Both hMutSα and hMutSß DNA mismatch repair complexes participate in 5-fluorouracil cytotoxicity

PLoS One. 2011;6(12):e28117. doi: 10.1371/journal.pone.0028117. Epub 2011 Dec 2.

Abstract

Background: Patients with advanced microsatellite unstable colorectal cancers do not show a survival benefit from 5-fluorouracil (5-FU)-based chemotherapy. We and others have shown that the DNA mismatch repair (MMR) complex hMutSα binds 5-FU incorporated into DNA. Although hMutSß is known to interact with interstrand crosslinks (ICLs) induced by drugs such as cisplatin and psoralen, it has not been demonstrated to interact with 5-FU incorporated into DNA. Our aim was to examine if hMutSß plays a role in 5-FU recognition.

Methods: We compared the normalized growth of 5-FU treated cells containing either or both mismatch repair complexes using MTT and clonogenic assays. We utilized oligonucleotides containing 5-FU and purified baculovirus-synthesized hMutSα and hMutSß in electromobility shift assays (EMSA) and further analyzed binding using surface plasmon resonance.

Results: MTT and clonogenic assays after 5-FU treatment demonstrated the most cytotoxicity in cells with both hMutSα and hMutSß, intermediate cytotoxicity in cells with hMutSα alone, and the least cytotoxicity in cells with hMutSß alone, hMutSß binds 5-FU-modified DNA, but its relative binding is less than the binding of 5-FU-modified DNA by hMutSα.

Conclusion: Cytotoxicity induced by 5-FU is dependent on intact DNA MMR, with relative cell death correlating directly with hMutSα and/or hMutSß 5-FU binding ability (hMutSα>hMutSß). The MMR complexes provide a hierarchical chemosensitivity for 5-FU cell death, and may have implications for treatment of patients with certain MMR-deficient tumors.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antimetabolites, Antineoplastic / pharmacology
  • Antineoplastic Agents / pharmacology*
  • Baculoviridae / genetics
  • Cell Line, Tumor
  • Colonic Neoplasms / drug therapy
  • Cross-Linking Reagents / pharmacology
  • DNA Mismatch Repair*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / physiology
  • Fluorouracil / pharmacology*
  • Humans
  • Microsatellite Repeats / genetics*
  • Oligonucleotides / chemistry
  • Recombinant Proteins / chemistry
  • Surface Plasmon Resonance
  • Tetrazolium Salts / pharmacology
  • Thiazoles / pharmacology
  • Treatment Outcome

Substances

  • Antimetabolites, Antineoplastic
  • Antineoplastic Agents
  • Cross-Linking Reagents
  • DNA-Binding Proteins
  • G-T mismatch-binding protein
  • Oligonucleotides
  • Recombinant Proteins
  • Tetrazolium Salts
  • Thiazoles
  • thiazolyl blue
  • Fluorouracil