title: Induction of apoptosis in human leukemia cells through an intrinsic pathway by cathachunine, a unique alkaloid isolated from Catharanthus roseus.
abstract: BACKGROUND
  Catharanthus roseus (L.) G. Don consists of a range of dimeric indole alkaloids with significant antitumor activities. These alkaloids have been found to possess apoptosis-inducing activity against tumor cells in vitro and in vivo mediated by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and c-Jun N-terminal kinase (JNK) pathways, in which DNA damage and mitochondrial dysfunction play important roles. In this study, a unique bisindole alkaloid named cathachunine, along with five known dimeric indole alkaloids, was obtained from C. roseus and investigated in vitro.
  
  
  OBJECTIVE
  The aim of this study was to investigate the antitumor activity of isolated alkaloids and the mechanism through which cathachunine exerts its antitumor effect.
  
  
  METHODS
  Cell growth inhibition was assessed by WST-1 and lactate dehydrogenase (LDH) assays in HL60, K562 leukemia cells and EA.hy926 umbilical vein cells. Induction of apoptosis in HL60 cells was confirmed by observation of nuclear morphology, a caspase-3 activity assay and annexin V-fluorescein isothiocyanate/propidium iodide (FITC/PI) double staining. The intrinsic apoptotic pathway induced by cathachunine was evidenced by B-cell lymphoma 2/Bcl-2-associated X protein (Bcl-2/Bax) dysregulation, loss of mitochondrial membrane potential, translocation of cytochrome c, and cleavage of caspase-3 and poly-ADP ribose polymerase (PARP). Reactive oxygen species (ROS) production after cathachunine treatment was determined by 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining. Cell cycle arrest of the S phase was also observed in HL60 cells after cathachunine treatment.
  
  
  RESULTS
  The WST-1 and LDH assays showed that Catharanthus alkaloids were cytotoxic toward human leukemia cells to a greater extent than toward normal human endothelial cells, and the anti-proliferation and pro-apoptosis abilities of cathachunine were much more potent than other previously reported alkaloids. The induction of apoptosis by cathachunine occurred through an ROS-dependent mitochondria-mediated intrinsic pathway rather than an extrinsic pathway, and was regulated by the Bcl-2 protein family.
  
  
  CONCLUSIONS
  An unprecedented bisindole alkaloid cathachunine which lost C-18' and C-19' was isolated from C. roseus. It exerted a potent antitumor effect toward human leukemia cells through the induction of apoptosis via an intrinsic pathway. Thus, this study provides evidence for a new lead compound from a natural source for anti-cancer investigations.
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Wang
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Chen-Yang
authors.initials: CY
authors.lastname: Li
authors.affiliation: Tianjin Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin 300193, PR China.
authors.forename: Miao-Miao
authors.initials: MM
authors.lastname: Jiang
authors.affiliation: Tianjin Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin 300193, PR China.
authors.forename: Dong
authors.initials: D
authors.lastname: Li
authors.affiliation: Business Technology Department, Shenzhen Institute for Drug Control, Shenzhen 518057, Guangdong, PR China.
authors.forename: Ping
authors.initials: P
authors.lastname: Wen
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Xun
authors.initials: X
authors.lastname: Song
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Jun-Da
authors.initials: JD
authors.lastname: Chen
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Li-Xuan
authors.initials: LX
authors.lastname: Guo
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Xiao-Peng
authors.initials: XP
authors.lastname: Hu
authors.affiliation: Experiment and Technology Center, Jinan University, Guangzhou 510632, Guangdong, PR China.
authors.forename: Guo-Qiang
authors.initials: GQ
authors.lastname: Li
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China.
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Tianjin Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin 300193, PR China. Electronic address: pharmwch@126.com.
authors.forename: Chun-Hua
authors.initials: CH
authors.lastname: Wang
authors.affiliation: Department of Pharmacy, School of Medicine, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Institute of Biotherapy, Shenzhen University, Shenzhen 518060, Guangdong, PR China; Engineering Laboratory of Shenzhen Natural Micromolecule Innovative Drugs, Shenzhen University, Shenzhen 518060, Guangdong, PR China. Electronic address: hezhendan@126.com.
authors.forename: Zhen-Dan
authors.initials: ZD
authors.lastname: He
mesh_terms: D000470:Alkaloids / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D030008:Catharanthus / Q000737:chemistry; D002681:China; D006801:Humans; D020014:K562 Cells / Q000187:drug effects; D007938:Leukemia / Q000188:drug therapy*; D010936:Plant Extracts / Q000494:pharmacology*
chemical_list: D000470:Alkaloids; D000970:Antineoplastic Agents; D010936:Plant Extracts
keywords: Apoptosis; Cathachunine; Intrinsic pathway; ROS