title: PBOX-15, a novel microtubule targeting agent, induces apoptosis, upregulates death receptors, and potentiates TRAIL-mediated apoptosis in multiple myeloma cells.
abstract: BACKGROUND
  In recent years, much progress has been made in the treatment of multiple myeloma. However, a major limitation of existing chemotherapeutic drugs is the eventual emergence of resistance; hence, the development of novel agents with new mechanisms of action is pertinent. Here, we describe the activity and mechanism of action of pyrrolo-1,5-benzoxazepine-15 (PBOX-15), a novel microtubule-targeting agent, in multiple myeloma cells.
  
  
  METHODS
  The anti-myeloma activity of PBOX-15 was assessed using NCI-H929, KMS11, RPMI8226, and U266 cell lines, and primary myeloma cells. Cell cycle distribution, apoptosis, cytochrome c release, and mitochondrial inner membrane depolarisation were analysed by flow cytometry; gene expression analysis was carried out using TaqMan Low Density Arrays; and expression of caspase-8 and Bcl-2 family of proteins was assessed by western blot analysis.
  
  
  RESULTS
  Pyrrolo-1,5-benzoxazepine-15 induced apoptosis in ex vivo myeloma cells and in myeloma cell lines. Death receptor genes were upregulated in both NCI-H929 and U266 cell lines, which displayed the highest and lowest apoptotic responses, respectively, following treatment with PBOX-15. The largest increase was detected for the death receptor 5 (DR5) gene, and cotreatment of both cell lines with tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), the DR5 ligand, potentiated the apoptotic response. In NCI-H929 cells, PBOX-15-induced apoptosis was shown to be caspase-8 dependent, with independent activation of extrinsic and intrinsic apoptotic pathways. A caspase-8-dependent decrease in expression of Bim(EL) preceded downregulation of other Bcl-2 proteins (Bid, Bcl-2, Mcl-1) in PBOX-15-treated NCI-H929 cells.
  
  
  CONCLUSIONS
  PBOX-15 induces apoptosis and potentiates TRAIL-induced cell death in multiple myeloma cells. Thus, PBOX-15 represents a promising agent, with a distinct mechanism of action, for the treatment of this malignancy.
authors.affiliation: John Durkan Leukaemia Laboratories, Institute of Molecular Medicine, Trinity College Dublin, St James's Hospital, Dublin 8, Ireland.
authors.forename: E N
authors.initials: EN
authors.lastname: Maginn
authors.forename: P V
authors.initials: PV
authors.lastname: Browne
authors.forename: P
authors.initials: P
authors.lastname: Hayden
authors.forename: E
authors.initials: E
authors.lastname: Vandenberghe
authors.forename: B
authors.initials: B
authors.lastname: MacDonagh
authors.forename: P
authors.initials: P
authors.lastname: Evans
authors.forename: M
authors.initials: M
authors.lastname: Goodyer
authors.forename: P
authors.initials: P
authors.lastname: Tewari
authors.forename: G
authors.initials: G
authors.lastname: Campiani
authors.forename: S
authors.initials: S
authors.lastname: Butini
authors.forename: D C
authors.initials: DC
authors.lastname: Williams
authors.forename: D M
authors.initials: DM
authors.lastname: Zisterer
authors.forename: M P
authors.initials: MP
authors.lastname: Lawler
authors.forename: A M
authors.initials: AM
authors.lastname: McElligott
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D015536:Down-Regulation / Q000187:drug effects; D006801:Humans; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000187:drug effects*; D009101:Multiple Myeloma / Q000473:pathology*; D010077:Oxazepines / Q000494:pharmacology*; D011758:Pyrroles / Q000494:pharmacology*; D053218:Receptors, Death Domain / Q000378:metabolism*; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000502:physiology*; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: C514160:4-acetoxy-5-(1-(naphthyl))naphtho(2,3-b)pyrrolo(2,1-d)(1,4)oxazepine; D010077:Oxazepines; D011758:Pyrroles; D053218:Receptors, Death Domain; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human