title: Effects of valproic acid on the cell cycle and apoptosis through acetylation of histone and tubulin in a scirrhous gastric cancer cell line.
abstract: BACKGROUND
  Management of peritoneal dissemination is the most critical problem in gastric cancer. This study was performed to investigate the inhibitory effects of valproic acid (VPA) on a highly peritoneal-seeding cell line of human scirrhous gastric cancer, OCUM-2MD3, and to explore the mechanism and the potential of VPA.
  
  
  METHODS
  The effects of VPA on the growth of OCUM-2MD3 cells were assessed by MTT assay. In addition, paclitaxel (PTX) was combined with VPA to evaluate their synergistic effects. HDAC1 and HDAC2 expression were evaluated by western blotting in OCUM-2MD3 cells and other gastric cancer cell lines (TMK-1, MKN-28). The acetylation status of histone H3 and α-tubulin after exposure to VPA were analyzed by western blotting. The activities of cell cycle regulatory proteins and apoptosis-modulating proteins were also examined by western blotting. The effects of VPA in vivo were evaluated in a xenograft model, and apoptotic activity was assessed by TUNEL assay.
  
  
  RESULTS
  OCUM-2MD3 cells showed high levels of HDAC1 and HDAC2 expression compared with TMK-1 and MKN-28. The concentration of VPA required for significant inhibition of cell viability (P < 0.05) was 5 mM at 24 h and 0.5 mM at 48 h and 72 h. The inhibition of VPA with PTX showed dose-dependent and combinatorial effects. VPA increased acetyl-histone H3, acetyl-α-tubulin, and p21WAF1 levels accompanied by upregulation of p27, caspase 3, and caspase 9, and downregulation of bcl-2, cyclin D1, and survivin. In the xenograft model experiment, the mean tumor volume of the VPA-treated group was significantly reduced by 36.4%, compared with that of the control group at 4 weeks after treatment (P < 0.01). The apoptotic index was significantly higher in the VPA-treated group (42.3% ± 3.5%) than in the control group (7.7% ± 2.5%) (P < 0.001).
  
  
  CONCLUSIONS
  VPA induced dynamic modulation of histone H3 and α-tubulin acetylation in relation with the anticancer effect and the enhancement of PTX in the OCUM-2MD3 cell line. Therefore, VPA in combination with PTX is expected to be a promising therapy for peritoneal dissemination of scirrhous gastric cancer.
authors.affiliation: Department of Gastroenterologic Surgery, Division of Cancer Medicine, Graduate School of Medical Science, Kanazawa University, Ishikawa 920-8641, Japan. y-yagi@live.jp
authors.forename: Yasumichi
authors.initials: Y
authors.lastname: Yagi
authors.forename: Sachio
authors.initials: S
authors.lastname: Fushida
authors.forename: Shinichi
authors.initials: S
authors.lastname: Harada
authors.forename: Jun
authors.initials: J
authors.lastname: Kinoshita
authors.forename: Isamu
authors.initials: I
authors.lastname: Makino
authors.forename: Katsunobu
authors.initials: K
authors.lastname: Oyama
authors.forename: Hidehiro
authors.initials: H
authors.lastname: Tajima
authors.forename: Hideto
authors.initials: H
authors.lastname: Fujita
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Takamura
authors.forename: Itasu
authors.initials: I
authors.lastname: Ninomiya
authors.forename: Takashi
authors.initials: T
authors.lastname: Fujimura
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Ohta
authors.forename: Masakazu
authors.initials: M
authors.lastname: Yashiro
authors.forename: Kosei
authors.initials: K
authors.lastname: Hirakawa
mesh_terms: D000107:Acetylation; D002293:Adenocarcinoma, Scirrhous / Q000378:metabolism / Q000473:pathology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D005260:Female; D056284:Histone Deacetylase 1 / Q000378:metabolism; D056464:Histone Deacetylase 2 / Q000378:metabolism; D006657:Histones / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D051379:Mice; D008819:Mice, Nude; D009361:Neoplasm Invasiveness; D013274:Stomach Neoplasms / Q000378:metabolism* / Q000473:pathology; D014404:Tubulin / Q000187:drug effects* / Q000378:metabolism; D014635:Valproic Acid / Q000494:pharmacology*; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D006657:Histones; D014404:Tubulin; D014635:Valproic Acid; C110550:HDAC1 protein, human; C533423:HDAC2 protein, human; D056284:Histone Deacetylase 1; D056464:Histone Deacetylase 2