Intrinsic mutagenic properties of 5-chlorocytosine: A mechanistic connection between chronic inflammation and cancer

Proc Natl Acad Sci U S A. 2015 Aug 18;112(33):E4571-80. doi: 10.1073/pnas.1507709112. Epub 2015 Aug 4.


During chronic inflammation, neutrophil-secreted hypochlorous acid can damage nearby cells inducing the genomic accumulation of 5-chlorocytosine (5ClC), a known inflammation biomarker. Although 5ClC has been shown to promote epigenetic changes, it has been unknown heretofore if 5ClC directly perpetrates a mutagenic outcome within the cell. The present work shows that 5ClC is intrinsically mutagenic, both in vitro and, at a level of a single molecule per cell, in vivo. Using biochemical and genetic approaches, we have quantified the mutagenic and toxic properties of 5ClC, showing that this lesion caused C→T transitions at frequencies ranging from 3-9% depending on the polymerase traversing the lesion. X-ray crystallographic studies provided a molecular basis for the mutagenicity of 5ClC; a snapshot of human polymerase β replicating across a primed 5ClC-containing template uncovered 5ClC engaged in a nascent base pair with an incoming dATP analog. Accommodation of the chlorine substituent in the template major groove enabled a unique interaction between 5ClC and the incoming dATP, which would facilitate mutagenic lesion bypass. The type of mutation induced by 5ClC, the C→T transition, has been previously shown to occur in substantial amounts both in tissues under inflammatory stress and in the genomes of many inflammation-associated cancers. In fact, many sequence-specific mutational signatures uncovered in sequenced cancer genomes feature C→T mutations. Therefore, the mutagenic ability of 5ClC documented in the present study may constitute a direct functional link between chronic inflammation and the genetic changes that enable and promote malignant transformation.

Keywords: 5-chloro-deoxycytidine; carcinogenesis; hypochlorite; inflammatory bowel disease; myeloperoxidase.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural

MeSH terms

  • Biomarkers, Tumor / metabolism
  • Carcinogenesis
  • Chromatography, High Pressure Liquid
  • Cytosine / analogs & derivatives*
  • Cytosine / chemistry
  • DNA Mutational Analysis
  • Humans
  • Hypochlorous Acid / chemistry
  • Inflammation / metabolism
  • Inflammatory Bowel Diseases / metabolism
  • Models, Molecular
  • Mutagenesis*
  • Mutagens*
  • Mutation
  • Neoplasms / metabolism*
  • Oligonucleotides / chemistry
  • Oligonucleotides / genetics
  • Peroxidase / metabolism
  • Sequence Analysis, DNA


  • Biomarkers, Tumor
  • Mutagens
  • Oligonucleotides
  • Hypochlorous Acid
  • Cytosine
  • Peroxidase
  • 5-chlorocytosine

Associated data

  • PDB/5BOL
  • PDB/5BOM
  • PDB/5BPC