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Assessment of DNA Double Strand Break Repair Activity Using High-throughput and Quantitative Luminescence-Based Reporter Assays
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In vitro evidence for homologous recombinational repair in resistance to melphalan

Z M Wang1, Z P Chen, Z Y Xu

  • 1Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montreal, PQ, Canada.

Journal of the National Cancer Institute
|October 5, 2001
PubMed
Abstract

Insights

Melphalan resistance in epithelial tumors is linked to homologous recombinational repair. Higher Xrcc3 protein levels and more Rad51 foci indicate increased melphalan resistance, suggesting this repair pathway is key.

Area of Science:

  • Molecular Biology
  • Cancer Research
  • DNA Repair

Background:

  • Nitrogen mustards like melphalan generate DNA interstrand cross-links, crucial for their antitumor activity.
  • Cell lines with faulty recombinational repair are sensitive to nitrogen mustards.
  • Melphalan resistance may involve accelerated DNA repair via homologous recombination (Rad51-related proteins) or nonhomologous endjoining (DNA-PK, Ku proteins).

Purpose of the Study:

  • To investigate the role of DNA repair mechanisms in melphalan resistance.
  • To determine if homologous recombination or nonhomologous endjoining pathways contribute to resistance in epithelial tumor cell lines.

Main Methods:

  • Assessed melphalan cytotoxicity using the sulforhodamine assay in 14 epithelial tumor cell lines.
  • Investigated homologous recombinational repair by measuring Rad51, Rad52, and Xrcc3 protein levels and melphalan-induced Rad51 foci.
  • Examined nonhomologous endjoining by quantifying Ku70, Ku86 protein levels, and DNA-PK activity.
  • Used linear regression to correlate protein levels, DNA-PK activity, and Rad51 foci with melphalan cytotoxicity.

Main Results:

  • Melphalan resistance showed a significant correlation with Xrcc3 protein levels (r =.587; P =.027).
  • A strong correlation was observed between melphalan resistance and the density of melphalan-induced Rad51 foci (r =.848; P =.008).
  • No significant correlation was found between melphalan resistance and Rad51, Rad52, Ku protein levels, or DNA-PK activity.

Conclusions:

  • The findings suggest that homologous recombinational repair plays a role in melphalan resistance.
  • Elevated Xrcc3 protein levels and increased Rad51 foci density are indicators of melphalan resistance in epithelial tumor cells.
  • This highlights the importance of the homologous recombinational repair pathway in the cellular response to melphalan treatment.

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