Mode of action study of the anti-inflammatory sesquiterpene lactone parthenolide

Biologically active natural products continue to be useful in the exploration and control of intracellular signaling processes. For example, the sesquiterpene lactone parthenolide from the anti-inflammatory medicinal herb Feverfew ( Tanacetum parthenium) has been shown to inhibit the pro-inflammatory signaling pathway. However, its mode of action has not been elucidated and no direct target has been identified. In this study, I used a biotinylated derivative of parthenolide as an affinity reagent to identify its molecular targets. I demonstrated that parthenolide directly binds to and inhibits IκB kinase β (IKKβ), the catalytic subunit of the IKK complex known to play a critical role in cytokine-mediated signaling. I found that mutating cysteine 179 in the activation loop of IKKβ renders it insensitive to parthenolide. Consistent with this finding, I showed that parthenolide prevents the degradation of unphosphorylated IκBα, in contrast to the accumulation of phosphorylated and polyubiquitinated IκBα caused by the treatment with proteasome inhibitors. Furthermore, I generated reduced parthenolide and showed that parthenolide's in vitro and in vivo anti-inflammatory activity is mediated through the unsaturated γ-lactone moiety shared by other sesquiterpene lactones. These results provide a possible molecular basis for the anti-inflammatory properties of sesquiterpene lactone-containing herbal medicines. In addition, these results may aid in the development of a new class of anti-inflammatory agents.; In the process of finding candidate parthenolide binding proteins, I identified a putative mouse phosphatase, which we termed p34. Sequence analysis reveals that it shares high homology to two yeast proteins, PHO2 and PHO13, which have been designated para-nitrophenylphosphatases. Characterization experiments performed by Dr. Ndubuisi confirm P34 as a putative alkaline phosphatase. In addition, its activity appears to be independent of TNF-α induction and is not affected by parthenolide treatment. Although it does seem to play a role in pro-inflammatory cytokine signaling, the characterization of p34 serves as a starting point for further investigation.