title: Antiangiogenic and antihepatocellular carcinoma activities of the Juniperus chinensis extract.
abstract: BACKGROUND
  To identify a novel therapeutic agent for hepatocellular carcinoma (HCC), for which no promising therapeutic agent exists, we screened a panel of plants and found that Juniperus chinensis exhibited potential antiangiogenic and anti-HCC activities. We further investigated the antiangiogenic and anti-HCC effects of the active ingredient of J. chinensis extract, CBT-143-S-F6F7, both in vitro and in vivo.
  
  
  METHODS
  A tube formation assay conducted using human umbilical vein endothelial cells (HUVECs) was first performed to identify the active ingredient of CBT-143-S-F6F7. A series of angiogenesis studies, including HUVEC migration, Matrigel plug, and chorioallantoic membrane (CAM) assays, were then performed to confirm the effects of CBT-143-S-F6F7 on angiogenesis. The effects of CBT-143-S-F6F7 on tumor growth were investigated using a subcutaneous and orthotopic mouse model of HCC. In vitro studies were performed to investigate the effects of CBT-143-S-F6F7 on the cell cycle and apoptosis in HCC cells. Moreover, protein arrays for angiogenesis and apoptosis were used to discover biomarkers that may be influenced by CBT-143-S-F6F7. Finally, nuclear magnetic resonance analysis was conducted to identify the compounds of CBT-143-S-F6F7.
  
  
  RESULTS
  CBT-143-S-F6F7 showed significantly antiangiogenic activity in various assays, including HUVEC tube formation and migration, CAM, and Matrigel plug assays. In in vivo studies, gavage with CBT-143-S-F6F7 significantly repressed subcutaneous Huh7 tumor growth in severe combined immunodeficient (SCID) mice, and prolonged the survival of orthotopic Huh7 tumor-bearing SCID mice (a 40 % increase in median survival duration compared with the vehicle-treated mice). Immunohistochemical staining of subcutaneous Huh7 tumors in CBT-143-S-F6F7-treated mice showed a significantly decrease in the cell cycle regulatory protein cyclin D1, cellular proliferation marker Ki-67, and endothelial marker CD31. CBT-143-S-F6F7 caused arrest of the G2/M phase and induced Huh7 cell apoptosis, possibly contributing to the inhibition of HCC tumors. Protein array analysis revealed that several angiogenic and antiapoptotic factors were suppressed in CBT-143-S-F6F7-treated Huh7 cells. Finally, five compounds from CBT-143-S-F6F7 were identified.
  
  
  CONCLUSIONS
  According to these results, we report for the first time the antiangiogenic and anti-HCC activities of CBT-143-S-F6F7, the active fractional extract of J. chinensis. We believe that CBT-143-S-F6F7 warrants further evaluation as a new anti-HCC drug.
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Zong-Keng
authors: ZK
authors: Kuo
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Mei-Wei
authors: MW
authors: Lin
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: I-Huang
authors: IH
authors: Lu
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Hsin-Jan
authors: HJ
authors: Yao
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Hsin-Chieh
authors: HC
authors: Wu
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Chun-Chung
authors: CC
authors: Wang
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Shyh-Horng
authors: SH
authors: Lin
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Si-Yuan
authors: SY
authors: Wu
authors: Greenhouse System Technology Center, Industrial Technology Research Institute, Central Region Campus, No.2 Wenxian Rd., Nantou City, 54041, Taiwan.
authors: Tien-Soung
authors: TS
authors: Tong
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Yi-Cheng
authors: YC
authors: Cheng
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Jui-Hung
authors: JH
authors: Yen
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Ching-Huai
authors: CH
authors: Ko
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan.
authors: Shu-Jiau
authors: SJ
authors: Chiou
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan. I-HorngPan@itri.org.tw.|, Present address: Bldg 13, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan. I-HorngPan@itri.org.tw.
authors: I-Horng
authors: IH
authors: Pan
authors: Center of Excellence for Drug Development, Biomedical Technology and Device Research Labs, Industrial Technology Research Institute, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan. tsenghw@itri.org.tw.|, Present address: Bldg 13, No. 321, Sec 2, Kuangfu Rd, Hsinchu City, 30011, Taiwan. tsenghw@itri.org.tw.
authors: Hsiang-Wen
authors: HW
authors: Tseng
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D002460:Cell Line; D005260:Female; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D018653:Juniperus / Q000737:chemistry*; D008099:Liver / Q000187:drug effects; D008113:Liver Neoplasms / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID; D009389:Neovascularization, Pathologic / Q000378:metabolism; D010936:Plant Extracts / Q000494:pharmacology*
chemical_list: D020533:Angiogenesis Inhibitors; D000970:Antineoplastic Agents; D010936:Plant Extracts
keywords: Antiangiogenesis; Hepatocellular carcinoma; Juniperus chinensis; Therapeutic agent