title: Niclosamide suppresses Hepatoma cell proliferation via the Wnt pathway.
abstract: BACKGROUND
  The Wnt pathway plays an important role in Hepatocarcinogenesis. We analyzed the association of the Wnt pathway with the proliferation of hepatoma cells using Wnt3a and niclosamide, a drug used to treat tapeworm infection.
  
  
  METHODS
  We performed an MTS assay to determine whether Wnt3a stimulated proliferation of Huh-6 and Hep3B human hepatoma cell lines after 72 hours of incubation with Wnt3a in serum-free medium. The cells were subjected to hematoxylin and eosin staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) after 48 hours of incubation. RNA was isolated 48 hours after addition of Wnt3a or niclosamide, and cyclin D1 expression levels were analyzed by real-time quantitative polymerase chain reaction. The promoter activity of T-cell factor was analyzed by luciferase assay 48 hours after transfection of TOPflash. Western blot analysis was performed with antibodies against β-catenin, dishevelled 2, and cyclin D1.
  
  
  RESULTS
  Cell proliferation increased with Wnt3a. Niclosamide suppressed proliferation with or without Wnt3a. Hematoxylin and eosin and TUNEL staining suggested that apoptosis occurred in cells with niclosamide. Cyclin D1 was upregulated in the presence of Wnt3a and downregulated with addition of niclosamide. The promoter activity of T-cell factor increased with Wnt3a, whereas T-cell factor promoter activity decreased with niclosamide. Western blot analysis showed that Wnt3a upregulated β-catenin, dishevelled 2, and cyclin D1, while niclosamide downregulated them.
  
  
  CONCLUSIONS
  Niclosamide is a potential candidate for the treatment of hepatoma.
authors.affiliation: Department of Gastroenterology, National Hospital Organization Shimoshizu Hospital, Yotsukaido City, Chiba, Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Tomizawa
authors.forename: Fuminobu
authors.initials: F
authors.lastname: Shinozaki
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Motoyoshi
authors.forename: Takao
authors.initials: T
authors.lastname: Sugiyama
authors.forename: Shigenori
authors.initials: S
authors.lastname: Yamamoto
authors.forename: Makoto
authors.initials: M
authors.lastname: Sueishi
authors.forename: Takanobu
authors.initials: T
authors.lastname: Yoshida
keywords: T-cell factor; Wnt3a; cyclin D1; luciferase assay; niclosamide