title: Silvestrol induces early autophagy and apoptosis in human melanoma cells.
abstract: BACKGROUND
  Silvestrol is a cyclopenta[b]benzofuran that was isolated from the fruits and twigs of Aglaia foveolata, a plant indigenous to Borneo in Southeast Asia. The purpose of the current study was to determine if inhibition of protein synthesis caused by silvestrol triggers autophagy and apoptosis in cultured human cancer cells derived from solid tumors.
  
  
  METHODS
  In vitro cell viability, flow cytometry, fluorescence microscopy, qPCR and immunoblot was used to study the mechanism of action of silvestrol in MDA-MB-435 melanoma cells.
  
  
  RESULTS
  By 24 h, a decrease in cyclin B and cyclin D expression was observed in silvestrol-treated cells relative to control. In addition, silvestrol blocked progression through the cell cycle at the G2-phase. In silvestrol-treated cells, DAPI staining of nuclear chromatin displayed nucleosomal fragments. Annexin V staining demonstrated an increase in apoptotic cells after silvestrol treatment. Silvestrol induced caspase-3 activation and apoptotic cell death in a time- and dose-dependent manner. Furthermore, both silvestrol and SAHA enhanced autophagosome formation in MDA-MB-435 cells. MDA-MB-435 cells responded to silvestrol treatment with accumulation of LC3-II and time-dependent p62 degradation. Bafilomycin A, an autophagy inhibitor, resulted in the accumulation of LC3 in cells treated with silvestrol. Silvestrol-mediated cell death was attenuated in ATG7-null mouse embryonic fibroblasts (MEFs) lacking a functional autophagy protein.
  
  
  CONCLUSIONS
  Silvestrol potently inhibits cell growth and induces cell death in human melanoma cells through induction of early autophagy and caspase-mediated apoptosis. Silvestrol represents a natural product scaffold that exhibits potent cytotoxic activity and could be used for the further study of autophagy and its relationship to apoptosis in cancer cells.
authors.affiliation: Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, Chicago, IL, 60607, USA. wchen54@uic.edu.
authors.forename: Wei-Lun
authors.initials: WL
authors.lastname: Chen
authors.affiliation: Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, Ohio State University, Columbus, OH, 43210, USA. xnplus@hotmail.com.
authors.forename: Li
authors.initials: L
authors.lastname: Pan
authors.affiliation: Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, Ohio State University, Columbus, OH, 43210, USA. kinghorn.4@osu.edu.
authors.forename: A Douglas
authors.initials: AD
authors.lastname: Kinghorn
authors.affiliation: Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, Chicago, IL, 60607, USA. swanson@pharmacy.wisc.edu.|School of Pharmacy, University of Wisconsin-Madison, Madison, WI, 53705, USA. swanson@pharmacy.wisc.edu.
authors.forename: Steven M
authors.initials: SM
authors.lastname: Swanson
authors.affiliation: Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, Chicago, IL, 60607, USA. joannab@uic.edu.
authors.forename: Joanna E
authors.initials: JE
authors.lastname: Burdette
mesh_terms: D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000008:administration & dosage; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D053148:Caspase 3 / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019926:Cyclin B / Q000096:biosynthesis; D056741:Cyclin D / Q000096:biosynthesis; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D018942:Macrolides / Q000008:administration & dosage; D008545:Melanoma / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D051379:Mice; D014315:Triterpenes / Q000008:administration & dosage*; D001572:Benzofurans
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D053148:Caspase 3; D019926:Cyclin B; D056741:Cyclin D; D018942:Macrolides; D014315:Triterpenes; C489897:silvestrol; D001572:Benzofurans; C057620:bafilomycin A