A high through-put screen for small molecules modulating MCM2 phosphorylation identifies Ryuvidine as an inducer of the DNA damage response

  • PLoS One. 2014 Jun 5;9(6):e98891. doi: 10.1371/journal.pone.0098891.
Jennifer FitzGerald  1 ,  Laura S Murillo  2 ,  Gemma O'Brien  1 ,  Enda O'Connell  3 ,  Aisling O'Connor  1 ,  Kevin Wu  1 ,  Guan-Nan Wang  1 ,  Michael D Rainey  1 ,  Alessandro Natoni  2 ,  Sandra Healy  1 ,  Michael O'Dwyer  4 ,  Corrado Santocanale  1
Affiliations
  • 1. Centre for Chromosome Biology and National Centre for Biomedical Engineering Science, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
  • 2. Centre for Chromosome Biology and National Centre for Biomedical Engineering Science, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland; School of Medicine, National University of Ireland Galway, Galway, Ireland.
  • 3. Screening Core, National Centre for Biomedical Engineering Science, Biosciences, National University of Ireland Galway, Galway, Ireland.
  • 4. School of Medicine, National University of Ireland Galway, Galway, Ireland.
PMID: 24902048 DOI: 10.1371/journal.pone.0098891
Abstract

DNA replication is an essential process for cell division and as such it is a process that is directly targeted by several Anticancer drugs. CDC7 plays an essential role in the activation of replication origins and has recently been proposed as a novel target for drug discovery. The MCM DNA helicase complex (MCM2-7) is a key target of the CDC7 kinase, and MCM phosphorylation status at specific sites is a reliable biomarker of CDC7 cellular activity. In this work we describe a cell-based assay that utilizes the "In Cell Western Technique" (ICW) to identify compounds that affect cellular CDC7 activity. By screening a library of approved drugs and kinase inhibitors we found several compounds that can affect CDC7-dependent phosphorylation of MCM2 in HeLa cells. Among these, Mitoxantrone, a Topoisomerase Inhibitor, and Ryuvidine, previously described as a CDK4 Inhibitor, cause a reduction in phosphorylated MCM2 levels and a sudden blockade of DNA synthesis that is accompanied by an ATM-dependent checkpoint response. This study sheds light on the previously observed cytotoxity of Ryuvidine, strongly suggesting that it is related to its effect of causing DNA damage.

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