Clemizole is an H1 histamine receptor antagonist, can inhibit NS4B's RNA binding and hepatitis C virus (HCV) replication.

H1 (RNA binding by NS4B):24±1 nM

Clemizole hydrochloride 能够抑制 HCV RNA 在细胞培养中的复制，这一作用是通过其对 NS4B RNA 结合活性的抑制实现的，对宿主细胞的毒性很小。Clemizole 对 W55R 突变型 J6/JFH RNA 的半抑制浓度（EC50）约为 18 μM（是野生型 RNA EC50 的 2.25 倍）[1]。Clemizole 是 TRPC5 通道的新型抑制剂，能够在低μM范围内有效阻断 TRPC5 电流和 Ca2+ 入口（IC50=1.0-1.3 μM）。Clemizole 对 TRPC5 的选择性是对其最接近的结构相似体 TRPC4β（IC50=6.4 μM）的六倍，对 TRPC3（IC50=9.1 μM）和 TRPC6（IC50=11.3 μM）的选择性几乎为十倍。作为 TRPC5 的新型阻断剂，Clemizole hydrochloride 的IC50 为 1.1 μM。浓度-反应曲线确认 Clemizole 对 TRPC5 的阻断作用是浓度依赖性的，并显示出明显的 IC50 为 1.1±0.04 μM[2]。

Clemizole hydrochloride 在 C57BL/6J 小鼠体内的血浆半衰期异常短（测定为0.15小时），并且非常迅速地转化为一种葡萄糖醛酸苷代谢物（M14）和一个脱烷基化代谢物（M12），以及多种较小的代谢物[3]。

Clemizole hydrochloride is dissolved in DMSO and stored, and then diluted with appropriate media before use[1]. Huh7.5 cells are maintained in DMEM supplemented with 1% L-glutamine, 1% penicillin, 1% streptomycin, 1× nonessential amino acids and 10% FBS. Cell lines are passaged twice weekly after treatment with 0.05% trypsin-0.02% EDTA and seeding at a dilution of 1:5. Subconfluent Huh7.5 cells are trypsinized and collected by centrifugation at 700 g for 5 min. The cells are then washed three times in ice-cold RNase-free PBS and resuspended at 1.5×107 cells/mL in PBS. Wild-type or mutant FL-J6/JFH-5′C19Rluc2AUbi RNA for electroporation is generated by transcription of XbaI linearized DNA templates using the T7 MEGAscript kit, followed by purification (RNA transcription and fluorescent labeling). We mixed 5 μg of RNA with 400 μL of washed Huh7.5 cells in a 2-mm-gap cuvette (BTX) and immediately pulsed (0.82 kV, five 99 μs pulses) with a BTX-830 electroporator. After a 10 min recovery at 25°C, pulsed cells are diluted into 10 mL of prewarmed growth medium. Cells from several electroporations are pooled to a common stock and seeded in 6-well plates (5×105 cells per well). After 24 h, medium is replaced and cells are grown in the presence of serial dilutions of the various inhibitory compounds (e.g., Clemizole hydrochloride) identified in the screen. Seventeen commercially available compounds, out of the 18 identified, are analyzed. Untreated cells are used as a negative control for water-soluble compounds. For compounds (e.g., Clemizole hydrochloride) solubilized in DMSO, untreated cells are grown in the presence of corresponding concentrations of the solvent as a negative control. Medium is changed daily. After 72 h of treatment cells are subjected to an Alamar Blue-based viability assay and luciferase assay. After 72 h of treatment cells are incubated for 3 h at 37°C in the presence of 10% Alamar Blue reagent.Plates are then scanned and fluorescence is detected by using FLEXstation II 384. Depending on the inhibitory compound's solvent (e.g., Clemizole hydrochloride), water or DMSO, signal is normalized relatively to untreated samples or samples grown in the presence of DMSO, respectively[1].