title: Biological activity of dihydroartemisinin in canine osteosarcoma cell lines.
abstract: OBJECTIVE
  To evaluate the biological activity of dihydroartemisinin on canine osteosarcoma cell lines in vitro.
  
  
  METHODS
  4 canine osteosarcoma cell lines.
  
  
  METHODS
  Cell viability assays were performed on canine osteosarcoma cell lines OSCA2, OSCA16, OSCA50, and D17 after 24, 48, and 72 hours of treatment with dihydroartemisinin at concentrations of 0.1 to 100 microM. Apoptosis was assessed by use of an ELISA for free nuclosomal DNA fragmentation and by western blot analysis for cleavage of caspase 3. Cell cycle analysis was performed by use of staining with propidium iodide and flow cytometry. Detection of reactive oxygen species (ROS) was conducted in the D17 cell line by use of 6-carboxy-2',7'-dihydrofluorescein diacetate and flow cytometry.
  
  
  RESULTS
  The concentration of dihydroartemisinin required for 50% inhibition of cell viability (IC50) was achieved in all 4 canine osteosarcoma cell lines and ranged from 8.7 to 43.6 microM. Induction of apoptosis was evident as an increase in nucleosomal DNA fragmentation, cleavage of caspase 3, and an increase in the population in the sub G0/G1 phase of the cell cycle detected by flow cytometry. Exposure to dihydroartemisinin also resulted in a decrease in the G0/G1 population. Iron-dependent generation of ROS was detected in dihydroartemisinin-treated D17 cells; ROS generation increased in a dose-dependent manner.
  
  
  CONCLUSIONS
  Incubation with dihydroartemisinin resulted in biological activity against canine osteosarcoma cell lines, which included induction of apoptosis and arrest of the cell cycle. Clinical trials of dihydroartemisinin in dogs with osteosarcoma should be conducted.
authors.affiliation: Department of Veterinary Clinical Sciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210, USA.
authors.forename: Kenji
authors.initials: K
authors.lastname: Hosoya
authors.forename: Sridhar
authors.initials: S
authors.lastname: Murahari
authors.forename: Albert
authors.initials: A
authors.lastname: Laio
authors.forename: Cheryl A
authors.initials: CA
authors.lastname: London
authors.forename: C Guillermo
authors.initials: CG
authors.lastname: Couto
authors.forename: William C
authors.initials: WC
authors.lastname: Kisseberth
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D037621:Artemisinins / Q000494:pharmacology*; D015153:Blotting, Western / Q000662:veterinary; D001859:Bone Neoplasms / Q000188:drug therapy* / Q000473:pathology / Q000662:veterinary*; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D053938:DNA Fragmentation; D004283:Dog Diseases / Q000188:drug therapy* / Q000473:pathology; D004285:Dogs; D004354:Drug Screening Assays, Antitumor; D005434:Flow Cytometry / Q000662:veterinary; D020128:Inhibitory Concentration 50; D012516:Osteosarcoma / Q000188:drug therapy* / Q000473:pathology / Q000662:veterinary*; D017382:Reactive Oxygen Species / Q000378:metabolism; D012717:Sesquiterpenes / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D037621:Artemisinins; D017382:Reactive Oxygen Species; D012717:Sesquiterpenes; C039060:artenimol; D053148:Caspase 3