Ciprofloxacin-induced G2 arrest and apoptosis in TK6 lymphoblastoid cells is not dependent on DNA double-strand break formation

Cancer Biol Ther. 2008 Jan;7(1):113-9. doi: 10.4161/cbt.7.1.5136. Epub 2007 Oct 8.

Abstract

Drugs developed for the treatment of conditions other than neoplasia can also show promise as potential antitumor agents. The fluoroquinolone antibiotic ciprofloxacin (CPFX) is known to modulate cycle cell progression and apoptosis in cancer cells, and is thought to induce DNA double-strand breaks (DSBs) via topoisomerase II (topo II) inhibition and stabilized cleavage complex (SCC) formation. DSBs trigger Ser-139 phosphorylation of histone H2AX (gammaH2AX) by PI-3-like kinases including ATM; gammaH2AX can serve as a marker of DNA damage when measured in situ using immunocytochemistry and flow cytometry. The aim of the present study was to investigate the relationship between CPFX-mediated DNA damage and induction of apoptosis in human lymphoblastoid cells and phytohaemagglutinin (PHA)-stimulated lymphocytes (Lymphs). Treatment of TK6 cells (wild-type p53) with 100 microg/ml CPFX for 2-10 h produced no increase in gammaH2AX; to the contrary, its level in S phase cells was reduced at 10 h compared to controls. Nevertheless, stabilization of topo IIalpha, ATM Ser-1981 phosphorylation and G(2) arrest was observed in TK6 cells exposed to CPFX for > or = 4 h. However, following 24 h treatment, gammaH2AX was dramatically increased in a sub-population of cells indicating the onset of apoptosis (confirmed by presence of activated caspase 3). CPFX had a similar lack of effect on induction of gammaH2AX at early time points in WTK1 and NH32 cells (devoid of functional p53) and proliferating Lymphs, however, induction of apoptosis was less pronounced than in TK6 cells. Formation of SCC and activation of ATM (but lack of gammaH2AX induction) indicates topo II-mediated chromatin or DNA changes in the absence of DSBs; ATM activation apparently triggers the G(2)M checkpoint leading to G(2) arrest. The subsequent induction of apoptosis appears to be facilitated by functional p53. CPFX may therefore have a potential use as a chemotherapeutic agent in the treatment of lymphoblast-derived cancer.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Apoptosis / drug effects*
  • Ataxia Telangiectasia Mutated Proteins
  • Caspase 3 / metabolism
  • Cell Cycle Proteins / metabolism
  • Cells, Cultured
  • Ciprofloxacin / pharmacology*
  • DNA Breaks, Double-Stranded / drug effects*
  • DNA-Binding Proteins / metabolism
  • G2 Phase / drug effects*
  • Histones / biosynthesis
  • Humans
  • Lymphocytes / cytology
  • Lymphocytes / drug effects*
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / metabolism
  • Tumor Suppressor Proteins / metabolism

Substances

  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • H2AX protein, human
  • Histones
  • Tumor Suppressor Proteins
  • Ciprofloxacin
  • ATM protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • Protein-Serine-Threonine Kinases
  • Caspase 3