title: AMD3100 combined with triptolide inhibit proliferation, invasion and metastasis and induce apoptosis of human U2OS osteosarcoma cells.
abstract: BACKGROUND
  Osteosarcoma (OS) mainly occurs in children and adolescents, and has a high propensity for lung metastasis. Little is known about the role of SDF-1/CXCR4 axis in OS progression. AMD3100 is a specific CXCR4 antagonist. Triptolide can induce apoptosis and proliferation inhibition in various cancer cell lines.
  
  
  OBJECTIVE
  This work aimed to investigate the effects of AMD3100 plus triptolide on the proliferation, apoptosis, invasion and metastasis of OS cells.
  
  
  METHODS
  The expression levels of SDF-1 and CXCR4 in five OS cell lines was analyzed by qRT-PCR, western blotting and ELISA assays. The effect of AMD3100 and triptolide on the proliferation, apoptosis and invasion of U2OS cells was evaluated by CCK-8, flow cytometry and transwell assay, respectively. Orthotopic intra-tibial growth and lung metastasis mouse model of OS were employed to evaluate the inhibition effect of AMD3100 and triptolide on primary OS growth and lung metastasis.
  
  
  RESULTS
  CXCR4 protein expression was detected in HOS-8603, MG-63, U2OS and 143B but not Saos2 cells, and all these cell lines expressed SDF-1. AMD3100 plus triptolide induced proliferation inhibition and apoptosis of U2OS cells, which was attributed to the downregulation of c-Myc, survivin, cyclin D1 and increased cleaved caspase-3 and PARP. AMD3100 and triptolide also suppressed SDF-1 induced invasion of CXCR4+ U2OS cells, which was validated by decreased expression of MMP-2 and 9, VEGF, m-Calpain and β-catenin. Moreover, the phosphorylation levels of Erk1/2, Akt and STAT3, as well as the nuclear translocation and phosphorylation of NF-κB p65 in U2OS cells were also reduced by AMD3100 and triptolide. In vivo, AMD3100 and triptolide significantly reduced primary tumor growth and lung metastasis of U2OS cells.
  
  
  CONCLUSIONS
  AMD3100 combined with triptolide can reduce proliferation and metastasis, and induce apoptosis of U2OS cells, which may be related to the Erk1/2, Akt, STAT3 and NF-κB pathways.
authors.affiliation: Department of Pediatrics, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Chunming
authors.initials: C
authors.lastname: Jiang
authors.affiliation: Department of Clinical Laboratory, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Fang
authors.affiliation: Department of Ophthalmology, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Hongxu
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Xuepeng
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Maoqiang
authors.initials: M
authors.lastname: Li
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Wu
authors.initials: W
authors.lastname: Jiang
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Tian
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China. Electronic address: sixdrazhu@yeah.net.
authors.forename: Liulong
authors.initials: L
authors.lastname: Zhu
authors.affiliation: Department of Orthopedic Surgery, Hangzhou First People's Hospital, Nanjing Medical University, Hangzhou 310006, Zhejiang, China. Electronic address: bianzydoct@yeah.net.
authors.forename: Zhenyu
authors.initials: Z
authors.lastname: Bian
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001596:Benzylamines; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects*; D000080027:Cyclams; D004224:Diterpenes / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects; D004852:Epoxy Compounds / Q000494:pharmacology; D005260:Female; D006571:Heterocyclic Compounds / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D008809:Mice, Inbred C3H; D009361:Neoplasm Invasiveness / Q000473:pathology*; D009362:Neoplasm Metastasis / Q000188:drug therapy* / Q000473:pathology; D012516:Osteosarcoma / Q000188:drug therapy* / Q000473:pathology; D010616:Phenanthrenes / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D001596:Benzylamines; D000080027:Cyclams; D004224:Diterpenes; D004852:Epoxy Compounds; D006571:Heterocyclic Compounds; D010616:Phenanthrenes; C001899:triptolide; C088327:plerixafor
keywords: AMD3100; Akt; Erk1/2; Osteosarcoma; STAT3; Triptolide