5-ASA Affects Cell Cycle Progression in Colorectal Cells by Reversibly Activating a Replication Checkpoint

Gastroenterology. 2007 Jan;132(1):221-35. doi: 10.1053/j.gastro.2006.10.016. Epub 2006 Oct 12.

Abstract

Background & aims: Individuals with inflammatory bowel disease are at risk of developing colorectal cancer (CRC). Epidemiologic, animal, and laboratory studies suggest that 5-amino-salicylic acid (5-ASA) protects from the development of CRC by altering cell cycle progression and by inducing apoptosis. Our previous results indicate that 5-ASA improves replication fidelity in colorectal cells, an effect that is active in reducing mutations. In this study, we hypothesized that 5-ASA restrains cell cycle progression by activating checkpoint pathways in colorectal cell lines, which would prevent tumor development and improve genomic stability.

Methods: CRC cells with different genetic backgrounds such as HT29, HCT116, HCT116(p53-/-), HCT116+chr3, and LoVo were treated with 5-ASA for 2-96 hours. Cell cycle progression, phosphorylation, and DNA binding of cell cycle checkpoint proteins were analyzed.

Results: We found that 5-ASA at concentrations between 10 and 40 mmol/L affects cell cycle progression by inducing cells to accumulate in the S phase. This effect was independent of the hMLH1, hMSH2, and p53 status because it was observed to a similar extent in all cell lines under investigation. Moreover, wash-out experiments demonstrated reversibility within 48 hours. Although p53 did not have a causative role, p53 Ser15 was strongly phosphorylated. Proteins involved in the ATM-and-Rad3-related kinase (ATR)-dependent S-phase checkpoint response (Chk1 and Rad17) were also phosphorylated but not ataxia telengectasia mutated kinase.

Conclusions: Our data demonstrate that 5-ASA causes cells to reversibly accumulate in S phase and activate an ATR-dependent checkpoint. The activation of replication checkpoint may slow down DNA replication and improve DNA replication fidelity, which increases the maintenance of genomic stability and counteracts carcinogenesis.

Publication types

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

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism
  • Anti-Inflammatory Agents, Non-Steroidal / pharmacology*
  • Apoptosis / drug effects
  • Ataxia Telangiectasia Mutated Proteins
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Cell Cycle / drug effects
  • Cell Cycle Proteins / metabolism
  • Cell Division / drug effects
  • Checkpoint Kinase 1
  • Colorectal Neoplasms / pathology
  • Colorectal Neoplasms / prevention & control*
  • DNA Mismatch Repair / drug effects
  • DNA Polymerase III / metabolism
  • DNA Replication / drug effects*
  • DNA Replication / physiology
  • DNA-Binding Proteins / metabolism
  • Flow Cytometry
  • HT29 Cells
  • Humans
  • Mesalamine / pharmacology*
  • Minichromosome Maintenance Complex Component 2
  • Minichromosome Maintenance Complex Component 7
  • Mitosis / drug effects
  • MutL Protein Homolog 1
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Protein Kinases / metabolism
  • Protein-Serine-Threonine Kinases / metabolism
  • S Phase / drug effects
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism

Substances

  • Adaptor Proteins, Signal Transducing
  • Anti-Inflammatory Agents, Non-Steroidal
  • CDC45 protein, human
  • CLSPN protein, human
  • Carrier Proteins
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • MLH1 protein, human
  • Nuclear Proteins
  • Rad17 protein, human
  • Tumor Suppressor Protein p53
  • Mesalamine
  • Protein Kinases
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • CHEK1 protein, human
  • Checkpoint Kinase 1
  • Protein-Serine-Threonine Kinases
  • DNA Polymerase III
  • MutL Protein Homolog 1
  • MCM2 protein, human
  • MCM7 protein, human
  • Minichromosome Maintenance Complex Component 2
  • Minichromosome Maintenance Complex Component 7