title: Resistance to bleomycin in cancer cell lines is characterized by prolonged doubling time, reduced DNA damage and evasion of G2/M arrest and apoptosis.
abstract: BACKGROUND
  To establish, characterize and elucidate potential mechanisms of acquired bleomycin (BLM) resistance using human cancer cell lines. Seven BLM-resistant cell lines were established by exposure to escalating BLM concentrations over a period of 16-24 months. IC50 values and cell doubling times were quantified using a real time cytotoxicity assay. COMET and γ-H2AX assays, cell cycle analysis, and apoptosis assessment further investigated the mechanisms of BLM resistance in these cell lines.
  
  
  RESULTS
  Compared with parental cell lines, real time cytotoxicity assays revealed 7 to 49 fold increases in IC50 and a mean doubling time increase of 147 % (range 64 %-352%) in BLM-resistant sub-clones (p<0.05 for both). Higher maintenance BLM concentrations were associated with higher IC50 and increased doubling times (p<0.05). Significantly reduced DNA damage (COMET and γ-H2AX assays), G2/M arrest, and apoptosis (p<0.05 for each set of comparison) following high-dose acute BLM exposure was observed in resistant sub-clones, compared with their BLM-sensitive parental counterparts. Three weeks of BLM-free culturing resulted in a partial return to BLM sensitivity in 3/7 BLM-resistant sub-clones (p<0.05).
  
  
  CONCLUSIONS
  Bleomycin resistance may be associated with reduced DNA damage after bleomycin exposure, resulting in reduced G2/M arrest, and reduced apoptosis.
authors.affiliation: Department of Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Qi
authors.initials: Q
authors.lastname: Wang
authors.affiliation: Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Kangping
authors.initials: K
authors.lastname: Cui
authors.affiliation: Department of Statistics and Actuarial Science, University of Waterloo, Waterloo, Ontario, Canada ; Department of Biostatistics, Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Osvaldo
authors.initials: O
authors.lastname: Espin-Garcia
authors.affiliation: Department of Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Dangxiao
authors.initials: D
authors.lastname: Cheng
authors.affiliation: Institute of Virology, Medical School of Wuhan University, Wuhan, Hubei, China.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Qiu
authors.affiliation: Department of Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Department of Medicine and Pharmacology, University of Toronto, Toronto, Ontario, Canada ; Division of Medical Oncology and Hematology, Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Malcolm
authors.initials: M
authors.lastname: Moore
authors.affiliation: Department of Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada ; Departments of Medical Biophysics and Radiation Oncology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Robert G
authors.initials: RG
authors.lastname: Bristow
authors.affiliation: Department of Biostatistics, Princess Margaret Hospital, Toronto, Ontario, Canada ; Dalla Lana School of Public Health, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Wei
authors.initials: W
authors.lastname: Xu
authors.affiliation: Department of Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada.
authors.forename: Sandy
authors.initials: S
authors.lastname: Der
authors.affiliation: Division of Medical Oncology and Hematology, Princess Margaret Hospital, Toronto, Ontario, Canada ; Dalla Lana School of Public Health, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Geoffrey
authors.initials: G
authors.lastname: Liu
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D001761:Bleomycin / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002999:Clone Cells; D020552:Comet Assay; D004249:DNA Damage; D004261:DNA Replication / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D005434:Flow Cytometry; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects*; D006657:Histones / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D013997:Time Factors
chemical_list: C468783:H2AX protein, human; D006657:Histones; D001761:Bleomycin