Targeting NF-κB p65 with a helenalin inspired bis-electrophile

John C. Widen, Aaron M. Kempema, Peter W. Villalta, Daniel A. Harki

Research output: Contribution to journalArticlepeer-review

20 Scopus citations


The canonical NF-κB signaling pathway is a mediator of the cellular inflammatory response and a target for developing therapeutics for multiple human diseases. The furthest downstream proteins in the pathway, the p50/p65 transcription factor heterodimer, have been recalcitrant toward small molecule inhibition despite the substantial number of compounds known to inhibit upstream proteins in the activation pathway. Given the roles of many of these upstream proteins in multiple biochemical pathways, targeting the p50/p65 heterodimer offers an opportunity for enhanced on-target specificity. Toward this end, the p65 protein presents two nondisulfide cysteines, Cys38 and Cys120, at its DNA-binding interface that are amenable to targeting by covalent molecules. The natural product helenalin, a sesquiterpene lactone, has been previously shown to target Cys38 on p65 and ablate its DNA-binding ability. Using helenalin as inspiration, simplified helenalin analogues were designed, synthesized, and shown to inhibit induced canonical NF-κB signaling in cell culture. Moreover, two simplified helenalin probes were proficient at forming covalent protein adducts, binding to Cys38 on recombinant p65, and targeting p65 in HeLa cells without engaging canonical NF-κB signaling proteins IκBα, p50, and IKKα/β. These studies further support that targeting the p65 transcription factor-DNA interface with covalent small molecule inhibitors is a viable approach toward regulating canonical NF-κB signaling.

Original languageEnglish (US)
Pages (from-to)102-113
Number of pages12
JournalACS Chemical Biology
Issue number1
StatePublished - 2017

Bibliographical note

Funding Information:
The work was generously supported by the University of Minnesota (Academic Health Center Seed Grant #2010.01), The V Foundation for Cancer Research (V Scholar Award to D. A. Harki), Hyundai Hope on Wheels (Hope Grant), and the NIH (R21-CA194661). J. C. Widen acknowledges the University of Minnesota, College of Pharmacy for a Bighley Graduate Fellowship. Mass spectrometry was performed at the Analytical Biochemistry Core Facility of the Masonic Cancer Center, which is supported by the NIH (P30-CA77598 and S10 RR-024618). We gratefully acknowledge V. G. Young, Jr. (University of Minnesota, Department of Chemistry, X-ray Crystallographic Laboratory) for solving the structure of 11 and funding from the NSF/MRI (#1229400) and the University of Minnesota for the purchase of the Bruker-AXS D8 Venture diffractometer. The Minnesota Supercomputing Institute (MSI) at the University of Minnesota is acknowledged for molecular modeling resources.

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