Epigenetically regulated clonal heritability of CTA expression profiles in human melanoma

J Cell Physiol. 2010 May;223(2):352-8. doi: 10.1002/jcp.22040.


The intratumoral heterogeneity of cancer testis antigens (CTA) expression, which is driven by promoter methylation status, may hamper the effectiveness of CTA-directed vaccination of melanoma patients. Thus, we investigated whether the intratumoral heterogeneity of CTA expression is inherited at cellular level, or evolves throughout cellular replication, leading to a phenotypically unstable tumor cell population with reduced immunogenicity and/or able to escape immune control. Utilizing a previously characterized ex vivo clonal model of intratumoral heterogeneity of CTA expression in melanoma, Mel 313 MAGE-A3-low clone 5 (clone 5(M3-low)) and MAGE-A3-high clone 14 (clone 14(M3-high)) were sub-cloned and analyzed for CTA profile. Molecular assays demonstrated that levels of MAGE-A3 expression were highly conserved among generated sub-clones, as compared to parental clones. A similar behavior was identified for an extensive panel of other CTA investigated. Inherited levels of MAGE-A3 expression correlated with the extent of promoter methylation among clone 5(M3-low) and clone 14(M3-high) sub-clones analyzed. Treatment of clone 5(M3-low) with a DNA hypomethylating agent (DHA) resulted in an up-regulated expression of MAGE-A3, which was inherited at single cell level, being still detectable at day 60 in its sub-clones. Bisulfite sequencing demonstrated that also MAGE-A3 promoter methylation status was inherited among sub-clones generated from DHA-treated clone 5(M3-low) and strictly correlated with MAGE-A3 expression levels in investigated sub-clones. Similar results were obtained for additional CTA studied. Altogether our findings demonstrate that constitutive and DHA-modified CTA profiles of melanoma cells are clonally inherited throughout cellular replications, thus providing relevant insights to improve the effectiveness of CTA-based immunotherapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antigens, Neoplasm / genetics*
  • Antimetabolites, Antineoplastic / pharmacology
  • Azacitidine / analogs & derivatives
  • Azacitidine / pharmacology
  • Cell Division / genetics
  • Clone Cells / drug effects
  • Clone Cells / metabolism*
  • Cloning, Molecular / methods
  • DNA Methylation / drug effects
  • Decitabine
  • Epigenesis, Genetic / drug effects
  • Epigenesis, Genetic / genetics*
  • Gene Expression Regulation, Neoplastic / drug effects
  • Gene Expression Regulation, Neoplastic / genetics*
  • Humans
  • Inheritance Patterns / genetics*
  • Melanoma / genetics*
  • Melanoma / metabolism*
  • Neoplasm Proteins / genetics
  • Promoter Regions, Genetic / genetics
  • Tumor Cells, Cultured


  • Antigens, Neoplasm
  • Antimetabolites, Antineoplastic
  • MAGEA3 protein, human
  • Neoplasm Proteins
  • Decitabine
  • Azacitidine