title: Glycogen synthase kinase-3β, NF-κB signaling, and tumorigenesis of human osteosarcoma.
abstract: BACKGROUND
  Glycogen synthase kinase-3β (GSK-3β), a serine/threonine protein kinase, may function as a tumor suppressor or an oncogene, depending on the tumor type. We sought to determine the biological function of GSK-3β in osteosarcoma, a rare pediatric cancer for which the identification of new therapeutic targets is urgent.
  
  
  METHODS
  We used cell viability assays, colony formation assays, and apoptosis assays to analyze the effects of altered GSK-3β expression in U2OS, MG63, SAOS2, U2OS/MTX300, and ZOS osteosarcoma cell lines. Nude mice (n = 5-8 mice per group) were injected with U2OS/MTX300, and ZOS cells to assess the role of GSK-3β in osteosarcoma growth in vivo and to evaluate the effects of inhibitors and/or anticancer drugs on tumor growth. We used an antibody array, polymerase chain reaction, western blotting, and a luciferase reporter assay to establish the effect of GSK-3β inhibition on the nuclear factor-κB (NF-κB) pathway. Immunochemistry was performed on primary tumor specimens from osteosarcoma patients (n = 74) to determine the relationship of GSK-3β activity with overall survival.
  
  
  RESULTS
  Osteosarcoma cells with low levels of inactive p-Ser9-GSK-3β formed colonies in vitro and tumors in vivo more readily than cells with higher levels and cells in which GSK-3β had been silenced formed fewer colonies and smaller tumors than parental cells. Silencing or pharmacological inhibition of GSK-3β resulted in apoptosis of osteosarcoma cells. Inhibition of GSK-3β resulted in inhibition of the NF-κB pathway and reduction of NF-κB-mediated transcription. Combination treatments with GSK-3β inhibitors, NF-κB inhibitors, and chemotherapy drugs increased the effectiveness of chemotherapy drugs in vitro and in vivo. Patients whose osteosarcoma specimens had hyperactive GSK-3β, and nuclear NF-κB had a shorter median overall survival time (49.2 months) compared with patients whose tumors had inactive GSK-3β and NF-κB (109.2 months).
  
  
  CONCLUSIONS
  GSK-3β activity may promote osteosarcoma tumor growth, and therapeutic targeting of the GSK-3β and/or NF-κB pathways may be an effective way to enhance the therapeutic activity of anticancer drugs against osteosarcoma.
authors.affiliation: State Key Laboratory of Oncology in South China, Sun Yat-Sen University Cancer Center, Guangzhou 510060, China.
authors.forename: Qing-Lian
authors.initials: QL
authors.lastname: Tang
authors.forename: Xian-Biao
authors.initials: XB
authors.lastname: Xie
authors.forename: Jin
authors.initials: J
authors.lastname: Wang
authors.forename: Qiong
authors.initials: Q
authors.lastname: Chen
authors.forename: An-Jia
authors.initials: AJ
authors.lastname: Han
authors.forename: Chang-Ye
authors.initials: CY
authors.lastname: Zou
authors.forename: Jun-Qiang
authors.initials: JQ
authors.lastname: Yin
authors.forename: Da-Wei
authors.initials: DW
authors.lastname: Liu
authors.forename: Yi
authors.initials: Y
authors.lastname: Liang
authors.forename: Zhi-Qiang
authors.initials: ZQ
authors.lastname: Zhao
authors.forename: Bi-Cheng
authors.initials: BC
authors.lastname: Yong
authors.forename: Ru-Hua
authors.initials: RH
authors.lastname: Zhang
authors.forename: Qi-Sheng
authors.initials: QS
authors.lastname: Feng
authors.forename: Wu-Guo
authors.initials: WG
authors.lastname: Deng
authors.forename: Xiao-Feng
authors.initials: XF
authors.lastname: Zhu
authors.forename: Binhua P
authors.initials: BP
authors.lastname: Zhou
authors.forename: Yi-Xin
authors.initials: YX
authors.lastname: Zeng
authors.forename: Jing-Nan
authors.initials: JN
authors.lastname: Shen
authors.forename: Tiebang
authors.initials: T
authors.lastname: Kang
mesh_terms: D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D001859:Bone Neoplasms / Q000378:metabolism*; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival; D020868:Gene Silencing; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007150:Immunohistochemistry; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D016328:NF-kappa B / Q000378:metabolism*; D009857:Oncogenes; D012516:Osteosarcoma / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014183:Transplantation, Heterologous; D014410:Tumor Stem Cell Assay
chemical_list: D016328:NF-kappa B; D008156:Luciferases; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3; D053148:Caspase 3