Cross-Resistance to Ionizing Radiation in a Murine Leukemic Cell Line Resistant to cis-Dichlorodiammineplatinum(II): Role of Ku Autoantigen

  1. Philippe Frit1,
  2. Yvan Canitrot1,
  3. Catherine Muller1,
  4. Nicolas Foray3,
  5. Patrick Calsou1,
  6. Elisabetta Marangoni2,
  7. Jean Bourhis2 and
  8. Bernard Salles1
  1. 1Institut de Pharmacologie et de Biologie Structurale, Centre National de la Recherche Scientifique, Unité Propre de Recherche 9062, Toulouse, France (P.F., Y.C., C.M., P.C., B.S.);2Unité Propre de l’Enseignement Supérieur, Pr Eschwege, Laboratoire de Radiobiologie, Institut Gustave Roussy, Villejuif, France (E.M., J.B.); and 3Unité Mixte de Recherche 1599 Centre National de la Recherche Scientifique, Institut Gustave Roussy, Villejuif, France (N.F.)

    Abstract

    cis-Dichlorodiammineplatinum(II) (CDDP; cisplatin) is commonly used in combination with ionizing radiation (IR) in the treatment of various malignancies. In vitro, many observations suggest that acquisition of CDDP resistance in cell lines confers cross-resistance to IR, but the molecular mechanisms involved have not been well documented yet. We report here the selection and characterization of a murine CDDP-resistant L1210 cell line (L1210/3R) that exhibits cross-resistance to IR because of an increased capacity to repair double-strand breaks compared with parental cells (L1210/P). In resistant cells, electrophoretic mobility shift assays revealed an increased DNA-end binding activity that could be ascribed, by supershifting the retardation complexes with antibodies, to the autoantigen Ku. The heterodimeric Ku protein, composed of 86-kDa (Ku80) and 70-kDa (Ku70) subunits, is the DNA-targeting component of DNA-dependent protein kinase (DNA-PK), which plays a critical role in mammalian DNA double-strand breaks repair. The increased Ku-binding activity in resistant cells was associated with an overexpression affecting specifically the Ku80 subunit. These data strongly suggest that the increase in Ku activity is responsible for the phenotype of cross-resistance to IR. In addition, these observations, along with previous results from DNA-PK− mutant cells, provide evidence in favor of a role of Ku/DNA-PK in resistance to CDDP. These results suggest that Ku activity may be an important molecular target in cancer therapy at the crossroad between cellular responses to CDDP and IR.

    Footnotes

    • Send reprint requests to: Dr. Bernard Salles, Institut de Pharmacologie et de Biologie Structurale, Centre National de la Recherche Scientifique, Unité Propre de Recherche 9062, 205 route de Narbonne, 31077 Toulouse Cedex, France. E-mail:salles{at}ipbs.fr

    • This work was supported by the Association pour la Recherche contre le Cancer and the Ligue Nationale contre le Cancer.

    • Abbreviations:
      CDDP
      cis-dichlorodiammineplatinum(II) (cisplatin)
      DNA-PK
      DNA-dependent protein kinase
      DNA-PKcs
      DNA-dependent protein kinase catalytic subunit
      IR
      ionizing radiation
      DSB
      double-strand break
      NM
      nitrogen mustard
      NER
      nucleotide excision repair
      ICL
      interstrand cross links or linking
      DEB
      DNA-end binding
      CHO
      Chinese hamster ovary
      TBST
      Tris-buffered saline/Tween 20
      FAR
      fraction of activity released
      EMSA
      electrophoretic mobility shift assay
      • Received October 9, 1998.
      • Accepted March 25, 1999.
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    1. Molecular Pharmacology July 1, 1999 vol. 56 no. 1 141-146
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