Withaferin A Strongly Elicits IκB Kinase β Hyperphosphorylation Concomitant with Potent Inhibition of Its Kinase Activity

Abstract

The transcription factor NFB plays a critical role in normal and pathophysiological immune responses. Therefore, NFB and the signaling pathways that regulate its activation have become a major focus of drug development programs. Withania somnifera (WS) is a medicinal plant that is widely used in Palestine for the treatment of various inflammatory disorders. In this study we show that the leave extract of WS, as well as its major constituent withaferin A (WA), potently inhibits NFB activation by preventing the tumor necrosis factor-induced activation of IB kinase ␤ via a thioalkylation-sensitive redox mechanism, whereas other WS-derived steroidal lactones, such as withanolide A and 12-deoxywithastramonolide, are far less effective. To our knowledge, this is the first communication of IB kinase ␤ inhibition by a plant-derived inhibitor, coinciding with MEK1/ ERK-dependent Ser-181 hyperphosphorylation. This prevents IB phosphorylation and degradation, which subsequently blocks NFB translocation, NFB/DNA binding, and gene transcription. Taken together, our results indicate that pure WA or WA-enriched WS extracts can be considered as a novel class of NFB inhibitors, which hold promise as novel anti-inflammatory agents for treatment of various inflammatory disorders and/or cancer.

Anti-cancer activity of withaferin A in B-cell lymphoma

“…al. suggested that WA inhibits NF-kB by inhibiting the IKKb kinase activity to phosphorylate IkB, the inhibitor of NF-kB, and thus preventing the translocation of the p65 subunit to the nucleus. 22 This idea was further expanded by Heyninck et. al., suggesting WA targets the Cys179 residue of IKKb and inhibits the IKK complex directly.…”

“…19 In most cancer cell lines WA inhibits the proliferation of tumor cells through a G 2 /M phase cell cycle arrest, 20 and inhibits the activation of NF-kB via interaction with the IKKg subunit preventing IkB phosphorylation. 21,22 In pancreatic cancer cells, Yu et. al.…”

Withaferin A sensitizes TRAIL-induced apoptosis through reactive oxygen species-mediated up-regulation of death receptor 5 and down-regulation of c-FLIP

“…It is generally recognized that c-FLIP protein levels can be transcriptionally regulated through the nuclear factor-κB or c-fos pathway [9,39,40] or by ubiquitin-and caspase-mediated degradation [37,[41][42][43]. Recent reports showed that Wit A has a strong ability not only to block completely the binding of the transcription factor to DNA but also to induce the cleavage of NF-κB [4,10]. In our system, c-FLIP L and c-FLIP S mRNA expression was down-regulated by Wit A treatment through inhibition of the NF-κB signaling pathways.…”

“…More recently, Wit A induced early reactive oxygen species (ROS) generation that caused the failure of nuclear factor-κB (NF-κB) binding to DNA as well as nuclear cleavage by activated caspase-3, leading to apoptosis [8]. Wit A was also shown to suppress NF-κB activation and its regulated genes' expression, including COX-2, intercellular adhesion molecule, inhibitor of apoptosis protein 1, and cellular FADD-like IL-1β-converting enzyme inhibitory protein (c-FLIP), in cancer cells [9,10], although its underlying molecular mechanisms have not yet been fully elucidated.…”

Nf-κb-Dependent Inhibition of HIV-1 Transcription by Withaferin A

“…The roots and leaves of W. somnifera are commonly prescribed for tumors, inflammation, arthritis, asthma, and hypertension [9][10][11][12][13][14][15]. W. somnifera is the source of the largest and structurally most diversified set of withanolides, of which withaferin A (WA) is among the first of the group to be isolated and to show significant pharmacological properties, such as anti-tumor, anti-inflammatory, and immunomodulatory activity [9].…”