- >1000 folds).

v    A156S, D168N, V170: EC₅₀ = 5.0-9.7 nM (2.6-5.1 folds).

●    GT 1a in transient assays: R155, A156, D168: 21.5-346 folds.

●    GT 1b in transient assays:

v    V36, Q41, F43, T54, T55, Y56, R109, I153: 0.3-9.9 folds.

v    R155, A156, D168, V170: 32-463 folds.

v    A156S, V163I, V170I: 0.4-3.7 folds.

●    HCV from clinical:

v    GT 1a/1b mutation: EC₅₀ = 2.9-27 nM.

In Vivo Efficacy

The effects of vaniprevir on HCV-infected chimpanzee:

●    HCV genotype 1a infection.

v    Decrease 5 Log IU/mL in plasma at 5 mg/kg BID and 2-3 Log at 1 mg/kg BID.

●    HCV genotype 1b infection:

v    Reduced viral load for 2-3 Log IU/mL at 1 mg/kg BID.

HCV genotype 3a infection: The viral load was reduced to 10% at 5 mg/kg.

Absorption of Vaniprevir

Exhibited a non-linear pharmacokinetics in rats, dogs, monkeys and humans following oral dosing.  The increases in C_max and AUC appeared to be more than dose-proportional in the dose range of 5 to 500 mg/kg (rats), 5-30 mg/kg (dogs), 5-100 mg/kg (monkeys), and 40-1000 mg (humans) of vaniprevir.

Had low oral bioavailability in rats (0-5%), dogs (12%) and monkeys (2%).

Was absorbed rapidly with the T_max occurring 0.5 to 1.3 h in rats, dogs and monkeys after oral dose of 5 mg/kg administration, and T_max increased as the dose increased.  T_max in humans was 1.01-4.0 h after oral administration.

Showed a half-life ranging between 0.9-1.3 h in non-clinical species after intravenous administration, and the half-life in humans was 4.13-6.89 h after oral administration.

Had high clearance in rats (74 mL/min/kg), but moderate in dogs (11 mL/min/kg) and monkeys (18 mL/min/kg).

Exhibited a confined tissue distribution in rats, dogs and monkeys, with apparent volumes of distribution at 1.9, 0.3 and.0.4 L/kg, respectively, after intravenous administration.

Distribution of Vaniprevir

Exhibited very high plasma protein binding in mice (99.0%), rats (99.5%), rabbits (99.0%) and dogs (99.3%), and high in humans (97.7%-98.8%) and monkeys (98.2%-98.7%).  Note that vaniprevir was mainly bound to HSA.

Had a C_b:C_p ratio of 0.8 in humans, suggesting little penetration into red blood cells.

SD and Long Evans male rats after a single oral administration:

●    Concentrations of radioactivity were measurable in the medulla of the brain at one sampling time, 4 h post-dose, all other lobes of the brain had no quantifiable radioactivity, suggesting that [¹⁴C]vaniprevir has limited penetration of the blood brain barrier.

●    Relatively higher concentration levels were observed in GI tract contents, kidneys, stomach, small intestine and liver.  Vaniprevir was not measurable at any collection time point in cerebellum, cerebrum, eye lens, olfactory lobe, and spinal cord.

●    Elimination of radioactivity from tissues was complete in the majority of tissues by 24 h post-dose.

●    The amount of radioactive equivalents was similar and low in both pigmented and non-pigmented rats indicating the vaniprevir was not bound to melanin.

Vaniprevir demonstrated good liver exposure in rats, dogs, and monkeys after an oral dose administration.  The AUC ratios of vaniprevir in liver and plasma of rats, dogs, and monkeys after a 5 mg/kg oral dose were >874, 102, and 455, respectively.  In rats, the liver exposure increased more than dose proportionally between 5 and 30 mg/kg, but less than dose proportionally between 30 and 300 mg/kg.  The dog liver exposure was more than dose proportional between 2 and 5 mg/kg.

Metabolism of Vaniprevir

Several metabolites of vaniprevir (M6, M7, M8, M9, M10) were observed in liver microsomes or S9 and hepatocytes of humans, rats, dogs and mice.

Overall, the parent drug represented the most abundant component in human and non-clinical species plasma, and metabolites were trace or below the limited quantitation.

Vaniprevir was mainly metabolized by CYP3A. ^[7]

Excretion of Vaniprevir

Was predominantly excreted in feces in humans, with parent as the major component in human feces.

Approximately 23.1% of the excreted [¹⁴C]radioactivity was vaniprevir, followed by 19.7% as M10 in human feces after oral administration.

About 52.1% and 53.0% of radioactivities were recovered via biliary excretion in bile duct-cannulated (BDC) rats and dogs.

Drug-Drug Interaction

Vaniprevir was an inhibitor of CYP3A (IC₅₀ = 18.78 μM) and UGT1A1 (IC₅₀ = 19 μM).

Vaniprevir was not an inducer of CYP450 (CYP3A4, 2B6 or 1A2).

Vaniprevir was a substrate of P-gp, OATP1B1 and OATP1B3, but not of BCRP.

Vaniprevir had inhibition for P-gp (IC₅₀ = 38 μM), OATP1B1 (IC₅₀ = 0.3 μM), OATP1B3 (IC₅₀ = 0.3 μM), BESP (IC₅₀ = 1.3 μM), BCRP (IC₅₀ = 13 μM), MRP2 (IC₅₀ = 42.2 μM), MRP3 (IC₅₀ = 9.9 μM) and MRP4 (IC₅₀ = 8.0 μM).

Single-Dose Toxicity

Acute toxicity by the oral route in rodents:

●    Mouse MTD: 2000 mg/kg.

Repeated-Dose Toxicity

Sub- and chronic toxicological potential by the oral route in rats (up to 6 months) and dogs (up to 9 months):

●    As the longest studies employed, the NOAEL in rats was 120 mg/kg/day (8.3 × MRHD), while that in dogs was 15 mg/kg (2.9 × MRHD).

●    The cross-species adverse effects concentrated on liver and biliary system disorders which can be monitored clinically.  Furthermore, all lesions were identified mild, and given that both liver and gall bladder epithelium were regenerative. Hypernomic concern should not be required.

Safety Pharmacology

No apparent toxicity was observed in the core battery of safety pharmacology studies.

Genotoxicity

No genetic risk was verified by the standard battery of genotoxicity assays.

Reproductive and Developmental Toxicity

Fertility and early embryonic development in rats:

●    The NOAEL was 250 mg/kg/day (HD) for both males and females.

Embryo-fetal development in rats and rabbits:

●    The maternal NOAEL was 120 mg/kg/day (3.8 × MRHD) in rats and 120 mg/kg/day (0.37 × MRHD) in rabbits.  The NOAEL for fetus was 120 mg/kg/day (3.8 × MRHD) in rats and 240 mg/kg/day (2.0 × MRHD) in rabbits.

Pre- and postnatal development in rats:

●    The NOAEL was determined as 120 and 180 mg/kg/day for F₀ and F₁ in rats.

Vaniprevir can be transferred through placenta and excreted through milk.

Carcinogenicity

There was no effect on associated non-neoplastic findings up to 26 weeks in transgenic mice or up to 104 weeks in rats.