Abstract

The nuclear factor κB (NF-κB) is a promising target for drug discovery. NF-κB is a heterodimeric complex of RelA and p50 subunits that interact with the DNA, regulating the expression of several genes; its dysregulation can trigger diverse diseases including inflammation, immunodeficiency, and cancer. There is some experimental evidence, based on whole cells studies, that natural sesquiterpene lactones (Sls) can inhibit the interaction of NF-κB with DNA, by alkylating the RelA subunit via a Michael addition. In the present work, 28 natural and semisynthetic pseudoguianolides were screened as potential inhibitors of NF-κB in a biochemical assay that was designed using pure NF-κB heterodimer, pseudoguianolides and a ~1000 bp palindromic DNA fragment harboring two NF-κB recognition sequences. By comparing the relative amount of free DNA fragment to the NF-κB – DNA complex, in a routine agarose gel electrophoresis, the destabilizing effect of a compound on the complex is estimated. The results of the assay and the following structure-activity relationship study, allowed the identification of several relevant structural features in the pseudoguaianolide skeleton, which are necessary to enhance the dissociating capacity of NF-κB–DNA complex. The most active compounds are substituted at C-3 (α-carbonyl), in addition to having the α-methylene-γ-lactone moiety which is essential for the alkylation of RelA.

Highlights

  • The nuclear factor kB (NF-kB) is considered to be a promising target for drug discovery

  • We developed a biochemical assay to study the effects of sesquiterpene lactones (SLs) on the interaction between a designed DNA-recognition target, and the canonical NF-kB heterodimer (RelA/p50) (Fig. 1)

  • The results show that DMSO had a strongly negative effect on the stability of the NF-kB:DNA complex and complete dissociation was observed at 30% DMSO concentration (Fig. 5b)

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Summary

Introduction

The nuclear factor kB (NF-kB) is considered to be a promising target for drug discovery. It regulates the transcription of pro-inflammatory and anti-apoptotic proteins, among others; dysregulation can lead to the development of chronic inflammation, immunodeficiency and cancer. Two major pharmaceutical approaches have received most attention: a) inhibition of the proteolytic activity of proteasome 26S, and b) inhibition of kB protein kinase (IKK). Both enzyme activities are indispensable for the degradation and phosphorylation of inhibitor-kB protein (IkB), and for the activation of NF-kB [10,11,12]

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