Dataset Information

Development of novel NEMO-binding domain mimetics for inhibiting IKK/NF-?B activation.

ABSTRACT: Nuclear factor ?B (NF-?B) is a transcription factor important for regulating innate and adaptive immunity, cellular proliferation, apoptosis, and senescence. Dysregulation of NF-?B and its upstream regulator I?B kinase (IKK) contributes to the pathogenesis of multiple inflammatory and degenerative diseases as well as cancer. An 11-amino acid peptide containing the NF-?B essential modulator (NEMO)-binding domain (NBD) derived from the C-terminus of ? subunit of IKK, functions as a highly selective inhibitor of the IKK complex by disrupting the association of IKK? and the IKK? subunit NEMO. A structure-based pharmacophore model was developed to identify NBD mimetics by in silico screening. Two optimized lead NBD mimetics, SR12343 and SR12460, inhibited tumor necrosis factor ? (TNF-?)- and lipopolysaccharide (LPS)-induced NF-?B activation by blocking the interaction between IKK? and NEMO and suppressed LPS-induced acute pulmonary inflammation in mice. Chronic treatment of a mouse model of Duchenne muscular dystrophy (DMD) with SR12343 and SR12460 attenuated inflammatory infiltration, necrosis and muscle degeneration, demonstrating that these small-molecule NBD mimetics are potential therapeutics for inflammatory and degenerative diseases.

SUBMITTER: Zhao J

PROVIDER: S-EPMC6013238 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Development of novel NEMO-binding domain mimetics for inhibiting IKK/NF-κB activation.

Zhao Jing J Zhang Lei L Mu Xiaodong X Doebelin Christelle C Nguyen William W Wallace Callen C Reay Daniel P DP McGowan Sara J SJ Corbo Lana L Clemens Paula R PR Wilson Gabriela Mustata GM Watkins Simon C SC Solt Laura A LA Cameron Michael D MD Huard Johnny J Niedernhofer Laura J LJ Kamenecka Theodore M TM Robbins Paul D PD

PLoS biology 20180611 6

Nuclear factor κB (NF-κB) is a transcription factor important for regulating innate and adaptive immunity, cellular proliferation, apoptosis, and senescence. Dysregulation of NF-κB and its upstream regulator IκB kinase (IKK) contributes to the pathogenesis of multiple inflammatory and degenerative diseases as well as cancer. An 11-amino acid peptide containing the NF-κB essential modulator (NEMO)-binding domain (NBD) derived from the C-terminus of β subunit of IKK, functions as a highly selectiv ...[more]

PMID: 29889904

Similar Datasets

Project description:Background contextNonphysiological mechanical loading and inflammation are both critically involved in intervertebral disc (IVD) degeneration, which is characterized by an increase in cytokines and matrix metalloproteases (MMPs) in the nucleus pulposus (NP). This process is known to be mediated by the NF-κB pathway.Clinical significanceCurrent clinical treatments for IVD degeneration focus on the alleviation of symptoms rather than targeting the underlying mechanism. Injection of an NF-κB inhibitor may attenuate the progression of IVD degeneration.PurposeTo investigate the ability of the NF-κB inhibitor, NEMO binding domain peptide (NBD), to alter IVD degeneration processes by reducing IL-1β- and mechanically-induced cytokine and MMP levels in human nucleus pulposus cells in vitro, and by attenuating IVD degeneration in an in vivo rat model for disc degeneration.Study designExperimental in vitro and animal model.Patient sampleDiscarded specimens of lumbar disc from 21 patients, and 12 Sprague Dawley rats.Outcome measuresGene and protein expression, cell viability, µMRI and histology.MethodsIL-1β-prestimulated human nucleus pulposus cells embedded into fibrin constructs were loaded in the Flexcell FX-5000 compression system at 5 kPa and 1 Hz for 48 hours in the presence and absence of NBD. Unloaded hNPC/fibrin constructs served as controls. Cell viability in loaded and unloaded constructs was quantified, and gene and protein expression levels determined. For in vivo testing, a rat needle disc puncture model was employed. Experimental groups included injured discs with and without NBD injection and uninjured controls. Levels of disc degeneration were determined via µMRI, qPCR and histology. Funding sources include $48,874 NASS Young Investigator Research Grant and $119,174 NIH 5K01AR071512-02. There were no applicable financial relationships or conflicts of interest.ResultsMechanical compression of hNPC/fibrin constructs resulted in upregulation of MMP-3 and IL-8. Supplementation of media with 10 μM NBD during loading increased cell viability, and decreased MMP-3 gene and protein levels. IVD injury in rat resulted in an increase in MMP-3, IL-1β and IL-6 gene expression. Injections of 250 µg of NBD during disc injury resulted in decreased IL-6 gene expression. µMRI analysis demonstrated a reduction of disc hydration in response to disc needle injury, which was attenuated in NBD-treated IVDs. Histological evaluation showed NP and AF lesion in injured discs, which was attenuated by NBD injection.ConclusionsThe results of this study show NBD peptide's capacity to reduce IL-1β- and loading-induced MMP-3 levels in hNPC/fibrin constructs while increasing the cells' viability, and to attenuate IVD degeneration in rat, involving downregulation of IL-6. Therefore, NBD may be a potential therapeutic agent to treat IVD degeneration.

Project description:Nuclear Factor kappa B (NF-?B) is a transcription factor involved in the regulation of cell signaling responses and is a key regulator of cellular processes involved in the immune response, differentiation, cell proliferation, and apoptosis. The constitutive activation of NF-?B contributes to multiple cellular outcomes and pathophysiological conditions such as rheumatoid arthritis, asthma, inflammatory bowel disease, AIDS and cancer. Thus there lies a huge therapeutic potential beneath inhibition of NF-?B signalling pathway for reducing these chronic ailments. Withania somnifera, a reputed herb in ayurvedic medicine, comprises a large number of steroidal lactones known as withanolides which show plethora of pharmacological activities like anti- inflammatory, antitumor, antibacterial, antioxidant, anticonvulsive, and immunosuppressive. Though a few studies have been reported depicting the effect of WA (withaferin A) on suppression of NF-?B activation, the mechanism behind this is still eluding the researchers. The study conducted here is an attempt to explore NF-?B signalling pathway modulating capability of Withania somnifera's major constituent WA and to elucidate its possible mode of action using molecular docking and molecular dynamics simulations studies.Formation of active IKK (I?B kinase) complex comprising NEMO (NF-?B Essential Modulator) and IKK? subunits is one of the essential steps for NF-?B signalling pathway, non-assembly of which can lead to prevention of the above mentioned vulnerable disorders. As observed from our semi-flexible docking analysis, WA forms strong intermolecular interactions with the NEMO chains thus building steric as well as thermodynamic barriers to the incoming IKK? subunits, which in turn pave way to naive complex formation capability of NEMO with IKK?. Docking of WA into active NEMO/IKK? complex using flexible docking in which key residues of the complex were kept flexible also suggest the disruption of the active complex. Thus the molecular docking analysis of WA into NEMO and active NEMO/IKK? complex conducted in this study provides significant evidence in support of the proposed mechanism of NF-?B activation suppression by inhibition or disruption of active NEMO/IKK? complex formation being accounted by non-assembly of the catalytically active NEMO/IKK? complex. Results from the molecular dynamics simulations in water show that the trajectories of the native protein and the protein complexed with WA are stable over a considerably long time period of 2.6 ns.NF-?B is one of the most attractive topics in current biological, biochemical, and pharmacological research, and in the recent years the number of studies focusing on its inhibition/regulation has increased manifolds. Small ligands (both natural and synthetic) are gaining particular attention in this context. Our computational analysis provided a rationalization of the ability of naturally occurring withaferin A to alter the NF-?B signalling pathway along with its proposed mode of inhibition of the pathway. The absence of active IKK multisubunit complex would prevent degradation of I?B proteins, as the I?B proteins would not get phosphorylated by IKK. This would ultimately lead to non-release of NF-?B and its further translocation to the nucleus thus arresting its nefarious acts. Conclusively our results strongly suggest that withaferin A is a potent anticancer agent as ascertained by its potent NF-?B modulating capability. Moreover the present MD simulations made clear the dynamic structural stability of NEMO/IKK? in complex with the drug WA, together with the inhibitory mechanism.

Dataset Information

Development of novel NEMO-binding domain mimetics for inhibiting IKK/NF-?B activation.

Publications

Development of novel NEMO-binding domain mimetics for inhibiting IKK/NF-κB activation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure