Inhibition of inflammatory cytokine production and lymphocyte proliferation by structurally different sesquiterpene lactones correlates with their effect on activation of NF-kappaB.

Many sesquiterpene lactones (Sls) are known to possess anti-inflammatory activities. To gain further insight into their structure-activity relationships and the molecular mechanism of action, four germacranolide sesquiterpene lactones which differ in the skeleton and the number of reactive centers (4beta,15-epoxy-miller-9E-enolide (1), 15-acetoxy-eremantholide B (2), a mixture of 15-(isovaleroyl)/15-(2-methyl-butyryl)-2alpha-acetoxy-miguanin (3), and 15-(2-hydroxy)-isobutyryloxy-micrantholide (4)) were investigated for their effect on production of proinflammatory cytokines (interleukin-1beta [IL-1beta], IL-6, and tumor necrosis factor-alpha [TNF-alpha]) as well as proliferation of concanavalin A (Con A) and lipopolysaccharide (LPS)-stimulated mouse lymphocytes. Compounds 1 and 3 which possess an alpha-methylene-gamma-lactone function and a conjugated carbonyl group induced a half-maximal inhibition of cytokine synthesis in adherent mouse peritoneal exudate cells at micromolar concentrations (IC(50) 0.69-1.70 microM), while compound 4 which contains only an alpha-methylene-gamma-lactone residue was less active (IC(50) > or 38 microM). Interestingly, compound 2, which carries only a conjugated keto group, displayed a potency similar to those of the bifunctional compounds 1 and 3. All four Sls suppressed proliferation of murine lymphocyte at IC(50) concentrations between 0.22 and 5.03 microM. The rank order of potency was 1 = 2 > 3 > 4. Generally, the growth of LPS-stimulated cells was more strongly influenced than those of Con A-activated lymphocytes. This effect was particularly pronounced with 4. Inhibitory concentrations correlated well with those necessary for inhibition of the transcription factor nuclear factor kappaB (NF-kappaB) observed in a previous investigation. Therefore, it can be assumed that NF-kappaB may be involved in the suppressive effect of Sls on cytokine production and lymphocyte proliferation.

Inhibition of transcription factor NF-kappaB by sesquiterpene lactones: a proposed molecular mechanism of action.

Many sesquiterpene lactones (SLs) possess considerable anti-inflammatory activity. They inhibit the transcription factor NF-kappaB by selectively alkylating its p65 subunit probably by reacting with cysteine residues. Here we assayed 28 sesquiterpene lactones for their ability to inhibit NF-kappaB. The majority of the potent NF-kappaB inhibitors possess two reactive centers in form of an alpha-methylene-gamma-lactone group and an alpha,beta- or alpha,beta,gamma,delta-unsaturated carbonyl group. Based on computer molecular modelling we propose a molecular mechanism of action, which is able to explain the p65 selectivity of the SLs and the observed correlation of high activity with alkylant bifunctionality. A single bifunctional SL molecule can alkylate the cysteine residue (Cys 38) in the DNA binding loop 1 (L1) and a further cysteine (Cys 120) in the nearby E' region. This cross link alters the position of tyrosine 36 and additional amino acids in such a way that their specific interactions with the DNA become impossible. We also created a model for monofunctional SLs.

Study of three sesquiterpene lactones from Tithonia diversifolia on their anti-inflammatory activity using the transcription factor NF-kappa B and enzymes of the arachidonic acid pathway as targets.

In Central America leaf extracts from the Asteraceae Tithonia diversifolia are used externally for the treatment of haematomas and wounds. Therefore, the main sesquiterpene lactones (Sls) of this species growing in Costa Rica, diversifolin (1), diversifolin methyl ether (2), and tirotundin (3), were studied for their anti-inflammatory activity. We determined whether these compounds inhibit cyclooxygenase-I, phospholipase A2, or the transcription factor NF-kappa B. Here we show that these Sls do not influence the enzymes of the arachidonic acid pathway, but inhibit the activation of NF-kappa B. Thereby, the synthesis of inflammatory mediators such as cytokines and chemokines is reduced. Our results indicate that the inhibitory activity of compounds 1-3 is due to alkylation of cysteine residues, which are probably located in the DNA binding domain of NF-kappa B. The Sls were also studied for their antibacterial activity, but only Sl 1 was moderately active against Bacillus subtilis in the agar plate diffusion test.