Combinatorial treatment of DNA and chromatin-modifying drugs cause cell death in human and canine osteosarcoma cell lines

PLoS One. 2012;7(9):e43720. doi: 10.1371/journal.pone.0043720. Epub 2012 Sep 5.

Abstract

Downregulation of microRNAs (miRNAs) at the 14q32 locus stabilizes the expression of cMYC, thus significantly contributing to osteosarcoma (OS) pathobiology. Here, we show that downregulation of 14q32 miRNAs is epigenetically regulated. The predicted promoter regions of miRNA clusters at 14q32 locus showed no recurrent patterns of differential methylation, but Saos2 cells showed elevated histone deacetylase (HDAC) activity. Treatment with 4-phenylbutyrate increased acetylation of histones associated with 14q32 miRNAs, but interestingly, robust restoration of 14q32 miRNA expression, attenuation of cMYC expression, and induction of apoptosis required concomitant treatment with 5-Azacytidine, an inhibitor of DNA methylation. These events were associated with genome-wide gene expression changes including induction of pro-apoptotic genes and downregulation of cell cycle genes. Comparable effects were achieved in human and canine OS cells using the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA/Vorinostat) and the DNA methylation inhibitor Zebularine (Zeb), with significantly more pronounced cytotoxicity in cells whose molecular phenotypes were indicative of aggressive biological behavior. These results suggested that the combination of these chromatin-modifying drugs may be a useful adjuvant in the treatment of rapidly progressive OS.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Chloromethyl Ketones / pharmacology
  • Animals
  • Antineoplastic Combined Chemotherapy Protocols / therapeutic use*
  • Apoptosis
  • Cell Line, Tumor
  • Chromatin / metabolism*
  • Cytidine / analogs & derivatives
  • Cytidine / pharmacology
  • DNA / metabolism*
  • DNA Methylation
  • Dogs
  • Drug Screening Assays, Antitumor
  • Epigenesis, Genetic
  • Gene Expression Profiling
  • Gene Expression Regulation, Neoplastic
  • Histone Deacetylase Inhibitors / pharmacology
  • Humans
  • Hydroxamic Acids / pharmacology
  • Osteosarcoma / drug therapy*
  • Vorinostat

Substances

  • Amino Acid Chloromethyl Ketones
  • Chromatin
  • Histone Deacetylase Inhibitors
  • Hydroxamic Acids
  • benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone
  • Vorinostat
  • Cytidine
  • pyrimidin-2-one beta-ribofuranoside
  • DNA

Grant support

This study was supported by a grant from the University of Minnesota Academic Health Center, by a translational research grant from the Masonic Cancer Center, University of Minnesota, and by grants from the Wyckoff Rein in Sarcoma Foundation, the AKC Canine Health Foundation, the Van Sloun Foundation, and the Kate Koogler Canine Cancer Fund (JFM and SS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.