DNA damage signaling in response to 5-fluorouracil in three colorectal cancer cell lines with different mismatch repair and TP53 status

Int J Oncol. 2011 Sep;39(3):673-82. doi: 10.3892/ijo.2011.1080. Epub 2011 Jun 14.


We studied patterns of DNA damage signaling and cell cycle response to clinically-relevant (bolus) and high doses of 5-fluorouracil (5-FU) in three colorectal cancer cell lines with differing MMR and TP53 status in an attempt to better understand how 5-FU exerts its cytotoxicity. The ATM/CHEK2/ CHEK1 signaling pathway was not activated in response to bolus 5-FU in the MMR-deficient cell lines HCT116 (TP53-proficient or TP53-depleted) and HCT15 (TP53-deficient), consistent with negligible/reparable DNA damage and no cell death. The pattern of DNA damage checkpoint activation in bolus 5-FU-treated HT29 (TP53-deficient/MMR-proficient) cultures suggested SSB formation (CHEK1 activation) followed by DSB formation (CHEK2 activation and increased phospho-H2AX levels), but no cell death suggested that DNA repair capacity was not overwhelmed. High-dose 5-FU treatment led to activation of ATM/CHEK2/TP53 (not CHEK1) in TP53-proficient and TP53-depleted HCT116 (later CHEK2 activation relative to TP53-proficient) cultures; HCT15 cultures had ATM activation only. These data and increased phospho-H2AX levels indicated DSB formation; apoptosis was induced in both cell lines indicating irreparable DNA damage. TP53-depleted HCT116 cultures also had DSBs after high-dose 5-FU treatment but experienced a (transient) G1/S cell cycle arrest that protected them from apoptosis. TP53 phosphorylation at Ser20/33/37 was seen in TP53-proficient HCT116 cultures regardless of 5-FU concentration at ≥4 h following treatment, indicating TP53 stabilization/transcriptional activation. Overall, activation of ATM, CHEK1 and/or CHEK2 and phospho-H2AX levels reflected the nature of 5-FU-induced DNA damage and indi-cated when DNA damage was significant (5-FU-dose-dependent). DNA repair and cell cycle responses to 5-FU-induced DNA damage were distinctly affected by MMR and TP53 (role in BER/NER) functionalities, but MMR deficiency especially seemed to confer less overall sensitivity to 5-FU.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antimetabolites, Antineoplastic / pharmacology*
  • Cell Cycle / drug effects
  • Cell Growth Processes / drug effects
  • Cell Line, Tumor
  • Colorectal Neoplasms / drug therapy*
  • Colorectal Neoplasms / genetics*
  • Colorectal Neoplasms / metabolism
  • Colorectal Neoplasms / pathology
  • DNA Damage*
  • DNA Mismatch Repair*
  • Female
  • Fluorouracil / pharmacology*
  • Genes, p53
  • HCT116 Cells
  • HT29 Cells
  • Humans
  • Male
  • Signal Transduction / drug effects
  • Transfection


  • Antimetabolites, Antineoplastic
  • Fluorouracil