Ethyl pyruvate inhibits nuclear factor-kappaB-dependent signaling by directly targeting p65

J Pharmacol Exp Ther. 2005 Mar;312(3):1097-105. doi: 10.1124/jpet.104.079707. Epub 2004 Nov 3.


Ethyl pyruvate has been shown to have anti-inflammatory properties in numerous cell culture and animal studies. In this series of experiments, we tested the hypothesis that ethyl pyruvate inhibits signaling by the pro-inflammatory transcription factor, NF-kappaB. Ethyl pyruvate inhibited luciferase expression in lipopolysaccharide-stimulated murine macrophage-like RAW 264.7 cells transfected with an NF-kappaB-dependent luciferase reporter vector. Ethyl pyruvate also decreased NF-kappaB DNA-binding activity in lipopolysaccharide-stimulated RAW 264.7 cells and decreased lipopolysaccharide-induced expression of an NF-kappaB-dependent gene, inducible nitric oxide synthase. Ethyl pyruvate had no effect on the degradation of IkappaBalpha or IkappaBbeta in lipopolysaccharide-stimulated RAW 264.7 cells, suggesting that ethyl pyruvate acts distally to this step in the activation of NF-kappaB. In a cell-free system, binding of p50 homodimers to an NF-kappaB consensus oligonucleotide sequence was unaffected by ethyl pyruvate over a wide range of concentrations, indicating that ethyl pyruvate probably does not modify or interact with the p50 subunit of NF-kappaB. In contrast, ethyl pyruvate inhibited DNA binding by ectopically overexpressed wild-type p65 homodimers. However, ethyl pyruvate failed to inhibit the DNA-binding activity of homodimers of an overexpressed mutant form of a p65 with substitution of serine for cysteine 38. Taken together, these results suggest that ethyl pyruvate inhibits DNA-binding by covalently modifying p65 at Cys(38). We conclude that some of the beneficial anti-inflammatory effects of ethyl pyruvate may be due to modification of p65, thereby inhibiting signaling via the NF-kappaB pathway.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Line
  • DNA / metabolism
  • Dimerization
  • I-kappa B Proteins / metabolism
  • Lipopolysaccharides / pharmacology
  • Mice
  • NF-KappaB Inhibitor alpha
  • NF-kappa B / antagonists & inhibitors*
  • NF-kappa B / metabolism
  • Nitric Oxide Synthase / genetics
  • Nitric Oxide Synthase Type II
  • Pyruvates / pharmacology*
  • Signal Transduction / drug effects*
  • Transcription Factor RelA
  • Transcription, Genetic / drug effects


  • I kappa B beta protein
  • I-kappa B Proteins
  • Lipopolysaccharides
  • NF-kappa B
  • Nfkbia protein, mouse
  • Pyruvates
  • Transcription Factor RelA
  • ethyl pyruvate
  • NF-KappaB Inhibitor alpha
  • DNA
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type II
  • Nos2 protein, mouse