Lomibuvir (VCH-222) is a selective, non-nucleoside allosteric inhibitor of HCV NS5B polymerase (RdRp) with a Kd of 17 nM. Lomibuvir inhibits the 1b/Con1 HCV subgenomic replicon with an EC 50 of 5.2 nM. Lomibuvir preferentially inhibits elongative RNA synthesis rather than de novo -initiated RNA synthesis [1].

HCV NS5B 1a:0.94-1.2 μM

Lomibuvir (VX-222) 以EC50为5.2 nM的效力抑制WT HCV 1b/Con1复制子。对于突变复制子M423T、L419M和I482L，Lomibuvir的EC50分别为79.8 nM、563.1 nM和45.3 nM。Lomibuvir轻微减少了de novo启动，但强烈抑制引物延伸。Lomibuvir对引物延伸的RNA合成的IC50为31 nM [1]。Lomibuvir是一种非核苷类、与位点异源的丙型肝炎病毒NS5B聚合酶抑制剂，已证明其在临床上有效 [2]。

在大鼠和犬类动物中，Lomibuvir展示了良好的药代动力学特性，包括低总体清除率和优异的口服生物利用度（高于30%）以及良好的ADME特性。Lomibuvir经由数个酶（CYP1A1、2A6、2B6、2C8、CYP3A4、UGT1A3）生物转化，预计在肝脏中主动转运，并主要以完整形式或作为糖苷结合物通过胆汁排泄。[3]

Anti-NS5B activity assay: The inhibitory effect of VX-222 on HCV NS5B activity is measured by evaluating the amount of radiolabeled UTP incorporated by the C-terminal ?21 truncated version of enzyme in a newly synthesized RNA using a homopolymeric RNA template / primer namely poly rA / oligo dT. Quantitative detection of incorporated radioactivity is done using a liquid scintillation counter. The in vitro kinetics of inhibition of HCV NS5B from genotype 1b strain BK by VX-222 are determined using the C-terminal ?21 truncated version of NS5B. VX-222 (1 to 1.5 μM) is tested in the presence of 10 to 75 μM nonradioactive UTP mixed with 0.89 to 6.70 μCi of [α-33P]-labeled UTP. RNA-dependent-RNA polymerase reactions are allowed to proceed for 18 min at 22 °C.

Huh7.5 cells harboring HCV RNA replicons are trypsinized and plated into 48-well plates at a concentration of 4 × 104 cells/well. The next day the medium is changed and VX-222 is added in 200 μL of complete medium. After 48 hours, total RNA is extracted and viral RNAs are quanti?ed by real-time reverse transcription-PCR (RT-PCR). The effective drug concentrations that reduced HCV RNA replicon levels by 50% (EC50) are calculated by nonlinear regression analysis with log curve ?tting.(Only for Reference)