Cepharanthin (CEP) inhibited DNA‐binding activity, nuclear translocation, and expression of nuclear factor‐kappa B (NF‐κB) in CCA cells. KKU‐M213 cells were treated with or without 10 μg/mL of CEP for 6, 12, 24, and 48 h, and the effects of CEP on NF‐κB activity determined. (a) Inhibitions of constitutive NF‐κB binding activity were examined in nuclear extracts by electrophoretic mobility shift assay (EMSA). Reaction without cell lysate was used as control and reaction with unlabeling oligonucleotide (× 200 of labeling probe) was used as a competitor. (b) CEP inhibited expression of NF‐κB p50/p105, p52/p100, and p65 in whole cell lysate of KKU‐M213, as determined by western blotting. (c) CEP reduced nuclear translocations of NF‐κB p50, p52, and p65. (d) Amount of IKK, IκB, and their phosphorylated forms after CEP treatment. Intensity of each band was normalized with actin or Histone H1, and compared relative to those without CEP (=1), as indicated by the number shown on the top of each band.